Novel MreB Inhibitors with Antibacterial Activity Against Gram (-) Bacteria

Novel MreB Inhibitors with Antibacterial Activity Against Gram (-) Bacteria | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article Novel MreB Inhibitors with Antibacterial Activity Against Gram (-) Bacteria Hye Yeon Sagong, Jesus D Rosado-Lugo, Eric J Bryan, Edgar Ferrer-Gonzalez, and 5 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-1824695/v1 This work is licensed under a CC BY 4.0 License Status: Under Review Version 1 posted 5 You are reading this latest preprint version Abstract MreB is a cytoskeleton protein present in rod-shaped bacteria that is both essential for bacterial cell division and highly conserved. Because most Gram (-) bacteria require MreB for cell division, chromosome segregation, cell wall morphogenesis, and cell polarity, it is an attractive target for antibacterial drug discovery. As MreB modulation is not associated with the activity of antibiotics in clinical use, acquired resistance to MreB inhibitors is also unlikely. Compounds, such as A22 and CBR-4830, are known to disrupt MreB function by inhibition of ATPase activity. However, the toxicity of these compounds has hindered efforts to assess the in vivo efficacy of these MreB inhibitors. The present study further examines the structure-activity of analogs related to CBR-4830 as it relates to relative antibiotic activity and improved drug properties. These data reveal that certain analogs have enhanced antibiotic activity. In addition, we evaluated several representative analogs ( 9 , 10 , 14 , 26 , and 31 ) for their abilities to target purified E. coli MreB (EcMreB) and inhibit its ATPase activity. Except for 14 , all these analogs were more potent than CBR-4830 as inhibitors of the ATPase activity of EcMreB with corresponding IC 50 values ranging from 6.3 ± 2.0 to 29.4 ± 8.8 uM. MreB inhibitors Gram (-) Antibiotics E. coli MreB ATPase Inhibition Structure Activity Relationships Figures Figure 1 Introduction MreB is the prokaryotic homolog of actin in eukaryotes. The function of MreB within bacteria is associated with its ability to form long filaments, a process similar to actin in eukaryotes [1-5]. These filaments are critical for to the retention of the structure of rod-shaped bacteria, which is dominant among many Gram (-) bacteria [1, 2, 4-6]. MreB is also essential for bacterial cell division [7]. This structural protein is an attractive target for antibacterial drug discovery as it is highly conserved and present in almost all rod-shaped bacteria [8]. Acquired resistance to such novel antibiotics would also be unlikely, as MreB modulation is not associated with the modes of action of antibiotics in clinical use. Polymerization of MreB requires ATP. Several benzyl thioureas such as A22 and MP265 ( 1, 2 ) as well as CBR-4830 and several of its analogs (3-6 ) have been identified as inhibitors of MreB function (Figure 1) [1, 2, 5, 8-15]. Current theories on the mode of action of these MreB inhibitors suggest that they act by noncompetitively binding to this structural protein, thereby affecting the ATP binding pocket. MreB loss of function is both lethal and pleiotropic [7, 16]. Multiple cellular processes are disrupted including cell shape determination, polar protein localization, and cell division [16-18]. MreB also has a direct role in the segregation of a specific region of the chromosome [18-21]. The MreB inhibitor, A22, is lethal at relatively low doses when administered iv to either rats or mice. Lynch et al reported in 2007 the identification and characterization of CBR-4830 (Figure 1) as a novel antimicrobial agent with activity against efflux-compromised P. aeruginosa [22]. Combined genetic, biochemical and cell morphology studies were performed that demonstrated that the cellular target for CBR-4830 is MreB. A limited structure-activity study was performed with analogs of CBR-4830, 3 - 6 . However, CBR-4830 proved to be the most active analog within this series. CBR-4830 proved not only to be difficult to formulate for iv administration because of poor solubility properties in aqueous vehicles, but it also proved to be lethal at relatively low concentration in mice. Recently, we reported on the enhanced activity of TXH11106 ( 7 ) relative to CBR-4830 [23, 24]. In the present study, we initiated studies to develop additional analogs related to CBR-4830 to further improve antibacterial potency and formulation properties relative to CBR-4830. Chemistry Our initial studies were focused on examining the impact of changing the fused alicyclic ring of the tetrahydrocarbazole pharmacophore of CBR-4830 to its next higher homologue, a cycloheptane moiety fused to the indole. 2-Bromo-5,6,7,8,9,10-hexahydrocyclohepta[ b ]indole-6-amine, 8 , and the dichloro analogs 9 and 10 were synthesized as outlined in Scheme 1 . Commercially available cycloheptanone was condensed with ethyl formate to form 2-(hydroxymethylene)cycloheptan-1-one. Reaction of this intermediate with the appropriate pre-formed diazonium salts provide the hydrazineylidene intermediates, which were converted to 8a and a mixture of 9a and 10a . The mixture of 9a and 10a was converted to their N-Boc derivatives and separated chromatographically, followed by removal of the Boc groups to provide the pure isomers. Reductive amination using ammonium acetate and NaBH 3 CN provided the 6-amino-5,6,7,8,9,10-hexahydro-cyclohepta[ b ]indoles 8 , 9 and 10 . We also synthesized similarly substituted 1,2,3,4-tetrahydrocyclopenta[ b ]indol-3-amines, 11 – 13 , which represent analogs that are a one carbon lower homologue of the tetrahydrocarbazole pharmacophore of CBR-4830. The methods used for these syntheses are outlined in Scheme 2 . Cyclopentanone was condensed with ethyl formate to provide 2-(hydroxymethylene)cyclopentan-1-one. This intermediate was reacted with the appropriate diazonium salt to give the desired hydrazineylidene intermediates, which were converted under acidic condition to the 1,4-dihydrocyclopenta[ b ]indole-3(2 H )-ones, 11a as well as a mixture of 12a and 13a . The mixture of 12a and 13a was resolved by formation of their N-Boc derivatives ( 12b and 13b ), chromatographic separation, followed by removal of their Boc groups. The impact of removing two methylene groups and opening the tetrahydrocyclohexyl ring of CBR-4830 on relative antimicrobial activity was also evaluated. The varied 2-(1-aminoethyl)indoles 14 – 21 were prepared as outlined in Scheme 3 . The Weinreb amide derivatives 14b - 21b were formed from the various 2-carboxyindole derivatives and then converted using methyllithium to the methyl ketones 14c - 21c . Reductive amination of 14c - 21c using ammonium acetate and sodium cyanoborohydride provide 14 – 21 . The influence of mono- or dimethylation on the amino group as well as methylation of the 1-position of 1-(5-bromo-1-methyl-1H-indol-2-yl)ethan-1-amine was examined. Compounds 22 , 23 , and 24 were synthesized as outlined in Scheme 4 . Reductive amination was performed using N-methylamine or N,N-dimethylamine, followed by reduction with sodium cyanoborohydride was used to prepare 22 or 23 . Methylation of 14c to form the 1-methylindole derivative 24a was accomplished using potassium carbonate in DMF and methyl iodide. Intermediate 24a was then converted 24 using ammonium acetate and sodium cyanoborohydride. We also examined the impact on activity of having the primary amine of 1-(5-bromo-1-methyl-1 H -indol-2-yl)ethan-1-amine attached to a primary carbon as opposed to a secondary carbon. We synthesized (5-bromo-1 H -indol-2-yl)methanamine, 25 , as outlined in Scheme 5 and evaluated its antibacterial properties. The preparation of the methylamine derivative 25 was performed starting from the 2-carboxy indole intermediate 14a . This acid was converted to its methyl ester, 25a , using methanol and concentrated HCl. The ester was reduced with lithium borohydride in THF to provide the hydroxymethyl intermediate, 25b . Using Dess-Martin periodinane in dichloromethane, this hydroxymethyl derivative was oxidized to the 2-formylindole derivative, 25c . Reductive amination with ammonium acetate and sodium cyanoborohydride provide the desired 2-aminomethyl indole derivative 25 . The N-(3-aminopropyl) derivatives of 8 , 11 , and 14 were also synthesized and their relative antibacterial activity compared. The synthetic methods used for their preparation are outlined in Scheme 6 . The ketone intermediates 8a , 11a , and 14c were each treated with N-Boc propylenediamine and then subject to reduction using sodium cyanoborohydride in ethanol. Excellent yields were obtained of each of the N-Boc propylenediamine derivatives, 26a , 27a and 28a . Removal of the N-Boc protecting groups with 4N HCl gave 26 – 28 . A series of N-acyl derivatives of 28 were also prepared. These included N-acylheteroaryl, N-acyl(3,4-dichlorophenyl), as well as N-acylcyclohexyl derivatives, as illustrated in Scheme 7 . The synthesis of a series carboxamides derived from 28 were prepared by condensation of various carboxylic acids using EDC, HOBt and DIPEA at room temperature. The commercially available carboxylic acids employed for the preparation of 29 – 33 were 5-chlorofuran-2-crboxylic acid, 5-chlorothiophene-2-carboxylic acid, 4,5-dichlorothiophene-2-carboxylic acid, 3,4-dichlorobenzoic acid and cyclohexane carboxylic acid. We also prepared the revered amide analog, 34 , that is related to 32 , as well as the aryloxy analog 35 , as illustrated in Scheme 8 and 9 . The reversed amide 34 was prepared by first preparing the 4-amino-N-(3,4-dichlorophenyl)butanamide, which was then reacted with 14c and the resulting imine reduce using sodium cyanoborohydride in ethanol to give 34 as outlined in Scheme 8 . The N-[4-(3,4-dichlorophenoxy)butyl] derivative of 14 was prepared from 3-(3,4-dichlorophenoxy)propylamine as illustrated in Scheme 9 . Reaction of this amine with 14c , followed by reduction with sodium cyanoborohydride provide 35 . Results Table 1 provides summarize the intrinsic activity of antibacterial activity against P. aeruginosa , E. coli , K. pneumoniae and A. baumannii for the cyclohepta analogs 8 - 10 , the cyclopenta analogs 11 - 13 , and the ring-opened analogs 14 - 21 , relative to CBR-4830. P. aeruginosa proved relatively resistant in the absence of a bacterial efflux pump inhibitor (EPI). Therefore, the MICs of all of these compounds against P. aeruginosa was also evaluated in the presence of 12.5 µg/ml of an EPI, namely N-(((2S,4R)-4-(aminomethyl)pyrrolidin-2-yl)methyl)-6-(4-fluorophenyl)-1H-indole-2-carboxamide [24]. A similar MIC (1.0 µg/ml) was observed for CBR-4830 against P. aeruginosa PAO1 in the presence of the EPI to that reported against the PAO1 mutant efflux defective strain CB392 ∆( mexAB-oprM ) ∆( mexCD-oprJ ) [24]. The more potent of the cyclohepta analogs evaluated against P. aeruginosa in the presence of an EPI was 10 with an MIC of 1.0 µg/ml. Among the cyclopenta analogs, the most potent was 13 , which had its dichloro substituents at the same relative sites on the fused indole moiety and also exhibited the same MIC value as 10 . These two analogs also exhibited the lower MICs against E. coli , with 10 and 13 having MICs of 8 and 16 µg/ml as compared to 8 ug/ml for CBR-4830. Compound 10 had an MIC of 2 µg/ml against K. pneumoniae , which was lower than that observed for 13 and CBR-4830, which had MICs of 8 µg/ml. The MICs of 10 and 13 against A. baumannii were 8 µg/m; which were lower than that observed for CBR-4830 (16 ug/ml). Both 10 and 13 proved to have significantly improved formulation properties and were also less toxic when administered intravenously to mice relative to CBR-4830 (data not shown). The ring-opened analogs 14-21 also had dramatically improved solubility in aqueous vehicles suitable for intravenous administration and were well tolerated when administered by this route to mice (data not shown). However, their relative intrinsic MICs against P. aeruginosa , E. coli , K. pneumoniae or A. baumanni were disappointing as none of these analogs had MIC values that were less than 32 µg/ml. Only in the presence of an EPI, were notable MICs observed against P. aeruginosa of 8 µg/ml for 14 and 21 and 16 ug/ml for 16 , 17 and 20 . The improve physicochemical properties of the ring-opened analogs prompted further studies on their structure-activity relationships. We synthesized and evaluated the N-methyl and N,N-dimethyl amino analogs, 22 , and 23 , as well as the N 1 -methyl derivative, 24, of compound 14 . We also prepared the methylamino analog, 25 , wherein the primary amine was attached to a primary carbon. The antimicrobial activities of these compounds are summarized in Table 2. Only in the case of 25 was notable activity observed against P. aeruginosa in the presence of an EPI. The potency observed was comparable to 14 . A series of N-(3-aminopropane) derivatives were prepared that incorporated the cycloheptyl, cyclopentyl and open-ring analogs of CBR-4830, specifically 26 , 27 and 28 (Table 3). Compound 26 had a lower MIC than 8 in terms of its intrinsic MIC against P. aeruginosa (32 versus 128 µg/ml), in the presence of an EPI, the observed MIC was much higher (32 versus 4 µg/ml). The differences between 8 and 26 were not that significant when evaluated in E. coli , K. pneumonia e and A. baumannii . Both 27 and 28 have either similar activity or are less active than their parent primary amines, 11 and 14 against E. coli , K. pneumonia e or A. baumannii . However, both 27 and 28 were notably less active than their parent primary amines when evaluated against P. aeruginosa in the presence of an EPI. Against E. coli , 26 had the same MIC as 8 (16 µg/ml), but it was slightly more active against K. pneumoniae (8 versus 16 µg/ml) and slightly less active against A. baumannii (32 versus 16 µg/ml). Compound 27 did not have remarkable activity against E. coli , K. pneumoniae , and A. baumannii . There were no major differences in the MICs observed for 14 and 28 observed in E. coli , K. pneumonia e or A. baumannii . We extended these studies to include a several N-(3-acylpropanamides) derived from the N-(3-aminopropane), 29 - 33 . The relative antimicrobial activity of these hydrophobic derivatives are summarized in Table 3. None of these analogs exhibited intrinsic activity against P. aeruginosa. In the presence of an EPI, these analogs failed to exhibit potency against P. aeruginosa comparable to 14 or 22 under these assay conditions. There were notable improvements in activity for 29 - 33 against E. coli , K. pneumoniae and A. baumanni relative to 22 . Most notably, 31 had MICs against E. coli , K. pneumoniae and A. baumanni that were 16-, 64-, and 8-fold lower than for 14 . The cyclohexyl amide, 33 , proved to be less activity than the aryl or heteroaryl amides, 29-32 , that were evaluated. We also prepared 34 , the reversed amide analog of 32 , as well as the 4-(3,4-dichlorophenoxy)butan-1-amine derivative, 35 . Similar activity was observed between 32 and its reversed amide, 34 , derivatives. The phenoxy derivative 35 also had similar activity to 34 . We evaluated six representative analogs ( 8 , 9 , 10 , 14 , 26 , and 31 ) for their abilities to target purified E. coli MreB (EcMreB) and inhibit its ATPase activity. All six compounds inhibited the ATPase activity of EcMreB (Figure 1), with corresponding IC 50 values ranging from 6.3 ± 2.0 µM for analog 31 to 120 ± 17.0 µM for analog 14 (Table 4). Based on IC 50 values, the EcMreB-inhibiting potency of the six analogs tested followed the hierarchy: 31 > 9 ≈ 26 ≈ 10 > 8 > 14 . Table 4. IC 50 values for inhibition of the ATPase activity of EcMreB by select analogs. Compound IC 50 (μM) 8 44.1 ± 7.8 9 24.5 ± 4.7 10 29.4 ± 8.8 14 120 ± 17.0 26 25.1 ± 7.0 31 6.3 ± 2.0 EcMreB = E. coli MreB; IC 50 = the compound concentration that inhibits the ATPase activity of EcMreB by 50%. IC 50 values were derived from fits of the velocity vs. [compound] plots shown in Figure 1, with the indicated uncertainties reflecting the standard deviation of the fitted curves from the experimental data points. Discussion The data in Table 1 indicate that cyclohepta and cyclopenta analogs related to CBR-4830 retain significant antibacterial activity. CBR-4830 proved difficult to formulate (requiring 40% propylene glycol to achieve a concentration of 2 mg/ml) and did produce severe toxic effects when administered to mice b.i.d. at a dose of 400 ug/mouse (data not shown). Several of the promising cyclohepa and cyclopenta analogs could be readily formulated using 10 mM citrate in water and were well tolerated when administered at more than twice this dose. The actual significance of these differences, however, needs to be examined in the context of comparative pharmacokinetic studies which have not been performed. The ring-opened analogs of CBR-4830 were significantly less active and only exhibited modest activity when evaluated against P. aeruginosa in the presence of a bacterial efflux pump inhibitor, such as N-(((2S,4R)-4-(aminomethyl)pyrrolidin-2-yl)methyl)-6-(4-fluorophenyl)-1 H -indole-2-carboxamide. The ring-open series of compounds, like the cyclohepta and cyclopenta analogs, had favorable formulation properties and were well tolerated in mice. Efforts to improve antibacterial activity among the ring-opened analog 8 , by either mono- or demethylation of the amino substituent as in 22 and 23 , or methylation of the indole nitrogen as in the case of 24 , failed to enhance activity (Table 2 ). Changing the 1-aminoethyl of 8 to a aminomethyl group as in 25 did restore activity against P. aeruginosa in the presence of the EPI, N-(((2S,4R)-4-(aminomethyl)pyrrolidin-2-yl)methyl)-6-(4-fluorophenyl)-1 H -indole-2-carboxamide. However, no significant activity was observed against E. coli , K. pneumoniae or A. baumannii (Table 2 ). The impact of attaching a 3-aminopropyl linkage to the amine of cycloheptyl, cyclopentyl or ring-opened analogs of CBR-4830 on antibacterial activity was also evaluated (Table 3 ). In the case of the cycloheptyl analog 26 , there was an enhancement in intrinsic activity against P. aeruginosa and a modest increase in activity against K. pneumonia e relative to 8 . In the case of the cyclopenta analog 27 , there was a notable loss in activity observed against P. aeruginosa when evaluated in the presence of an EPI and no increase in activity against E. coli , K. pneumoniae or A. baumanni relative to 11 . Modification of the ring-opened analog by attaching this linkage as in 28 primarily resulted in a loss of the activity that was previously observed with 14 in the presence of an EPI against P. aeruginosa . We examined the effect of modifying the terminal amine of 28 by forming various hydrophobic amide derivatives as a possible means of enhancing the antibacterial properties of these ring-opened analogs. While these analogs 29–33 did not exhibit significant intrinsic activity against P. aeruginosa , their activity in the presence of an EPI was improved relative to 28 . More significantly, their intrinsic antibacterial activity was increased against E. coli and K. pneumoniae , and to a less extent against A. baumannii . Similar improvements in activity were observed with the reversed amide derivative 34 and the oxyphenyl analog 35 . We also sought to validate MreB as the antibacterial target of our analogs. To this end, we selected six representative analogs and evaluated the compounds for their abilities to inhibit the catalytic (ATPase) activity of purified E. coli MreB (EcMreB). Significantly, all six compounds tested exhibited concentration-dependent inhibition of EcMreB ATPase activity (Fig. 1 ), establishing their antibacterial actions as MreB inhibitors. In addition, the relative EcMreB-inhibiting potencies of the compounds (as reflected by the magnitudes of the IC 50 values shown in Table 4 ) were generally correlated with their corresponding antibacterial activities, further validating MreB as the antibacterial target of our compounds. Among the analogs tested, 31 was the most potent inhibitor of EcMreB activity. Significantly, this analog also exhibited the best antibacterial activity against K. pneumoniae of all compounds evaluated here and was among the best agents versus E. coli and A. baumannii as well. Analog 31 may thus represent an important reference compound for the development of next-generation MreB inhibitors. Conclusions The structure activity studies performed with these varied analogs of CBR-4830 indicate that in many instances significant antibacterial activity can be retained. In addition, it is possible to reduce the toxicity associated with CBR-4830 and improve its formulation properties. The extreme toxicity of A22 and CBR-4830 has hindered efforts to demonstrate the in vivo efficacy of these MreB inhibitors. In view of their unique mechanism of action, such agents would be anticipated to be useful against multidrug-resistant bacteria. The data on the structure-activity of these CBR-4830 derivatives suggest that the development of next-generation MreB inhibitors could overcome this limitation and provide a means for demonstrating the potential clinical efficacy and utility of MreB inhibitors as antibiotics. Evaluation of comparative pharmacokinetic parameters and efficacy in vivo remain essential for assessing the potential of this novel class of antibiotics. Experimental Chemistry: General Methods All reactions, unless otherwise stated, were done under nitrogen atmosphere. Reaction monitoring and follow-up were done using aluminum backed Silica G TLC plates with UV254 (Sorbent Technologies), visualizing with ultraviolet light. Flash column chromatography was done on a Combi Flash Rf Teledyne ISCO using hexane, ethyl acetate, dichloromethane, and methanol. The 1 H (400 MHz) and 13 C (100 MHz) NMR spectra were done in CDCl 3 , Methanol-d 4 , and DMSO-d 6 and recorded on a Bruker Avance III (400 MHz) Multinuclear NMR Spectrometer. Data is expressed in parts per million relative to the residual nondeuterated solvent signals, spin multiplicities are given as s (singlet), d (doublet), dd (doublet of doublets), t (triplet), dt (doublet of triplets), q (quartet), m (multiplet), and bs (broad singlet), and coupling constants ( J ) are reported in Hertz. Analytical HPLC was performed on a Shimadzu LC-20AT Prominence liquid chromatograph using a 150 x 4.6 mm Princeton SPHER-100 RP C18 1000A 5 um column using 0% water for 2 minutes and a 0-100% water/methanol gradient over a 5-minute period and 5 minutes at 100% methanol at a 2.0 ml/minute flow rate monitoring uv absorbance at 254 and 296 nm. Using this method of analysis, the purity of all compounds used in bioassays was determined to be ≥ 95%. Mass spectrometry (MS) was performed by electrospray (ESI) ionization using a Shimadzu 2020 LC-MS quadrupole mass spectrometer using a 0-100% water/methanol gradient over a 3-minute period. HRMS experiments were conducted using the Waters ACQUITY UPLC – Synapt G2 HRMS (Milford, MA) to determine the accurate mass values of final compounds. UPLC separation was performed on an ACQUITY UPLC BEH C18 column (2.1 mm × 50 mm, 1.7 μm) with 0.1% formic acid in water and 0.1% formic acid in methanol as mobile phase A and B, respectively. The column temperature was maintained at 40°C. The gradient was set from 5% to 95% B over 3 minutes at a flow rate of 0.25 mL/min. Extensive fragmentation under the conditions used for these HRMS analyses did not allow for the detection of parent ions. HRMS data obtained under these conditions are provided for several of the compounds evaluated in Supplemental Materials. 2-Bromo-5,6,7,8,9,10-hexahydrocyclohepta[ b ]indol-6-amine (8) 2-Bromo-7,8,9,10-tetrahydrocyclohepta[ b ]indol-6(5 H )-one (100 mg, 0.36 mmol), ammonium acetate (277 mg, 3.60 mmol) and sodium cyanoborohydride (113 mg, 1.80 mmol) were dissolved in ethanol (10 mL). The mixture was stirred for overnight at 60 o C. The reaction mixture was acidified with 6N HCl, and it was washed with ethyl acetate. Then, the aqueous layer was basified with NaOH, and it was extracted with ethyl acetate. The organic layer was washed with brine, and it was dried over Na 2 SO 4 . The organic layer was concentrated under reduced pressure, and the residue was purified on an ISCO chromatograph (0-10% methanol/ dichloromethane + 0.1% NH 4 OH) to give the product as a white solid (34 mg, 34%); 1 H NMR (300 MHz) (CD 3 OD) δ 10.76 (bs, 1H), 7.51 (d, J = 2 Hz, 1H), 7.24 (d, J = 8 Hz, 1H), 7.03 (dd, J = 8 Hz, J = 2 Hz, 1H), 4.01-3.97 (m, 1H), 2.86-2.81 (m, 1H), 2.58-2.56 (m, 1H), 1.99-1.84 (m, 2H), 1.71-1.50 (m, 4H); LC/MS RT = 2.57 (M-H - : 277/279). (E) -2-(Hydroxymethylene)cycloheptan-1-one Cycloheptanone (2.95 mL, 25 mmol) was dissolved in anhydrous tetrahydrofuran (10 mL), and it was cooled to 0 o C. A solution of 1.0 M LiHMDS in tetrahydrofuran (30 mL, 30 mmol) was slowly added, and it was stirred for 5 minutes at 0 o C. Ethyl formate (2.42 mL, 30 mmol) was slowly added, and it was stirred for 2 hours at 0 o C. The reaction mixture was diluted with ethyl acetate, and it was washed with 6N HCl and brine. The organic layer was dried over Na 2 SO 4 . The organic layer was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-20% ethyl acetate/hexane) to give product as a colorless oil (1.30 g, 37%); 1 H NMR (300 MHz) (CDCl 3 ) δ 7.59 (d, J = 9 Hz, 1H), 2.51-2.47 (m, 2H), 2.23-2.19 (m, 2H), 1.73-1.54 (m, 6H). (E) -2-(2-(4-Bromophenyl)hydrazineylidene)cycloheptan-1-one To a solution of 4-bromoaniline (1.60 g, 9.30 mmol) in concentrated hydrochloric acid (2 mL), a solution of sodium nitrite (642 mg, 9.30 mmol) in water (4 mL) was added slowly at 0 o C. The mixture was stirred for 30 minutes at 0 o C. In a separate round bottom flask, (E) -2-(hydroxymethylene)cycloheptan-1-one (1.30 g, 9.30 mmol) was dissolve in methanol (12 mL). To the mixture, a solution of sodium acetate (1.91 g, 23.3 mmol) in water (5 mL) was added slowly at 0 o C. The mixture was stirred for 20 minutes at 0 o C. Then, the freshly prepared diazonium salt solution was slowly added. The mixture was stirred for additional 30 minutes at 0 o C, and the formed yellow suspension was filtered to give the product as a yellow solid (2.17 g, 79%); 1 H NMR (300 MHz) (DMSO-d 6 ) δ 13.27 (s, 1H), 7.43 (d, J = 9 Hz, 2H), 7.23 (d, J = 9 Hz, 2H), 2.62-2.56 (m, 4H), 1.68 (m, 6H). 2-Bromo-7,8,9,10-tetrahydrocyclohepta[ b ]indol-6(5 H )-one (8a) (E) -2-(2-(4-Bromophenyl)hydrazineylidene)cycloheptan-1-one (2.17 g, 7.35 mmol) was dissolved in a mixture of concentrated hydrochloric acid (2 mL) and acetic acid (8 mL), and it was stirred for 30 minutes at 130 o C. The resulting dark brown suspension was diluted with ethyl acetate, and it was washed with 10% NaOH and brine. The organic layer was dried over Na 2 SO 4 , and it was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-20% ethyl acetate/hexane) to give product as a yellow solid (472 mg, 23%); 1 H NMR (300 MHz) (CDCl 3 ) δ 8.90 (bs, 1H), 7.79 (s, 1H), 7.41 (dd, J = 9 Hz, J = 2 Hz, 1H), 7.26-7.23 (m, 1H), 3.11-3.07 (m, 2H), 2.86-2.82 (m, 2H), 2.11-1.99 (m, 4H). 1,2-Dichloro-5,6,7,8,9,10-hexahydrocyclohepta[ b ]indol-6-amine (9) . 1,2-Dichloro-7,8,9,10-tetrahydrocyclohepta[ b ]indol-6(5 H )-one (99 mg, 0.37 mmol), ammonium acetate (285 mg, 3.70 mmol) and sodium cyanoborohydride (116 mg, 1.85 mmol) were dissolved in ethanol (10 mL). The reaction mixture was stirred for overnight at 60 o C. After removal of solvent, the reaction mixture was diluted with ethyl acetate. The organic layer was washed with 10% sodium hydroxide and brine, and it was dried over sodium sulfate. The organic layer was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-10% methanol/dichloromethane + 0.1% NH 4 OH) to give product as a white solid. The solid was then treated with 4N hydrochloric acid in dioxane, and it was stirred for 30 minutes at room temperature. The white suspension was concentrated under reduced pressure, and the resulting residue was suspended in ethyl acetate. The suspension was filtered to give hydrochloric acid salt of product as a white solid (42 mg, 37%); 1 H NMR (300 MHz) (DMSO-d 6 ) δ 11.65 (bs, 1H), 8.42 (bs, 3H), 7.37 (d, J = 8 Hz, 1H), 7.25 (d, J = 9 Hz, 1H), 4.58 (m, 1H), 3.65-3.60 (m, 2H), 3.12-2.98 (m, 2H), 1.97-1.89 (m, 4H); LC/MS RT = 2.90 (M-H - : 267/269). tert- Butyl 1,2-dichloro-6-oxo-7,8,9,10-tetrahydrocyclohepta[ b ]indole-5(6 H )-carboxylate (9a) and tert -Butyl 2,3-dichloro-6-oxo-7,8,9,10-tetrahydrocyclohepta[ b ]indole-5(6 H )-carboxylate (10a) To a solution of 3,4-dichloroaniline (1.53 g, 9.42 mmol) in concentrated hydrochloric acid (10 mL), a solution of sodium nitrite (650 mg, 9.42 mmol) in water (20 mL) was added slowly at 0 o C. The mixture was stirred for 30 minutes at 0 o C. In a separate round bottom flask, (E)-2-(hydroxymethylene)cycloheptan-1-one (1.32 g, 9.42 mmol) was dissolve in methanol (12 mL). To the mixture, a solution of sodium acetate (1.93 g, 23.55 mmol) in water (5 mL) was added slowly at 0 o C. The mixture was stirred for 20 minutes at 0 o C. Then, the freshly prepared diazonium salt solution was slowly added. The mixture was stirred for additional 30 minutes at 0 o C. The reaction mixture was diluted with ethyl acetate, and the organic layer was washed with saturated sodium bicarbonate followed brine. The organic layer was concentrated under reduced pressure and the resulted dark brown oil was carried next step without further purification. The residue was dissolved in formic acid (10 mL), and it was stirred for 2 hours at 100 o C. The resulting dark brown suspension was diluted with ethyl acetate, and it was washed with 10% NaOH and brine. The organic layer was dried over sodium sulfate, and it was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-20% ethyl acetate/hexane) to give a mixture of two regioisomers, 1,2-dichloro-7,8,9,10-tetrahydrocyclohepta[ b ]indol-6(5 H )-one and 2,3-dichloro-7,8,9,10-tetrahydrocyclohepta[ b ]indol-6(5 H )-one as a beige solid (212 mg, 8%). To a solution of a mixture of the two regioisomers (212 mg, 0.79 mmol) in tetrahydrofuran (10 mL), Boc anhydride (345 mg, 1.58 mmol) and DMAP (89 mg, 0.79 mmol) were added. The reaction mixture was stirred for 3 hours. The reaction mixture was diluted with ethyl acetate, and it was washed with saturated ammonium chloride, followed by brine. The organic layer was dried over sodium sulfate, and it was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-10% ethyl acetate/hexane) to give tert- butyl 1,2-dichloro-6-oxo-7,8,9,10-tetrahydrocyclohepta[ b ]indole-5(6 H )-carboxylate (152 mg, 49%). 1 H NMR (300 MHz) (CDCl 3 ) δ 7.93 (d, J = 9 Hz, 1H), 7.41 (d, J = 9 Hz, 1H), 3.42-3.38 (m, 2H), 2.87-2.83 (m, 2H), 1.97-1.95 (m, 4H), 1.55 (s, 9H). along with tert -butyl 2,3-dichloro-6-oxo-7,8,9,10-tetrahydrocyclohepta[ b ]indole-5(6 H )-carboxylate (59 mg, 20%). 1 H NMR (300 MHz) (CDCl 3 ) δ 8.20 (s, 1H), 7.62 (s, 1H), 2.91 (m, 2H), 2.85 (m, 2H), 2.02-1.98 (m, 4H), 1.57 (s, 9H). 1,2-Dichloro-7,8,9,10-tetrahydrocyclohepta[ b ]indol-6(5 H )-one (9a) To a solution of tert- butyl 1,2-dichloro-6-oxo-7,8,9,10-tetrahydrocyclohepta[ b ]indole-5(6 H )-carboxylate (152 mg, 0.41 mmol) in dichloromethane (5 mL), trifluoroacetic acid (1 mL) was added. The reaction mixture was stirred for an hour at room temperature. After removal of solvent, the mixture was diluted with ethyl acetate, and it was washed with 10% sodium hydroxide and brine. The organic layer was dried over sodium sulfate, and it was concentrated under reduced pressure to give product as a white solid (99 mg, 90%); 1 H NMR (300 MHz) (CDCl 3 ) δ 9.08 (bs, 1H), 7.35 (d, J = 9 Hz, 1H), 7.20 (d, J = 9 Hz, 1H), 3.58-3.54 (m, 2H), 2.87-2.83 (m, 2H), 2.11-2.04 (m, 2H), 1.99-1.91 (m, 2H). 2,3-Dichloro-7,8,9,10-tetrahydrocyclohepta[ b ]indol-6(5 H )-one (10a) To a solution of tert -butyl 2,3-dichloro-6-oxo-7,8,9,10-tetrahydrocyclohepta[ b ]indole-5(6 H )-carboxylate (943 mg, 2.64 mmol) in dichloromethane (25 mL), trifluoroacetic acid (5 mL) was added. The reaction mixture was stirred for 2 hours at room temperature. After removal of solvent, the mixture was diluted with ethyl acetate, and it was washed with 10% sodium hydroxide and brine. The organic layer was dried over sodium sulfate, and it was concentrated under reduced pressure to give product as a white solid (675 mg, 95%); 1 H NMR (300 MHz) (CDCl 3 ) δ 8.90 (bs, 1H), 7.74 (s, 1H), 7.48 (s, 1H), 3.10-3.06 (m, 2H), 2.87-2.83 (m, 2H), 2.11-2.05 (m, 2H), 2.01-1.99 (m, 2H). 2,3-Dichloro-5,6,7,8,9,10-hexahydrocyclohepta[ b ]indol-6-amine (10). 2,3-Dichloro-7,8,9,10-tetrahydrocyclohepta[ b ]indol-6(5 H )-one (100 mg, 0.37 mmol), ammonium acetate (285 mg, 3.70 mmol) and sodium cyanoborohydride (116 mg, 1.85 mmol) were dissolved in ethanol (10 mL). The reaction mixture was stirred for overnight at 60 o C. After removal of solvent, the reaction mixture was diluted with ethyl acetate. The organic layer was washed with 10% sodium hydroxide and brine, and it was dried over sodium sulfate. The organic layer was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-10% methanol/dichloromethane + 0.1% NH 4 OH) to give product as a white solid. The solid was then treated with 4N hydrochloric acid in dioxane, and it was stirred for 30 minutes at room temperature. The white suspension was concentrated under reduced pressure, and the resulting residue was suspended in ethyl acetate. The suspension was filtered to give hydrochloric acid salt of product as a white solid (76 mg, 67%); 1 H NMR (300 MHz) (DMSO-d 6 ) δ 11.36 (bs, 1H), 8.44 (bs, 3H), 7.78 (s, 1H), 7.62 (s, 1H), 4.56 (m, 1H), 3.55-3.26 (m, 2H), 2.89-2.71 (m, 4H), 1.93-1.76 (m, 2H); LC/MS RT = 2.91 (M-H - : 267/269). 7-Bromo-1,2,3,4-tetrahydrocyclopenta[ b ]indol-3-amine (11) 7-Bromo-1,4-dihydrocyclopenta[ b ]indol-3(2 H )-one (100 mg, 0.40 mmol), ammonium acetate (308 mg, 4.00 mmol) and sodium cyanoborohydride (126 mg, 2.00 mmol) were dissolved in ethanol (10 mL). The mixture was stirred for overnight at 60 o C. An additional ammonium acetate (308 mg, 4.00 mmol) and sodium cyanoborohydride (126 mg, 2.00 mmol) along with catalytic amount of acetic acid were added. The mixture was stirred for 5 hours at 85 o C. The reaction mixture was acidified with 6N HCl, and it was washed with ethyl acetate. Then, the aqueous layer was basified with NaOH, and it was extracted with ethyl acetate. The organic layer was washed with brine, and it was dried over Na 2 SO 4 . The organic layer was concentrated under reduced pressure, and the residue was purified on an ISCO chromatograph (0-10% methanol/dichloromethane + 0.1% NH 4 OH) to give the product as a white solid (17 mg, 17%); 1 H NMR (300 MHz) (CD 3 OD) δ 7.48 (d, J = 1 Hz, 1H), 7.22 (d, J = 9 Hz, 1H), 7.12 (dd, J = 9 H, J = 2 Hz, 1H), 4.43-4.41 (m, 1H), 2.94-2.83 (m, 2H), 2.78-2.62 (m, 1H), 2.21-2.12 (m, 1H); LC/MS RT = 2.57 (M-H - : 249/251). 5 (E) -2-(Hydroxymethylene)cyclopentan-1-one Cyclopentanone (2.21 mL, 25 mmol) was dissolved in anhydrous tetrahydrofuran (10 mL), and it was cooled to 0 o C. A solution of 1.0 M LiHMDS in tetrahydrofuran (30 mL, 30 mmol) was slowly added, and it was stirred for 5 minutes at 0 o C. Ethyl formate (2.42 mL, 30 mmol) was slowly added, and it was stirred for 2 hours at 0 o C. The reaction mixture was diluted with ethyl acetate, and it was washed with 6N HCl and brine. The organic layer was dried over Na 2 SO 4 , and it was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-30% ethyl acetate/hexane) to give product as a white solid (772 mg, 28%); 1 H NMR (300 MHz) (CDCl 3 ) δ 7.21 (s, 1H), 2.56-2.51 (m, 2H), 2.43-2.37 (m, 2H), 2.01-1.94 (m, 2H). (E) -2-(2-(4-Bromophenyl)hydrazineylidene)cyclopentan-1-one To a solution of 4-bromoaniline (1.07 g, 6.23 mmol) in concentrated hydrochloric acid (2 mL), a solution of sodium nitrite (430 mg, 6.23 mmol) in water (4 mL) was added slowly at 0 o C. The mixture was stirred for 30 minutes at 0 o C. In a separate round bottom flask, 5 (E) -2-(hydroxymethylene)cyclopentan-1-one (700 mg, 6.23 mmol) was dissolve in methanol (12 mL). To the mixture, a solution of sodium acetate (1.28 g, 15.58 mmol) in water (5 mL) was added slowly at 0 o C. The mixture was stirred for 20 minutes at the temperature. Then, the freshly prepared diazonium salt solution was slowly added. The mixture was stirred for additional 30 minutes at the temperature, and the formed yellow suspension was filtered to give the product as a yellow solid (1.47 g, 89%); 1 H NMR (300 MHz) (DMSO-d 6 ) δ 10.03 (s, 1H), 7.42 (d, J = 9 Hz, 2H), 7.19 (d, J = 9 Hz, 2H), 2.65-2.60 (m, 2H), 2.34-2.29 (m, 2H), 1.99-1.95 (m, 2H). 7-Bromo-1,4-dihydrocyclopenta[ b ]indol-3(2 H )-one (11a) (E) -2-(2-(4-Bromophenyl)hydrazineylidene)cyclopentan-1-one (1.47 g, 5.50 mmol) was dissolved in a mixture of concentrated hydrochloric acid (2 mL) and acetic acid (8 mL), and it was stirred for 30 minutes at 130 o C. The resulting dark brown suspension was diluted with ethyl acetate, and it was washed with 10% NaOH and brine. The organic layer was dried over Na 2 SO 4 , and it was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-20% ethyl acetate/hexane) to give product as a beige solid (116 mg, 8%); 1 H NMR (300 MHz) (CDCl 3 ) δ 8.80 (bs, 1H), 7.87 (d, J = 2 Hz, 1H), 7.48 (dd, J = 9 H, J = 2 Hz, 1H), 7.35 (d, J = 9 Hz, 1H), 3.10-3.07 (m, 2H), 3.03-3.00 (m, 2H). 6,7-Dichloro-1,2,3,4-tetrahydrocyclopenta[ b ]indol-3-amine. (12). 6,7-Dichloro-1,4-dihydrocyclopenta[ b ]indol-3(2 H )-one (148 mg, 0.62 mmol), ammonium acetate (956 mg, 12.4 mmol) and sodium cyanoborohydride (195 mg, 3.1 mmol) were dissolved in ethanol (20 mL). The reaction mixture was stirred for overnight at 60 o C. After removal of solvent, the reaction mixture was diluted with ethyl acetate. The organic layer was washed with 10% sodium hydroxide and brine, and it was dried over sodium sulfate. The organic layer was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-10% methanol/dichloromethane + 0.1% NH 4 OH) to give product as a white solid (93 mg, 62%); 1 H NMR (300 MHz) (MeOD) δ 7.46 (s, 1H), 7.45 (s, 1H), 4.43-4.39 (m, 1H), 2.93-2.79 (m, 2H), 2.71-2.61 (m, 1H), 2.21-2.12 (m, 1H); LC/MS RT = 2.74 (M-H - : 239/241). 6,7-Dichloro-1,4-dihydrocyclopenta[ b ]indol-3(2 H )-one (12a) To a solution of tert- butyl 6,7-dichloro-3-oxo-2,3-dihydrocyclopenta[ b ]indole-4(1 H )-carboxylate (571 mg, 1.68 mmol) in dichloromethane (25 mL), trifluoroacetic acid (5 mL) was added. The reaction mixture was stirred for an hour at room temperature. After removal of solvent, the mixture was diluted with ethyl acetate, and it was washed with 10% sodium hydroxide and brine. The organic layer was dried over sodium sulfate, and it was concentrated under reduced pressure to give product as a white solid (148 mg, 37%); 1 H NMR (300 MHz) (DMSO-d 6 ) δ 11.98 (bs, 1H), 8.06 (s, 1H), 7.64 (s, 1H), 3.02-2.99 (m, 2H), 2.90-2.87 (m, 2H). 7,8-Dichloro-1,2,3,4-tetrahydrocyclopenta[ b ]indol-3-amine (13) 7,8-Dichloro-1,4-dihydrocyclopenta[ b ]indol-3(2 H )-one (100 mg, 0.42 mmol), ammonium acetate (647 mg, 8.40 mmol) and sodium cyanoborohydride (132 mg, 2.10 mmol) were dissolved in ethanol (10 mL). The reaction mixture was stirred for overnight at 60 o C. After removal of solvent, the reaction mixture was diluted with ethyl acetate. The organic layer was washed with 10% sodium hydroxide and brine, and it was dried over sodium sulfate. The organic layer was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-10% methanol/dichloromethane + 0.1% NH 4 OH) to give product as a white solid (41 mg, 41%); 1 H NMR (300 MHz) (MeOD) δ 7.22 (d, J = 9 Hz, 1H), 7.09 (d, J = 9 Hz, 1H), 4.47-4.44 (m, 1H), 3.16-3.07 (m, 1H), 2.95-2.83 (m, 2H), 2.24-2.16 (m, 1H); LC/MS RT = 2.74 (M-H - : 239/241). 7,8-Dichloro-1,4-dihydrocyclopenta[ b ]indol-3(2 H )-one (13a) To a solution of tert- butyl 7,8-dichloro-3-oxo-2,3-dihydrocyclopenta[ b ]indole-4(1 H )-carboxylate (121 mg, 0.36 mmol) in dichloromethane (5 mL), trifluoroacetic acid (1 mL) was added. The reaction mixture was stirred for an hour at room temperature. After removal of solvent, the mixture was diluted with ethyl acetate, and it was washed with 10% sodium hydroxide and brine. The organic layer was dried over sodium sulfate, and it was concentrated under reduced pressure to give product as a white solid (27 mg, 31%); 1 H NMR (300 MHz) (CDCl 3 ) δ 9.61 (bs, 1H), 7.41 (d, J = 9 Hz, 1H), 7.36 (d, J = 9 Hz, 1H), 3.35-3.32 (m, 2H), 3.09-3.03 (m, 2H). tert- Butyl 7,8-dichloro-3-oxo-2,3-dihydrocyclopenta[ b ]indole-4(1 H )-carboxylate (12b) and tert -Butyl 6,7-dichloro-3-oxo-2,3-dihydrocyclopenta[ b ]indole-4(1 H )-carboxylate (13b) To a suspension of 3,4-dichloroaniline (8.10 g, 50.00 mmol) in water (30 mL), concentrated hydrochloric acid (12.5 mL) was slowly added at 0 o C. Then, a solution of sodium nitrite (3.45 g, 50.00 mmol) in water (35 mL) was slowly added at 0 o C. The mixture was stirred for 30 minutes at 0 o C. This freshly prepared diazonium salt solution was slowly added to a solution of 2-oxocyclopentane-1-carboxylic acid (6.40 g, 50.00 mmol) in concentrated hydrochloric acid (4.58 mL). The mixture was stirred for additional 30 minutes at 0 o C to give a yellow suspension. The suspension was filtered to give intermediate, (E)-2-(2-(3,4-dichlorophenyl)hydrazineylidene)-cyclopentan-1-one, as an orange solid. The intermediate was then dissolved in acetonitrile (50 mL), and 1.8M sulfuric acid was added. The reaction mixture was stirred for overnight at 75 o C. The resulting dark brown suspension was diluted with ethyl acetate, and it was washed with 10% NaOH and brine. The organic layer was dried over sodium sulfate, and it was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-20% ethyl acetate/hexane) to give a mixture of two regioisomers, 7,8-dichloro-1,4-dihydrocyclopenta[ b ]indol-3(2 H )-one and 6,7-dichloro-1,4-dihydrocyclopenta[ b ]indol-3(2 H )-one as a dark brown solid (493 mg, 10%). To a solution of a mixture of the two regioisomers (493 mg, 2.05 mmol) in tetrahydrofuran (25 mL), Boc anhydride (895 mg, 4.10 mmol) and DAMP (230 mg, 2.05 mmol) were added. The reaction mixture was stirred for 3 hours. The reaction mixture was diluted with ethyl acetate, and it was washed with saturated ammonium chloride, followed by brine. The organic layer was dried over sodium sulfate, and it was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-10% ethyl acetate/hexane) to give tert- butyl 7,8-dichloro-3-oxo-2,3-dihydrocyclopenta[ b ]indole-4(1 H )-carboxylate (121 mg, 17%). 1 H NMR (300 MHz) (CDCl 3 ) δ 8.15 (d, J = 9 Hz, 1H), 7.48 (d, J = 9 Hz, 1H), 3.26-3.22 (m, 2H), 3.02-2.99 (m, 2H), 1.68 (s, 9H). along with tert -butyl 6,7-dichloro-3-oxo-2,3-dihydrocyclopenta[ b ]indole-4(1 H )-carboxylate (160 mg, 23%). 1 H NMR (300 MHz) (CDCl 3 ) δ 8.47 (s, 1H), 7.73 (s, 1H), 3.01-3.00 (m, 4H), 1.69 (s, 9H). 1-(5-Bromo-1 H -indol-2-yl)ethan-1-amine (14) 1-(5-Bromo-1 H -indol-2-yl)ethan-1-one (95 mg, 0.40 mmol), ammonium acetate (308 mg, 4.00 mmol) and sodium cyanoborohydride (126 mg, 2.00 mmol) were dissolved in ethanol (5 mL). The mixture was stirred for overnight at 60 o C. The reaction mixture was diluted with ethyl acetate. The organic layer was washed with 10% NaOH and brine, and it was dried over Na 2 SO 4 . The organic layer was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-10% methanol/dichloromethane + 0.1% NH 4 OH) to give the product as a yellow oil (32 mg, 33%); 1 H NMR (300 MHz) (DMSO-d 6 ) δ 11.07 (bs, 1H), 7.57 (d, J = 2 Hz, 1H), 7.24 (d, J = 9 Hz, 1H), 7.08 (dd, J = 8 Hz, J = 2 Hz, 1H), 6.18 (s, 1H), 4.11-4.04 (m, 1H), 1.34 (d, J = 7 Hz, 3H); LC/MS RT = 2.50 (M-H - : 237/239). 5-Bromo-N-methoxy-N-methyl-1 H -indole-2-carboxamide (14b) 5-Bromo-1 H -indole-2-carboxylic acid (1.0 g, 4.17 mmol), N,O-dimethylhydroxylamine hydrochloride (611 mg, 6.26 mmol), HOBt (563 mg, 4.17 mmol), EDC hydrochloride (1.68 g, 8.76 mmol) and triethylamine (2.32 mL, 16.68 mmol) were dissolved in anhydrous DMF (40 mL). The mixture was stirred for overnight at room temperature. The reaction mixture was diluted with ethyl acetate, and the organic layer was washed with water, saturated NH 4 Cl, 10% NaOH and brine. The organic layer was dried over Na 2 SO 4 , and it was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-100% ethyl acetate/hexane) to give product as a white solid (800 mg, 68%); 1 H NMR (300 MHz) (CDCl 3 ) δ 9.33 (bs, 1H), 7.83 (s, 1H), 7.38 (dd, J = 9 Hz, J = 1 Hz, 1H), 7.31 (d, J = 9 Hz, 1H), 7.15 (s, 1H), 3.85 (s, 3H), 3.43 (s, 3H). 1-(5-Bromo-1 H -indol-2-yl)ethan-1-one (14c) 5-Bromo-N-methoxy-N-methyl-1 H -indole-2-carboxamide (600 mg, 2.12 mmol) was dissolved in anhydrous tetrahydrofuran (50 mL). The mixture was cooled to -78 o C, then, a solution of 1.6 M methyllithium in diethyl ether (4.00 mL, 6.36 mmol) was added. The mixture was stirred for 2 hours at -78 o C. An additional solution of 1.6 M methyllithium in diethyl ether (4.00 mL, 6.36 mmol) was added. The mixture was stirred for an hour at -78 o C. The reaction was stopped by addition of water (10 mL). After removal of the solvent, the reaction mixture was diluted with ethyl acetate. The organic layer was washed with saturated NH 4 Cl and brine, and it was dried over Na 2 SO 4 . The organic layer was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-20% ethyl acetate/hexane) to give product as a white solid (344 mg, 68%); 1 H NMR (300 MHz) (CDCl 3 ) δ 9.19 (bs, 1H), 7.85 (s, 1H), 7.42 (dd, J = 9 Hz, J = 2 Hz, 1H), 7.31 (d, J = 8 Hz, 1H), 7.12 (s, 1H), 2.60 (s, 3H). 1-(1 H -indol-2-yl)ethan-1-amine (15). 1-(1 H -Indol-2-yl)ethan-1-one (100 mg, 0.63 mmol), ammonium acetate (486 mg, 6.30 mmol) and sodium cyanoborohydride (198 mg, 3.15 mmol) were dissolved in ethanol (10 mL). The mixture was stirred for overnight at 60 o C. The reaction mixture was diluted with ethyl acetate. The organic layer was washed with 10% NaOH and brine, and it was dried over Na 2 SO 4 . The organic layer was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (100% dichloromethane followed by 10% methanol/dichloromethane + 0.1% NH 4 OH) to give the product as a colorless oil (85 mg, 84%); 1 H NMR (300 MHz) (CD 3 OD) δ 7.44 (d, J = 8 Hz, 1H), 7.29 (d, J = 8 Hz, 1H), 7.03 (t, J = 8 Hz, 1H), 6.94 (t, J = 7 Hz, 1H), 6.29 (s, 1H), 4.24-4.17 (m, 1H), 1.51 (d, J = 7 Hz, 3H); LC/MS RT = 2.19 (M-H - : 159). N-Methoxy-N-methyl-1 H -indole-2-carboxamide (15b) 1 H -Indole-2-carboxylic acid (1.0 g, 6.21 mmol), N,O-dimethylhydroxylamine hydrochloride (909 mg, 9.32 mmol), HOBt (839 mg, 6.21 mmol), EDC hydrochloride (2.5 g, 13.04 mmol) and triethylamine (3.46 mL, 24.84 mmol) were dissolved in anhydrous DMF (40 mL). The mixture was stirred for overnight at room temperature. The reaction mixture was diluted with ethyl acetate, and the organic layer was washed with water, saturated NH 4 Cl, 10% NaOH and brine. The organic layer was dried over Na 2 SO 4 , and it was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-30% ethyl acetate/hexane) to give product as a white solid (450 mg, 35%); 1 H NMR (300 MHz) (CDCl 3 ) δ 9.30 (bs, 1H), 7.70 (d, J = 8 Hz, 1H), 7.44 (d, J = 8 Hz, 1H), 7.31 (t, J = 8 Hz, 1H), 7.25 (s, 1H), 7.14 (t, J = 8 Hz, 1H), 3.85 (s, 3H), 3.44 (s, 3H). 1-(1 H -Indol-2-yl)ethan-1-one (15c) N-Methoxy-N-methyl-1 H -indole-2-carboxamide (450 mg, 2.20 mmol) was dissolved in anhydrous tetrahydrofuran (50 mL). The mixture was cooled to -78 o C, then, a solution of 1.6 M methyllithium in diethyl ether (4.13 mL, 6.60 mmol) was added. The mixture was stirred for 2 hours at -78 o C. An additional solution of 1.6 M methyllithium in diethyl ether (4.13 mL, 6.60 mmol) was added. The mixture was stirred for an hour at -78 o C. The reaction was stopped by addition of water (10 mL). After removal of the solvent, the reaction mixture was diluted with ethyl acetate. The organic layer was washed with saturated NH 4 Cl and brine, and it was dried over Na 2 SO 4 . The organic layer was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-20% ethyl acetate/hexane) to give product as a white solid (290 mg, 83%); 1 H NMR (300 MHz) (CDCl 3 ) δ 9.37 (bs, 1H), 7.72 (d, J = 8 Hz, 1H), 7.45 (d, J = 8 Hz, 1H), 7.38-7.33 (m, 1H), 7.22-7.14 (m, 2H), 2.62 (s, 3H). 1-(5-Fluoro-1 H -indol-2-yl)ethan-1-amine (16) 1-(5-Fluoro-1 H -indol-2-yl)ethan-1-one (100 mg, 0.56 mmol), ammonium acetate (432 mg, 5.60 mmol) and sodium cyanoborohydride (176 mg, 2.80 mmol) were dissolved in ethanol (10 mL). The mixture was stirred for overnight at 60 o C. The reaction mixture was acidified with 6N HCl, and it was washed with ethyl acetate. Then, the aqueous layer was basified with NaOH, and it was extracted with ethyl acetate. The organic layer was washed with brine, and it was dried over Na 2 SO 4 . The organic layer was concentrated under reduced pressure, and the residue was purified on an ISCO chromatograph (0-10% methanol/dichloromethane + 0.1% NH 4 OH) to give the product as a white solid (61 mg, 60%); 1 H NMR (300 MHz) (CD 3 OD) δ 7.25-7.21 (m, 1H), 7.10 (d, J = 8 Hz, 1H), 6.79 (t, J = 8 Hz, 1H), 6.28 (s, 1H), 4.22-4.17 (m, 1H), 1.49 (d, J = 6 Hz, 3H); LC/MS RT = 2.33 (M-H - : 177). 5-Fluoro-N-methoxy-N-methyl-1 H -indole-2-carboxamide (16b) 5-Fluoro-1 H -indole-2-carboxylic acid (1.0 g, 5.58 mmol), N, O-dimethylhydroxylamine hydrochloride (816 mg, 8.37 mmol), HOBt (754 mg, 5.58 mmol), EDC hydrochloride (2.25 g, 11.7 mmol) and triethylamine (3.11 mL, 22.3 mmol) were dissolved in anhydrous DMF (50 mL). The mixture was stirred for overnight at room temperature. The reaction mixture was diluted with ethyl acetate, and the organic layer was washed with water, saturated NH 4 Cl, 10% NaOH and brine. The organic layer was dried over Na 2 SO 4 , and it was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-50% ethyl acetate/hexane) to give product as a white solid (770 mg, 62%); 1 H NMR (300 MHz) (CDCl 3 ) δ 9.94 (bs, 1H), 7.41-7.36 (m, 1H), 7.32 (dd, J = 9 Hz, J = 2 Hz, 1H), 7.19 (s, 1H), 7.06 (td, J = 9 Hz, J = 3 Hz, 1H), 3.85 (s, 3H), 3.46 (s, 3H). 1-(5-Fluoro-1 H -indol-2-yl)ethan-1-one (16c) 5-Fluoro-N-methoxy-N-methyl-1 H -indole-2-carboxamide (300 mg, 1.35 mmol) was dissolved in anhydrous tetrahydrofuran (25 mL). The mixture was cooled to -78 o C, then, a solution of 1.6 M methyllithium in diethyl ether (2.53 mL, 4.05 mmol) was added. The mixture was stirred for 2 hours at -78 o C. The reaction was stopped by addition of water (10 mL). After removal of the solvent, the reaction mixture was diluted with ethyl acetate. The organic layer was washed with saturated NH 4 Cl and brine, and it was dried over Na 2 SO 4 . The organic layer was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-20% ethyl acetate/hexane) to give product as a white solid (220 mg, 92%); 1 H NMR (300 MHz) (CDCl 3 ) δ 9.18 (bs, 1H), 7.39-7.32 (m, 2H), 7.16-7.08 (m, 2H), 2.60 (s, 3H). 1-(5-(Trifluoromethyl)-1 H -indol-2-yl)ethan-1-amine (17). 1-(5-Trifluoromethyl-1 H -indol-2-yl)ethan-1-one (100 mg, 0.44 mmol), ammonium acetate (339 mg, 4.40 mmol) and sodium cyanoborohydride (138 mg, 2.20 mmol) were dissolved in ethanol (10 mL). The mixture was stirred for overnight at 60 o C. The reaction mixture was acidified with 6N HCl, and it was washed with ethyl acetate. Then, the aqueous layer was basified with NaOH, and it was extracted with ethyl acetate. The organic layer was washed with brine, and it was dried over Na 2 SO 4 . The organic layer was concentrated under reduced pressure, and the residue was purified on an ISCO chromatograph (0-10% methanol/dichloromethane + 0.1% NH 4 OH) to give the product as a white solid (42 mg, 42%); 1 H NMR (300 MHz) (DMSO-d 6 ) δ 11.34 (bs, 1H), 7.79 (s, 1H), 7.46 (d, J = 9 Hz, 1H), 7.27 (d, J = 8 Hz, 1H), 6.36 (s, 1H), 4.15-4.09 (m, 1H), 1.37 (d, J = 6 Hz, 3H); LC/MS RT = 2.50 (M-H - : 227). 5-(Trifluoromethyl)-1 H -indole-2-carboxylic acid (17a) 2-Iodo-4-(trifluoromethyl)aniline (1.0 g, 3.48 mmol), pyruvic acid (0.74 mL, 10.44 mmol), DABCO (1.17 g, 10.44 mmol) and Pd(OAc) 2 (79 mg, 0.35 mmol) were dissolved in anhydrous DMF (10 mL). The mixture was purged with nitrogen, and it was stirred for 4 hours at 110 o C. The reaction mixture was diluted with ethyl acetate, and the organic layer was washed with 1N HCl and brine. The organic layer was dried over Na 2 SO 4 , and it was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-50% ethyl acetate/hexane) to give product as a beige solid (484 mg, 54%); 1 H NMR (300 MHz) (CD 3 OD) δ 11.66 (bs, 1H), 7.98 (s, 1H), 7.57 (d, J = 9 Hz, 1H), 7.47 (d, J = 9 Hz, 1H), 7.26 (s, 1H). N-Methoxy-N-methyl-5-(trifluoromethyl)-1 H -indole-2-carboxamide (17b) 5-(Trifluoromethyl)-1 H -indole-2-carboxylic acid (484 mg, 2.11 mmol), N, O-dimethyl-hydroxylamine hydrochloride (309 mg, 3.17 mmol), HOBt (285 mg, 2.11 mmol), EDC hydrochloride (849 mg, 4.43 mmol) and triethylamine (1.18 mL, 8.44 mmol) were dissolved in anhydrous DMF (50 mL). The mixture was stirred for overnight at room temperature. The reaction mixture was diluted with ethyl acetate, and the organic layer was washed with water, saturated NH 4 Cl, 10% NaOH and brine. The organic layer was dried over Na 2 SO 4 , and it was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-50% ethyl acetate/hexane) to give product as a white solid (402 mg, 70%); 1 H NMR (300 MHz) (CDCl 3 ) δ 9.97 (bs, 1H), 8.01 (s, 1H), 7.53 (m, 2H), 7.32 (s, 1H), 3.86 (s, 3H), 3.45 (s, 3H). 1-(5-(Trifluoromethyl)-1 H -indol-2-yl)ethan-1-one (17c) N-Methoxy-N-methyl-5-(trifluoromethyl)-1 H -indole-2-carboxamide (300 mg, 1.10 mmol) was dissolved in anhydrous tetrahydrofuran (20 mL). The mixture was cooled to -78 o C, then, a solution of 1.6 M methyllithium in diethyl ether (2.06 mL, 3.30 mmol) was added. The mixture was stirred for 2 hours at -78 o C. The reaction was stopped by addition of water (10 mL). After removal of the solvent, the reaction mixture was diluted with ethyl acetate. The organic layer was washed with saturated NH 4 Cl and brine, and it was dried over Na 2 SO 4 . The organic layer was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-20% ethyl acetate/hexane) to give product as a white solid (197 mg, 80%); 1 H NMR (300 MHz) (CDCl 3 ) δ 9.40 (bs, 1H), 8.03 (s, 1H), 7.59-7.51 (m, 2H), 7.28 (s, 1H), 2.64 (s, 3H). 1-(5-Methoxy-1 H -indol-2-yl)ethan-1-amine (18). 1-(5-Methoxy-1 H -indol-2-yl)ethan-1-one (100 mg, 0.42 mmol), ammonium acetate (324 mg, 4.20 mmol) and sodium cyanoborohydride (132 mg, 2.10 mmol) were dissolved in ethanol (10 mL). The mixture was stirred for overnight at 60 o C. The reaction mixture was acidified with 6N HCl, and it was washed with ethyl acetate. Then, the aqueous layer was basified with NaOH, and it was extracted with ethyl acetate. The organic layer was washed with brine, and it was dried over Na 2 SO 4 . The organic layer was concentrated under reduced pressure, and the residue was purified on an ISCO chromatograph (0-10% methanol/dichloromethane + 0.1% NH 4 OH) to give the product as a white solid (58 mg, 58%); 1 H NMR (300 MHz) (DMSO-d 6 ) δ 10.66 (bs, 1H), 7.15 (d, J = 9 Hz, 1H), 6.90 (d, J = 2 Hz, 1H), 6.61 (dd, J = 9 Hz, J = 2 Hz, 1H), 6.09 (s, 1H), 4.08-4.02 (m, 1H), 3.69 (s, 3H), 1.33 (d, J = 6 Hz, 3H); LC/MS RT = 2.17 (M-H - : 189). N,5-Dimethoxy-N-methyl-1 H -indole-2-carboxamide (18b) 5-Methoxy-1 H -indole-2-carboxylic acid (1.0 g, 5.23 mmol), N, O-dimethylhydroxylamine hydrochloride (766 mg, 7.85 mmol), HOBt (707 mg, 5.23 mmol), EDC hydrochloride (2.10 g, 10.98 mmol) and triethylamine (2.92 mL, 20.92 mmol) were dissolved in anhydrous DMF (50 mL). The mixture was stirred for overnight at room temperature. The reaction mixture was diluted with ethyl acetate, and the organic layer was washed with water, saturated NH 4 Cl, 10% NaOH and brine. The organic layer was dried over Na 2 SO 4 , and it was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-50% ethyl acetate/hexane) to give product as a white solid (743 mg, 60%); 1 H NMR (300 MHz) (CDCl 3 ) δ 9.78 (bs, 1H), 7.35 (d, J = 9 Hz, 1H), 7.17 (s, 1H), 7.1 (d, J = 2 Hz, 1H), 6.98 (dd, J = 9 Hz, J = 2 Hz, 1H), 3.85 (s, 3H), 3.84 (s, 3H), 3.46 (s, 3H). 1-(5-Methoxy-1 H -indol-2-yl)ethan-1-one (18c) N,5-Dimethoxy-N-methyl-1 H -indole-2-carboxamide (300 mg, 1.28 mmol) was dissolved in anhydrous tetrahydrofuran (20 mL). The mixture was cooled to -78 o C, then, a solution of 1.6 M methyllithium in diethyl ether (2.40 mL, 3.84 mmol) was added. The mixture was stirred for 2 hours at -78 o C. The reaction was stopped by addition of water (10 mL). After removal of the solvent, the reaction mixture was diluted with ethyl acetate. The organic layer was washed with saturated NH 4 Cl and brine, and it was dried over Na 2 SO 4 . The organic layer was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-20% ethyl acetate/hexane) to give product as a white solid (180 mg, 74%); 1 H NMR (300 MHz) (CDCl 3 ) δ 8.91 (bs, 1H), 7.31 (d, J = 9 Hz, 1H), 7.12-7.08 (m, 2H), 7.03 (dd, J = 9 Hz, J = 3 Hz, 1H), 3.85 (s, 3H), 2.58 (s, 3H). 1-(5-(Trifluoromethoxy)-1 H -indol-2-yl)ethan-1-amine (19) 1-(5-(Trifluoromethoxy)-1 H -indol-2-yl)ethan-1-one (100 mg, 0.41 mmol), ammonium acetate (316 mg, 4.10 mmol) and sodium cyanoborohydride (129 mg, 2.05 mmol) were dissolved in ethanol (10 mL). The mixture was stirred for overnight at 60 o C. The reaction mixture was acidified with 6N HCl, and it was washed with ethyl acetate. Then, the aqueous layer was basified with NaOH, and it was extracted with ethyl acetate. The organic layer was washed with brine, and it was dried over Na 2 SO 4 . The organic layer was concentrated under reduced pressure, and the residue was purified on an ISCO chromatograph (0-10% methanol/dichloromethane + 0.1% NH 4 OH) to give the product as a white solid (48 mg, 48%); 1 H NMR (300 MHz) (DMSO-d 6 ) δ 11.15 (bs, 1H), 7.38 (s, 1H), 7.35 (d, J = 9 Hz, 1H), 6.94 (d, J = 8 Hz, 1H), 6.27 (s. 1H), 4.14-4.07 (m, 1H), 1.36 (d, J = 7 Hz, 3H); LC/MS RT = 2.55 (M-H - : 243). 5-(Trifluoromethoxy)-1 H -indole-2-carboxylic acid (19a) 2-Iodo-4-(trifluoromethoxy)aniline (1.0 g, 3.30 mmol), pyruvic acid (0.70 mL, 9.90 mmol), DABCO (1.11 g, 9.90 mmol) and Pd(OAc) 2 (74 mg, 0.33 mmol) were dissolved in anhydrous DMF (10 mL). The mixture was purged with nitrogen, and it was stirred for 4 hours at 110 o C. The reaction mixture was diluted with ethyl acetate, and the organic layer was washed with 1N HCl and brine. The organic layer was dried over Na 2 SO 4 , and it was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-50% ethyl acetate/hexane) to give product as a beige solid (592 mg, 73%); 1 H NMR (300 MHz) (DMSO-d 6 ) δ 12.02 (bs, 1H), 7.64 (s, 1H), 7.49 (d, J = 9 Hz, 1H), 7.20 (d, J = 6 Hz, 1H), 7.13 (s, 1H). N-Methoxy-N-methyl-5-(trifluoromethoxy)-1 H -indole-2-carboxamide (19b) 5-(Trifluoromethoxy)-1 H -indole-2-carboxylic acid (592 mg, 2.41 mmol), N, O-dimethyl-hydroxylamine hydrochloride (353 mg, 3.62 mmol), HOBt (326 mg, 2.41 mmol), EDC hydrochloride (970 mg, 5.06 mmol) and triethylamine (1.18 mL, 8.44 mmol) were dissolved in anhydrous DMF (50 mL). The mixture was stirred for overnight at room temperature. The reaction mixture was diluted with ethyl acetate, and the organic layer was washed with water, saturated NH 4 Cl, 10% NaOH and brine. The organic layer was dried over Na 2 SO 4 , and it was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-70% ethyl acetate/hexane) to give product as a white solid (378 mg, 54%); 1 H NMR (300 MHz) (CDCl 3 ) δ 9.33 (bs, 1H), 7.55 (s, 1H), 7.42 (d, J = 9 Hz, 1H), 7.24 (s, 1H), 7.18 (d, J = 9 Hz, 1H), 3.85 (s, 3H), 3.44 (s, 3H). 1-(5-(Trifluoromethoxy)-1 H -indol-2-yl)ethan-1-one (19c) N-Methoxy-N-methyl-5-(trifluoromethoxy)-1 H -indole-2-carboxamide (378 mg, 1.31 mmol) was dissolved in anhydrous tetrahydrofuran (20 mL). The mixture was cooled to -78 o C, then, a solution of 1.6 M methyllithium in diethyl ether (2.46 mL, 3.93 mmol) was added. The mixture was stirred for 2 hours at -78 o C. The reaction was stopped by addition of water (10 mL). After removal of the solvent, the reaction mixture was diluted with ethyl acetate. The organic layer was washed with saturated NH 4 Cl and brine, and it was dried over Na 2 SO 4 . The organic layer was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-20% ethyl acetate/hexane) to give product as a white solid (259 mg, 81%); 1 H NMR (300 MHz) (CDCl 3 ) δ 9.35 (bs, 1H), 7.57 (s, 1H), 7.44 (d, J = 9 Hz, 1H), 7.26-7.20 (m, 2H), 2.62 (s, 3H). 1-(4,6-Dichloro-1 H -indol-2-yl)ethan-1-amine (20). 1-(4,6-Dichloro-1 H -indol-2-yl)ethan-1-one (100 mg, 0.44 mmol), ammonium acetate (339 mg, 4.40 mmol) and sodium cyanoborohydride (138 mg, 2.20 mmol) were dissolved in ethanol (10 mL). The mixture was stirred for overnight at 60 o C. The reaction mixture was acidified with 6N HCl, and it was washed with ethyl acetate. Then, the aqueous layer was basified with NaOH, and it was extracted with ethyl acetate. The organic layer was washed with brine, and it was dried over Na 2 SO 4 . The organic layer was concentrated under reduced pressure, and the residue was purified on an ISCO chromatograph (0-10% methanol/dichloromethane + 0.1% NH 4 OH) to give the product as a white solid (55 mg, 54%); 1 H NMR (300 MHz) (DMSO-d 6 ) δ 7.32 (s, 1H), 7.06 (s, 1H), 6.27 (s, 1H), 4.12-4.01 (m, 1H), 1.35 (d, J = 8 Hz, 3H); LC/MS RT = 2.61 (M-H - : 227/229). 4,6-Dichloro-N-methoxy-N-methyl-1 H -indole-2-carboxamide (20b) 4,6-Dichloro-1 H -indole-2-carboxylic acid (500 mg, 2.17 mmol), N, O-dimethylhydroxylamine hydrochloride (423 mg, 4.34 mmol), HOBt (332 mg, 2.17 mmol), EDC hydrochloride (874 mg, 4.56 mmol) and triethylamine (1.32 mL, 8.68 mmol) were dissolved in anhydrous DMF (40 mL). The mixture was stirred for overnight at room temperature. The reaction mixture was diluted with ethyl acetate, and the organic layer was washed with water, saturated NH 4 Cl, 10% NaOH and brine. The organic layer was dried over Na 2 SO 4 , and it was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-50% ethyl acetate/hexane) to give product as a white solid (505 mg, 85%); 1 H NMR (300 MHz) (DMSO-d 6 ) δ 12.10 (bs, 1H), 7.45 (d, J = 1 Hz, 1H), 7.24 (d, J = 1 Hz, 1H), 7.09 (s, 1H), 3.80 (s, 3H), 3.33 (s, 3H). 1-(4,6-Dichloro-1 H -indol-2-yl)ethan-1-one (20c) 4,6-Dichloro-N-methoxy-N-methyl-1 H -indole-2-carboxamide (500 mg, 1.83 mmol) was dissolved in anhydrous tetrahydrofuran (25 mL). The mixture was cooled to -78 o C, and, then, a solution of 1.6 M methyllithium in diethyl ether (3.43 mL, 5.49 mmol) was added. The mixture was stirred for 2 hours at -78 o C. The reaction was stopped by addition of water (10 mL). After removal of the solvent, the reaction mixture was diluted with ethyl acetate. The organic layer was washed with saturated NH 4 Cl and brine, and it was dried over Na 2 SO 4 . The organic layer was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-20% ethyl acetate/hexane) to give product as a white solid (278 mg, 67%); 1 H NMR (300 MHz) (DMSO-d 6 ) δ 12.20 (bs, 1H), 7.41 (s, 1H), 7.39 (s, 1H), 7.27 (s, 1H), 2.57 (s, 3H). 1-(5,6-Difluoro-1 H -indol-2-yl)ethan-1-amine (21). 1-(5,6-Difluoro-1 H -indol-2-yl)ethan-1-one (150 mg, 0.77 mmol), ammonium acetate (594 mg, 7.70 mmol) and sodium cyanoborohydride (242 mg, 3.85 mmol) were dissolved in ethanol (10 mL). The mixture was stirred for overnight at 60 o C. The reaction mixture was acidified with 6N HCl, and it was washed with ethyl acetate. Then, the aqueous layer was basified with NaOH, and it was extracted with ethyl acetate. The organic layer was washed with brine, and it was dried over Na 2 SO 4 . The organic layer was concentrated under reduced pressure, and the residue was purified on an ISCO chromatograph (0-10% methanol/dichloromethane + 0.1% NH 4 OH) to give the product as a white solid (69 mg, 46%); 1 H NMR (300 MHz) (DMSO-d 6 ) δ 11.49 (bs, 1H), 8.49 (bs, 2H), 7.57-7.50 (m, 1H), 7.44-7.38 (m, 1H), 6.50 (s, 1H), 4.58-4.51 (m, 1H), 1.58 (d, J = 6 Hz, 3H); LC/MS RT = 2.47 (M-H - : 195). 5,6-Difluoro-N-methoxy-N-methyl-1 H -indole-2-carboxamide (21b) 5,6-Difluoro-1 H -indole-2-carboxylic acid (500 mg, 2.54 mmol), N, O-dimethylhydroxylamine hydrochloride (496 mg, 5.08 mmol), HOBt (389 mg, 2.54 mmol), EDC hydrochloride (1.02 g, 5.33 mmol) and triethylamine (1.54 mL, 10.2 mmol) were dissolved in anhydrous DMF (20 mL). The mixture was stirred for overnight at room temperature. The reaction mixture was diluted with ethyl acetate, and the organic layer was washed with water, saturated NH 4 Cl, 10% NaOH and brine. The organic layer was dried over Na 2 SO 4 , and it was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-50% ethyl acetate/hexane) to give product as a white solid (367 mg, 60%); 1 H NMR (300 MHz) (CDCl 3 ) δ 9.33 (bs, 1H), 7.45-7.38 (m, 1H), 7.23-7.17 (m, 2H), 3.85 (s, 3H), 3.42 (s, 3H). 1-(5,6-Difluoro-1 H -indol-2-yl)ethan-1-one (21c) 5,6-Difluoro-N-methoxy-N-methyl-1 H -indole-2-carboxamide (350 mg, 1.46 mmol) was dissolved in anhydrous tetrahydrofuran (20 mL). The mixture was cooled to -78 o C, then, a solution of 1.6 M methyllithium in diethyl ether (2.73 mL, 4.38 mmol) was added. The mixture was stirred for 2 hours at -78 o C. The reaction was stopped by addition of water (10 mL). After removal of the solvent, the reaction mixture was diluted with ethyl acetate. The organic layer was washed with saturated NH 4 Cl and brine, and it was dried over Na 2 SO 4 . The organic layer was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-20% ethyl acetate/hexane) to give product as a white solid (207 mg, 73%); 1 H NMR (300 MHz) (CDCl 3 ) δ 9.24 (bs, 1H), 7.47-7.41 (m, 1H), 7.26-7.14 (m, 2H), 2.58 (s, 3H). 1-(5-Bromo-1 H -indol-2-yl)-N-methylethan-1-amine (22). To a mixture of 1-(5-bromo-1 H -indol-2-yl)ethan-1-one (50 mg, 0.21 mmol) in ethanol (5 mL), a solution of 2.0 M methylamine in tetrahydrofuran (1.05 mL, 2.10 mmol) was added. The mixture was treated with catalytic amount of acetic acid, and it was stirred for an hour at 60 o C. Then, the mixture was treated with sodium cyanoborohydride (66 mg, 1.05 mmol), and it was stirred for overnight at 60 o C. The reaction mixture was acidified with 6N HCl, and it was washed with ethyl acetate. Then, the aqueous layer was basified with NaOH, and it was extracted with ethyl acetate. The organic layer was washed with brine, and it was dried over Na 2 SO 4 . The organic layer was concentrated under reduced pressure, and the residue was purified on an ISCO chromatograph (0-10% methanol/dichloromethane + 0.1% NH 4 OH) to give the product as a colorless oil (53 mg, 100%); 1 H NMR (300 MHz) (CDCl 3 ) δ 9.79 (bs, 1H), 7.67 (s, 1H), 7.27-7.26 (m, 2H), 6.41 (s, 1H), 4.35-4.32 (m, 1H), 2.43 (s, 3H), 1.71 (d, J = 7 Hz, 3H); LC/MS RT = 2.60 (M-H - : 251/253). 1-(5-Bromo-1 H -indol-2-yl)-N,N-dimethylethan-1-amine (23). To a mixture of 1-(5-bromo-1 H -indol-2-yl)ethan-1-one (75 mg, 0.32 mmol) in ethanol (10 mL), a solution of 2.0 M N,N-dimethylamine in tetrahydrofuran (1.60 mL, 3.20 mmol) was added. The mixture was treated with catalytic amount of acetic acid followed by sodium cyanoborohydride (101 mg, 1.69 mmol), and it was stirred for overnight at 60 o C. The reaction mixture was acidified with 6N HCl, and it was washed with ethyl acetate. Then, the aqueous layer was basified with NaOH, and it was extracted with ethyl acetate. The organic layer was washed with brine, and it was dried over Na 2 SO 4 . The organic layer was concentrated under reduced pressure, and the residue was purified on an ISCO chromatograph (0-10% methanol/dichloromethane + 0.1% NH 4 OH) to give the product as a colorless oil (12 mg, 14%); 1 H NMR (300 MHz) (CDCl 3 ) δ 8.81 (bs, 1H), 7.66 (s, 1H), 7.26-7.20 (m, 2H), 6.24 (s, 1H), 3.82-3.80 (m, 1H), 2.25 (s, 6H), 1.40 (d, J = 7 Hz, 3H); LC/MS RT = 2.62 (M-H - : 265/267). 1-(5-Bromo-1-methyl-1 H -indol-2-yl)ethan-1-amine (24). 1-(5-Bromo-1-methyl-1 H -indol-2-yl)ethan-1-one (91 mg, 0.36 mmol), ammonium acetate (277 mg, 3.60 mmol) and sodium cyanoborohydride (113 mg, 1.80 mmol) were dissolved in ethanol (10 mL). Then, catalytic amount of acetic acid was added. The mixture was stirred for overnight at 60 o C. The reaction mixture was diluted with ethyl acetate. The organic layer was washed with 10% NaOH and brine, and it was dried over Na 2 SO 4 . The organic layer was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-10% methanol/dichloromethane + 0.1% NH 4 OH) to give the product as a white solid (57 mg, 63%); 1 H NMR (300 MHz) (CD 3 OD) δ 7.62 (d, J = 2 Hz, 1H), 7.28 (d, J = 9 Hz, 1H), 7.21 (dd, J = 9 Hz, J = 2 Hz, 1H), 6.44 (s, 1H), 4.44-4.38 (m, 1H), 3.75(s, 3H), 1.56 (d, J = 7 Hz, 3H); LC/MS RT = 2.49 (M-NH 2 + :236/238). 1-(5-Bromo-1-methyl-1 H -indol-2-yl)ethan-1-one (24a) 1-(5-Bromo-1 H -indol-2-yl)ethan-1-one (100 mg, 0.42 mmol), K 2 CO 3 (116 mg, 0.84 mmol) and methyl iodide (52 µL, 0.84 mmol) were dissolved in anhydrous DMF (5 mL). The mixture was stirred for 3 hours at 60 o C. After the mixture was cooled to room temperature, it was diluted with ethyl acetate. The organic layer was washed with water and brine, and it was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-50% ethyl acetate/hexane) to give product as a white solid (91 mg, 86%); 1 H NMR (300 MHz) (CDCl 3 ) δ 7.82 (d, J = 2 Hz, 1H), 7.45 (dd, J = 9 Hz, J = 2 Hz, 1H), 7.27 (d, J = 9 Hz, 1H), 7.20 (s, 1H), 4.05 (s, 3H), 2.61 (s, 3H). (5-Bromo-1 H -indol-2-yl)methanamine (25). 5-Bromo-1 H -indole-2-carbaldehyde (55 mg, 0.25 mmol) was dissolved in ethanol (25 mL), and it was treated with ammonium acetate (193 mg, 2.50 mmol). The mixture was stirred for an hour at room temperature. Then, sodium cyanoborohydride (79 mg, 1.25 mmol) was added, and it was stirred for 3 hours at room temperature. The reaction mixture was diluted with ethyl acetate, and it was washed with saturated NaHCO 3 and brine. The organic layer was dried over Na 2 SO 4 , and it was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-10% methanol/dichloromethane + 0.1% NH 4 OH) to give (5-bromo-1 H -indol-2-yl)methanamine as a white solid (26 mg, 46%). 1 H NMR (300 MHz) (DMSO-d 6 ) δ 11.25 (bs, 1H), 7.72 (d, J = 2 Hz, 1H), 7.36 (d, J = 8 Hz, 1H), 7.19 (dd, J = 9 Hz, J = 2 Hz, 1H), 6.46 (s, 1H), 4.11 (s, 2H); LC/MS RT = 2.52 (M-H - : 223/225). along with bis((5-bromo-1 H -indol-2-yl)methyl)amine as a white solid (26 mg, 49%). 1 H NMR (300 MHz) (CDCl 3 ) δ 7.65 (s, 2H), 7.26-7.21 (m, 4H), 6.31 (s, 2H), 3.97 (s, 4H); LC/MS RT = 2.97 (M-H - : 430/432/434). Methyl 5-bromo-1 H -indole-2-carboxylat e (25a) 5-Bromo-1 H -indole-2-carboxylic acid (1.5 g, 6.25 mmol) was dissolved in methanol (100 mL), and it was treated with catalytic amount of concentrated sulfuric acid. The mixture was refluxed overnight. After removal of solvent, it was diluted with ethyl acetate. The organic layer was washed with 10% NaOH and brine, and it was dried over Na 2 SO 4 . The organic layer was concentrated under reduced pressure, and the residue was purified on an ISCO chromatograph (0-50% ethyl acetate/hexane) to give product as a white solid (1.19 g, 75%); 1 H NMR (300 MHz) (CDCl 3 ) δ 8.97 (bs, 1H), 7.83 (s, 1H), 7.40 (dd, J = 9 Hz, J = 2 Hz, 1H), 7.30 (d, J = 9 Hz, 1H), 7.14 (s, 1H), 3.95 (s, 3H). (5-Bromo-1 H -indol-2-yl)methanol (25b) Methyl 5-bromo-1 H -indole-2-carboxylate (200 mg, 0.79 mmol) and lithium borohydride (86 mg, 3.95 mmol) were dissolved in anhydrous tetrahydrofuran (10 mL) at 0 o C, and it was stirred for overnight at room temperature. The reaction mixture was diluted with ethyl acetate, and it was washed with saturated NH 4 Cl and brine. The organic layer was dried over Na 2 SO 4 , and it was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-50% ethyl acetate/hexane) to give product as a white solid (149 mg, 83 %); 1 H NMR (300 MHz) (CDCl 3 ) δ 8.37 (bs, 1H), 7.69 (s, 1H), 7.28-7.21 (m, 2H), 6.34 (d, J = 1 Hz, 1H), 4.84 (s, 2H). 5-Bromo-1 H -indole-2-carbaldehyde (25c) (5-Bromo-1 H -indol-2-yl)methanol (100 mg, 0.44 mmol) and Dess-Martin periodinane (280 mg, 0.66 mmol) were dissolved in dichloromethane (10 mL), and it was stirred for 15 minutes at room temperature. The reaction mixture was diluted with ethyl acetate, and it was washed with 10% sodium thiosulfate, saturated NaHCO 3 and brine. The organic layer was dried over Na 2 SO 4 , and it was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-10% ethyl acetate/hexane) to give product as a white solid (55 mg, 56%); 1 H NMR (300 MHz) (CDCl 3 ) δ 9.85 (s, 1H), 9.05 (bs, 1H), 7.90 (s, 1H), 7.47 (dd, J = 9 Hz, J = 2 Hz, 1H), 7.34 (d, J = 9 Hz, 1H), 7.21 (s, 1H). N 1 -(2-bromo-5,6,7,8,9,10-hexahydrocyclohepta[ b ]indol-6-yl)propane-1,3-diamine (26). To a solution of tert -butyl (3-((2-bromo-5,6,7,8,9,10-hexahydrocyclohepta[ b ]indol-6-yl)amino)propyl)carbamate (614 mg, 1.41 mmol) in methanol (3 mL), 4N HCl in dioxane (6 mL, 24 mmol) was added. The reaction mixture was stirred for 1.0 hour at room temperature. After removal of solvent, the reaction mixture was diluted with ethyl acetate. The organic layer was washed with 10% sodium hydroxide and brine, and it was dried over sodium sulfate. The organic layer was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-10% methanol/dichloromethane + 0.1% NH 4 OH) to give the product as yellow oil (202 mg, 43%); 1 H NMR (300 MHz) (CDCl 3 ) δ 10.02 (bs, 1H), 7.58 (s, 1H), 7.15-7.14 (m, 2H), 3.87-3.83 (m, 1H), 2.96-2.75 (m, 6H), 2.63-2.51 (m, 1H), 2.12-2.02 (m, 2H), 1.94-1.88 (m, 1H), 1.74-1.62 (m, 4H); LC/MS RT = 2.52 (M+H + : 336/338). tert- Butyl (3-((2-bromo-5,6,7,8,9,10-hexahydrocyclohepta[ b ]indol-6-yl)amino)propyl)carbamate (26a) 2-Bromo-7,8,9,10-tetrahydrocyclohepta[ b ]indol-6(5 H )-one (500 mg, 1.80 mmol), N-Boc propylenediamine (941 mg, 5.40 mmol) and sodium cyanoborohydride (566 mg, 9.00 mmol) were dissolved in ethanol (20 mL). Catalytic amount of acetic acid was added. The reaction mixture was stirred for overnight at 60 o C. After removal of solvent, the reaction mixture was diluted with ethyl acetate. The organic layer was washed with 10% sodium hydroxide and brine, and it was dried over sodium sulfate. The organic layer was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-100% ethyl acetate/hexane) to give the product as yellow oil (614 mg, 78%); 1 H NMR (300 MHz) (CDCl 3 ) δ 9.51 (bs, 1H), 7.61 (s, 1H), 7.36-7.25 (m, 2H), 5.14-5.12 (m, 1H), 4.62-4.61 (m, 1H), 3.25-2.80 (m, 6H), 2.45-2.43 (m, 1H), 2.12-1.82 (m, 7H), 1.39 (s, 9H); LC/MS RT = 3.13 (M+H + : 436/438). N 1 -(7-bromo-1,2,3,4-tetrahydrocyclopenta[ b ]indol-3-yl)propane-1,3-diamine (27). To a solution of tert -butyl (3-((7-bromo-1,2,3,4-tetrahydrocyclopenta[ b ]indol-3-yl)amino)-propyl)carbamate (817 mg, 2.00 mmol) in methanol (3 mL), 4N HCl in dioxane (6 mL, 24 mmol) was added. The reaction mixture was stirred for an hour at room temperature. After removal of solvent, the reaction mixture was diluted with ethyl acetate. The organic layer was washed with 10% sodium hydroxide and brine, and it was dried over sodium sulfate. The organic layer was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-10% methanol/dichloromethane + 0.1% NH 4 OH) to give the product as a white solid (616 mg, 100%); 1 H NMR (300 MHz) (CDCl 3 ) δ 9.25 (bs, 1H), 7.58 (s, 1H), 7.26-7.18 (m, 2H), 4.37-4.33 (m, 1H), 2.88-2.65 (m, 8H), 2.12-2.04 (m, 1H), 2.04-1.97 (m, 2H); LC/MS RT = 2.50 (M+H + : 308/310). tert -Butyl (3-((7-bromo-1,2,3,4-tetrahydrocyclopenta[ b ]indol-3-yl)amino)propyl)carbamate (27a) 7-Bromo-1,4-dihydrocyclopenta[ b ]indol-3(2 H )-one (500 mg, 2.00 mmol), N-Boc propylenediamine (1.05 g, 6.00 mmol) and sodium cyanoborohydride (628 mg, 10.00 mmol) were dissolved in ethanol (20 mL). A catalytic amount of acetic acid was added. The reaction mixture was stirred for overnight at 60 o C. After removal of solvent, the reaction mixture was diluted with ethyl acetate. The organic layer was washed with 10% sodium hydroxide and brine, and it was dried over sodium sulfate. The organic layer was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-100% ethyl acetate/hexane) to give the product as a foamy yellow solid (817 mg, 100%); 1 H NMR (300 MHz) (CDCl 3 ) δ 9.16 (bs, 1H), 7.63 (s, 1H), 7.33-7.32 (m, 2H), 5.07-5.03 (m, 1H), 4.83 (m, 1H), 3.26-3.24 (m, 2H), 3.04-2.85 (m, 5H), 2.60-2.54 (m, 1H), 1.98-1.96 (m, 2H), 1.39 (s, 9H); LC/MS RT = 3.08 (M+H + : 408/410). N 1 -(1-(5-Bromo-1 H -indol-2-yl)ethyl)propane-1,3-diamine (28). To a solution of tert- butyl (3-((1-(5-bromo-1H-indol-2-yl)ethyl)amino)propyl)carbamate (817 mg, 2.06 mmol) in methanol (3 mL), 4N HCl in dioxane (6 mL, 24 mmol) was added. The reaction mixture was stirred for an hour at room temperature. After removal of solvent, the reaction mixture was diluted with ethyl acetate. The organic layer was washed with 10% sodium hydroxide and brine, and it was dried over sodium sulfate. The organic layer was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-10% methanol/dichloromethane + 0.1% NH 4 OH) to give the product as colorless oil (332 mg, 54%); 1 H NMR (300 MHz) (CDCl 3 ) δ 9.22 (bs, 1H), 7.65 (s, 1H), 7.26-7.20 (m, 3H), 6.24 (s, 1H), 4.05-4.01 (m, 1H), 2.83-2.54 (m, 4H), 1.67-1.60 (m, 2H), 1.45 (d, J = 6 Hz, 3H); LC/MS RT = 2.49 (M+H + : 296/298). tert- Butyl (3-((1-(5-bromo-1 H -indol-2-yl)ethyl)amino)propyl)carbamate (28a) 1-(5-Bromo-1 H -indol-2-yl)ethan-1-one (500 mg, 2.10 mmol), N-Boc propylenediamine (1.10 g, 6.30 mmol) and sodium cyanoborohydride (314 mg, 10.50 mmol) were dissolved in ethanol (20 mL). Catalytic amount of acetic acid was added. The reaction mixture was stirred for overnight at 60 o C. After removal of solvent, the reaction mixture was diluted with ethyl acetate. The organic layer was washed with 10% sodium hydroxide and brine, and it was dried over sodium sulfate. The organic layer was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-100% ethyl acetate/hexane) to give the product as colorless oil (817 mg, 99%); 1 H NMR (300 MHz) (CDCl 3 ) δ 9.50 (bs, 1H), 7.68 (s, 1H), 7.37-7.26 (m, 3H), 6.43 (s, 1H), 5.04 (m, 1H), 4.35-4.33 (m, 1H), 3.30-3.28 (m, 2H), 2.80-2.76 (m, 2H), 1.85-1.80 (m, 2H), 1.75 (d, J = 7 Hz, 3H), 1.46 (s, 9H). N-(3-((1-(5-bromo-1 H -indol-2-yl)ethyl)amino)propyl)-4,5-dichlorothiophene-2-carboxamide (29). 5-Chlorofuran-2-carboxylic acid (120 mg, 0.82 mmol), EDC hydrochloric acid (157 mg, 0.82 mmol), HOBt (55 mg, 0.41 mmol) and DIPEA (0.23 mL, 1.26 mmol) were dissolved in DMF (5 mL). After 5 minutes of stirring, N 1 -(1-(5-bromo-1 H -indol-2-yl)ethyl)propane-1,3-diamine (120 mg, 0.41 mmol) was added, and it was stirred overnight at room temperature. The reaction mixture was diluted with ethyl acetate, and it was washed with 1N hydrochloric acid, 10% sodium hydroxide and brine. The organic layer was dried over sodium sulfate. The organic layer was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-10% methanol/dichloromethane + 0.1% NH 4 OH) to give the product as a white solid (54 mg, 31%); 1 H NMR (300 MHz) (CDCl 3 ) δ 9.02 (bs, 1H), 7.63 (s, 1H), 7.18 (m, 2H), 7.11 (d, J = 3 Hz, 1H), 7.05 (bs, 1H), 6.32 (d, J = 3 Hz, 1H), 6.24 (s, 1H), 4.03-3.98 (m, 1H), 3.76-3.69 (m, 1H), 3.51-3.35 (m, 1H), 2.73-2.68 (m, 1H), 2.60-2.54 (m, 1H), 1.72-1.69 (m, 2H), 1.49 (d, J = 6 Hz, 3H); LC/MS RT = 2.95 (M+H + : 424/426/428). N-(3-((1-(5-bromo-1 H -indol-2-yl)ethyl)amino)propyl)-5-chlorothiophene-2-carboxamide (30). 5-Chlorothiophene-2-carboxylic acid (249 mg, 1.53 mmol), EDC hydrochloric acid (196 mg, 1.02 mmol), HOBt (69 mg, 0.51 mmol) and DIPEA (0.27 mL, 1.53 mmol) were dissolved in DMF (2 mL). After 5 minutes of stirring, N 1 -(1-(5-bromo-1 H -indol-2-yl)ethyl)propane-1,3-diamine (150 mg, 0.51 mmol) was added, and it was stirred overnight at room temperature. The reaction mixture was diluted with ethyl acetate, and it was washed with 1N hydrochloric acid, 10% sodium hydroxide and brine. The organic layer was dried over sodium sulfate. The organic layer was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-10% methanol/dichloromethane + 0.1% NH 4 OH) to give the product as a white solid (29 mg, 13%); 1 H NMR (300 MHz) (CDCl 3 ) δ 9.07 (bs, 1H), 7.64 (s, 1H), 7.26-7.06 (m, 2H), 6.79-6.78 (m, 2H), 6.26 (s, 1H), 4.05-4.00 (m, 1H), 3.66-3.60 (m, 1H), 3.51-3.39 (m, 1H), 2.75-2.71 (m, 1H), 2.62-2.54 (m, 1H) 1.73-1.70 (m, 2H), 1.51 (d, J = 7 Hz, 3H); LC/MS RT = 3.05 (M+H + : 440/442/444). N-(3-((1-(5-bromo-1 H -indol-2-yl)ethyl)amino)propyl)-4,5-dichlorothiophene-2-carboxamide (31). 4,5-Dichlorothiophene-2-carboxylic acid (162 mg, 0.82 mmol), EDC hydrochloric acid (157 mg, 0.82 mmol), HOBt (55 mg, 0.41 mmol) and DIPEA (0.23 mL, 1.26 mmol) were dissolved in DMF (5 mL). After 5 minutes of stirring, N 1 -(1-(5-bromo-1 H -indol-2-yl)ethyl)propane-1,3-diamine (120 mg, 0.41 mmol) was added, and it was stirred overnight at room temperature. The reaction mixture was diluted with ethyl acetate, and it was washed with 1N hydrochloric acid, 10% sodium hydroxide and brine. The organic layer was dried over sodium sulfate. The organic layer was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-10% methanol/dichloromethane + 0.1% NH 4 OH) to give the product as a white solid (63 mg, 32%); 1 H NMR (300 MHz) (CDCl 3 ) δ 8.88 (bs, 1H), 7.64 (s, 1H), 7.26-7.12 (m, 3H), 7.06 (bs, 1H), 6.26 (s, 1H), 4.04-3.98 (m, 1H), 3.65-3.58 (m, 1H), 3.49-3.40 (m, 1H), 2.75-2.69 (m, 1H), 2.63-2.57 (m, 1H) 1.74-1.68 (m, 2H), 1.49 (d, J = 7 Hz, 3H); LC/MS RT = 3.05 (M+H + : 474/476/478). N-(3-((1-(5-bromo-1 H -indol-2-yl)ethyl)amino)propyl)-3,4-dichlorobenzamide (32). 3,4-Dichlorobenzoic acid (32 mg, 0.17 mmol), EDC hydrochloric acid (65 mg, 0.34 mmol), HOBt (23 mg, 0.17 mmol) and DIPEA (0.09 mL, 0.51 mmol) were disolved in dichloromethane (5 mL). After 5 minutes of stirring, N 1 -(1-(5-bromo-1 H -indol-2-yl)ethyl)propane-1,3-diamine (50 mg, 0.17 mmol) was added, and it was stirred overnight at room temperature. The reaction mixture was diluted with ethyl acetate, and it was washed with 1N hydrochloric acid, 10% sodium hydroxide and brine. The organic layer was dried over sodium sulfate. The organic layer was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-10% methanol/dichloromethane + 0.1% NH 4 OH) to give the product as colorless oil (14 mg, 18%); 1 H NMR (300 MHz) (CDCl 3 ) δ 9.97 (bs, 1H), 7.88 (d, J = 2 Hz, 1H), 7.64 (s, 1H), 7.53 (dd, J = 8 Hz, J = 2Hz, 2H), 7.42 (d, J = 8 Hz, 1H), 7.25-7.18 (m, 2H), 4.35-4.28 (m, 1H), 3.63-3.48 (m, 2H), 2.81-2.69 (m, 2H), 1.94-1.90 (m, 2H), 1.72 (d, J = 7 Hz, 3H); LC/MS RT = 3.12 (M+H + : 468/470/472). N-(3-((1-(5-bromo-1 H -indol-2-yl)ethyl)amino)propyl)-cyclohexanecarboxamide (33) Cyclohexane carboxylic acid (22 mg, 0.17 mmol), EDC hydrochloric acid (65 mg, 0.34 mmol), HOBt (23 mg, 0.17 mmol) and DIPEA (0.09 mL, 0.51 mmol) were dissolved in dichloromethane (5 mL). After 5 minutes of stirring, N 1 -(1-(5-bromo-1 H -indol-2-yl)ethyl)propane-1,3-diamine (50 mg, 0.17 mmol) was added, and it was stirred overnight at room temperature. The reaction mixture was diluted with ethyl acetate, and it was washed with 1N hydrochloric acid, 10% sodium hydroxide and brine. The organic layer was dried over sodium sulfate. The organic layer was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-10% methanol/dichloromethane + 0.1% NH 4 OH) to give the product as colorless oil (55 mg, 80%); 1 H NMR (300 MHz) (CDCl 3 ) δ 9.28 (bs, 1H), 7.64 (d, J = 2 Hz, 1H), 7.26 (d, J = 9 Hz, 1H), 7.19 (dd, J = 8 Hz, J = 2 Hz, 1H), 6.23 (d, J = 1 Hz, 1H), 5.68 (bs, 1H), 4.00-3.96 (m, 1H), 3.54-3.47 (m, 1H), 3.27-3.20 (m, 1H), 2.62-2.54 (m, 1H), 2.49-2.41 (m, 1H), 2.04-2.00 (m, 1H), 1.79-1.57 (m, 6H), 1.47 (d, J = 7 Hz, 3H), 1.38-1.14 (m, 6H); LC/MS RT = 2.88 (M+H + : 406/408). 4-((1-(5-Bromo-1 H -indol-2-yl)ethyl)amino)-N-(3,4-dichlorophenyl)butanamide (34). To a solution of 1-(5-bromo-1 H -indol-2-yl)ethan-1-one (243 mg, 1.02 mmol) in ethanol (10 mL), 4-amino- N -(3,4-dichlorophenyl)butanamide (253 mg, 1.02 mmol) and sodium cyanoborohydride (320 mg, 5.10 mmol) were added. The reaction mixture was stirred for 60 o C for 24 hours. The mixture was diluted with ethyl acetate, and the organic layer was washed with 10% sodium hydroxide, saturated ammonium chloride, and brine. The organic layer was then dried over sodium sulfate, and it was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (10% methanol/dichloromethane + 1% NH 4 OH) to give product as colorless oil (73 mg, 15%); 1 H NMR (300 MHz, DMSO-d 6 ) d 11.30 (bs, 1H), 10.25 (bs, 1H), 8.00-7.99 (m, 1H), 7.72 (s, 1H), 7.60-7.55 (m, 1H), 7.49-7.44 (m, 1H), 7.38-7.36 (m, 1H), 7.23-7.21 (m, 1H), 6.48 (s, 1H), 4.33 (m, 1H), 2.76-2.60 (m, 2H), 2.42 (m, 2H), 1.84 (m, 2H), 1.57 (m, 3H). tert -Butyl (4-((3,4-dichlorophenyl)amino)-4-oxobutyl)carbamate To a solution of 4-(( tert -butoxy carbonyl)amino)butanoic acid (1.0 g, 4.9 mmol) in dichloromethane (50 mL), EDC hydrochloric acid (1.98 g, 10.3 mmol), HOBt (665 mg, 4.9 mmol) and DIPEA (1.80 mL, 10.33 mmol) were added. After the mixture stirred for 5 minutes, 3,4-dichloroaniline (797 mg, 4.92 mmol) was added. The mixture was then stirred for overnight at room temperature. The mixture was then diluted with dichloromethane, and the organic layer was washed with 1N hydrochloric acid, 10% sodium hydroxide, saturated ammonium chloride and brine. The organic layer was then dried over sodium sulfate, and it was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-30% ethyl acetate/hexane) to give product as a white solid (757 mg, 44%); 1 H NMR (300 MHz, CDCl 3 ) d 9.47 (bs, 1H), 7.92 (s, 1H), 7.48 (d, J = 9 Hz, 1H), 7.37 (d, J = 9 Hz, 1H), 4.87 (m, 1H), 3.27-3.24 (m, 2H), 2.41-2.37 (m, 2H), 1.89-1.87 (m, 2H), 1.49 (s, 9H). 4-Amino- N -(3,4-dichlorophenyl)butanamide To a solution of tert -butyl (4-((3,4-dichlorophenyl)amino)-4-oxobutyl)carbamate (500 mg, 1.44 mmol) in methanol (0.36 mL), 4N hydrochloric acid (3.60 mL, 14.40 mmol) was added. The reaction mixture was stirred for 2 hours at room temperature. The mixture was diluted with ethyl acetate, and the organic layer was washed with 10% sodium hydroxide saturated ammonium chloride and brine. The organic layer was then dried over sodium sulfate, and it was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-100% ethyl acetate/hexane followed by 10% methanol/dichloromethane + 1% NH 4 OH) to give product as colorless oil (253 mg, 71%); 1 H NMR (300 MHz, CDCl 3 ) d 9.85 (bs, 1H),7.78 (d, J = 2Hz, 1H), 7.38-7.32 (m, 2H), 2.90-2.86 (m, 2H), 2.54-2.49 (m, 2H), 1.80-1.84 (m, 2H), 1.64 (bs, 2H). N-(1-(5-Bromo-1 H -indol-2-yl)ethyl)-4-(3,4-dichlorophenoxy)butan-1-amine (35). To a solution of 1-(5-bromo-1 H -indol-2-yl)ethan-1-one (190 mg, 0.80 mmol) in ethanol (10 mL), 4-(3,4-dichlorophenoxy)butan-1-amine (187 mg, 0.80 mmol) and sodium cyanoborohydride (251 mg, 4.00 mmol) were added. The reaction mixture was stirred for 60 o C for 24 hours. The mixture was diluted with ethyl acetate, and the organic layer was washed with 10% sodium hydroxide, saturated ammonium chloride and brine. The organic layer was then dried over sodium sulfate, and it was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (10% methanol/dichloromethane + 1% NH 4 OH) to give product as colorless oil (47 mg, 13%); 1 H NMR (300 MHz, CDCl 3 ) d 9.11 (bs, 1H), 7.71 (s, 1H), 7.31-7.28 (m, 3H), 6.91 (s, 1H), 6.62 (d, J = 9 Hz, 1H), 6.44 (s, 1H), 4.11-4.39 (m, 1H), 3.87 (m, 2H), 2.78 (m, 2H), 1.81 (m, 4H), 1.72 (d, J = 7 Hz, 3H). 2-(4-(3,4-Dichlorophenoxy)butyl)isoindoline-1,3-dione To a solution of 2-(4-bromobutyl)isoindoline-1,3-dione (500 mg, 1.77 mmol) in DMF (5 mL), 3,4-dichlorophenol (289 mg, 1.77 mmol) and potassium carbonate (245 mg, 1.77 mmol) were added. The reaction mixture was stirred for 5 hours at room temperature. The mixture was diluted with ethyl acetate, the organic layer was washed with water and brine. The organic layer was then dried over sodium sulfate, and it was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-30% ethyl acetate/hexane) to give product as colorless oil (545 mg, 84%); 1 H NMR (300 MHz, CDCl 3 ) d 7.89-7.86 (m, 2H), 7.76-7.73 (m, 2H), 7.29 (d, J = 9 Hz, 1H), 6.98 (d, J = 3 Hz, 1H), 6.75 (dd, J = 9 Hz, J = 3 Hz, 1H), 4.00-3.96 (m, 2H), 3.81-3.77 (m, 2H), 1.89-1.87 (m, 4H). 4-(3,4-Dichlorophenoxy)butan-1-amine To a solution of 2-(4-(3,4-dichlorophenoxy)butyl)isoindoline-1,3-dione (545 mg, 1.50 mmol) in methanol (15 mL), hydrazine monohydrate (0.15 mL, 3.00 mmol) was added at room temperature. The reaction mixture was stirred for 2 hours at 60 o C. The white suspension was formed, and the suspension was filtered. The suspension was concentrated under reduced pressure, and the residue was diluted with ethyl acetate. The organic layer was washed with 10% sodium hydroxide and brine. The organic layer was then dried over sodium sulfate, and it was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-100% ethyl acetate/hexane followed by 10% methanol/dichloromethane + 1% NH 4 OH) to give product as colorless oil (187 mg, 53%); 1 H NMR (300 MHz, CDCl 3 ) d 7.31 (d, J = 9 Hz, 1H), 6.99 (d, J = 3 Hz, 1H), 6.75 (dd, J = 9 Hz, J = 3 Hz, 1H), 3.97-3.92 (m, 2H), 2.80-2.75 (m, 2H), 1.85-1.80 (m, 2H), 1.66-1.58 (m, 2H), 1.23 (bs, 2H). Intrinsic MIC Assays MIC assays were conducted in accordance with Clinical and Laboratory Standards Institute (CLSI) guidelines for broth microdilution. A 96-well plate containing cation-adjusted Mueller-Hinton (CAMH) broth with 2-fold serial dilution of compounds was inoculated with log-phase bacteria at 5x10 5 CFU/mL. The final volume in each well was 100 µL. Each compound was tested in duplicate. The microtiter plates were incubated in an aerobic environment for 18 hours at 37 °C. Then the bacterial growth was tested by reading the plate with a VersaMax plate reader (Molecular Devices, Inc.) at 600 nm. The MIC was defined as the lowest compound concentration that inhibited 90% of bacterial growth. The intrinsic MIC of the experimental EPIs was tested with the method described above. The 2-fold serial dilution begins with 100 µg/mL of tested compound in the first column of the 96-well plates. The following Gram-negative bacterial strains were included in these assays: Escherichia coli ATCC 25922 Klebsiella pneumoniae ATCC 13883 and ATCC 10031 Pseudomonas aeruginosa PAO1 Acinetobacter baumannii ATCC 19606 MIC Assays in the Presence of a Bacterial Efflux Inhibitor The EPI assay for the purposes of these studies represents a MIC assay in which the MIC of the antibiotic against the bacteria is tested in the presence of an experimental efflux pump inhibitor (EPI), N-(((2S,4R)-4-(aminomethyl)pyrrolidin-2-yl)methyl)-6-(4-fluorophenyl)-1H-indole-2-carboxamide. The highest concentration of the EPI present in the assay typically is ½ of the intrinsic MIC of the compound. If the intrinsic MIC of the EPI is greater than 100 µg/mL, the EPI assay was tested with 50 µg/mL. Using serial dilutions of the EPI, its enhancement of antibiotic activity was then evaluated. The relative EPI activity was decided by comparing the MIC of the antibiotic in the presence of the EPI compound with the intrinsic MIC of the antibiotic alone. For comparative purposes, we used this EPI at concentration of 12.5 µg/mL against varying concentration of our test compounds. ATPase Assays with E. coli MreB (EcMreB) The cloning and expression of E. coli MreB as well as all ATPase assays were conducted as described previously (23). Declarations Notes The authors declare the following competing financial interest(s): Dr. LaVoie and Dr. Pilch are co-founders of TAXIS Pharmaceuticals and Dr. Parhi is a shareholder and therefore have a financial interest in the company. Acknowledgement This research was supported in part by Research Agreement between Rutgers University and TAXIS Pharmaceuticals, Inc. References Figge, RM, Divakaruni, AV, Gober, JW. MreB, the cell shape-determining bacterial actin homologue, co-ordinates cell wall morphogenesis in Caulobacter crescentus . Mol. Microbiol. 2004; 51 (5): 1321-1332. Strahl, H, Bürmann, F, Hamoen. LW. The actin homologue MreB organizes the bacterial cell membrane. Nat. Commun. 2013; 5 (3442): 1-11. Wang, H, Xie, L, Luo, H, Xie, J. Bacterial cytoskeleton and implications for new antibiotic targets. J. Drug Target. 2016;2 4 (5): 392-398. Busiek, K, Margolin, W. Bacterial Actin and Tubulin Homologs in Cell Growth and Division. Curr. Biol . 2015; 25 (6); R243-R254. Foss, MH, Eun, Y-J, Weibel, DB. Chemical-biological studies of subcellular organization in bacteria. Biochemistry 2011; 50 :7719-7734. White, CL, Gober, JW. MreB: pilot or passenger of cell wall synthesis? Trends Microbiol . 2012; 20 (2), 74-79. Fenton, AK, Gerdes, K. Direct Interaction of FtsZ and MreB is required for septum synthesis and cell division in Escherichia coli . EMBO J. 2013; 32 (13): 1953-1965. Vollmer, W. The prokaryotic cytoskeleton: a putative target for inhibitors and antibiotics? Appl. Microbiol. Biotechnol. 2006;73: 37-47. Noguchi, N, Yanagimoto, K, Nakaminami, H, Wakabayashi, M, Iwai, N, Wachi, M, Sasatsu, M. Anti-infectious effect of S-benzylisothiourea compound A22, which inhibits the actin-like protein, MreB, in Shigella flexneri . Biol. Pharm. Bull. 2008; 31 (7): 1327-1332. Barker, CA, Allison, SE, Zlitni, S, Nguyen, ND, Das, R, Melacini, G, Capretta, AA, Brown, ED. Degradation of MAC13243 and studies of the interaction of resulting thiourea compounds with the lipoprotein targeting chaperone LolA. Bioorg. Med. Chem. Lett. 2013; 23: 2426-2431. Bonez, PC, Ramos, AP, Nascimento, K, Copetti, PM, Souza, ME, Rossi, GG, Agertt, VA, Sagrillo, MR, Santos, RCV, Campos, MMA. Antibacterial, cyto and genotoxic activities of A22 compound ((S-3, 4-dichlorobenzyl) isothiourea hydrochloride). Microb. Pathog. 2016;99: 14-18. Iwai, N, Fujii, T, Nagura, H, Wachi, M, Kitazume, T. Structure-activity relationship study of the bacterial actin-like protein MreB inhibitors: Effects of substitution of benzyl group in S-benzylisothiourea. Biosci. Biotechnol. Biochem. 2007; 71 (1): 246-248. Iwai, N, Ebata, T, Nagura, H, Kitazume, T, Nagai, K, Wachi, M. Structure-activity relationship of S-benzylisothiourea derivatives to induce spherical cells in Escherichia coli. Biosci. Biotechnol. Biochem . , 2004; 68 (11): 2265-2269. Nicholson, A, Perry, JD, James, AL, Stanforth, SP, Carnell, S, Wilkinson, K, Kahn, CMA, De Soyza, A, Gould, FK. In vitro activity of S-(3,4-dichlorobenzyl)isothiourea hydrochloride and novel structurally related compounds against multidrug-resistant bacteria, including Pseudomonas aeruginosa and Burkholderia cepacia complex. Int. J. Antimicrob. Agents 2012; 39 : 27-32. Bean, GJ, Flickinger, ST, Westler, WM, McCully, ME, Sept, D, Weibel, DB, Amann, KJ. A22 disrupts the bacterial actin cytoskeleton by directly binding and inducing a low-affinity state in MreB. Biochemistry , 2009; 48 (22): 4852-4857. Takacs, CN, Poggio, S, Charbon, G, Pucheault, M, Vollmer, W, Wagner, CJ. MreB drives de novo rod morphogenesis in Caulobacter crescentus via remodeling of the cell wall. J. Bacteriol. 2010: 1671-1684. Gerdes, K, Møller-Jensen, J, Ebersbach, G, Kruse, T, Nordström, K. Bacterial mitotic machineries. Cell 2004;116(3): 359-366. Kruse, T, Gerdes, K. Bacterial DNA segregation by the actin-like MreB protein. Trends Cell Biol. 2005;15(7): 343-345. Kruse, T, Blagoev, B, Lobner-Olesen, A, Wachi, M, Sasaki, K, Iwai, N, Mann, M, Gerdes, K. Actin homolog MreB and RNA polymerase interact and are both required for chromosome segregation in Escherichia coli . Genes Dev. 2006;20: 113-124. Kruse, T, Moller-Jensen, J, Lobner-Olesen, A, Gerdes, K. Dysfunctional MreB inhibits chromosome segregation in Escherichia coli . EMBO J. 2003; 22 (19): 5283-5292. Gitai, Z, Dye, NA, Reisenauer, A, Wachi, M, Shapiro, L. MreB actin-mediated segregation of a specific region of a bacterial chromosome. Cell 2005;120: 329-341. Robertson, GT, Doyle, TB, Du, Q, Duncan, L, Mdluli, KE, Lynch, AS. A novel indole compound that inhibits Pseudomonas aeruginosa growth by targeting MreB is a substrate for MexAB-OprM. J. Bacteriol . 2007;189(19): 6870-6881. Bryan, EJ, Sagong HY, Parhi AK, Grier MC, Roberge JC, LaVoie EJ, Pilch, DS. TXH11106: A third-generation MreB inhibitor with enhanced activity against a broad range of Gram-negative bacterial pathogens. Antibiotics 2022;11: 693-709. LaVoie, E, Parhi, A, Sagong, HY, inventors; Therapeutic compounds and methods to treat infection.US 0155507 A1, 2020 May 21. Tables Tables 1 to 3 are available in the Supplementary Files section scheme Scheme 1 to 9 are available in the Supplementary Files section. Supplementary Files MreBInhibitorsSupplementaryMaterial.docx scheme1.png Scheme 1. Reagents and conditions: a) LiHMDA, ethyl formate, 0 °C (37%); b) NaNO 2 , conc. HCl, 0 °C; c) MeOH, NaOAc, 0 °C (79%); d) conc. HCl, AcOH, 130 °C (23% for 8a), (9a and 10a (8%); e) Boc anhydride, DMAP, THF, rt, (49% 9b and 20% 10b); f) TFA in DCM (1:5), rt, (90% for 9a and 95% for 10a); g) NaBH 3 CN, NH 4 OAc, EtOH, 60 °C, 34% for 8, 37% for 9, and 67% for 10. scheme2.png Scheme 2. Reagents and conditions: a) LiHMDA, ethyl formate, 0 °C (37%); b) NaNO 2 , conc. HCl, 0 °C; c) MeOH, NaOAc, 0 °C (28%); d) conc. HCl, AcOH, 130 °C (8% for 11a), (12a and 13a (10%); e) Boc anhydride, DMAP, THF, rt, (17% 12b and 23% 13b); f) TFA in DCM (1:5), rt, (31% for 12a and 37% for 13a); g) NaBH 3 CN, NH 4 OAc, EtOH, 60 °C, 17% for 11, 41% for 12, and 62% for 13. scheme3.png Scheme 3. Reagents and conditions: a) CH 3 ONHCH 3 ·HCl, EDC·HCl; b) CH 3 Li, THF/EtO; c) NH 4 OAc, NaBH 3 CN, EtOH. scheme4.png Scheme 4. Reagents and conditions: a) i (CH 3 NH, THF, AcOH cat., 60 °C, ii NaBH 3 CN, 60 °C, (100% for 22); i (CH 3 ) 2 N, THF, AcOH cat., 60 °C, ii NaBH 3 CN, 60 °C (14% for 23), c) MeI, K 2 CO 3 , DMF (86% for 24a); d) NaBH 3 CN, NH 4 OAc, cat. AcOH, EtOH, 60 °C (63% for 24). scheme5.png Scheme 5. Reagents and conditions: a) MeOH, conc. HCl, reflux, (75%); b) LiBH 4 , THF, rt, (83%); c) DMP, DCM, rt, (56%); d) NH 4 OAc, EtOH, NaBH 3 CN, rt, (46%). scheme6.png Scheme 6. Reagents and conditions: a) N-Boc Propylenediamine, NaBH 3 CN, cat. AcOH, EtOH (78% for 26a, 100% for 27a and 99% for 28a); b) 4N HCl in dioxane/MeOH (43% for 26, 100% for 27 and 54% for 28). scheme7.png Scheme 7. Reagents and conditions: a) EDC·HCl, HOBt, DIPEA, rt, (31% for 29, 13% for 30, 22% for 31, 18% for 32 and 80% for 33). scheme8.png Scheme 8. Reagents and conditions: a) MeOH, 4N HCl in dioxane (71%); b) NaBH 3 CN, EtOH, 60 °C, (15%). scheme9.png Scheme 9. Reagents and conditions: a) Hydrazine, MeOH, 60 °C (53%); b) NaBH 3 CN, EtOH, 60 °C, (13%). GraphicalAbstract.png Graphical Abstract Tables.docx Cite Share Download PDF Status: Under Review Version 1 posted Editorial decision: Minor Revisions 08 Aug, 2022 Reviewers agreed at journal 28 Jul, 2022 Reviewers invited by journal 12 Jul, 2022 Editor assigned by journal 07 Jul, 2022 First submitted to journal 04 Jul, 2022 You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-1824695","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":120580519,"identity":"e1c7f251-827b-4e9f-aa67-5b3f3d1a7a12","order_by":0,"name":"Hye Yeon Sagong","email":"","orcid":"","institution":"TAXIS Pharmaceuticals","correspondingAuthor":false,"prefix":"","firstName":"Hye","middleName":"Yeon","lastName":"Sagong","suffix":""},{"id":120580520,"identity":"fbe01a67-57db-4e71-b4ef-047f0a80ef84","order_by":1,"name":"Jesus D Rosado-Lugo","email":"","orcid":"","institution":"TAXIS Pharmaceuticals, Inc.","correspondingAuthor":false,"prefix":"","firstName":"Jesus","middleName":"D","lastName":"Rosado-Lugo","suffix":""},{"id":120580521,"identity":"fe94dc9d-3f09-4b02-8746-b8543de9f1a5","order_by":2,"name":"Eric J Bryan","email":"","orcid":"","institution":"Rutgers Robert Wood Johnson Medical School","correspondingAuthor":false,"prefix":"","firstName":"Eric","middleName":"J","lastName":"Bryan","suffix":""},{"id":120580522,"identity":"d4f25578-bdc4-4ea0-b71b-e9a765f72976","order_by":3,"name":"Edgar Ferrer-Gonzalez","email":"","orcid":"","institution":"UMDNJ RWJMS: Rutgers Robert Wood Johnson Medical School","correspondingAuthor":false,"prefix":"","firstName":"Edgar","middleName":"","lastName":"Ferrer-Gonzalez","suffix":""},{"id":120580523,"identity":"fa4d1fcc-66e6-4377-a68f-914630cb0825","order_by":4,"name":"Yiling Wang","email":"","orcid":"","institution":"Rutgers University Ernest Mario School of Pharmacy","correspondingAuthor":false,"prefix":"","firstName":"Yiling","middleName":"","lastName":"Wang","suffix":""},{"id":120580524,"identity":"ed1c5505-b195-405d-b97c-e1bf34162aec","order_by":5,"name":"Yanlu Cao","email":"","orcid":"","institution":"TAXIS Pharmaceuticals, Inc","correspondingAuthor":false,"prefix":"","firstName":"Yanlu","middleName":"","lastName":"Cao","suffix":""},{"id":120580525,"identity":"916e5ca1-d680-4aa4-ab3c-8310b32a3d5d","order_by":6,"name":"Ajit K Parhi","email":"","orcid":"","institution":"TAXIS Pharmaceuticals, Inc","correspondingAuthor":false,"prefix":"","firstName":"Ajit","middleName":"K","lastName":"Parhi","suffix":""},{"id":120580526,"identity":"c3c69d40-fb25-445c-9bb3-ec9f3d544782","order_by":7,"name":"Daniel S Pilch","email":"","orcid":"","institution":"UMDNJ RWJMS: Rutgers Robert Wood Johnson Medical School","correspondingAuthor":false,"prefix":"","firstName":"Daniel","middleName":"S","lastName":"Pilch","suffix":""},{"id":120580527,"identity":"b37203fa-976d-4520-aad7-37b026e4be44","order_by":8,"name":"Edmond Joseph LaVoie","email":"data:image/png;base64,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","orcid":"https://orcid.org/0000-0001-9244-1249","institution":"Rutgers University New Brunswick","correspondingAuthor":true,"prefix":"","firstName":"Edmond","middleName":"Joseph","lastName":"LaVoie","suffix":""}],"badges":[],"createdAt":"2022-07-04 17:11:33","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-1824695/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-1824695/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":24095194,"identity":"03c325cd-1b6e-418f-bf93-d8e5f6956122","added_by":"auto","created_at":"2022-07-20 14:57:28","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":142950,"visible":true,"origin":"","legend":"\u003cp\u003eImpact of increasing concentrations of selected analogs on the velocity (v) of the ATPase reaction of \u003cem\u003eE. coli\u003c/em\u003e MreB (EcMreB). Each experimental data point represents an average of two replicates, with the indicated error bars reflecting the standard deviation from the mean. The solid curves represent nonlinear least squares fits of the experimental data points as previously described (23).\u0026nbsp;\u003c/p\u003e\u003cp\u003e\u003cbr\u003e\u003c/p\u003e","description":"","filename":"11.png","url":"https://assets-eu.researchsquare.com/files/rs-1824695/v1/99203d849b5f4740b1c248d1.png"},{"id":24096870,"identity":"4f2cf4bd-c41b-40ca-a599-58d090435f71","added_by":"auto","created_at":"2022-07-20 15:17:33","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":1445567,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-1824695/v1/6de8d2e4-1f7b-4821-80f9-3a593c507b1a.pdf"},{"id":24096865,"identity":"5f60b454-9c21-492d-aeff-d840553d75a4","added_by":"auto","created_at":"2022-07-20 15:17:29","extension":"docx","order_by":1,"title":"","display":"","copyAsset":false,"role":"supplement","size":21009,"visible":true,"origin":"","legend":"","description":"","filename":"MreBInhibitorsSupplementaryMaterial.docx","url":"https://assets-eu.researchsquare.com/files/rs-1824695/v1/581c099b850f8ce653364a39.docx"},{"id":24095195,"identity":"7ac5f6b1-c85d-46dd-b328-e5dd15214238","added_by":"auto","created_at":"2022-07-20 14:57:28","extension":"png","order_by":2,"title":"","display":"","copyAsset":false,"role":"supplement","size":50974,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eScheme 1.\u0026nbsp;\u003c/strong\u003eReagents and conditions: a) LiHMDA, ethyl formate, 0 °C (37%); b) NaNO\u003csub\u003e2\u003c/sub\u003e, conc. HCl, 0 °C; c) MeOH, NaOAc, 0 °C (79%); d) conc. HCl, AcOH, 130 °C (23% for \u003cstrong\u003e8a\u003c/strong\u003e), (\u003cstrong\u003e9a\u003c/strong\u003e and \u003cstrong\u003e10a\u003c/strong\u003e (8%); e) Boc anhydride, DMAP, THF, rt, (49% \u003cstrong\u003e9b\u003c/strong\u003e and 20% \u003cstrong\u003e10b\u003c/strong\u003e); f) TFA in DCM (1:5), rt, (90% for \u003cstrong\u003e9a\u003c/strong\u003e and 95% for \u003cstrong\u003e10a\u003c/strong\u003e); g) NaBH\u003csub\u003e3\u003c/sub\u003eCN, NH\u003csub\u003e4\u003c/sub\u003eOAc, EtOH, 60 °C, 34% for \u003cstrong\u003e8\u003c/strong\u003e, 37% for \u003cstrong\u003e9\u003c/strong\u003e, and 67% for \u003cstrong\u003e10\u003c/strong\u003e.\u003c/p\u003e\u003cp\u003e\u003cbr\u003e\u003c/p\u003e","description":"","filename":"scheme1.png","url":"https://assets-eu.researchsquare.com/files/rs-1824695/v1/2346e292315d1dda690dd5e2.png"},{"id":24096230,"identity":"2ffbc28d-633e-48c1-bda4-d45ec5e9dcb8","added_by":"auto","created_at":"2022-07-20 15:12:28","extension":"png","order_by":3,"title":"","display":"","copyAsset":false,"role":"supplement","size":67797,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eScheme 2.\u0026nbsp;\u003c/strong\u003eReagents and conditions: a) LiHMDA, ethyl formate, 0 °C (37%); b) NaNO\u003csub\u003e2\u003c/sub\u003e, conc. HCl, 0 °C; c) MeOH, NaOAc, 0 °C (28%); d) conc. HCl, AcOH, 130 °C (8% for \u003cstrong\u003e11a\u003c/strong\u003e), (\u003cstrong\u003e12a\u003c/strong\u003e and \u003cstrong\u003e13a\u003c/strong\u003e (10%); e) Boc anhydride, DMAP, THF, rt, (17% \u003cstrong\u003e12b\u003c/strong\u003e and 23% \u003cstrong\u003e13b\u003c/strong\u003e); f) TFA in DCM (1:5), rt, (31% for \u003cstrong\u003e12a\u003c/strong\u003e and 37% for \u003cstrong\u003e13a\u003c/strong\u003e); g) NaBH\u003csub\u003e3\u003c/sub\u003eCN, NH\u003csub\u003e4\u003c/sub\u003eOAc, EtOH, 60 °C, 17% for \u003cstrong\u003e11\u003c/strong\u003e, 41% for \u003cstrong\u003e12\u003c/strong\u003e, and 62% for \u003cstrong\u003e13\u003c/strong\u003e.\u003c/p\u003e","description":"","filename":"scheme2.png","url":"https://assets-eu.researchsquare.com/files/rs-1824695/v1/62b7d6f767c276d6d36562a4.png"},{"id":24096232,"identity":"9b024dfc-7b7d-4f69-b01f-0f4151c2f608","added_by":"auto","created_at":"2022-07-20 15:12:29","extension":"png","order_by":4,"title":"","display":"","copyAsset":false,"role":"supplement","size":34319,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eScheme 3.\u0026nbsp;\u003c/strong\u003eReagents and conditions: a) CH\u003csub\u003e3\u003c/sub\u003eONHCH\u003csub\u003e3\u003c/sub\u003e·HCl, EDC·HCl; b) CH\u003csub\u003e3\u003c/sub\u003eLi, THF/EtO; c) NH\u003csub\u003e4\u003c/sub\u003eOAc, NaBH\u003csub\u003e3\u003c/sub\u003eCN, EtOH.\u003c/p\u003e","description":"","filename":"scheme3.png","url":"https://assets-eu.researchsquare.com/files/rs-1824695/v1/1bd444e942209dde6a1cb5b9.png"},{"id":24095763,"identity":"da08f3cd-3c16-4e0c-a4f6-cd2fdd1d1718","added_by":"auto","created_at":"2022-07-20 15:02:28","extension":"png","order_by":5,"title":"","display":"","copyAsset":false,"role":"supplement","size":42045,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eScheme 4.\u0026nbsp;\u003c/strong\u003eReagents and conditions: a) i (CH\u003csub\u003e3\u003c/sub\u003eNH, THF, AcOH cat., 60 °C, ii NaBH\u003csub\u003e3\u003c/sub\u003eCN, 60 °C, (100% for \u003cstrong\u003e22\u003c/strong\u003e); i (CH\u003csub\u003e3\u003c/sub\u003e)\u003csub\u003e2\u003c/sub\u003eN, THF, AcOH cat., 60 °C, ii NaBH\u003csub\u003e3\u003c/sub\u003eCN, 60 °C (14% for \u003cstrong\u003e23\u003c/strong\u003e), c) MeI, K\u003csub\u003e2\u003c/sub\u003eCO\u003csub\u003e3\u003c/sub\u003e, DMF (86% for \u003cstrong\u003e24a\u003c/strong\u003e); d) NaBH\u003csub\u003e3\u003c/sub\u003eCN, NH\u003csub\u003e4\u003c/sub\u003eOAc, cat. AcOH, EtOH, 60 °C (63% for \u003cstrong\u003e24\u003c/strong\u003e).\u003c/p\u003e","description":"","filename":"scheme4.png","url":"https://assets-eu.researchsquare.com/files/rs-1824695/v1/3f93898b65371b6d881b4064.png"},{"id":24096068,"identity":"ae51be3a-d518-47fd-8708-9f8aecbc2dac","added_by":"auto","created_at":"2022-07-20 15:07:28","extension":"png","order_by":6,"title":"","display":"","copyAsset":false,"role":"supplement","size":18554,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eScheme 5.\u0026nbsp;\u003c/strong\u003eReagents and conditions: a) MeOH, conc. HCl, reflux, (75%); b) LiBH\u003csub\u003e4\u003c/sub\u003e, THF, rt, (83%); c) DMP, DCM, rt, (56%); d) NH\u003csub\u003e4\u003c/sub\u003eOAc, EtOH, NaBH\u003csub\u003e3\u003c/sub\u003eCN, rt, (46%).\u003c/p\u003e","description":"","filename":"scheme5.png","url":"https://assets-eu.researchsquare.com/files/rs-1824695/v1/971361dc8e1ab8960a157a31.png"},{"id":24095767,"identity":"54a99e2d-a975-49b0-acf8-c9151b8d03eb","added_by":"auto","created_at":"2022-07-20 15:02:29","extension":"png","order_by":7,"title":"","display":"","copyAsset":false,"role":"supplement","size":56539,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eScheme 6.\u0026nbsp;\u003c/strong\u003eReagents and conditions: a) N-Boc Propylenediamine, NaBH\u003csub\u003e3\u003c/sub\u003eCN, cat. AcOH, EtOH (78% for \u003cstrong\u003e26a\u003c/strong\u003e, 100% for \u003cstrong\u003e27a\u003c/strong\u003e and 99% for \u003cstrong\u003e28a\u003c/strong\u003e); b) 4N HCl in dioxane/MeOH (43% for \u003cstrong\u003e26\u003c/strong\u003e, 100% for \u003cstrong\u003e27\u003c/strong\u003e and 54% for \u003cstrong\u003e28\u003c/strong\u003e).\u003c/p\u003e","description":"","filename":"scheme6.png","url":"https://assets-eu.researchsquare.com/files/rs-1824695/v1/24dc18c69daefa5e76faf350.png"},{"id":24095205,"identity":"8ee51677-16ce-43ad-b7c1-1f0f96456783","added_by":"auto","created_at":"2022-07-20 14:57:29","extension":"png","order_by":8,"title":"","display":"","copyAsset":false,"role":"supplement","size":51105,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eScheme 7.\u0026nbsp;\u003c/strong\u003eReagents and conditions: a) EDC·HCl, HOBt, DIPEA, rt, (31% for \u003cstrong\u003e29\u003c/strong\u003e, 13% for \u003cstrong\u003e30\u003c/strong\u003e, 22% for \u003cstrong\u003e31\u003c/strong\u003e, 18% for \u003cstrong\u003e32\u003c/strong\u003e and 80% for \u003cstrong\u003e33\u003c/strong\u003e).\u003c/p\u003e","description":"","filename":"scheme7.png","url":"https://assets-eu.researchsquare.com/files/rs-1824695/v1/7ff7069038170b5d00d0e598.png"},{"id":24095201,"identity":"7c667918-70d9-40e7-8bad-56333ba53017","added_by":"auto","created_at":"2022-07-20 14:57:28","extension":"png","order_by":9,"title":"","display":"","copyAsset":false,"role":"supplement","size":26088,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eScheme 8. \u003c/strong\u003eReagents and conditions: a) MeOH, 4N HCl in dioxane (71%); b) NaBH\u003csub\u003e3\u003c/sub\u003eCN, EtOH, 60 °C, (15%).\u0026nbsp;\u003c/p\u003e","description":"","filename":"scheme8.png","url":"https://assets-eu.researchsquare.com/files/rs-1824695/v1/d9f513bb520ee60b2e877241.png"},{"id":24095202,"identity":"eadd591d-0b8e-4bbf-8f16-b73ac9032c27","added_by":"auto","created_at":"2022-07-20 14:57:29","extension":"png","order_by":10,"title":"","display":"","copyAsset":false,"role":"supplement","size":27222,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eScheme 9. \u003c/strong\u003eReagents and conditions: a) Hydrazine, MeOH, 60 °C (53%); b) NaBH\u003csub\u003e3\u003c/sub\u003eCN, EtOH, 60 °C, (13%).\u003c/p\u003e","description":"","filename":"scheme9.png","url":"https://assets-eu.researchsquare.com/files/rs-1824695/v1/acd44a2f7a6a65995efc482e.png"},{"id":24095769,"identity":"49383937-790b-48ca-b232-f341a964701f","added_by":"auto","created_at":"2022-07-20 15:02:29","extension":"png","order_by":11,"title":"","display":"","copyAsset":false,"role":"supplement","size":75060,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eGraphical Abstract\u003c/strong\u003e\u003c/p\u003e","description":"","filename":"GraphicalAbstract.png","url":"https://assets-eu.researchsquare.com/files/rs-1824695/v1/aed8bf4f64a6e7f4180e7994.png"},{"id":24095204,"identity":"c8839126-eab6-4d43-aa93-536546495e8f","added_by":"auto","created_at":"2022-07-20 14:57:29","extension":"docx","order_by":12,"title":"","display":"","copyAsset":false,"role":"supplement","size":99604,"visible":true,"origin":"","legend":"","description":"","filename":"Tables.docx","url":"https://assets-eu.researchsquare.com/files/rs-1824695/v1/23591e7821383c4736bc812a.docx"}],"financialInterests":"","formattedTitle":"Novel MreB Inhibitors with Antibacterial Activity Against Gram (-) Bacteria","fulltext":[{"header":"Introduction","content":"\u003cp\u003eMreB is the prokaryotic homolog of actin in eukaryotes. \u0026nbsp;The function of MreB within bacteria is associated with its ability to form long filaments, a process similar to actin in eukaryotes [1-5]. \u0026nbsp;These filaments are critical for to the retention of the structure of rod-shaped bacteria, which is dominant among many Gram (-) bacteria [1, 2, 4-6]. \u0026nbsp;MreB is also essential for bacterial cell division [7]. \u0026nbsp; This structural protein is an attractive target for antibacterial drug discovery as it is highly conserved and present in almost all rod-shaped bacteria [8]. \u0026nbsp;Acquired resistance to such novel antibiotics would also be unlikely, as MreB modulation is not associated with the modes of action of antibiotics in clinical use.\u003c/p\u003e\n\u003cp\u003ePolymerization of MreB requires ATP. \u0026nbsp;Several benzyl thioureas such as A22 and MP265 (\u003cstrong\u003e1, 2\u003c/strong\u003e) as well as CBR-4830 and several of its analogs \u003cstrong\u003e(3-6\u003c/strong\u003e) have been identified as inhibitors of MreB function (Figure 1) [1, 2, 5, 8-15]. \u0026nbsp;Current theories on the mode of action of these MreB inhibitors suggest that they act by noncompetitively binding to this structural protein, thereby affecting the ATP binding pocket. \u0026nbsp;MreB loss of function is both lethal and pleiotropic [7, 16]. \u0026nbsp;Multiple cellular processes are disrupted including cell shape determination, polar protein localization, and cell division [16-18]. \u0026nbsp;MreB also has a direct role in the segregation of a specific region of the chromosome [18-21]. \u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cimg src=\"data:image/png;base64,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\"\u003e\u003c/p\u003e\n\u003cp\u003eThe MreB inhibitor, A22, is lethal at relatively low doses when administered iv to either rats or mice. \u0026nbsp;Lynch et al reported in 2007 the identification and characterization of CBR-4830 (Figure 1) as a novel antimicrobial agent with activity against efflux-compromised \u003cem\u003eP.\u003c/em\u003e \u003cem\u003eaeruginosa\u003c/em\u003e [22]. \u0026nbsp;Combined genetic, biochemical and cell morphology studies were performed that demonstrated that the cellular target for CBR-4830 is MreB. \u0026nbsp;A limited structure-activity study was performed with analogs of CBR-4830, \u003cstrong\u003e3\u003c/strong\u003e-\u003cstrong\u003e6\u003c/strong\u003e. \u0026nbsp;However, CBR-4830 proved to be the most active analog within this series. \u0026nbsp;CBR-4830 proved not only to be difficult to formulate for iv administration because of poor solubility properties in aqueous vehicles, but it also proved to be lethal at relatively low concentration in mice. \u0026nbsp; Recently, we reported on the enhanced activity of TXH11106 (\u003cstrong\u003e7\u003c/strong\u003e) relative to CBR-4830 [23, 24]. \u0026nbsp;In the present study, we initiated studies to develop additional analogs related to CBR-4830 to further improve antibacterial potency and formulation properties relative to CBR-4830.\u0026nbsp;\u003c/p\u003e"},{"header":"Chemistry","content":"\u003cp\u003eOur initial studies were focused on examining the impact of changing the fused alicyclic ring of the tetrahydrocarbazole pharmacophore of CBR-4830 to its next higher homologue, a cycloheptane moiety fused to the indole. 2-Bromo-5,6,7,8,9,10-hexahydrocyclohepta[\u003cem\u003eb\u003c/em\u003e]indole-6-amine, \u003cb\u003e8\u003c/b\u003e, and the dichloro analogs \u003cb\u003e9\u003c/b\u003e and \u003cb\u003e10\u003c/b\u003e were synthesized as outlined in Scheme \u003cspan refid=\"Sch1\" class=\"InternalRef\"\u003e1\u003c/span\u003e. Commercially available cycloheptanone was condensed with ethyl formate to form 2-(hydroxymethylene)cycloheptan-1-one. Reaction of this intermediate with the appropriate pre-formed diazonium salts provide the hydrazineylidene intermediates, which were converted to \u003cb\u003e8a\u003c/b\u003e and a mixture of \u003cb\u003e9a\u003c/b\u003e and \u003cb\u003e10a\u003c/b\u003e. The mixture of \u003cb\u003e9a\u003c/b\u003e and \u003cb\u003e10a\u003c/b\u003e was converted to their N-Boc derivatives and separated chromatographically, followed by removal of the Boc groups to provide the pure isomers. Reductive amination using ammonium acetate and NaBH\u003csub\u003e3\u003c/sub\u003eCN provided the 6-amino-5,6,7,8,9,10-hexahydro-cyclohepta[\u003cem\u003eb\u003c/em\u003e]indoles \u003cb\u003e8\u003c/b\u003e, \u003cb\u003e9\u003c/b\u003e and \u003cb\u003e10\u003c/b\u003e.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eWe also synthesized similarly substituted 1,2,3,4-tetrahydrocyclopenta[\u003cem\u003eb\u003c/em\u003e]indol-3-amines, \u003cb\u003e11\u003c/b\u003e\u0026ndash;\u003cb\u003e13\u003c/b\u003e, which represent analogs that are a one carbon lower homologue of the tetrahydrocarbazole pharmacophore of CBR-4830. The methods used for these syntheses are outlined in Scheme \u003cspan refid=\"Sch2\" class=\"InternalRef\"\u003e2\u003c/span\u003e. Cyclopentanone was condensed with ethyl formate to provide 2-(hydroxymethylene)cyclopentan-1-one. This intermediate was reacted with the appropriate diazonium salt to give the desired hydrazineylidene intermediates, which were converted under acidic condition to the 1,4-dihydrocyclopenta[\u003cem\u003eb\u003c/em\u003e]indole-3(2\u003cem\u003eH\u003c/em\u003e)-ones, \u003cb\u003e11a\u003c/b\u003e as well as a mixture of \u003cb\u003e12a\u003c/b\u003e and \u003cb\u003e13a\u003c/b\u003e. The mixture of \u003cb\u003e12a\u003c/b\u003e and \u003cb\u003e13a\u003c/b\u003e was resolved by formation of their N-Boc derivatives (\u003cb\u003e12b\u003c/b\u003e and \u003cb\u003e13b\u003c/b\u003e), chromatographic separation, followed by removal of their Boc groups.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eThe impact of removing two methylene groups and opening the tetrahydrocyclohexyl ring of CBR-4830 on relative antimicrobial activity was also evaluated. The varied 2-(1-aminoethyl)indoles \u003cb\u003e14\u003c/b\u003e\u0026ndash;\u003cb\u003e21\u003c/b\u003e were prepared as outlined in Scheme \u003cspan refid=\"Sch3\" class=\"InternalRef\"\u003e3\u003c/span\u003e.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eThe Weinreb amide derivatives \u003cb\u003e14b\u003c/b\u003e-\u003cb\u003e21b\u003c/b\u003e were formed from the various 2-carboxyindole derivatives and then converted using methyllithium to the methyl ketones \u003cb\u003e14c\u003c/b\u003e-\u003cb\u003e21c\u003c/b\u003e. Reductive amination of \u003cb\u003e14c\u003c/b\u003e-\u003cb\u003e21c\u003c/b\u003e using ammonium acetate and sodium cyanoborohydride provide \u003cb\u003e14\u003c/b\u003e\u0026ndash;\u003cb\u003e21\u003c/b\u003e.\u003c/p\u003e \u003cp\u003eThe influence of mono- or dimethylation on the amino group as well as methylation of the 1-position of 1-(5-bromo-1-methyl-1H-indol-2-yl)ethan-1-amine was examined. Compounds \u003cb\u003e22\u003c/b\u003e, \u003cb\u003e23\u003c/b\u003e, and \u003cb\u003e24\u003c/b\u003e were synthesized as outlined in Scheme \u003cspan refid=\"Sch4\" class=\"InternalRef\"\u003e4\u003c/span\u003e.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eReductive amination was performed using N-methylamine or N,N-dimethylamine, followed by reduction with sodium cyanoborohydride was used to prepare \u003cb\u003e22\u003c/b\u003e or \u003cb\u003e23\u003c/b\u003e. Methylation of \u003cb\u003e14c\u003c/b\u003e to form the 1-methylindole derivative \u003cb\u003e24a\u003c/b\u003e was accomplished using potassium carbonate in DMF and methyl iodide. Intermediate \u003cb\u003e24a\u003c/b\u003e was then converted \u003cb\u003e24\u003c/b\u003e using ammonium acetate and sodium cyanoborohydride.\u003c/p\u003e \u003cp\u003eWe also examined the impact on activity of having the primary amine of 1-(5-bromo-1-methyl-1\u003cem\u003eH\u003c/em\u003e-indol-2-yl)ethan-1-amine attached to a primary carbon as opposed to a secondary carbon. We synthesized (5-bromo-1\u003cem\u003eH\u003c/em\u003e-indol-2-yl)methanamine, \u003cb\u003e25\u003c/b\u003e, as outlined in Scheme \u003cspan refid=\"Sch5\" class=\"InternalRef\"\u003e5\u003c/span\u003e and evaluated its antibacterial properties.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eThe preparation of the methylamine derivative \u003cb\u003e25\u003c/b\u003e was performed starting from the 2-carboxy indole intermediate \u003cb\u003e14a\u003c/b\u003e. This acid was converted to its methyl ester, \u003cb\u003e25a\u003c/b\u003e, using methanol and concentrated HCl. The ester was reduced with lithium borohydride in THF to provide the hydroxymethyl intermediate, \u003cb\u003e25b\u003c/b\u003e. Using Dess-Martin periodinane in dichloromethane, this hydroxymethyl derivative was oxidized to the 2-formylindole derivative, \u003cb\u003e25c\u003c/b\u003e. Reductive amination with ammonium acetate and sodium cyanoborohydride provide the desired 2-aminomethyl indole derivative \u003cb\u003e25\u003c/b\u003e.\u003c/p\u003e \u003cp\u003eThe N-(3-aminopropyl) derivatives of \u003cb\u003e8\u003c/b\u003e, \u003cb\u003e11\u003c/b\u003e, and \u003cb\u003e14\u003c/b\u003e were also synthesized and their relative antibacterial activity compared. The synthetic methods used for their preparation are outlined in Scheme \u003cspan refid=\"Sch6\" class=\"InternalRef\"\u003e6\u003c/span\u003e.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eThe ketone intermediates \u003cb\u003e8a\u003c/b\u003e, \u003cb\u003e11a\u003c/b\u003e, and \u003cb\u003e14c\u003c/b\u003e were each treated with N-Boc propylenediamine and then subject to reduction using sodium cyanoborohydride in ethanol. Excellent yields were obtained of each of the N-Boc propylenediamine derivatives, \u003cb\u003e26a\u003c/b\u003e, \u003cb\u003e27a\u003c/b\u003e and \u003cb\u003e28a\u003c/b\u003e. Removal of the N-Boc protecting groups with 4N HCl gave \u003cb\u003e26\u003c/b\u003e\u0026ndash;\u003cb\u003e28\u003c/b\u003e.\u003c/p\u003e \u003cp\u003eA series of N-acyl derivatives of \u003cb\u003e28\u003c/b\u003e were also prepared. These included N-acylheteroaryl, N-acyl(3,4-dichlorophenyl), as well as N-acylcyclohexyl derivatives, as illustrated in Scheme \u003cspan refid=\"Sch7\" class=\"InternalRef\"\u003e7\u003c/span\u003e.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eThe synthesis of a series carboxamides derived from \u003cb\u003e28\u003c/b\u003e were prepared by condensation of various carboxylic acids using EDC, HOBt and DIPEA at room temperature. The commercially available carboxylic acids employed for the preparation of \u003cb\u003e29\u003c/b\u003e\u0026ndash;\u003cb\u003e33\u003c/b\u003e were 5-chlorofuran-2-crboxylic acid, 5-chlorothiophene-2-carboxylic acid, 4,5-dichlorothiophene-2-carboxylic acid, 3,4-dichlorobenzoic acid and cyclohexane carboxylic acid.\u003c/p\u003e \u003cp\u003eWe also prepared the revered amide analog, \u003cb\u003e34\u003c/b\u003e, that is related to \u003cb\u003e32\u003c/b\u003e, as well as the aryloxy analog \u003cb\u003e35\u003c/b\u003e, as illustrated in Scheme \u003cspan refid=\"Sch8\" class=\"InternalRef\"\u003e8\u003c/span\u003e and \u003cspan refid=\"Sch9\" class=\"InternalRef\"\u003e9\u003c/span\u003e. The reversed amide \u003cb\u003e34\u003c/b\u003e was prepared by first preparing the 4-amino-N-(3,4-dichlorophenyl)butanamide, which was then reacted with \u003cb\u003e14c\u003c/b\u003e and the resulting imine reduce using sodium cyanoborohydride in ethanol to give \u003cb\u003e34\u003c/b\u003e as outlined in Scheme \u003cspan refid=\"Sch8\" class=\"InternalRef\"\u003e8\u003c/span\u003e.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eThe N-[4-(3,4-dichlorophenoxy)butyl] derivative of \u003cb\u003e14\u003c/b\u003e was prepared from 3-(3,4-dichlorophenoxy)propylamine as illustrated in Scheme \u003cspan refid=\"Sch9\" class=\"InternalRef\"\u003e9\u003c/span\u003e. Reaction of this amine with \u003cb\u003e14c\u003c/b\u003e, followed by reduction with sodium cyanoborohydride provide \u003cb\u003e35\u003c/b\u003e.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e"},{"header":"Results","content":"\u003cp\u003eTable 1 provides summarize the intrinsic activity of antibacterial activity against \u003cem\u003eP. aeruginosa\u003c/em\u003e, \u003cem\u003eE. coli\u003c/em\u003e,\u003cem\u003e\u0026nbsp;K. pneumoniae\u003c/em\u003e and \u003cem\u003eA. baumannii\u003c/em\u003e for the cyclohepta analogs \u003cstrong\u003e8\u003c/strong\u003e-\u003cstrong\u003e10\u003c/strong\u003e, the cyclopenta analogs \u003cstrong\u003e11\u003c/strong\u003e-\u003cstrong\u003e13\u003c/strong\u003e, and the ring-opened analogs \u003cstrong\u003e14\u003c/strong\u003e-\u003cstrong\u003e21\u003c/strong\u003e, relative to CBR-4830. \u003cem\u003e\u0026nbsp;P. aeruginosa\u003c/em\u003e proved relatively resistant in the absence of a bacterial efflux pump inhibitor (EPI). \u0026nbsp;Therefore, the MICs of all of these compounds against\u003cem\u003e\u0026nbsp;P. aeruginosa\u003c/em\u003e was also evaluated in the presence of 12.5 \u0026micro;g/ml of an EPI, namely N-(((2S,4R)-4-(aminomethyl)pyrrolidin-2-yl)methyl)-6-(4-fluorophenyl)-1H-indole-2-carboxamide [24]. \u0026nbsp; A similar MIC (1.0 \u0026micro;g/ml) was observed for CBR-4830 \u003cem\u003eagainst P. aeruginosa\u003c/em\u003e PAO1 in the presence of the EPI to that reported against the PAO1 mutant efflux defective strain CB392 ∆(\u003cem\u003emexAB-oprM\u003c/em\u003e)\u0026nbsp;∆(\u003cem\u003emexCD-oprJ\u003c/em\u003e) [24]. \u0026nbsp;The more potent of the cyclohepta analogs evaluated against \u003cem\u003eP. aeruginosa\u003c/em\u003e in the presence of an EPI was \u003cstrong\u003e10\u003c/strong\u003e with an MIC of 1.0\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u0026micro;g/ml. \u0026nbsp;Among the cyclopenta analogs, the most potent was \u003cstrong\u003e13\u003c/strong\u003e, which had its dichloro substituents at the same relative sites on the fused indole moiety and also exhibited the same MIC value as \u003cstrong\u003e10\u003c/strong\u003e. \u0026nbsp;These two analogs also exhibited the lower MICs against \u003cem\u003eE. coli\u003c/em\u003e, with\u003cem\u003e\u0026nbsp;\u003c/em\u003e\u003cstrong\u003e10\u003c/strong\u003e\u003cem\u003e\u0026nbsp;\u003c/em\u003eand \u003cstrong\u003e13\u0026nbsp;\u003c/strong\u003ehaving MICs of 8 and 16 \u0026micro;g/ml as compared to 8 ug/ml for CBR-4830. \u0026nbsp;Compound \u003cstrong\u003e10\u003c/strong\u003e had an MIC of 2 \u0026micro;g/ml against \u003cem\u003eK. pneumoniae\u003c/em\u003e, which was lower than that observed for \u003cstrong\u003e13\u003c/strong\u003e and CBR-4830, which had MICs of 8 \u0026micro;g/ml. \u0026nbsp;The MICs of \u003cstrong\u003e10\u003c/strong\u003e and \u003cstrong\u003e13\u003c/strong\u003e against \u003cem\u003eA. baumannii\u003c/em\u003e were 8 \u0026micro;g/m; which were lower than that observed for CBR-4830 (16 ug/ml). \u0026nbsp;Both \u003cstrong\u003e10\u003c/strong\u003e and \u003cstrong\u003e13\u003c/strong\u003e proved to have significantly improved formulation properties and were also less toxic when administered intravenously to mice relative to CBR-4830 (data not shown). \u0026nbsp;\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eThe ring-opened analogs \u003cstrong\u003e14-21\u003c/strong\u003e also had dramatically improved solubility in aqueous vehicles suitable for intravenous administration and were well tolerated when administered by this route to mice (data not shown). \u0026nbsp;However, their relative intrinsic MICs against\u003cem\u003e\u0026nbsp;P. aeruginosa\u003c/em\u003e, \u003cem\u003eE. coli\u003c/em\u003e, \u003cem\u003eK. pneumoniae\u003c/em\u003e or \u003cem\u003eA. baumanni\u0026nbsp;\u003c/em\u003ewere disappointing as none of these analogs had MIC values that were less than 32 \u0026micro;g/ml. \u0026nbsp;Only in the presence of an EPI, were notable MICs observed against \u003cem\u003eP. aeruginosa\u003c/em\u003e of 8 \u0026micro;g/ml for \u003cstrong\u003e14\u003c/strong\u003e and \u003cstrong\u003e21\u0026nbsp;\u003c/strong\u003eand\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e16 ug/ml for \u003cstrong\u003e16\u003c/strong\u003e, \u003cstrong\u003e17\u003c/strong\u003e and\u003cstrong\u003e\u0026nbsp;20\u003c/strong\u003e.\u003c/p\u003e\n\u003cp\u003eThe improve physicochemical properties of the ring-opened analogs prompted further studies on their structure-activity relationships. \u0026nbsp; We synthesized and evaluated the N-methyl and N,N-dimethyl amino analogs, \u003cstrong\u003e22\u003c/strong\u003e, and\u003cstrong\u003e\u0026nbsp;23\u003c/strong\u003e, as well as the N\u003csup\u003e1\u003c/sup\u003e-methyl derivative, \u003cstrong\u003e24,\u003c/strong\u003e of compound \u003cstrong\u003e14\u003c/strong\u003e. \u0026nbsp;We also prepared the methylamino analog, \u003cstrong\u003e25\u003c/strong\u003e, wherein the primary amine was attached to a primary carbon. \u0026nbsp;The antimicrobial activities of these compounds are summarized in Table 2. \u0026nbsp;Only in the case of \u003cstrong\u003e25\u003c/strong\u003e was notable activity observed against \u003cem\u003eP. aeruginosa\u0026nbsp;\u003c/em\u003ein the presence of an EPI. \u0026nbsp;The potency observed was comparable to \u003cstrong\u003e14\u003c/strong\u003e.\u003c/p\u003e\n\u003cp\u003eA series of N-(3-aminopropane) derivatives were prepared that incorporated the cycloheptyl, cyclopentyl and open-ring analogs of CBR-4830, specifically \u003cstrong\u003e26\u003c/strong\u003e, \u003cstrong\u003e27\u003c/strong\u003e and \u003cstrong\u003e28\u0026nbsp;\u003c/strong\u003e(Table 3). \u0026nbsp;Compound \u003cstrong\u003e26\u003c/strong\u003e had a lower MIC than \u003cstrong\u003e8\u003c/strong\u003e in terms of its intrinsic MIC against \u003cem\u003eP. aeruginosa\u003c/em\u003e (32 versus 128 \u0026micro;g/ml), in the presence of an EPI, the observed MIC was much higher (32 versus 4 \u0026micro;g/ml). \u0026nbsp;The differences between \u003cstrong\u003e8\u003c/strong\u003e and \u003cstrong\u003e26\u003c/strong\u003e were not that significant when evaluated in \u003cem\u003eE. coli\u003c/em\u003e, \u003cem\u003eK. pneumonia\u003c/em\u003ee and \u003cem\u003eA. baumannii\u003c/em\u003e. \u0026nbsp;Both \u003cstrong\u003e27\u003c/strong\u003e and \u003cstrong\u003e28\u003c/strong\u003e have either similar activity or are less active than their parent primary amines, \u003cstrong\u003e11\u003c/strong\u003e and \u003cstrong\u003e14\u003c/strong\u003e against \u003cem\u003eE. coli\u003c/em\u003e, \u003cem\u003eK. pneumonia\u003c/em\u003ee or \u003cem\u003eA. baumannii\u003c/em\u003e. \u0026nbsp;However, both \u003cstrong\u003e27\u003c/strong\u003e and \u003cstrong\u003e28\u003c/strong\u003e were notably less active than their parent primary amines when evaluated against \u003cem\u003eP. aeruginosa\u003c/em\u003e in the presence of an EPI. \u0026nbsp;Against \u003cem\u003eE. coli\u003c/em\u003e, \u003cstrong\u003e26\u003c/strong\u003e had the same MIC as \u003cstrong\u003e8\u003c/strong\u003e (16 \u0026micro;g/ml), but it was slightly more active against \u003cem\u003eK. pneumoniae\u0026nbsp;\u003c/em\u003e(8 versus 16 \u0026micro;g/ml) and slightly less active against \u003cem\u003eA. baumannii\u0026nbsp;\u003c/em\u003e(32 versus 16 \u0026micro;g/ml). \u0026nbsp;Compound \u003cstrong\u003e27\u003c/strong\u003e did not have remarkable activity against \u003cem\u003eE. coli\u003c/em\u003e, \u003cem\u003eK. pneumoniae\u003c/em\u003e, and \u003cem\u003eA. baumannii\u003c/em\u003e. \u0026nbsp;There were no major differences in the MICs observed for \u003cstrong\u003e14\u003c/strong\u003e and \u003cstrong\u003e28\u003c/strong\u003e observed in \u003cem\u003eE. coli\u003c/em\u003e, \u003cem\u003eK. pneumonia\u003c/em\u003ee or \u003cem\u003eA. baumannii\u003c/em\u003e. \u0026nbsp; We extended these studies to include a several N-(3-acylpropanamides) derived from the N-(3-aminopropane), \u003cstrong\u003e29\u003c/strong\u003e-\u003cstrong\u003e33\u003c/strong\u003e. \u0026nbsp; The relative antimicrobial activity of these hydrophobic derivatives are summarized in Table 3. None of these analogs exhibited intrinsic activity against \u003cem\u003eP. aeruginosa.\u003c/em\u003e\u0026nbsp; In the presence of an EPI, these analogs failed to exhibit potency against \u003cem\u003eP. aeruginosa\u003c/em\u003e comparable to \u003cstrong\u003e14\u003c/strong\u003e or \u003cstrong\u003e22\u0026nbsp;\u003c/strong\u003eunder these assay conditions. \u003cstrong\u003e\u0026nbsp;\u003c/strong\u003eThere were notable improvements in activity for \u003cstrong\u003e29\u003c/strong\u003e-\u003cstrong\u003e33\u003c/strong\u003e against \u003cem\u003eE. coli\u003c/em\u003e, \u003cem\u003eK. pneumoniae\u003c/em\u003e and \u003cem\u003eA. baumanni\u0026nbsp;\u003c/em\u003erelative to \u003cstrong\u003e22\u003c/strong\u003e. \u0026nbsp;Most notably, \u003cstrong\u003e31\u003c/strong\u003e had MICs against \u003cem\u003eE. coli\u003c/em\u003e, \u003cem\u003eK. pneumoniae\u003c/em\u003e and \u003cem\u003eA. baumanni\u0026nbsp;\u003c/em\u003ethat were 16-, 64-, and 8-fold lower than for \u003cstrong\u003e14\u003c/strong\u003e. \u0026nbsp;The cyclohexyl amide, \u003cstrong\u003e33\u003c/strong\u003e, proved to be less activity than the aryl or heteroaryl amides, \u003cstrong\u003e29-32\u003c/strong\u003e, that were evaluated.\u003c/p\u003e\n\u003cp\u003eWe also prepared\u003cstrong\u003e\u0026nbsp;34\u003c/strong\u003e, the reversed amide analog of \u003cstrong\u003e32\u003c/strong\u003e, as well as the 4-(3,4-dichlorophenoxy)butan-1-amine derivative, \u003cstrong\u003e35\u003c/strong\u003e.\u003cstrong\u003e\u0026nbsp;\u0026nbsp;\u003c/strong\u003eSimilar activity was observed between \u003cstrong\u003e32\u003c/strong\u003e and its reversed amide, \u003cstrong\u003e34\u003c/strong\u003e, derivatives. \u0026nbsp;The phenoxy derivative \u003cstrong\u003e35\u003c/strong\u003e also had similar activity to \u003cstrong\u003e34\u003c/strong\u003e.\u003c/p\u003e\n\u003cp\u003eWe evaluated six representative analogs (\u003cstrong\u003e8\u003c/strong\u003e, \u003cstrong\u003e9\u003c/strong\u003e, \u003cstrong\u003e10\u003c/strong\u003e, \u003cstrong\u003e14\u003c/strong\u003e, \u003cstrong\u003e26\u003c/strong\u003e, and \u003cstrong\u003e31\u003c/strong\u003e) for their abilities to target purified \u003cem\u003eE. coli\u003c/em\u003e MreB (EcMreB) and inhibit its ATPase activity. \u0026nbsp; All six compounds inhibited the ATPase activity of EcMreB (Figure 1), with corresponding IC\u003csub\u003e50\u003c/sub\u003e values ranging from 6.3 \u0026plusmn; 2.0 \u0026micro;M for analog \u003cstrong\u003e31\u003c/strong\u003e to 120 \u0026plusmn; 17.0 \u0026micro;M for analog \u003cstrong\u003e14\u003c/strong\u003e (Table 4). \u0026nbsp;Based on IC\u003csub\u003e50\u003c/sub\u003e values, the EcMreB-inhibiting potency of the six analogs tested followed the hierarchy: \u003cstrong\u003e31\u003c/strong\u003e \u0026gt; \u003cstrong\u003e9\u003c/strong\u003e \u0026asymp;\u003cstrong\u003e26\u003c/strong\u003e \u003cspan style='color: rgb(0, 0, 0); font-family: \"Times New Roman\"; font-size: medium; font-style: normal; font-variant-ligatures: normal; font-variant-caps: normal; font-weight: 400; letter-spacing: normal; orphans: 2; text-align: start; text-indent: 0px; text-transform: none; white-space: normal; widows: 2; word-spacing: 0px; -webkit-text-stroke-width: 0px; text-decoration-style: initial; text-decoration-color: initial; display: inline !important; float: none;'\u003e\u0026asymp;\u003c/span\u003e \u003cstrong\u003e10\u003c/strong\u003e \u0026gt; \u003cstrong\u003e8\u003c/strong\u003e \u0026gt; \u003cstrong\u003e14\u003c/strong\u003e.\u003c/p\u003e\n\u003cdiv align=\"center\"\u003e\n \u003ctable border=\"1\" cellpadding=\"0\" cellspacing=\"0\" width=\"0\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd colspan=\"2\" valign=\"top\" width=\"100%\"\u003e\n \u003cp\u003e\u003cstrong\u003eTable 4.\u003c/strong\u003e\u0026nbsp; IC\u003csub\u003e50\u003c/sub\u003e values for inhibition of the ATPase activity of EcMreB by select analogs.\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"50%\"\u003e\n \u003cp\u003e\u003cstrong\u003eCompound\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"50%\"\u003e\n \u003cp\u003e\u003cstrong\u003eIC\u003csub\u003e50\u003c/sub\u003e (\u0026mu;M)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"50%\"\u003e\n \u003cp\u003e\u003cstrong\u003e8\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"50%\"\u003e\n \u003cp\u003e44.1 \u0026plusmn; 7.8\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"50%\"\u003e\n \u003cp\u003e\u003cstrong\u003e9\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"50%\"\u003e\n \u003cp\u003e24.5 \u0026plusmn; 4.7\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"50%\"\u003e\n \u003cp\u003e\u003cstrong\u003e10\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"50%\"\u003e\n \u003cp\u003e29.4 \u0026plusmn; 8.8\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"50%\"\u003e\n \u003cp\u003e\u003cstrong\u003e14\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"50%\"\u003e\n \u003cp\u003e120 \u0026plusmn; 17.0\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"50%\"\u003e\n \u003cp\u003e\u003cstrong\u003e26\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"50%\"\u003e\n \u003cp\u003e25.1 \u0026plusmn; 7.0\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"50%\"\u003e\n \u003cp\u003e\u003cstrong\u003e31\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"50%\"\u003e\n \u003cp\u003e6.3 \u0026plusmn; 2.0\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd colspan=\"2\" width=\"100%\"\u003e\n \u003cp\u003eEcMreB = E. coli MreB; IC\u003csub\u003e50\u003c/sub\u003e = the compound concentration that inhibits the ATPase activity of EcMreB by 50%. \u0026nbsp; \u0026nbsp; IC\u003csub\u003e50\u003c/sub\u003e values were derived from fits of the velocity vs. [compound] plots shown in Figure 1, with the indicated uncertainties reflecting the standard deviation of the fitted curves from the experimental data points.\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n \u003c/table\u003e\n\u003c/div\u003e"},{"header":"Discussion","content":"\u003cp\u003eThe data in Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e indicate that cyclohepta and cyclopenta analogs related to CBR-4830 retain significant antibacterial activity. CBR-4830 proved difficult to formulate (requiring 40% propylene glycol to achieve a concentration of 2 mg/ml) and did produce severe toxic effects when administered to mice b.i.d. at a dose of 400 ug/mouse (data not shown). Several of the promising cyclohepa and cyclopenta analogs could be readily formulated using 10 mM citrate in water and were well tolerated when administered at more than twice this dose. The actual significance of these differences, however, needs to be examined in the context of comparative pharmacokinetic studies which have not been performed. The ring-opened analogs of CBR-4830 were significantly less active and only exhibited modest activity when evaluated against \u003cem\u003eP. aeruginosa\u003c/em\u003e in the presence of a bacterial efflux pump inhibitor, such as N-(((2S,4R)-4-(aminomethyl)pyrrolidin-2-yl)methyl)-6-(4-fluorophenyl)-1\u003cem\u003eH\u003c/em\u003e-indole-2-carboxamide. The ring-open series of compounds, like the cyclohepta and cyclopenta analogs, had favorable formulation properties and were well tolerated in mice. Efforts to improve antibacterial activity among the ring-opened analog \u003cb\u003e8\u003c/b\u003e, by either mono- or demethylation of the amino substituent as in \u003cb\u003e22\u003c/b\u003e and \u003cb\u003e23\u003c/b\u003e, or methylation of the indole nitrogen as in the case of \u003cb\u003e24\u003c/b\u003e, failed to enhance activity (Table\u0026nbsp;\u003cspan refid=\"Tab2\" class=\"InternalRef\"\u003e2\u003c/span\u003e). Changing the 1-aminoethyl of \u003cb\u003e8\u003c/b\u003e to a aminomethyl group as in \u003cb\u003e25\u003c/b\u003e did restore activity against \u003cem\u003eP. aeruginosa\u003c/em\u003e in the presence of the EPI, N-(((2S,4R)-4-(aminomethyl)pyrrolidin-2-yl)methyl)-6-(4-fluorophenyl)-1\u003cem\u003eH\u003c/em\u003e-indole-2-carboxamide. However, no significant activity was observed against \u003cem\u003eE. coli\u003c/em\u003e, \u003cem\u003eK. pneumoniae\u003c/em\u003e or \u003cem\u003eA. baumannii\u003c/em\u003e (Table\u0026nbsp;\u003cspan refid=\"Tab2\" class=\"InternalRef\"\u003e2\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eThe impact of attaching a 3-aminopropyl linkage to the amine of cycloheptyl, cyclopentyl or ring-opened analogs of CBR-4830 on antibacterial activity was also evaluated (Table\u0026nbsp;\u003cspan refid=\"Tab3\" class=\"InternalRef\"\u003e3\u003c/span\u003e). In the case of the cycloheptyl analog \u003cb\u003e26\u003c/b\u003e, there was an enhancement in intrinsic activity against \u003cem\u003eP. aeruginosa\u003c/em\u003e and a modest increase in activity against \u003cem\u003eK. pneumonia\u003c/em\u003ee relative to \u003cb\u003e8\u003c/b\u003e. In the case of the cyclopenta analog \u003cb\u003e27\u003c/b\u003e, there was a notable loss in activity observed against \u003cem\u003eP. aeruginosa\u003c/em\u003e when evaluated in the presence of an EPI and no increase in activity against \u003cem\u003eE. coli\u003c/em\u003e, \u003cem\u003eK. pneumoniae\u003c/em\u003e or \u003cem\u003eA. baumanni\u003c/em\u003e relative to \u003cb\u003e11\u003c/b\u003e. Modification of the ring-opened analog by attaching this linkage as in \u003cb\u003e28\u003c/b\u003e primarily resulted in a loss of the activity that was previously observed with \u003cb\u003e14\u003c/b\u003e in the presence of an EPI against \u003cem\u003eP. aeruginosa\u003c/em\u003e.\u003c/p\u003e \u003cp\u003eWe examined the effect of modifying the terminal amine of \u003cb\u003e28\u003c/b\u003e by forming various hydrophobic amide derivatives as a possible means of enhancing the antibacterial properties of these ring-opened analogs. While these analogs \u003cb\u003e29\u0026ndash;33\u003c/b\u003e did not exhibit significant intrinsic activity against \u003cem\u003eP. aeruginosa\u003c/em\u003e, their activity in the presence of an EPI was improved relative to \u003cb\u003e28\u003c/b\u003e. More significantly, their intrinsic antibacterial activity was increased against \u003cem\u003eE. coli\u003c/em\u003e and \u003cem\u003eK. pneumoniae\u003c/em\u003e, and to a less extent against \u003cem\u003eA. baumannii\u003c/em\u003e. Similar improvements in activity were observed with the reversed amide derivative \u003cb\u003e34\u003c/b\u003e and the oxyphenyl analog \u003cb\u003e35\u003c/b\u003e.\u003c/p\u003e \u003cp\u003eWe also sought to validate MreB as the antibacterial target of our analogs. To this end, we selected six representative analogs and evaluated the compounds for their abilities to inhibit the catalytic (ATPase) activity of purified \u003cem\u003eE. coli\u003c/em\u003e MreB (EcMreB). Significantly, all six compounds tested exhibited concentration-dependent inhibition of EcMreB ATPase activity (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003e), establishing their antibacterial actions as MreB inhibitors. In addition, the relative EcMreB-inhibiting potencies of the compounds (as reflected by the magnitudes of the IC\u003csub\u003e50\u003c/sub\u003e values shown in Table\u0026nbsp;\u003cspan refid=\"Tab4\" class=\"InternalRef\"\u003e4\u003c/span\u003e) were generally correlated with their corresponding antibacterial activities, further validating MreB as the antibacterial target of our compounds. Among the analogs tested, \u003cb\u003e31\u003c/b\u003e was the most potent inhibitor of EcMreB activity. Significantly, this analog also exhibited the best antibacterial activity against \u003cem\u003eK. pneumoniae\u003c/em\u003e of all compounds evaluated here and was among the best agents versus \u003cem\u003eE. coli\u003c/em\u003e and \u003cem\u003eA. baumannii\u003c/em\u003e as well. Analog \u003cb\u003e31\u003c/b\u003e may thus represent an important reference compound for the development of next-generation MreB inhibitors.\u003c/p\u003e"},{"header":"Conclusions","content":"\u003cp\u003eThe structure activity studies performed with these varied analogs of CBR-4830 indicate that in many instances significant antibacterial activity can be retained. In addition, it is possible to reduce the toxicity associated with CBR-4830 and improve its formulation properties. The extreme toxicity of A22 and CBR-4830 has hindered efforts to demonstrate the \u003cem\u003ein vivo\u003c/em\u003e efficacy of these MreB inhibitors. In view of their unique mechanism of action, such agents would be anticipated to be useful against multidrug-resistant bacteria. The data on the structure-activity of these CBR-4830 derivatives suggest that the development of next-generation MreB inhibitors could overcome this limitation and provide a means for demonstrating the potential clinical efficacy and utility of MreB inhibitors as antibiotics. Evaluation of comparative pharmacokinetic parameters and efficacy in vivo remain essential for assessing the potential of this novel class of antibiotics.\u003c/p\u003e"},{"header":"Experimental","content":"\u003cp\u003e\u003cstrong\u003eChemistry: General Methods\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eAll reactions, unless otherwise stated, were done under nitrogen atmosphere. Reaction monitoring and follow-up were done using aluminum backed Silica G TLC plates with UV254 (Sorbent Technologies), visualizing with ultraviolet light. Flash column chromatography was done on a Combi Flash Rf Teledyne ISCO using hexane, ethyl acetate, dichloromethane, and methanol. The \u003csup\u003e1\u003c/sup\u003eH (400 MHz) and \u003csup\u003e13\u003c/sup\u003eC (100 MHz) NMR spectra were done in CDCl\u003csub\u003e3\u003c/sub\u003e, Methanol-d\u003csub\u003e4\u003c/sub\u003e, and DMSO-d\u003csub\u003e6\u003c/sub\u003e and recorded on a Bruker Avance III (400 MHz) Multinuclear NMR Spectrometer. Data is expressed in parts per million relative to the residual nondeuterated solvent signals, spin multiplicities are given as s (singlet), d (doublet), dd (doublet of doublets), t (triplet), dt (doublet of triplets), q (quartet), m (multiplet), and bs (broad singlet), and coupling constants (\u003cem\u003eJ\u003c/em\u003e) are reported in Hertz. Analytical HPLC was performed on a Shimadzu LC-20AT Prominence liquid chromatograph using a 150 x 4.6 mm Princeton SPHER-100 RP C18 1000A 5 um column using 0% water for 2 minutes and a 0-100% water/methanol gradient over a 5-minute period and 5 minutes at 100% methanol at a 2.0 ml/minute flow rate monitoring uv absorbance at 254 and 296 nm. Using this method of analysis, the purity of all compounds used in bioassays was determined to be \u0026ge; 95%. Mass spectrometry (MS) was performed by electrospray (ESI) ionization using a Shimadzu 2020 LC-MS quadrupole mass spectrometer using a 0-100% water/methanol gradient over a 3-minute period. HRMS experiments were conducted using the Waters ACQUITY UPLC \u0026ndash; Synapt G2 HRMS (Milford, MA) to determine the accurate mass values of final compounds. UPLC separation was performed on an ACQUITY UPLC BEH C18 column (2.1 mm \u0026times; 50 mm, 1.7 \u0026mu;m) with 0.1% formic acid in water and 0.1% formic acid in methanol as mobile phase A and B, respectively. The column temperature was maintained at 40\u0026deg;C. The gradient was set from 5% to 95% B over 3 minutes at a flow rate of 0.25 mL/min. Extensive fragmentation under the conditions used for these HRMS analyses did not allow for the detection of parent ions. HRMS data obtained under these conditions are provided for several of the compounds evaluated in Supplemental Materials.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e2-Bromo-5,6,7,8,9,10-hexahydrocyclohepta[\u003cem\u003eb\u003c/em\u003e]indol-6-amine (8)\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e2-Bromo-7,8,9,10-tetrahydrocyclohepta[\u003cem\u003eb\u003c/em\u003e]indol-6(5\u003cem\u003eH\u003c/em\u003e)-one (100 mg, 0.36 mmol), ammonium acetate (277 mg, 3.60 mmol) and sodium cyanoborohydride (113 mg, 1.80 mmol) were dissolved in ethanol (10 mL). The mixture was stirred for overnight at 60 \u003csup\u003eo\u003c/sup\u003eC. The reaction mixture was acidified with 6N HCl, and it was washed with ethyl acetate. Then, the aqueous layer was basified with NaOH, and it was extracted with ethyl acetate. The organic layer was washed with brine, and it was dried over Na\u003csub\u003e2\u003c/sub\u003eSO\u003csub\u003e4\u003c/sub\u003e. The organic layer was concentrated under reduced pressure, and the residue was purified on an ISCO chromatograph (0-10% methanol/ dichloromethane + 0.1% NH\u003csub\u003e4\u003c/sub\u003eOH) to give the product as a white solid (34 mg, 34%); \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (CD\u003csub\u003e3\u003c/sub\u003eOD) \u0026delta; 10.76 (bs, 1H), 7.51 (d, \u003cem\u003eJ\u003c/em\u003e = 2 Hz, 1H), 7.24 (d, \u003cem\u003eJ\u003c/em\u003e = 8 Hz, 1H), 7.03 (dd, \u003cem\u003eJ\u003c/em\u003e = 8 Hz, \u003cem\u003eJ\u003c/em\u003e = 2 Hz, 1H), 4.01-3.97 (m, 1H), 2.86-2.81 (m, 1H), 2.58-2.56 (m, 1H), 1.99-1.84 (m, 2H), 1.71-1.50 (m, 4H); LC/MS RT = 2.57 (M-H\u003csup\u003e-\u003c/sup\u003e: 277/279).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u003cem\u003e(E)\u003c/em\u003e\u003c/strong\u003e\u003cstrong\u003e-2-(Hydroxymethylene)cycloheptan-1-one\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eCycloheptanone (2.95 mL, 25 mmol) was dissolved in anhydrous tetrahydrofuran (10 mL), and it was cooled to 0 \u003csup\u003eo\u003c/sup\u003eC. A solution of 1.0 M LiHMDS in tetrahydrofuran (30 mL, 30 mmol) was slowly added, and it was stirred for 5 minutes at 0 \u003csup\u003eo\u003c/sup\u003eC. Ethyl formate (2.42 mL, 30 mmol) was slowly added, and it was stirred for 2 hours at 0 \u003csup\u003eo\u003c/sup\u003eC. The reaction mixture was diluted with ethyl acetate, and it was washed with 6N HCl and brine. The organic layer was dried over Na\u003csub\u003e2\u003c/sub\u003eSO\u003csub\u003e4\u003c/sub\u003e. The organic layer was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-20% ethyl acetate/hexane) to give product as a colorless oil (1.30 g, 37%); \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (CDCl\u003csub\u003e3\u003c/sub\u003e) \u0026delta; 7.59 (d, \u003cem\u003eJ\u003c/em\u003e = 9 Hz, 1H), 2.51-2.47 (m, 2H), 2.23-2.19 (m, 2H), 1.73-1.54 (m, 6H).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u003cem\u003e(E)\u003c/em\u003e\u003c/strong\u003e\u003cstrong\u003e-2-(2-(4-Bromophenyl)hydrazineylidene)cycloheptan-1-one\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eTo a solution of 4-bromoaniline (1.60 g, 9.30 mmol) in concentrated hydrochloric acid (2 mL), a solution of sodium nitrite (642 mg, 9.30 mmol) in water (4 mL) was added slowly at 0 \u003csup\u003eo\u003c/sup\u003eC. The mixture was stirred for 30 minutes at 0 \u003csup\u003eo\u003c/sup\u003eC. In a separate round bottom flask, \u003cem\u003e(E)\u003c/em\u003e-2-(hydroxymethylene)cycloheptan-1-one (1.30 g, 9.30 mmol)\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003ewas dissolve in methanol (12 mL). To the mixture, a solution of sodium acetate (1.91 g, 23.3 mmol) in water (5 mL) was added slowly at 0 \u003csup\u003eo\u003c/sup\u003eC. The mixture was stirred for 20 minutes at 0 \u003csup\u003eo\u003c/sup\u003eC. Then, the freshly prepared diazonium salt solution was slowly added. The mixture was stirred for additional 30 minutes at 0 \u003csup\u003eo\u003c/sup\u003eC, and the formed yellow suspension was filtered to give the product as a yellow solid (2.17 g, 79%); \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (DMSO-d\u003csub\u003e6\u003c/sub\u003e) \u0026delta; 13.27 (s, 1H), 7.43 (d, \u003cem\u003eJ\u003c/em\u003e = 9 Hz, 2H), 7.23 (d, \u003cem\u003eJ\u003c/em\u003e = 9 Hz, 2H), 2.62-2.56 (m, 4H), 1.68 (m, 6H).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e2-Bromo-7,8,9,10-tetrahydrocyclohepta[\u003cem\u003eb\u003c/em\u003e]indol-6(5\u003cem\u003eH\u003c/em\u003e)-one (8a)\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cem\u003e(E)\u003c/em\u003e-2-(2-(4-Bromophenyl)hydrazineylidene)cycloheptan-1-one (2.17 g, 7.35 mmol) was dissolved in a mixture of concentrated hydrochloric acid (2 mL) and acetic acid (8 mL), and it was stirred for 30 minutes at 130 \u003csup\u003eo\u003c/sup\u003eC. The resulting dark brown suspension was diluted with ethyl acetate, and it was washed with 10% NaOH and brine. The organic layer was dried over Na\u003csub\u003e2\u003c/sub\u003eSO\u003csub\u003e4\u003c/sub\u003e, and it was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-20% ethyl acetate/hexane) to give product as a yellow solid (472 mg, 23%); \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (CDCl\u003csub\u003e3\u003c/sub\u003e) \u0026delta; 8.90 (bs, 1H), 7.79 (s, 1H), 7.41 (dd, \u003cem\u003eJ\u003c/em\u003e = 9 Hz, \u003cem\u003eJ\u003c/em\u003e = 2 Hz, 1H), 7.26-7.23 (m, 1H), 3.11-3.07 (m, 2H), 2.86-2.82 (m, 2H), 2.11-1.99 (m, 4H).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e1,2-Dichloro-5,6,7,8,9,10-hexahydrocyclohepta[\u003cem\u003eb\u003c/em\u003e]indol-6-amine (9)\u003cem\u003e.\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e1,2-Dichloro-7,8,9,10-tetrahydrocyclohepta[\u003cem\u003eb\u003c/em\u003e]indol-6(5\u003cem\u003eH\u003c/em\u003e)-one (99 mg, 0.37 mmol), ammonium acetate (285 mg, 3.70 mmol) and sodium cyanoborohydride (116 mg, 1.85 mmol) were dissolved in ethanol (10 mL). The reaction mixture was stirred for overnight at 60 \u003csup\u003eo\u003c/sup\u003eC. After removal of solvent, the reaction mixture was diluted with ethyl acetate. The organic layer was washed with 10% sodium hydroxide and brine, and it was dried over sodium sulfate. The organic layer was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-10% methanol/dichloromethane + 0.1% NH\u003csub\u003e4\u003c/sub\u003eOH) to give product as a white solid. The solid was then treated with 4N hydrochloric acid in dioxane, and it was stirred for 30 minutes at room temperature. The white suspension was concentrated under reduced pressure, and the resulting residue was suspended in ethyl acetate. The suspension was filtered to give hydrochloric acid salt of product as a white solid (42 mg, 37%); \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (DMSO-d\u003csub\u003e6\u003c/sub\u003e) \u0026delta; 11.65 (bs, 1H), 8.42 (bs, 3H), 7.37 (d, \u003cem\u003eJ\u003c/em\u003e = 8 Hz, 1H), 7.25 (d, \u003cem\u003eJ\u003c/em\u003e = 9 Hz, 1H), 4.58 (m, 1H), 3.65-3.60 (m, 2H), 3.12-2.98 (m, 2H), 1.97-1.89 (m, 4H); LC/MS RT = 2.90 (M-H\u003csup\u003e-\u003c/sup\u003e: 267/269).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u003cem\u003etert-\u003c/em\u003e\u003c/strong\u003e\u003cstrong\u003eButyl 1,2-dichloro-6-oxo-7,8,9,10-tetrahydrocyclohepta[\u003cem\u003eb\u003c/em\u003e]indole-5(6\u003cem\u003eH\u003c/em\u003e)-carboxylate (9a) and \u003cem\u003etert\u003c/em\u003e-Butyl 2,3-dichloro-6-oxo-7,8,9,10-tetrahydrocyclohepta[\u003cem\u003eb\u003c/em\u003e]indole-5(6\u003cem\u003eH\u003c/em\u003e)-carboxylate (10a)\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eTo a solution of 3,4-dichloroaniline (1.53 g, 9.42 mmol) in concentrated hydrochloric acid (10 mL), a solution of sodium nitrite (650 mg, 9.42 mmol) in water (20 mL) was added slowly at 0 \u003csup\u003eo\u003c/sup\u003eC. The mixture was stirred for 30 minutes at 0 \u003csup\u003eo\u003c/sup\u003eC. In a separate round bottom flask, (E)-2-(hydroxymethylene)cycloheptan-1-one (1.32 g, 9.42 mmol) was dissolve in methanol (12 mL). To the mixture, a solution of sodium acetate (1.93 g, 23.55 mmol) in water (5 mL) was added slowly at 0 \u003csup\u003eo\u003c/sup\u003eC. The mixture was stirred for 20 minutes at 0 \u003csup\u003eo\u003c/sup\u003eC. Then, the freshly prepared diazonium salt solution was slowly added. The mixture was stirred for additional 30 minutes at 0 \u003csup\u003eo\u003c/sup\u003eC. The reaction mixture was diluted with ethyl acetate, and the organic layer was washed with saturated sodium bicarbonate followed brine. The organic layer was concentrated under reduced pressure and the resulted dark brown oil was carried next step without further purification.\u003c/p\u003e\n\u003cp\u003eThe residue was dissolved in formic acid (10 mL), and it was stirred for 2 hours at 100 \u003csup\u003eo\u003c/sup\u003eC. The resulting dark brown suspension was diluted with ethyl acetate, and it was washed with 10% NaOH and brine. The organic layer was dried over sodium sulfate, and it was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-20% ethyl acetate/hexane) to give a mixture of two regioisomers, 1,2-dichloro-7,8,9,10-tetrahydrocyclohepta[\u003cem\u003eb\u003c/em\u003e]indol-6(5\u003cem\u003eH\u003c/em\u003e)-one and 2,3-dichloro-7,8,9,10-tetrahydrocyclohepta[\u003cem\u003eb\u003c/em\u003e]indol-6(5\u003cem\u003eH\u003c/em\u003e)-one as a beige solid (212 mg, 8%).\u003c/p\u003e\n\u003cp\u003eTo a solution of a mixture of the two regioisomers (212 mg, 0.79 mmol) in tetrahydrofuran (10 mL), Boc anhydride (345 mg, 1.58 mmol) and DMAP (89 mg, 0.79 mmol) were added. The reaction mixture was stirred for 3 hours. The reaction mixture was diluted with ethyl acetate, and it was washed with saturated ammonium chloride, followed by brine. The organic layer was dried over sodium sulfate, and it was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-10% ethyl acetate/hexane) to give \u003cem\u003etert-\u003c/em\u003ebutyl 1,2-dichloro-6-oxo-7,8,9,10-tetrahydrocyclohepta[\u003cem\u003eb\u003c/em\u003e]indole-5(6\u003cem\u003eH\u003c/em\u003e)-carboxylate (152 mg, 49%). \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (CDCl\u003csub\u003e3\u003c/sub\u003e) \u0026delta; 7.93 (d, \u003cem\u003eJ\u003c/em\u003e = 9 Hz, 1H), 7.41 (d, \u003cem\u003eJ\u003c/em\u003e = 9 Hz, 1H), 3.42-3.38 (m, 2H), 2.87-2.83 (m, 2H), 1.97-1.95 (m, 4H), 1.55 (s, 9H). along with \u003cem\u003etert\u003c/em\u003e-butyl 2,3-dichloro-6-oxo-7,8,9,10-tetrahydrocyclohepta[\u003cem\u003eb\u003c/em\u003e]indole-5(6\u003cem\u003eH\u003c/em\u003e)-carboxylate (59 mg, 20%). \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (CDCl\u003csub\u003e3\u003c/sub\u003e) \u0026delta; 8.20 (s, 1H), 7.62 (s, 1H), 2.91 (m, 2H), 2.85 (m, 2H), 2.02-1.98 (m, 4H), 1.57 (s, 9H).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e1,2-Dichloro-7,8,9,10-tetrahydrocyclohepta[\u003cem\u003eb\u003c/em\u003e]indol-6(5\u003cem\u003eH\u003c/em\u003e)-one (9a)\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eTo a solution of \u003cem\u003etert-\u003c/em\u003ebutyl 1,2-dichloro-6-oxo-7,8,9,10-tetrahydrocyclohepta[\u003cem\u003eb\u003c/em\u003e]indole-5(6\u003cem\u003eH\u003c/em\u003e)-carboxylate (152 mg, 0.41 mmol) in dichloromethane (5 mL), trifluoroacetic acid (1 mL) was added. The reaction mixture was stirred for an hour at room temperature. After removal of solvent, the mixture was diluted with ethyl acetate, and it was washed with 10% sodium hydroxide and brine. The organic layer was dried over sodium sulfate, and it was concentrated under reduced pressure to give product as a white solid (99 mg, 90%); \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (CDCl\u003csub\u003e3\u003c/sub\u003e) \u0026delta; 9.08 (bs, 1H), 7.35 (d, \u003cem\u003eJ\u003c/em\u003e = 9 Hz, 1H), 7.20 (d, \u003cem\u003eJ\u003c/em\u003e = 9 Hz, 1H), 3.58-3.54 (m, 2H), 2.87-2.83 (m, 2H), 2.11-2.04 (m, 2H), 1.99-1.91 (m, 2H).\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e2,3-Dichloro-7,8,9,10-tetrahydrocyclohepta[\u003cem\u003eb\u003c/em\u003e]indol-6(5\u003cem\u003eH\u003c/em\u003e)-one (10a)\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eTo a solution of \u003cem\u003etert\u003c/em\u003e-butyl 2,3-dichloro-6-oxo-7,8,9,10-tetrahydrocyclohepta[\u003cem\u003eb\u003c/em\u003e]indole-5(6\u003cem\u003eH\u003c/em\u003e)-carboxylate (943 mg, 2.64 mmol) in dichloromethane (25 mL), trifluoroacetic acid (5 mL) was added. The reaction mixture was stirred for 2 hours at room temperature. After removal of solvent, the mixture was diluted with ethyl acetate, and it was washed with 10% sodium hydroxide and brine. The organic layer was dried over sodium sulfate, and it was concentrated under reduced pressure to give product as a white solid (675 mg, 95%); \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (CDCl\u003csub\u003e3\u003c/sub\u003e) \u0026delta; 8.90 (bs, 1H), 7.74 (s, 1H), 7.48 (s, 1H), 3.10-3.06 (m, 2H), 2.87-2.83 (m, 2H), 2.11-2.05 (m, 2H), 2.01-1.99 (m, 2H).\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e2,3-Dichloro-5,6,7,8,9,10-hexahydrocyclohepta[\u003cem\u003eb\u003c/em\u003e]indol-6-amine (10).\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e2,3-Dichloro-7,8,9,10-tetrahydrocyclohepta[\u003cem\u003eb\u003c/em\u003e]indol-6(5\u003cem\u003eH\u003c/em\u003e)-one (100 mg, 0.37 mmol), ammonium acetate (285 mg, 3.70 mmol) and sodium cyanoborohydride (116 mg, 1.85 mmol) were dissolved in ethanol (10 mL). The reaction mixture was stirred for overnight at 60 \u003csup\u003eo\u003c/sup\u003eC. After removal of solvent, the reaction mixture was diluted with ethyl acetate. The organic layer was washed with 10% sodium hydroxide and brine, and it was dried over sodium sulfate. The organic layer was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-10% methanol/dichloromethane + 0.1% NH\u003csub\u003e4\u003c/sub\u003eOH) to give product as a white solid. The solid was then treated with 4N hydrochloric acid in dioxane, and it was stirred for 30 minutes at room temperature. The white suspension was concentrated under reduced pressure, and the resulting residue was suspended in ethyl acetate. The suspension was filtered to give hydrochloric acid salt of product as a white solid (76 mg, 67%); \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (DMSO-d\u003csub\u003e6\u003c/sub\u003e) \u0026delta; 11.36 (bs, 1H), 8.44 (bs, 3H), 7.78 (s, 1H), 7.62 (s, 1H), 4.56 (m, 1H), 3.55-3.26 (m, 2H), 2.89-2.71 (m, 4H), 1.93-1.76 (m, 2H); LC/MS RT = 2.91 (M-H\u003csup\u003e-\u003c/sup\u003e: 267/269).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e7-Bromo-1,2,3,4-tetrahydrocyclopenta[\u003cem\u003eb\u003c/em\u003e]indol-3-amine (11)\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e7-Bromo-1,4-dihydrocyclopenta[\u003cem\u003eb\u003c/em\u003e]indol-3(2\u003cem\u003eH\u003c/em\u003e)-one (100 mg, 0.40 mmol), ammonium acetate (308 mg, 4.00 mmol) and sodium cyanoborohydride (126 mg, 2.00 mmol) were dissolved in ethanol (10 mL). The mixture was stirred for overnight at 60 \u003csup\u003eo\u003c/sup\u003eC. An additional ammonium acetate (308 mg, 4.00 mmol) and sodium cyanoborohydride (126 mg, 2.00 mmol) along with catalytic amount of acetic acid were added. The mixture was stirred for 5 hours at 85 \u003csup\u003eo\u003c/sup\u003eC. The reaction mixture was acidified with 6N HCl, and it was washed with ethyl acetate. Then, the aqueous layer was basified with NaOH, and it was extracted with ethyl acetate. The organic layer was washed with brine, and it was dried over Na\u003csub\u003e2\u003c/sub\u003eSO\u003csub\u003e4\u003c/sub\u003e. The organic layer was concentrated under reduced pressure, and the residue was purified on an ISCO chromatograph (0-10% methanol/dichloromethane + 0.1% NH\u003csub\u003e4\u003c/sub\u003eOH) to give the product as a white solid (17 mg, 17%); \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (CD\u003csub\u003e3\u003c/sub\u003eOD) \u0026delta; 7.48 (d, \u003cem\u003eJ\u003c/em\u003e = 1 Hz, 1H), 7.22 (d, \u003cem\u003eJ\u003c/em\u003e = 9 Hz, 1H), 7.12 (dd, \u003cem\u003eJ\u003c/em\u003e = 9 H, \u003cem\u003eJ\u003c/em\u003e = 2 Hz, 1H), 4.43-4.41 (m, 1H), 2.94-2.83 (m, 2H), 2.78-2.62 (m, 1H), 2.21-2.12 (m, 1H); LC/MS RT = 2.57 (M-H\u003csup\u003e-\u003c/sup\u003e: 249/251).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e5 \u003cem\u003e(E)\u003c/em\u003e-2-(Hydroxymethylene)cyclopentan-1-one\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eCyclopentanone (2.21 mL, 25 mmol) was dissolved in anhydrous tetrahydrofuran (10 mL), and it was cooled to 0 \u003csup\u003eo\u003c/sup\u003eC. A solution of 1.0 M LiHMDS in tetrahydrofuran (30 mL, 30 mmol) was slowly added, and it was stirred for 5 minutes at 0 \u003csup\u003eo\u003c/sup\u003eC. Ethyl formate (2.42 mL, 30 mmol) was slowly added, and it was stirred for 2 hours at 0 \u003csup\u003eo\u003c/sup\u003eC. The reaction mixture was diluted with ethyl acetate, and it was washed with 6N HCl and brine. The organic layer was dried over Na\u003csub\u003e2\u003c/sub\u003eSO\u003csub\u003e4\u003c/sub\u003e, and it was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-30% ethyl acetate/hexane) to give product as a white solid (772 mg, 28%); \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (CDCl\u003csub\u003e3\u003c/sub\u003e) \u0026delta; 7.21 (s, 1H), 2.56-2.51 (m, 2H), 2.43-2.37 (m, 2H), 2.01-1.94 (m, 2H).\u003cstrong\u003e\u003cem\u003e\u0026nbsp;\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u003cem\u003e(E)\u003c/em\u003e\u003c/strong\u003e\u003cstrong\u003e-2-(2-(4-Bromophenyl)hydrazineylidene)cyclopentan-1-one\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eTo a solution of 4-bromoaniline (1.07 g, 6.23 mmol) in concentrated hydrochloric acid (2 mL), a solution of sodium nitrite (430 mg, 6.23 mmol) in water (4 mL) was added slowly at 0 \u003csup\u003eo\u003c/sup\u003eC. The mixture was stirred for 30 minutes at 0 \u003csup\u003eo\u003c/sup\u003eC. In a separate round bottom flask, 5 \u003cem\u003e(E)\u003c/em\u003e-2-(hydroxymethylene)cyclopentan-1-one (700 mg, 6.23 mmol) was dissolve in methanol (12 mL). To the mixture, a solution of sodium acetate (1.28 g, 15.58 mmol) in water (5 mL) was added slowly at 0 \u003csup\u003eo\u003c/sup\u003eC. The mixture was stirred for 20 minutes at the temperature. Then, the freshly prepared diazonium salt solution was slowly added. The mixture was stirred for additional 30 minutes at the temperature, and the formed yellow suspension was filtered to give the product as a yellow solid (1.47 g, 89%); \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (DMSO-d\u003csub\u003e6\u003c/sub\u003e) \u0026delta; 10.03 (s, 1H), 7.42 (d, \u003cem\u003eJ\u003c/em\u003e = 9 Hz, 2H), 7.19 (d, \u003cem\u003eJ\u003c/em\u003e = 9 Hz, 2H), 2.65-2.60 (m, 2H), 2.34-2.29 (m, 2H), 1.99-1.95 (m, 2H).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e7-Bromo-1,4-dihydrocyclopenta[\u003cem\u003eb\u003c/em\u003e]indol-3(2\u003cem\u003eH\u003c/em\u003e)-one (11a)\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cem\u003e(E)\u003c/em\u003e-2-(2-(4-Bromophenyl)hydrazineylidene)cyclopentan-1-one (1.47 g, 5.50 mmol) was dissolved in a mixture of concentrated hydrochloric acid (2 mL) and acetic acid (8 mL), and it was stirred for 30 minutes at 130 \u003csup\u003eo\u003c/sup\u003eC. The resulting dark brown suspension was diluted with ethyl acetate, and it was washed with 10% NaOH and brine. The organic layer was dried over Na\u003csub\u003e2\u003c/sub\u003eSO\u003csub\u003e4\u003c/sub\u003e, and it was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-20% ethyl acetate/hexane) to give product as a beige solid (116 mg, 8%); \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (CDCl\u003csub\u003e3\u003c/sub\u003e) \u0026delta; 8.80 (bs, 1H), 7.87 (d, \u003cem\u003eJ\u003c/em\u003e = 2 Hz, 1H), 7.48 (dd, \u003cem\u003eJ\u003c/em\u003e = 9 H, \u003cem\u003eJ\u003c/em\u003e = 2 Hz, 1H), 7.35 (d, \u003cem\u003eJ\u003c/em\u003e = 9 Hz, 1H), 3.10-3.07 (m, 2H), 3.03-3.00 (m, 2H).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e6,7-Dichloro-1,2,3,4-tetrahydrocyclopenta[\u003cem\u003eb\u003c/em\u003e]indol-3-amine. (12).\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e6,7-Dichloro-1,4-dihydrocyclopenta[\u003cem\u003eb\u003c/em\u003e]indol-3(2\u003cem\u003eH\u003c/em\u003e)-one (148 mg, 0.62 mmol), ammonium acetate (956 mg, 12.4 mmol) and sodium cyanoborohydride (195 mg, 3.1 mmol) were dissolved in ethanol (20 mL). The reaction mixture was stirred for overnight at 60 \u003csup\u003eo\u003c/sup\u003eC. After removal of solvent, the reaction mixture was diluted with ethyl acetate. The organic layer was washed with 10% sodium hydroxide and brine, and it was dried over sodium sulfate. The organic layer was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-10% methanol/dichloromethane + 0.1% NH\u003csub\u003e4\u003c/sub\u003eOH) to give product as a white solid (93 mg, 62%); \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (MeOD) \u0026delta; 7.46 (s, 1H), 7.45 (s, 1H), 4.43-4.39 (m, 1H), 2.93-2.79 (m, 2H), 2.71-2.61 (m, 1H), 2.21-2.12 (m, 1H); LC/MS RT = 2.74 (M-H\u003csup\u003e-\u003c/sup\u003e: 239/241).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e6,7-Dichloro-1,4-dihydrocyclopenta[\u003cem\u003eb\u003c/em\u003e]indol-3(2\u003cem\u003eH\u003c/em\u003e)-one (12a)\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eTo a solution of \u003cem\u003etert-\u003c/em\u003ebutyl 6,7-dichloro-3-oxo-2,3-dihydrocyclopenta[\u003cem\u003eb\u003c/em\u003e]indole-4(1\u003cem\u003eH\u003c/em\u003e)-carboxylate (571 mg, 1.68 mmol) in dichloromethane (25 mL), trifluoroacetic acid (5 mL) was added. The reaction mixture was stirred for an hour at room temperature. After removal of solvent, the mixture was diluted with ethyl acetate, and it was washed with 10% sodium hydroxide and brine. The organic layer was dried over sodium sulfate, and it was concentrated under reduced pressure to give product as a white solid (148 mg, 37%); \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (DMSO-d\u003csub\u003e6\u003c/sub\u003e) \u0026delta; 11.98 (bs, 1H), 8.06 (s, 1H), 7.64 (s, 1H), 3.02-2.99 (m, 2H), 2.90-2.87 (m, 2H).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e7,8-Dichloro-1,2,3,4-tetrahydrocyclopenta[\u003cem\u003eb\u003c/em\u003e]indol-3-amine (13)\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e7,8-Dichloro-1,4-dihydrocyclopenta[\u003cem\u003eb\u003c/em\u003e]indol-3(2\u003cem\u003eH\u003c/em\u003e)-one (100 mg, 0.42 mmol), ammonium acetate (647 mg, 8.40 mmol) and sodium cyanoborohydride (132 mg, 2.10 mmol) were dissolved in ethanol (10 mL). The reaction mixture was stirred for overnight at 60 \u003csup\u003eo\u003c/sup\u003eC. After removal of solvent, the reaction mixture was diluted with ethyl acetate. The organic layer was washed with 10% sodium hydroxide and brine, and it was dried over sodium sulfate. The organic layer was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-10% methanol/dichloromethane + 0.1% NH\u003csub\u003e4\u003c/sub\u003eOH) to give product as a white solid (41 mg, 41%); \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (MeOD) \u0026delta; 7.22 (d, \u003cem\u003eJ\u003c/em\u003e = 9 Hz, 1H), 7.09 (d, \u003cem\u003eJ\u003c/em\u003e = 9 Hz, 1H), 4.47-4.44 (m, 1H), 3.16-3.07 (m, 1H), 2.95-2.83 (m, 2H), 2.24-2.16 (m, 1H); LC/MS RT = 2.74 (M-H\u003csup\u003e-\u003c/sup\u003e: 239/241).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e7,8-Dichloro-1,4-dihydrocyclopenta[\u003cem\u003eb\u003c/em\u003e]indol-3(2\u003cem\u003eH\u003c/em\u003e)-one (13a)\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eTo a solution of \u003cem\u003etert-\u003c/em\u003ebutyl 7,8-dichloro-3-oxo-2,3-dihydrocyclopenta[\u003cem\u003eb\u003c/em\u003e]indole-4(1\u003cem\u003eH\u003c/em\u003e)-carboxylate (121 mg, 0.36 mmol) in dichloromethane (5 mL), trifluoroacetic acid (1 mL) was added. The reaction mixture was stirred for an hour at room temperature. After removal of solvent, the mixture was diluted with ethyl acetate, and it was washed with 10% sodium hydroxide and brine. The organic layer was dried over sodium sulfate, and it was concentrated under reduced pressure to give product as a white solid (27 mg, 31%); \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (CDCl\u003csub\u003e3\u003c/sub\u003e) \u0026delta; 9.61 (bs, 1H), 7.41 (d, \u003cem\u003eJ\u003c/em\u003e = 9 Hz, 1H), 7.36 (d, \u003cem\u003eJ\u003c/em\u003e = 9 Hz, 1H), 3.35-3.32 (m, 2H), 3.09-3.03 (m, 2H).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u003cem\u003etert-\u003c/em\u003e\u003c/strong\u003e\u003cstrong\u003eButyl 7,8-dichloro-3-oxo-2,3-dihydrocyclopenta[\u003cem\u003eb\u003c/em\u003e]indole-4(1\u003cem\u003eH\u003c/em\u003e)-carboxylate (12b) and\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u003cem\u003etert\u003c/em\u003e\u003c/strong\u003e\u003cstrong\u003e-Butyl 6,7-dichloro-3-oxo-2,3-dihydrocyclopenta[\u003cem\u003eb\u003c/em\u003e]indole-4(1\u003cem\u003eH\u003c/em\u003e)-carboxylate (13b)\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eTo a suspension of 3,4-dichloroaniline (8.10 g, 50.00 mmol) in water (30 mL), concentrated hydrochloric acid (12.5 mL) was slowly added at 0 \u003csup\u003eo\u003c/sup\u003eC. Then, a solution of sodium nitrite (3.45 g, 50.00 mmol) in water (35 mL) was slowly added at 0 \u003csup\u003eo\u003c/sup\u003eC. The mixture was stirred for 30 minutes at 0 \u003csup\u003eo\u003c/sup\u003eC. This freshly prepared diazonium salt solution was slowly added to a solution of 2-oxocyclopentane-1-carboxylic acid (6.40 g, 50.00 mmol) in concentrated hydrochloric acid (4.58 mL). The mixture was stirred for additional 30 minutes at 0 \u003csup\u003eo\u003c/sup\u003eC to give a yellow suspension. The suspension was filtered to give intermediate, (E)-2-(2-(3,4-dichlorophenyl)hydrazineylidene)-cyclopentan-1-one, as an orange solid.\u003c/p\u003e\n\u003cp\u003eThe intermediate was then dissolved in acetonitrile (50 mL), and 1.8M sulfuric acid was added. The reaction mixture was stirred for overnight at 75 \u003csup\u003eo\u003c/sup\u003eC. The resulting dark brown suspension was diluted with ethyl acetate, and it was washed with 10% NaOH and brine. The organic layer was dried over sodium sulfate, and it was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-20% ethyl acetate/hexane) to give a mixture of two regioisomers, 7,8-dichloro-1,4-dihydrocyclopenta[\u003cem\u003eb\u003c/em\u003e]indol-3(2\u003cem\u003eH\u003c/em\u003e)-one and 6,7-dichloro-1,4-dihydrocyclopenta[\u003cem\u003eb\u003c/em\u003e]indol-3(2\u003cem\u003eH\u003c/em\u003e)-one as a dark brown solid (493 mg, 10%).\u003c/p\u003e\n\u003cp\u003eTo a solution of a mixture of the two regioisomers (493 mg, 2.05 mmol) in tetrahydrofuran (25 mL), Boc anhydride (895 mg, 4.10 mmol) and DAMP (230 mg, 2.05 mmol) were added. The reaction mixture was stirred for 3 hours. The reaction mixture was diluted with ethyl acetate, and it was washed with saturated ammonium chloride, followed by brine. The organic layer was dried over sodium sulfate, and it was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-10% ethyl acetate/hexane) to give \u003cem\u003etert-\u003c/em\u003ebutyl 7,8-dichloro-3-oxo-2,3-dihydrocyclopenta[\u003cem\u003eb\u003c/em\u003e]indole-4(1\u003cem\u003eH\u003c/em\u003e)-carboxylate (121 mg, 17%). \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (CDCl\u003csub\u003e3\u003c/sub\u003e) \u0026delta; 8.15 (d, \u003cem\u003eJ\u003c/em\u003e = 9 Hz, 1H), 7.48 (d, \u003cem\u003eJ\u003c/em\u003e = 9 Hz, 1H), 3.26-3.22 (m, 2H), 3.02-2.99 (m, 2H), 1.68 (s, 9H). along with \u003cem\u003etert\u003c/em\u003e-butyl 6,7-dichloro-3-oxo-2,3-dihydrocyclopenta[\u003cem\u003eb\u003c/em\u003e]indole-4(1\u003cem\u003eH\u003c/em\u003e)-carboxylate (160 mg, 23%). \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (CDCl\u003csub\u003e3\u003c/sub\u003e) \u0026delta; 8.47 (s, 1H), 7.73 (s, 1H), 3.01-3.00 (m, 4H), 1.69 (s, 9H).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e1-(5-Bromo-1\u003cem\u003eH\u003c/em\u003e-indol-2-yl)ethan-1-amine (14)\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e1-(5-Bromo-1\u003cem\u003eH\u003c/em\u003e-indol-2-yl)ethan-1-one (95 mg, 0.40 mmol), ammonium acetate (308 mg, 4.00 mmol) and sodium cyanoborohydride (126 mg, 2.00 mmol) were dissolved in ethanol (5 mL). The mixture was stirred for overnight at 60 \u003csup\u003eo\u003c/sup\u003eC. The reaction mixture was diluted with ethyl acetate. The organic layer was washed with 10% NaOH and brine, and it was dried over Na\u003csub\u003e2\u003c/sub\u003eSO\u003csub\u003e4\u003c/sub\u003e. The organic layer was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-10% methanol/dichloromethane + 0.1% NH\u003csub\u003e4\u003c/sub\u003eOH) to give the product as a yellow oil (32 mg, 33%);\u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (DMSO-d\u003csub\u003e6\u003c/sub\u003e) \u0026delta; 11.07 (bs, 1H), 7.57 (d, \u003cem\u003eJ\u003c/em\u003e = 2 Hz, 1H), 7.24 (d, \u003cem\u003eJ\u003c/em\u003e = 9 Hz, 1H), 7.08 (dd, \u003cem\u003eJ\u0026nbsp;\u003c/em\u003e= 8 Hz, \u003cem\u003eJ\u003c/em\u003e = 2 Hz, 1H), 6.18 (s, 1H), 4.11-4.04 (m, 1H), 1.34 (d, \u003cem\u003eJ\u003c/em\u003e = 7 Hz, 3H); LC/MS RT = 2.50 (M-H\u003csup\u003e-\u003c/sup\u003e: 237/239).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e5-Bromo-N-methoxy-N-methyl-1\u003cem\u003eH\u003c/em\u003e-indole-2-carboxamide (14b)\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e5-Bromo-1\u003cem\u003eH\u003c/em\u003e-indole-2-carboxylic acid (1.0 g, 4.17 mmol), N,O-dimethylhydroxylamine hydrochloride (611 mg, 6.26 mmol), HOBt (563 mg, 4.17 mmol), EDC hydrochloride (1.68 g, 8.76 mmol) and triethylamine (2.32 mL, 16.68 mmol) were dissolved in anhydrous DMF (40 mL). The mixture was stirred for overnight at room temperature. The reaction mixture was diluted with ethyl acetate, and the organic layer was washed with water, saturated NH\u003csub\u003e4\u003c/sub\u003eCl, 10% NaOH and brine. The organic layer was dried over Na\u003csub\u003e2\u003c/sub\u003eSO\u003csub\u003e4\u003c/sub\u003e, and it was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-100% ethyl acetate/hexane) to give product as a white solid (800 mg, 68%); \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (CDCl\u003csub\u003e3\u003c/sub\u003e) \u0026delta; 9.33 (bs, 1H), 7.83 (s, 1H), 7.38 (dd, \u003cem\u003eJ\u003c/em\u003e = 9 Hz, \u003cem\u003eJ\u003c/em\u003e = 1 Hz, 1H), 7.31 (d, \u003cem\u003eJ\u003c/em\u003e = 9 Hz, 1H), 7.15 (s, 1H), 3.85 (s, 3H), 3.43 (s, 3H).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e1-(5-Bromo-1\u003cem\u003eH\u003c/em\u003e-indol-2-yl)ethan-1-one (14c)\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e5-Bromo-N-methoxy-N-methyl-1\u003cem\u003eH\u003c/em\u003e-indole-2-carboxamide (600 mg, 2.12 mmol) was dissolved in anhydrous tetrahydrofuran (50 mL). The mixture was cooled to -78 \u003csup\u003eo\u003c/sup\u003eC, then, a solution of 1.6 M methyllithium in diethyl ether (4.00 mL, 6.36 mmol) was added. The mixture was stirred for 2 hours at -78 \u003csup\u003eo\u003c/sup\u003eC. An additional solution of 1.6 M methyllithium in diethyl ether (4.00 mL, 6.36 mmol) was added. The mixture was stirred for an hour at -78 \u003csup\u003eo\u003c/sup\u003eC. The reaction was stopped by addition of water (10 mL). After removal of the solvent, the reaction mixture was diluted with ethyl acetate. The organic layer was washed with saturated NH\u003csub\u003e4\u003c/sub\u003eCl and brine, and it was dried over Na\u003csub\u003e2\u003c/sub\u003eSO\u003csub\u003e4\u003c/sub\u003e. The organic layer was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-20% ethyl acetate/hexane) to give product as a white solid (344 mg, 68%); \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (CDCl\u003csub\u003e3\u003c/sub\u003e) \u0026delta; 9.19 (bs, 1H), 7.85 (s, 1H), 7.42 (dd, \u003cem\u003eJ\u003c/em\u003e = 9 Hz, \u003cem\u003eJ\u003c/em\u003e = 2 Hz, 1H), 7.31 (d, \u003cem\u003eJ\u003c/em\u003e = 8 Hz, 1H), 7.12 (s, 1H), 2.60 (s, 3H).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e1-(1\u003cem\u003eH\u003c/em\u003e-indol-2-yl)ethan-1-amine (15).\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e1-(1\u003cem\u003eH\u003c/em\u003e-Indol-2-yl)ethan-1-one (100 mg, 0.63 mmol), ammonium acetate (486 mg, 6.30 mmol) and sodium cyanoborohydride (198 mg, 3.15 mmol) were dissolved in ethanol (10 mL). The mixture was stirred for overnight at 60 \u003csup\u003eo\u003c/sup\u003eC. The reaction mixture was diluted with ethyl acetate. The organic layer was washed with 10% NaOH and brine, and it was dried over Na\u003csub\u003e2\u003c/sub\u003eSO\u003csub\u003e4\u003c/sub\u003e. The organic layer was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (100% dichloromethane followed by 10% methanol/dichloromethane + 0.1% NH\u003csub\u003e4\u003c/sub\u003eOH) to give the product as a colorless oil (85 mg, 84%);\u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (CD\u003csub\u003e3\u003c/sub\u003eOD) \u0026delta; 7.44 (d, \u003cem\u003eJ\u003c/em\u003e = 8 Hz, 1H), 7.29 (d, \u003cem\u003eJ\u003c/em\u003e = 8 Hz, 1H), 7.03 (t, \u003cem\u003eJ\u003c/em\u003e = 8 Hz, 1H), 6.94 (t, \u003cem\u003eJ\u003c/em\u003e = 7 Hz, 1H), 6.29 (s, 1H), 4.24-4.17 (m, 1H), 1.51 (d, \u003cem\u003eJ\u003c/em\u003e = 7 Hz, 3H); LC/MS RT = 2.19 (M-H\u003csup\u003e-\u003c/sup\u003e: 159).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eN-Methoxy-N-methyl-1\u003cem\u003eH\u003c/em\u003e-indole-2-carboxamide (15b)\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e1\u003cem\u003eH\u003c/em\u003e-Indole-2-carboxylic acid (1.0 g, 6.21 mmol), N,O-dimethylhydroxylamine hydrochloride (909 mg, 9.32 mmol), HOBt (839 mg, 6.21 mmol), EDC hydrochloride (2.5 g, 13.04 mmol) and triethylamine (3.46 mL, 24.84 mmol) were dissolved in anhydrous DMF (40 mL). The mixture was stirred for overnight at room temperature. The reaction mixture was diluted with ethyl acetate, and the organic layer was washed with water, saturated NH\u003csub\u003e4\u003c/sub\u003eCl, 10% NaOH and brine. The organic layer was dried over Na\u003csub\u003e2\u003c/sub\u003eSO\u003csub\u003e4\u003c/sub\u003e, and it was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-30% ethyl acetate/hexane) to give product as a white solid (450 mg, 35%); \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (CDCl\u003csub\u003e3\u003c/sub\u003e) \u0026delta; 9.30 (bs, 1H), 7.70 (d, \u003cem\u003eJ\u003c/em\u003e = 8 Hz, 1H), 7.44 (d, \u003cem\u003eJ\u003c/em\u003e = 8 Hz, 1H), 7.31 (t, \u003cem\u003eJ\u003c/em\u003e = 8 Hz, 1H), 7.25 (s, 1H), 7.14 (t, \u003cem\u003eJ\u003c/em\u003e = 8 Hz, 1H), 3.85 (s, 3H), 3.44 (s, 3H).\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e1-(1\u003cem\u003eH\u003c/em\u003e-Indol-2-yl)ethan-1-one (15c)\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eN-Methoxy-N-methyl-1\u003cem\u003eH\u003c/em\u003e-indole-2-carboxamide (450 mg, 2.20 mmol) was dissolved in anhydrous tetrahydrofuran (50 mL). The mixture was cooled to -78\u003csup\u003eo\u003c/sup\u003eC, then, a solution of 1.6 M methyllithium in diethyl ether (4.13 mL, 6.60 mmol) was added. The mixture was stirred for 2 hours at -78\u003csup\u003eo\u003c/sup\u003eC. An additional solution of 1.6 M methyllithium in diethyl ether (4.13 mL, 6.60 mmol) was added. The mixture was stirred for an hour at -78\u003csup\u003eo\u003c/sup\u003eC. The reaction was stopped by addition of water (10 mL). After removal of the solvent, the reaction mixture was diluted with ethyl acetate. The organic layer was washed with saturated NH\u003csub\u003e4\u003c/sub\u003eCl and brine, and it was dried over Na\u003csub\u003e2\u003c/sub\u003eSO\u003csub\u003e4\u003c/sub\u003e. The organic layer was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-20% ethyl acetate/hexane) to give product as a white solid (290 mg, 83%); \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (CDCl\u003csub\u003e3\u003c/sub\u003e) \u0026delta; 9.37 (bs, 1H), 7.72 (d, \u003cem\u003eJ\u003c/em\u003e = 8 Hz, 1H), 7.45 (d, \u003cem\u003eJ\u003c/em\u003e = 8 Hz, 1H), 7.38-7.33 (m, 1H), 7.22-7.14 (m, 2H), 2.62 (s, 3H).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e1-(5-Fluoro-1\u003cem\u003eH\u003c/em\u003e-indol-2-yl)ethan-1-amine (16)\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e1-(5-Fluoro-1\u003cem\u003eH\u003c/em\u003e-indol-2-yl)ethan-1-one (100 mg, 0.56 mmol), ammonium acetate (432 mg, 5.60 mmol) and sodium cyanoborohydride (176 mg, 2.80 mmol) were dissolved in ethanol (10 mL). The mixture was stirred for overnight at 60 \u003csup\u003eo\u003c/sup\u003eC. The reaction mixture was acidified with 6N HCl, and it was washed with ethyl acetate. Then, the aqueous layer was basified with NaOH, and it was extracted with ethyl acetate. The organic layer was washed with brine, and it was dried over Na\u003csub\u003e2\u003c/sub\u003eSO\u003csub\u003e4\u003c/sub\u003e. The organic layer was concentrated under reduced pressure, and the residue was purified on an ISCO chromatograph (0-10% methanol/dichloromethane + 0.1% NH\u003csub\u003e4\u003c/sub\u003eOH) to give the product as a white solid (61 mg, 60%);\u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (CD\u003csub\u003e3\u003c/sub\u003eOD) \u0026delta; 7.25-7.21 (m, 1H), 7.10 (d, \u003cem\u003eJ\u003c/em\u003e = 8 Hz, 1H), 6.79 (t, \u003cem\u003eJ\u003c/em\u003e = 8 Hz, 1H), 6.28 (s, 1H), 4.22-4.17 (m, 1H), 1.49 (d, \u003cem\u003eJ\u003c/em\u003e = 6 Hz, 3H); LC/MS RT = 2.33 (M-H\u003csup\u003e-\u003c/sup\u003e: 177).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e5-Fluoro-N-methoxy-N-methyl-1\u003cem\u003eH\u003c/em\u003e-indole-2-carboxamide (16b)\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e5-Fluoro-1\u003cem\u003eH\u003c/em\u003e-indole-2-carboxylic acid (1.0 g, 5.58 mmol), N, O-dimethylhydroxylamine hydrochloride (816 mg, 8.37 mmol), HOBt (754 mg, 5.58 mmol), EDC hydrochloride (2.25 g, 11.7 mmol) and triethylamine (3.11 mL, 22.3 mmol) were dissolved in anhydrous DMF (50 mL). The mixture was stirred for overnight at room temperature. The reaction mixture was diluted with ethyl acetate, and the organic layer was washed with water, saturated NH\u003csub\u003e4\u003c/sub\u003eCl, 10% NaOH and brine. The organic layer was dried over Na\u003csub\u003e2\u003c/sub\u003eSO\u003csub\u003e4\u003c/sub\u003e, and it was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-50% ethyl acetate/hexane) to give product as a white solid (770 mg, 62%); \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (CDCl\u003csub\u003e3\u003c/sub\u003e) \u0026delta; 9.94 (bs, 1H), 7.41-7.36 (m, 1H), 7.32 (dd, \u003cem\u003eJ\u003c/em\u003e = 9 Hz, \u003cem\u003eJ\u003c/em\u003e = 2 Hz, 1H), 7.19 (s, 1H), 7.06 (td, \u003cem\u003eJ\u003c/em\u003e = 9 Hz, \u003cem\u003eJ\u003c/em\u003e = 3 Hz, 1H), 3.85 (s, 3H), 3.46 (s, 3H).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e1-(5-Fluoro-1\u003cem\u003eH\u003c/em\u003e-indol-2-yl)ethan-1-one (16c)\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e5-Fluoro-N-methoxy-N-methyl-1\u003cem\u003eH\u003c/em\u003e-indole-2-carboxamide (300 mg, 1.35 mmol) was dissolved in anhydrous tetrahydrofuran (25 mL). The mixture was cooled to -78 \u003csup\u003eo\u003c/sup\u003eC, then, a solution of 1.6 M methyllithium in diethyl ether (2.53 mL, 4.05 mmol) was added. The mixture was stirred for 2 hours at -78 \u003csup\u003eo\u003c/sup\u003eC. The reaction was stopped by addition of water (10 mL). After removal of the solvent, the reaction mixture was diluted with ethyl acetate. The organic layer was washed with saturated NH\u003csub\u003e4\u003c/sub\u003eCl and brine, and it was dried over Na\u003csub\u003e2\u003c/sub\u003eSO\u003csub\u003e4\u003c/sub\u003e. The organic layer was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-20% ethyl acetate/hexane) to give product as a white solid (220 mg, 92%); \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (CDCl\u003csub\u003e3\u003c/sub\u003e) \u0026delta; 9.18 (bs, 1H), 7.39-7.32 (m, 2H), 7.16-7.08 (m, 2H), 2.60 (s, 3H).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e1-(5-(Trifluoromethyl)-1\u003cem\u003eH\u003c/em\u003e-indol-2-yl)ethan-1-amine (17).\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e1-(5-Trifluoromethyl-1\u003cem\u003eH\u003c/em\u003e-indol-2-yl)ethan-1-one (100 mg, 0.44 mmol), ammonium acetate (339 mg, 4.40 mmol) and sodium cyanoborohydride (138 mg, 2.20 mmol) were dissolved in ethanol (10 mL). The mixture was stirred for overnight at 60 \u003csup\u003eo\u003c/sup\u003eC. The reaction mixture was acidified with 6N HCl, and it was washed with ethyl acetate. Then, the aqueous layer was basified with NaOH, and it was extracted with ethyl acetate. The organic layer was washed with brine, and it was dried over Na\u003csub\u003e2\u003c/sub\u003eSO\u003csub\u003e4\u003c/sub\u003e. The organic layer was concentrated under reduced pressure, and the residue was purified on an ISCO chromatograph (0-10% methanol/dichloromethane + 0.1% NH\u003csub\u003e4\u003c/sub\u003eOH) to give the product as a white solid (42 mg, 42%);\u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (DMSO-d\u003csub\u003e6\u003c/sub\u003e) \u0026delta; 11.34 (bs, 1H), 7.79 (s, 1H), 7.46 (d, \u003cem\u003eJ\u003c/em\u003e = 9 Hz, 1H), 7.27 (d, \u003cem\u003eJ\u003c/em\u003e = 8 Hz, 1H), 6.36 (s, 1H), 4.15-4.09 (m, 1H), 1.37 (d, \u003cem\u003eJ\u003c/em\u003e = 6 Hz, 3H); LC/MS RT = 2.50 (M-H\u003csup\u003e-\u003c/sup\u003e: 227).\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e5-(Trifluoromethyl)-1\u003cem\u003eH\u003c/em\u003e-indole-2-carboxylic acid (17a)\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e2-Iodo-4-(trifluoromethyl)aniline (1.0 g, 3.48 mmol), pyruvic acid (0.74 mL, 10.44 mmol), DABCO (1.17 g, 10.44 mmol) and Pd(OAc)\u003csub\u003e2\u003c/sub\u003e (79 mg, 0.35 mmol) were dissolved in anhydrous DMF (10 mL). The mixture was purged with nitrogen, and it was stirred for 4 hours at 110 \u003csup\u003eo\u003c/sup\u003eC. The reaction mixture was diluted with ethyl acetate, and the organic layer was washed with 1N HCl and brine. The organic layer was dried over Na\u003csub\u003e2\u003c/sub\u003eSO\u003csub\u003e4\u003c/sub\u003e, and it was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-50% ethyl acetate/hexane) to give product as a beige solid (484 mg, 54%); \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (CD\u003csub\u003e3\u003c/sub\u003eOD) \u0026delta; 11.66 (bs, 1H), 7.98 (s, 1H), 7.57 (d, \u003cem\u003eJ\u003c/em\u003e = 9 Hz, 1H), 7.47 (d, \u003cem\u003eJ\u003c/em\u003e = 9 Hz, 1H), 7.26 (s, 1H).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eN-Methoxy-N-methyl-5-(trifluoromethyl)-1\u003cem\u003eH\u003c/em\u003e-indole-2-carboxamide (17b)\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e5-(Trifluoromethyl)-1\u003cem\u003eH\u003c/em\u003e-indole-2-carboxylic acid (484 mg, 2.11 mmol), N, O-dimethyl-hydroxylamine hydrochloride (309 mg, 3.17 mmol), HOBt (285 mg, 2.11 mmol), EDC hydrochloride (849 mg, 4.43 mmol) and triethylamine (1.18 mL, 8.44 mmol) were dissolved in anhydrous DMF (50 mL). The mixture was stirred for overnight at room temperature. The reaction mixture was diluted with ethyl acetate, and the organic layer was washed with water, saturated NH\u003csub\u003e4\u003c/sub\u003eCl, 10% NaOH and brine. The organic layer was dried over Na\u003csub\u003e2\u003c/sub\u003eSO\u003csub\u003e4\u003c/sub\u003e, and it was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-50% ethyl acetate/hexane) to give product as a white solid (402 mg, 70%); \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (CDCl\u003csub\u003e3\u003c/sub\u003e) \u0026delta; 9.97 (bs, 1H), 8.01 (s, 1H), 7.53 (m, 2H), 7.32 (s, 1H), 3.86 (s, 3H), 3.45 (s, 3H).\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e1-(5-(Trifluoromethyl)-1\u003cem\u003eH\u003c/em\u003e-indol-2-yl)ethan-1-one (17c)\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eN-Methoxy-N-methyl-5-(trifluoromethyl)-1\u003cem\u003eH\u003c/em\u003e-indole-2-carboxamide (300 mg, 1.10 mmol) was dissolved in anhydrous tetrahydrofuran (20 mL). The mixture was cooled to -78 \u003csup\u003eo\u003c/sup\u003eC, then, a solution of 1.6 M methyllithium in diethyl ether (2.06 mL, 3.30 mmol) was added. The mixture was stirred for 2 hours at -78 \u003csup\u003eo\u003c/sup\u003eC. The reaction was stopped by addition of water (10 mL). After removal of the solvent, the reaction mixture was diluted with ethyl acetate. The organic layer was washed with saturated NH\u003csub\u003e4\u003c/sub\u003eCl and brine, and it was dried over Na\u003csub\u003e2\u003c/sub\u003eSO\u003csub\u003e4\u003c/sub\u003e. The organic layer was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-20% ethyl acetate/hexane) to give product as a white solid (197 mg, 80%); \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (CDCl\u003csub\u003e3\u003c/sub\u003e) \u0026delta; 9.40 (bs, 1H), 8.03 (s, 1H), 7.59-7.51 (m, 2H), 7.28 (s, 1H), 2.64 (s, 3H).\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e1-(5-Methoxy-1\u003cem\u003eH\u003c/em\u003e-indol-2-yl)ethan-1-amine (18).\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e1-(5-Methoxy-1\u003cem\u003eH\u003c/em\u003e-indol-2-yl)ethan-1-one (100 mg, 0.42 mmol), ammonium acetate (324 mg, 4.20 mmol) and sodium cyanoborohydride (132 mg, 2.10 mmol) were dissolved in ethanol (10 mL). The mixture was stirred for overnight at 60 \u003csup\u003eo\u003c/sup\u003eC. The reaction mixture was acidified with 6N HCl, and it was washed with ethyl acetate. Then, the aqueous layer was basified with NaOH, and it was extracted with ethyl acetate. The organic layer was washed with brine, and it was dried over Na\u003csub\u003e2\u003c/sub\u003eSO\u003csub\u003e4\u003c/sub\u003e. The organic layer was concentrated under reduced pressure, and the residue was purified on an ISCO chromatograph (0-10% methanol/dichloromethane + 0.1% NH\u003csub\u003e4\u003c/sub\u003eOH) to give the product as a white solid (58 mg, 58%);\u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (DMSO-d\u003csub\u003e6\u003c/sub\u003e) \u0026delta; 10.66 (bs, 1H), 7.15 (d, \u003cem\u003eJ\u003c/em\u003e = 9 Hz, 1H), 6.90 (d, \u003cem\u003eJ\u003c/em\u003e = 2 Hz, 1H), 6.61 (dd, \u003cem\u003eJ\u003c/em\u003e = 9 Hz, \u003cem\u003eJ\u003c/em\u003e = 2 Hz, 1H), 6.09 (s, 1H), 4.08-4.02 (m, 1H), 3.69 (s, 3H), 1.33 (d, \u003cem\u003eJ\u003c/em\u003e = 6 Hz, 3H); LC/MS RT = 2.17 (M-H\u003csup\u003e-\u003c/sup\u003e: 189).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eN,5-Dimethoxy-N-methyl-1\u003cem\u003eH\u003c/em\u003e-indole-2-carboxamide (18b)\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e5-Methoxy-1\u003cem\u003eH\u003c/em\u003e-indole-2-carboxylic acid (1.0 g, 5.23 mmol), N, O-dimethylhydroxylamine hydrochloride (766 mg, 7.85 mmol), HOBt (707 mg, 5.23 mmol), EDC hydrochloride (2.10 g, 10.98 mmol) and triethylamine (2.92 mL, 20.92 mmol) were dissolved in anhydrous DMF (50 mL). The mixture was stirred for overnight at room temperature. The reaction mixture was diluted with ethyl acetate, and the organic layer was washed with water, saturated NH\u003csub\u003e4\u003c/sub\u003eCl, 10% NaOH and brine. The organic layer was dried over Na\u003csub\u003e2\u003c/sub\u003eSO\u003csub\u003e4\u003c/sub\u003e, and it was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-50% ethyl acetate/hexane) to give product as a white solid (743 mg, 60%); \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (CDCl\u003csub\u003e3\u003c/sub\u003e) \u0026delta; 9.78 (bs, 1H), 7.35 (d, \u003cem\u003eJ\u003c/em\u003e = 9 Hz, 1H), 7.17 (s, 1H), 7.1 (d, \u003cem\u003eJ\u003c/em\u003e = 2 Hz, 1H), 6.98 (dd, \u003cem\u003eJ\u003c/em\u003e = 9 Hz, \u003cem\u003eJ\u003c/em\u003e = 2 Hz, 1H), 3.85 (s, 3H), 3.84 (s, 3H), 3.46 (s, 3H).\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e1-(5-Methoxy-1\u003cem\u003eH\u003c/em\u003e-indol-2-yl)ethan-1-one (18c)\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eN,5-Dimethoxy-N-methyl-1\u003cem\u003eH\u003c/em\u003e-indole-2-carboxamide (300 mg, 1.28 mmol) was dissolved in anhydrous tetrahydrofuran (20 mL). The mixture was cooled to -78\u003csup\u003eo\u003c/sup\u003eC, then, a solution of 1.6 M methyllithium in diethyl ether (2.40 mL, 3.84 mmol) was added. The mixture was stirred for 2 hours at -78\u003csup\u003eo\u003c/sup\u003eC. The reaction was stopped by addition of water (10 mL). After removal of the solvent, the reaction mixture was diluted with ethyl acetate. The organic layer was washed with saturated NH\u003csub\u003e4\u003c/sub\u003eCl and brine, and it was dried over Na\u003csub\u003e2\u003c/sub\u003eSO\u003csub\u003e4\u003c/sub\u003e. The organic layer was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-20% ethyl acetate/hexane) to give product as a white solid (180 mg, 74%); \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (CDCl\u003csub\u003e3\u003c/sub\u003e) \u0026delta; 8.91 (bs, 1H), 7.31 (d, \u003cem\u003eJ\u003c/em\u003e = 9 Hz, 1H), 7.12-7.08 (m, 2H), 7.03 (dd, \u003cem\u003eJ\u003c/em\u003e = 9 Hz, \u003cem\u003eJ\u003c/em\u003e = 3 Hz, 1H), 3.85 (s, 3H), 2.58 (s, 3H).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e1-(5-(Trifluoromethoxy)-1\u003cem\u003eH\u003c/em\u003e-indol-2-yl)ethan-1-amine (19)\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e1-(5-(Trifluoromethoxy)-1\u003cem\u003eH\u003c/em\u003e-indol-2-yl)ethan-1-one (100 mg, 0.41 mmol), ammonium acetate (316 mg, 4.10 mmol) and sodium cyanoborohydride (129 mg, 2.05 mmol) were dissolved in ethanol (10 mL). The mixture was stirred for overnight at 60 \u003csup\u003eo\u003c/sup\u003eC. The reaction mixture was acidified with 6N HCl, and it was washed with ethyl acetate. Then, the aqueous layer was basified with NaOH, and it was extracted with ethyl acetate. The organic layer was washed with brine, and it was dried over Na\u003csub\u003e2\u003c/sub\u003eSO\u003csub\u003e4\u003c/sub\u003e. The organic layer was concentrated under reduced pressure, and the residue was purified on an ISCO chromatograph (0-10% methanol/dichloromethane + 0.1% NH\u003csub\u003e4\u003c/sub\u003eOH) to give the product as a white solid (48 mg, 48%);\u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (DMSO-d\u003csub\u003e6\u003c/sub\u003e) \u0026delta; 11.15 (bs, 1H), 7.38 (s, 1H), 7.35 (d, \u003cem\u003eJ\u003c/em\u003e = 9 Hz, 1H), 6.94 (d, \u003cem\u003eJ\u003c/em\u003e = 8 Hz, 1H), 6.27 (s. 1H), 4.14-4.07 (m, 1H), 1.36 (d, \u003cem\u003eJ\u003c/em\u003e = 7 Hz, 3H); LC/MS RT = 2.55 (M-H\u003csup\u003e-\u003c/sup\u003e: 243).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e5-(Trifluoromethoxy)-1\u003cem\u003eH\u003c/em\u003e-indole-2-carboxylic acid (19a)\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e2-Iodo-4-(trifluoromethoxy)aniline (1.0 g, 3.30 mmol), pyruvic acid (0.70 mL, 9.90 mmol), DABCO (1.11 g, 9.90 mmol) and Pd(OAc)\u003csub\u003e2\u003c/sub\u003e (74 mg, 0.33 mmol) were dissolved in anhydrous DMF (10 mL). The mixture was purged with nitrogen, and it was stirred for 4 hours at 110 \u003csup\u003eo\u003c/sup\u003eC. The reaction mixture was diluted with ethyl acetate, and the organic layer was washed with 1N HCl and brine. The organic layer was dried over Na\u003csub\u003e2\u003c/sub\u003eSO\u003csub\u003e4\u003c/sub\u003e, and it was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-50% ethyl acetate/hexane) to give product as a beige solid (592 mg, 73%); \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (DMSO-d\u003csub\u003e6\u003c/sub\u003e) \u0026delta; 12.02 (bs, 1H), 7.64 (s, 1H), 7.49 (d, \u003cem\u003eJ\u003c/em\u003e = 9 Hz, 1H), 7.20 (d, \u003cem\u003eJ\u003c/em\u003e = 6 Hz, 1H), 7.13 (s, 1H).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eN-Methoxy-N-methyl-5-(trifluoromethoxy)-1\u003cem\u003eH\u003c/em\u003e-indole-2-carboxamide (19b)\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e5-(Trifluoromethoxy)-1\u003cem\u003eH\u003c/em\u003e-indole-2-carboxylic acid (592 mg, 2.41 mmol), N, O-dimethyl-hydroxylamine hydrochloride (353 mg, 3.62 mmol), HOBt (326 mg, 2.41 mmol), EDC hydrochloride (970 mg, 5.06 mmol) and triethylamine (1.18 mL, 8.44 mmol) were dissolved in anhydrous DMF (50 mL). The mixture was stirred for overnight at room temperature. The reaction mixture was diluted with ethyl acetate, and the organic layer was washed with water, saturated NH\u003csub\u003e4\u003c/sub\u003eCl, 10% NaOH and brine. The organic layer was dried over Na\u003csub\u003e2\u003c/sub\u003eSO\u003csub\u003e4\u003c/sub\u003e, and it was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-70% ethyl acetate/hexane) to give product as a white solid (378 mg, 54%); \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (CDCl\u003csub\u003e3\u003c/sub\u003e) \u0026delta; 9.33 (bs, 1H), 7.55 (s, 1H), 7.42 (d, \u003cem\u003eJ\u003c/em\u003e = 9 Hz, 1H), 7.24 (s, 1H), 7.18 (d, \u003cem\u003eJ\u003c/em\u003e = 9 Hz, 1H), 3.85 (s, 3H), 3.44 (s, 3H).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e1-(5-(Trifluoromethoxy)-1\u003cem\u003eH\u003c/em\u003e-indol-2-yl)ethan-1-one (19c)\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eN-Methoxy-N-methyl-5-(trifluoromethoxy)-1\u003cem\u003eH\u003c/em\u003e-indole-2-carboxamide (378 mg, 1.31 mmol) was dissolved in anhydrous tetrahydrofuran (20 mL). The mixture was cooled to -78 \u003csup\u003eo\u003c/sup\u003eC, then, a solution of 1.6 M methyllithium in diethyl ether (2.46 mL, 3.93 mmol) was added. The mixture was stirred for 2 hours at -78 \u003csup\u003eo\u003c/sup\u003eC. The reaction was stopped by addition of water (10 mL). After removal of the solvent, the reaction mixture was diluted with ethyl acetate. The organic layer was washed with saturated NH\u003csub\u003e4\u003c/sub\u003eCl and brine, and it was dried over Na\u003csub\u003e2\u003c/sub\u003eSO\u003csub\u003e4\u003c/sub\u003e. The organic layer was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-20% ethyl acetate/hexane) to give product as a white solid (259 mg, 81%); \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (CDCl\u003csub\u003e3\u003c/sub\u003e) \u0026delta; 9.35 (bs, 1H), 7.57 (s, 1H), 7.44 (d, \u003cem\u003eJ\u003c/em\u003e = 9 Hz, 1H), 7.26-7.20 (m, 2H), 2.62 (s, 3H).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e1-(4,6-Dichloro-1\u003cem\u003eH\u003c/em\u003e-indol-2-yl)ethan-1-amine (20).\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e1-(4,6-Dichloro-1\u003cem\u003eH\u003c/em\u003e-indol-2-yl)ethan-1-one (100 mg, 0.44 mmol), ammonium acetate (339 mg, 4.40 mmol) and sodium cyanoborohydride (138 mg, 2.20 mmol) were dissolved in ethanol (10 mL). The mixture was stirred for overnight at 60 \u003csup\u003eo\u003c/sup\u003eC. The reaction mixture was acidified with 6N HCl, and it was washed with ethyl acetate. Then, the aqueous layer was basified with NaOH, and it was extracted with ethyl acetate. The organic layer was washed with brine, and it was dried over Na\u003csub\u003e2\u003c/sub\u003eSO\u003csub\u003e4\u003c/sub\u003e. The organic layer was concentrated under reduced pressure, and the residue was purified on an ISCO chromatograph (0-10% methanol/dichloromethane + 0.1% NH\u003csub\u003e4\u003c/sub\u003eOH) to give the product as a white solid (55 mg, 54%);\u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (DMSO-d\u003csub\u003e6\u003c/sub\u003e) \u0026delta; 7.32 (s, 1H), 7.06 (s, 1H), 6.27 (s, 1H), 4.12-4.01 (m, 1H), 1.35 (d, \u003cem\u003eJ\u003c/em\u003e = 8 Hz, 3H); LC/MS RT = 2.61 (M-H\u003csup\u003e-\u003c/sup\u003e: 227/229).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e4,6-Dichloro-N-methoxy-N-methyl-1\u003cem\u003eH\u003c/em\u003e-indole-2-carboxamide (20b)\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e4,6-Dichloro-1\u003cem\u003eH\u003c/em\u003e-indole-2-carboxylic acid (500 mg, 2.17 mmol), N, O-dimethylhydroxylamine hydrochloride (423 mg, 4.34 mmol), HOBt (332 mg, 2.17 mmol), EDC hydrochloride (874 mg, 4.56 mmol) and triethylamine (1.32 mL, 8.68 mmol) were dissolved in anhydrous DMF (40 mL). The mixture was stirred for overnight at room temperature. The reaction mixture was diluted with ethyl acetate, and the organic layer was washed with water, saturated NH\u003csub\u003e4\u003c/sub\u003eCl, 10% NaOH and brine. The organic layer was dried over Na\u003csub\u003e2\u003c/sub\u003eSO\u003csub\u003e4\u003c/sub\u003e, and it was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-50% ethyl acetate/hexane) to give product as a white solid (505 mg, 85%); \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (DMSO-d\u003csub\u003e6\u003c/sub\u003e) \u0026delta; 12.10 (bs, 1H), 7.45 (d, \u003cem\u003eJ\u003c/em\u003e = 1 Hz, 1H), 7.24 (d, \u003cem\u003eJ\u003c/em\u003e = 1 Hz, 1H), 7.09 (s, 1H), 3.80 (s, 3H), 3.33 (s, 3H).\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e1-(4,6-Dichloro-1\u003cem\u003eH\u003c/em\u003e-indol-2-yl)ethan-1-one (20c)\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e4,6-Dichloro-N-methoxy-N-methyl-1\u003cem\u003eH\u003c/em\u003e-indole-2-carboxamide (500 mg, 1.83 mmol) was dissolved in anhydrous tetrahydrofuran (25 mL). The mixture was cooled to -78 \u003csup\u003eo\u003c/sup\u003eC, and, then, a solution of 1.6 M methyllithium in diethyl ether (3.43 mL, 5.49 mmol) was added. The mixture was stirred for 2 hours at -78 \u003csup\u003eo\u003c/sup\u003eC. The reaction was stopped by addition of water (10 mL). After removal of the solvent, the reaction mixture was diluted with ethyl acetate. The organic layer was washed with saturated NH\u003csub\u003e4\u003c/sub\u003eCl and brine, and it was dried over Na\u003csub\u003e2\u003c/sub\u003eSO\u003csub\u003e4\u003c/sub\u003e. The organic layer was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-20% ethyl acetate/hexane) to give product as a white solid (278 mg, 67%); \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (DMSO-d\u003csub\u003e6\u003c/sub\u003e) \u0026delta; 12.20 (bs, 1H), 7.41 (s, 1H), 7.39 (s, 1H), 7.27 (s, 1H), 2.57 (s, 3H).\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e1-(5,6-Difluoro-1\u003cem\u003eH\u003c/em\u003e-indol-2-yl)ethan-1-amine (21).\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e1-(5,6-Difluoro-1\u003cem\u003eH\u003c/em\u003e-indol-2-yl)ethan-1-one (150 mg, 0.77 mmol), ammonium acetate (594 mg, 7.70 mmol) and sodium cyanoborohydride (242 mg, 3.85 mmol) were dissolved in ethanol (10 mL). The mixture was stirred for overnight at 60 \u003csup\u003eo\u003c/sup\u003eC. The reaction mixture was acidified with 6N HCl, and it was washed with ethyl acetate. Then, the aqueous layer was basified with NaOH, and it was extracted with ethyl acetate. The organic layer was washed with brine, and it was dried over Na\u003csub\u003e2\u003c/sub\u003eSO\u003csub\u003e4\u003c/sub\u003e. The organic layer was concentrated under reduced pressure, and the residue was purified on an ISCO chromatograph (0-10% methanol/dichloromethane + 0.1% NH\u003csub\u003e4\u003c/sub\u003eOH) to give the product as a white solid (69 mg, 46%);\u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (DMSO-d\u003csub\u003e6\u003c/sub\u003e) \u0026delta; 11.49 (bs, 1H), 8.49 (bs, 2H), 7.57-7.50 (m, 1H), 7.44-7.38 (m, 1H), 6.50 (s, 1H), 4.58-4.51 (m, 1H), 1.58 (d, \u003cem\u003eJ\u003c/em\u003e = 6 Hz, 3H); LC/MS RT = 2.47 (M-H\u003csup\u003e-\u003c/sup\u003e: 195).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e5,6-Difluoro-N-methoxy-N-methyl-1\u003cem\u003eH\u003c/em\u003e-indole-2-carboxamide (21b)\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e5,6-Difluoro-1\u003cem\u003eH\u003c/em\u003e-indole-2-carboxylic acid (500 mg, 2.54 mmol), N, O-dimethylhydroxylamine hydrochloride (496 mg, 5.08 mmol), HOBt (389 mg, 2.54 mmol), EDC hydrochloride (1.02 g, 5.33 mmol) and triethylamine (1.54 mL, 10.2 mmol) were dissolved in anhydrous DMF (20 mL). The mixture was stirred for overnight at room temperature. The reaction mixture was diluted with ethyl acetate, and the organic layer was washed with water, saturated NH\u003csub\u003e4\u003c/sub\u003eCl, 10% NaOH and brine. The organic layer was dried over Na\u003csub\u003e2\u003c/sub\u003eSO\u003csub\u003e4\u003c/sub\u003e, and it was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-50% ethyl acetate/hexane) to give product as a white solid (367 mg, 60%); \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (CDCl\u003csub\u003e3\u003c/sub\u003e) \u0026delta; 9.33 (bs, 1H), 7.45-7.38 (m, 1H), 7.23-7.17 (m, 2H), 3.85 (s, 3H), 3.42 (s, 3H).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e1-(5,6-Difluoro-1\u003cem\u003eH\u003c/em\u003e-indol-2-yl)ethan-1-one (21c)\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e5,6-Difluoro-N-methoxy-N-methyl-1\u003cem\u003eH\u003c/em\u003e-indole-2-carboxamide (350 mg, 1.46 mmol) was dissolved in anhydrous tetrahydrofuran (20 mL). The mixture was cooled to -78 \u003csup\u003eo\u003c/sup\u003eC, then, a solution of 1.6 M methyllithium in diethyl ether (2.73 mL, 4.38 mmol) was added. The mixture was stirred for 2 hours at -78 \u003csup\u003eo\u003c/sup\u003eC. The reaction was stopped by addition of water (10 mL). After removal of the solvent, the reaction mixture was diluted with ethyl acetate. The organic layer was washed with saturated NH\u003csub\u003e4\u003c/sub\u003eCl and brine, and it was dried over Na\u003csub\u003e2\u003c/sub\u003eSO\u003csub\u003e4\u003c/sub\u003e. The organic layer was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-20% ethyl acetate/hexane) to give product as a white solid (207 mg, 73%); \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (CDCl\u003csub\u003e3\u003c/sub\u003e) \u0026delta; 9.24 (bs, 1H), 7.47-7.41 (m, 1H), 7.26-7.14 (m, 2H), 2.58 (s, 3H).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e1-(5-Bromo-1\u003cem\u003eH\u003c/em\u003e-indol-2-yl)-N-methylethan-1-amine (22).\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eTo a mixture of 1-(5-bromo-1\u003cem\u003eH\u003c/em\u003e-indol-2-yl)ethan-1-one (50 mg, 0.21 mmol) in ethanol (5 mL), a solution of 2.0 M methylamine in tetrahydrofuran (1.05 mL, 2.10 mmol) was added. The mixture was treated with catalytic amount of acetic acid, and it was stirred for an hour at 60 \u003csup\u003eo\u003c/sup\u003eC. Then, the mixture was treated with sodium cyanoborohydride (66 mg, 1.05 mmol), and it was stirred for overnight at 60 \u003csup\u003eo\u003c/sup\u003eC. The reaction mixture was acidified with 6N HCl, and it was washed with ethyl acetate. Then, the aqueous layer was basified with NaOH, and it was extracted with ethyl acetate. The organic layer was washed with brine, and it was dried over Na\u003csub\u003e2\u003c/sub\u003eSO\u003csub\u003e4\u003c/sub\u003e. The organic layer was concentrated under reduced pressure, and the residue was purified on an ISCO chromatograph (0-10% methanol/dichloromethane + 0.1% NH\u003csub\u003e4\u003c/sub\u003eOH) to give the product as a colorless oil (53 mg, 100%); \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (CDCl\u003csub\u003e3\u003c/sub\u003e) \u0026delta; 9.79 (bs, 1H), 7.67 (s, 1H), 7.27-7.26 (m, 2H), 6.41 (s, 1H), 4.35-4.32 (m, 1H), 2.43 (s, 3H), 1.71 (d, \u003cem\u003eJ\u003c/em\u003e = 7 Hz, 3H); LC/MS RT = 2.60 (M-H\u003csup\u003e-\u003c/sup\u003e: 251/253).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e1-(5-Bromo-1\u003cem\u003eH\u003c/em\u003e-indol-2-yl)-N,N-dimethylethan-1-amine (23).\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eTo a mixture of 1-(5-bromo-1\u003cem\u003eH\u003c/em\u003e-indol-2-yl)ethan-1-one\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e(75 mg, 0.32 mmol) in ethanol (10 mL), a solution of 2.0 M N,N-dimethylamine in tetrahydrofuran (1.60 mL, 3.20 mmol) was added. The mixture was treated with catalytic amount of acetic acid followed by sodium cyanoborohydride (101 mg, 1.69 mmol), and it was stirred for overnight at 60 \u003csup\u003eo\u003c/sup\u003eC. The reaction mixture was acidified with 6N HCl, and it was washed with ethyl acetate. Then, the aqueous layer was basified with NaOH, and it was extracted with ethyl acetate. The organic layer was washed with brine, and it was dried over Na\u003csub\u003e2\u003c/sub\u003eSO\u003csub\u003e4\u003c/sub\u003e. The organic layer was concentrated under reduced pressure, and the residue was purified on an ISCO chromatograph (0-10% methanol/dichloromethane + 0.1% NH\u003csub\u003e4\u003c/sub\u003eOH) to give the product as a colorless oil (12 mg, 14%); \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (CDCl\u003csub\u003e3\u003c/sub\u003e) \u0026delta; 8.81 (bs, 1H), 7.66 (s, 1H), 7.26-7.20 (m, 2H), 6.24 (s, 1H), 3.82-3.80 (m, 1H), 2.25 (s, 6H), 1.40 (d, \u003cem\u003eJ\u003c/em\u003e = 7 Hz, 3H); LC/MS RT = 2.62 (M-H\u003csup\u003e-\u003c/sup\u003e: 265/267).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e1-(5-Bromo-1-methyl-1\u003cem\u003eH\u003c/em\u003e-indol-2-yl)ethan-1-amine (24).\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e1-(5-Bromo-1-methyl-1\u003cem\u003eH\u003c/em\u003e-indol-2-yl)ethan-1-one (91 mg, 0.36 mmol), ammonium acetate (277 mg, 3.60 mmol) and sodium cyanoborohydride (113 mg, 1.80 mmol) were dissolved in ethanol (10 mL). Then, catalytic amount of acetic acid was added. The mixture was stirred for overnight at 60 \u003csup\u003eo\u003c/sup\u003eC. The reaction mixture was diluted with ethyl acetate. The organic layer was washed with 10% NaOH and brine, and it was dried over Na\u003csub\u003e2\u003c/sub\u003eSO\u003csub\u003e4\u003c/sub\u003e. The organic layer was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-10% methanol/dichloromethane + 0.1% NH\u003csub\u003e4\u003c/sub\u003eOH) to give the product as a white solid (57 mg, 63%);\u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (CD\u003csub\u003e3\u003c/sub\u003eOD) \u0026delta; 7.62 (d, \u003cem\u003eJ\u003c/em\u003e = 2 Hz, 1H), 7.28 (d, \u003cem\u003eJ\u003c/em\u003e = 9 Hz, 1H), 7.21 (dd, J = 9 Hz, \u003cem\u003eJ\u003c/em\u003e = 2 Hz, 1H), 6.44 (s, 1H), 4.44-4.38 (m, 1H), 3.75(s, 3H), 1.56 (d, \u003cem\u003eJ\u003c/em\u003e = 7 Hz, 3H); LC/MS RT = 2.49 (M-NH\u003csub\u003e2\u003c/sub\u003e\u003csup\u003e+\u003c/sup\u003e:236/238).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e1-(5-Bromo-1-methyl-1\u003cem\u003eH\u003c/em\u003e-indol-2-yl)ethan-1-one (24a)\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e1-(5-Bromo-1\u003cem\u003eH\u003c/em\u003e-indol-2-yl)ethan-1-one (100 mg, 0.42 mmol), K\u003csub\u003e2\u003c/sub\u003eCO\u003csub\u003e3\u003c/sub\u003e (116 mg, 0.84 mmol) and methyl iodide (52 \u0026micro;L, 0.84 mmol) were dissolved in anhydrous DMF (5 mL). The mixture was stirred for 3 hours at 60 \u003csup\u003eo\u003c/sup\u003eC. After the mixture was cooled to room temperature, it was diluted with ethyl acetate. The organic layer was washed with water and brine, and it was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-50% ethyl acetate/hexane) to give product as a white solid (91 mg, 86%); \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (CDCl\u003csub\u003e3\u003c/sub\u003e) \u0026delta; 7.82 (d, \u003cem\u003eJ\u003c/em\u003e = 2 Hz, 1H), 7.45 (dd, \u003cem\u003eJ\u003c/em\u003e = 9 Hz, \u003cem\u003eJ\u003c/em\u003e = 2 Hz, 1H), 7.27 (d, \u003cem\u003eJ\u003c/em\u003e = 9 Hz, 1H), 7.20 (s, 1H), 4.05 (s, 3H), 2.61 (s, 3H).\u003cstrong\u003e\u003cu\u003e\u0026nbsp;\u003c/u\u003e\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e(5-Bromo-1\u003cem\u003eH\u003c/em\u003e-indol-2-yl)methanamine (25).\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e5-Bromo-1\u003cem\u003eH\u003c/em\u003e-indole-2-carbaldehyde (55 mg, 0.25 mmol) was dissolved in ethanol (25 mL), and it was treated with ammonium acetate (193 mg, 2.50 mmol). The mixture was stirred for an hour at room temperature. Then, sodium cyanoborohydride (79 mg, 1.25 mmol) was added, and it was stirred for 3 hours at room temperature. The reaction mixture was diluted with ethyl acetate, and it was washed with saturated NaHCO\u003csub\u003e3\u003c/sub\u003e and brine. The organic layer was dried over Na\u003csub\u003e2\u003c/sub\u003eSO\u003csub\u003e4\u003c/sub\u003e, and it was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-10% methanol/dichloromethane + 0.1% NH\u003csub\u003e4\u003c/sub\u003eOH) to give (5-bromo-1\u003cem\u003eH\u003c/em\u003e-indol-2-yl)methanamine as a white solid (26 mg, 46%). \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (DMSO-d\u003csub\u003e6\u003c/sub\u003e) \u0026delta; 11.25 (bs, 1H), 7.72 (d, \u003cem\u003eJ\u003c/em\u003e = 2 Hz, 1H), 7.36 (d, \u003cem\u003eJ\u003c/em\u003e = 8 Hz, 1H), 7.19 (dd, \u003cem\u003eJ\u003c/em\u003e = 9 Hz, \u003cem\u003eJ\u003c/em\u003e = 2 Hz, 1H), 6.46 (s, 1H), 4.11 (s, 2H); LC/MS RT = 2.52 (M-H\u003csup\u003e-\u003c/sup\u003e: 223/225). along with bis((5-bromo-1\u003cem\u003eH\u003c/em\u003e-indol-2-yl)methyl)amine as a white solid (26 mg, 49%). \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (CDCl\u003csub\u003e3\u003c/sub\u003e) \u0026delta; 7.65 (s, 2H), 7.26-7.21 (m, 4H), 6.31 (s, 2H), 3.97 (s, 4H); LC/MS RT = 2.97 (M-H\u003csup\u003e-\u003c/sup\u003e: 430/432/434).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eMethyl 5-bromo-1\u003cem\u003eH\u003c/em\u003e-indole-2-carboxylat\u003c/strong\u003ee\u003cstrong\u003e\u0026nbsp;(25a)\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e5-Bromo-1\u003cem\u003eH\u003c/em\u003e-indole-2-carboxylic acid (1.5 g, 6.25 mmol) was dissolved in methanol (100 mL), and it was treated with catalytic amount of concentrated sulfuric acid. The mixture was refluxed overnight. After removal of solvent, it was diluted with ethyl acetate. The organic layer was washed with 10% NaOH and brine, and it was dried over Na\u003csub\u003e2\u003c/sub\u003eSO\u003csub\u003e4\u003c/sub\u003e. The organic layer was concentrated under reduced pressure, and the residue was purified on an ISCO chromatograph (0-50% ethyl acetate/hexane) to give product as a white solid (1.19 g, 75%); \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (CDCl\u003csub\u003e3\u003c/sub\u003e) \u0026delta; 8.97 (bs, 1H), 7.83 (s, 1H), 7.40 (dd, \u003cem\u003eJ\u003c/em\u003e = 9 Hz, \u003cem\u003eJ\u003c/em\u003e = 2 Hz, 1H), 7.30 (d, \u003cem\u003eJ\u003c/em\u003e = 9 Hz, 1H), 7.14 (s, 1H), 3.95 (s, 3H).\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e(5-Bromo-1\u003cem\u003eH\u003c/em\u003e-indol-2-yl)methanol (25b)\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eMethyl 5-bromo-1\u003cem\u003eH\u003c/em\u003e-indole-2-carboxylate (200 mg, 0.79 mmol) and lithium borohydride (86 mg, 3.95 mmol) were dissolved in anhydrous tetrahydrofuran (10 mL) at 0 \u003csup\u003eo\u003c/sup\u003eC, and it was stirred for overnight at room temperature. The reaction mixture was diluted with ethyl acetate, and it was washed with saturated NH\u003csub\u003e4\u003c/sub\u003eCl and brine. The organic layer was dried over Na\u003csub\u003e2\u003c/sub\u003eSO\u003csub\u003e4\u003c/sub\u003e, and it was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-50% ethyl acetate/hexane) to give product as a white solid (149 mg, 83 %); \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (CDCl\u003csub\u003e3\u003c/sub\u003e) \u0026delta; 8.37 (bs, 1H), 7.69 (s, 1H), 7.28-7.21 (m, 2H), 6.34 (d, \u003cem\u003eJ\u003c/em\u003e = 1 Hz, 1H), 4.84 (s, 2H).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e5-Bromo-1\u003cem\u003eH\u003c/em\u003e-indole-2-carbaldehyde (25c)\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e(5-Bromo-1\u003cem\u003eH\u003c/em\u003e-indol-2-yl)methanol (100 mg, 0.44 mmol) and Dess-Martin periodinane (280 mg, 0.66 mmol) were dissolved in dichloromethane (10 mL), and it was stirred for 15 minutes at room temperature. The reaction mixture was diluted with ethyl acetate, and it was washed with 10% sodium thiosulfate, saturated NaHCO\u003csub\u003e3\u003c/sub\u003e and brine. The organic layer was dried over Na\u003csub\u003e2\u003c/sub\u003eSO\u003csub\u003e4\u003c/sub\u003e, and it was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-10% ethyl acetate/hexane) to give product as a white solid (55 mg, 56%); \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (CDCl\u003csub\u003e3\u003c/sub\u003e) \u0026delta; 9.85 (s, 1H), 9.05 (bs, 1H), 7.90 (s, 1H), 7.47 (dd, \u003cem\u003eJ\u003c/em\u003e = 9 Hz, \u003cem\u003eJ\u003c/em\u003e = 2 Hz, 1H), 7.34 (d, \u003cem\u003eJ\u003c/em\u003e = 9 Hz, 1H), 7.21 (s, 1H).\u003cstrong\u003e\u003cu\u003e\u0026nbsp;\u003c/u\u003e\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eN\u003csup\u003e1\u003c/sup\u003e-(2-bromo-5,6,7,8,9,10-hexahydrocyclohepta[\u003cem\u003eb\u003c/em\u003e]indol-6-yl)propane-1,3-diamine (26).\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eTo a solution of \u003cem\u003etert\u003c/em\u003e-butyl (3-((2-bromo-5,6,7,8,9,10-hexahydrocyclohepta[\u003cem\u003eb\u003c/em\u003e]indol-6-yl)amino)propyl)carbamate (614 mg, 1.41 mmol) in methanol (3 mL), 4N HCl in dioxane (6 mL, 24 mmol) was added. The reaction mixture was stirred for 1.0 hour at room temperature. After removal of solvent, the reaction mixture was diluted with ethyl acetate. The organic layer was washed with 10% sodium hydroxide and brine, and it was dried over sodium sulfate. The organic layer was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-10% methanol/dichloromethane + 0.1% NH\u003csub\u003e4\u003c/sub\u003eOH) to give the product as yellow oil (202 mg, 43%); \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (CDCl\u003csub\u003e3\u003c/sub\u003e) \u0026delta; 10.02 (bs, 1H), 7.58 (s, 1H), 7.15-7.14 (m, 2H), 3.87-3.83 (m, 1H), 2.96-2.75 (m, 6H), 2.63-2.51 (m, 1H), 2.12-2.02 (m, 2H), 1.94-1.88 (m, 1H), 1.74-1.62 (m, 4H); LC/MS RT = 2.52 (M+H\u003csup\u003e+\u003c/sup\u003e: 336/338).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u003cem\u003etert-\u003c/em\u003e\u003c/strong\u003e\u003cstrong\u003eButyl (3-((2-bromo-5,6,7,8,9,10-hexahydrocyclohepta[\u003cem\u003eb\u003c/em\u003e]indol-6-yl)amino)propyl)carbamate (26a)\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e2-Bromo-7,8,9,10-tetrahydrocyclohepta[\u003cem\u003eb\u003c/em\u003e]indol-6(5\u003cem\u003eH\u003c/em\u003e)-one (500 mg, 1.80 mmol), N-Boc propylenediamine (941 mg, 5.40 mmol) and sodium cyanoborohydride (566 mg, 9.00 mmol) were dissolved in ethanol (20 mL). Catalytic amount of acetic acid was added. The reaction mixture was stirred for overnight at 60 \u003csup\u003eo\u003c/sup\u003eC. After removal of solvent, the reaction mixture was diluted with ethyl acetate. The organic layer was washed with 10% sodium hydroxide and brine, and it was dried over sodium sulfate. The organic layer was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-100% ethyl acetate/hexane) to give the product as yellow oil (614 mg, 78%); \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (CDCl\u003csub\u003e3\u003c/sub\u003e) \u0026delta; 9.51 (bs, 1H), 7.61 (s, 1H), 7.36-7.25 (m, 2H), 5.14-5.12 (m, 1H), 4.62-4.61 (m, 1H), 3.25-2.80 (m, 6H), 2.45-2.43 (m, 1H), 2.12-1.82 (m, 7H), 1.39 (s, 9H); LC/MS RT = 3.13 (M+H\u003csup\u003e+\u003c/sup\u003e: 436/438).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eN\u003csup\u003e1\u003c/sup\u003e-(7-bromo-1,2,3,4-tetrahydrocyclopenta[\u003cem\u003eb\u003c/em\u003e]indol-3-yl)propane-1,3-diamine (27).\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eTo a solution of \u003cem\u003etert\u003c/em\u003e-butyl (3-((7-bromo-1,2,3,4-tetrahydrocyclopenta[\u003cem\u003eb\u003c/em\u003e]indol-3-yl)amino)-propyl)carbamate (817 mg, 2.00 mmol) in methanol (3 mL), 4N HCl in dioxane (6 mL, 24 mmol) was added. The reaction mixture was stirred for an hour at room temperature. After removal of solvent, the reaction mixture was diluted with ethyl acetate. The organic layer was washed with 10% sodium hydroxide and brine, and it was dried over sodium sulfate. The organic layer was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-10% methanol/dichloromethane + 0.1% NH\u003csub\u003e4\u003c/sub\u003eOH) to give the product as a white solid (616 mg, 100%); \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (CDCl\u003csub\u003e3\u003c/sub\u003e) \u0026delta; 9.25 (bs, 1H), 7.58 (s, 1H), 7.26-7.18 (m, 2H), 4.37-4.33 (m, 1H), 2.88-2.65 (m, 8H), 2.12-2.04 (m, 1H), 2.04-1.97 (m, 2H); LC/MS RT = 2.50 (M+H\u003csup\u003e+\u003c/sup\u003e: 308/310).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u003cem\u003etert\u003c/em\u003e\u003c/strong\u003e\u003cstrong\u003e-Butyl (3-((7-bromo-1,2,3,4-tetrahydrocyclopenta[\u003cem\u003eb\u003c/em\u003e]indol-3-yl)amino)propyl)carbamate (27a)\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e7-Bromo-1,4-dihydrocyclopenta[\u003cem\u003eb\u003c/em\u003e]indol-3(2\u003cem\u003eH\u003c/em\u003e)-one (500 mg, 2.00 mmol), N-Boc propylenediamine (1.05 g, 6.00 mmol) and sodium cyanoborohydride (628 mg, 10.00 mmol) were dissolved in ethanol (20 mL). A catalytic amount of acetic acid was added. The reaction mixture was stirred for overnight at 60 \u003csup\u003eo\u003c/sup\u003eC. After removal of solvent, the reaction mixture was diluted with ethyl acetate. The organic layer was washed with 10% sodium hydroxide and brine, and it was dried over sodium sulfate. The organic layer was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-100% ethyl acetate/hexane) to give the product as a foamy yellow solid (817 mg, 100%); \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (CDCl\u003csub\u003e3\u003c/sub\u003e) \u0026delta; 9.16 (bs, 1H), 7.63 (s, 1H), 7.33-7.32 (m, 2H), 5.07-5.03 (m, 1H), 4.83 (m, 1H), 3.26-3.24 (m, 2H), 3.04-2.85 (m, 5H), 2.60-2.54 (m, 1H), 1.98-1.96 (m, 2H), 1.39 (s, 9H); LC/MS RT = 3.08 (M+H\u003csup\u003e+\u003c/sup\u003e: 408/410).\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eN\u003csup\u003e1\u003c/sup\u003e-(1-(5-Bromo-1\u003cem\u003eH\u003c/em\u003e-indol-2-yl)ethyl)propane-1,3-diamine (28).\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eTo a solution of \u003cem\u003etert-\u003c/em\u003ebutyl (3-((1-(5-bromo-1H-indol-2-yl)ethyl)amino)propyl)carbamate (817 mg, 2.06 mmol) in methanol (3 mL), 4N HCl in dioxane (6 mL, 24 mmol) was added. The reaction mixture was stirred for an hour at room temperature. After removal of solvent, the reaction mixture was diluted with ethyl acetate. The organic layer was washed with 10% sodium hydroxide and brine, and it was dried over sodium sulfate. The organic layer was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-10% methanol/dichloromethane + 0.1% NH\u003csub\u003e4\u003c/sub\u003eOH) to give the product as colorless oil (332 mg, 54%); \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (CDCl\u003csub\u003e3\u003c/sub\u003e) \u0026delta; 9.22 (bs, 1H), 7.65 (s, 1H), 7.26-7.20 (m, 3H), 6.24 (s, 1H), 4.05-4.01 (m, 1H), 2.83-2.54 (m, 4H), 1.67-1.60 (m, 2H), 1.45 (d, \u003cem\u003eJ\u003c/em\u003e = 6 Hz, 3H); LC/MS RT = 2.49 (M+H\u003csup\u003e+\u003c/sup\u003e: 296/298).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u003cem\u003etert-\u003c/em\u003e\u003c/strong\u003e\u003cstrong\u003eButyl (3-((1-(5-bromo-1\u003cem\u003eH\u003c/em\u003e-indol-2-yl)ethyl)amino)propyl)carbamate (28a)\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e1-(5-Bromo-1\u003cem\u003eH\u003c/em\u003e-indol-2-yl)ethan-1-one (500 mg, 2.10 mmol), N-Boc propylenediamine (1.10 g, 6.30 mmol) and sodium cyanoborohydride (314 mg, 10.50 mmol) were dissolved in ethanol (20 mL). Catalytic amount of acetic acid was added. The reaction mixture was stirred for overnight at 60 \u003csup\u003eo\u003c/sup\u003eC. After removal of solvent, the reaction mixture was diluted with ethyl acetate. The organic layer was washed with 10% sodium hydroxide and brine, and it was dried over sodium sulfate. The organic layer was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-100% ethyl acetate/hexane) to give the product as colorless oil (817 mg, 99%); \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (CDCl\u003csub\u003e3\u003c/sub\u003e) \u0026delta; 9.50 (bs, 1H), 7.68 (s, 1H), 7.37-7.26 (m, 3H), 6.43 (s, 1H), 5.04 (m, 1H), 4.35-4.33 (m, 1H), 3.30-3.28 (m, 2H), 2.80-2.76 (m, 2H), 1.85-1.80 (m, 2H), 1.75 (d, \u003cem\u003eJ\u003c/em\u003e = 7 Hz, 3H), 1.46 (s, 9H).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eN-(3-((1-(5-bromo-1\u003cem\u003eH\u003c/em\u003e-indol-2-yl)ethyl)amino)propyl)-4,5-dichlorothiophene-2-carboxamide (29).\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e5-Chlorofuran-2-carboxylic acid (120 mg, 0.82 mmol), EDC hydrochloric acid (157 mg, 0.82 mmol), HOBt (55 mg, 0.41 mmol) and DIPEA (0.23 mL, 1.26 mmol) were dissolved in DMF (5 mL). After 5 minutes of stirring, N\u003csup\u003e1\u003c/sup\u003e-(1-(5-bromo-1\u003cem\u003eH\u003c/em\u003e-indol-2-yl)ethyl)propane-1,3-diamine (120 mg, 0.41 mmol) was added, and it was stirred overnight at room temperature. The reaction mixture was diluted with ethyl acetate, and it was washed with 1N hydrochloric acid, 10% sodium hydroxide and brine. The organic layer was dried over sodium sulfate. The organic layer was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-10% methanol/dichloromethane + 0.1% NH\u003csub\u003e4\u003c/sub\u003eOH) to give the product as a white solid (54 mg, 31%); \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (CDCl\u003csub\u003e3\u003c/sub\u003e) \u0026delta; 9.02 (bs, 1H), 7.63 (s, 1H), 7.18 (m, 2H), 7.11 (d, \u003cem\u003eJ\u003c/em\u003e = 3 Hz, 1H), 7.05 (bs, 1H), 6.32 (d, \u003cem\u003eJ\u003c/em\u003e = 3 Hz, 1H), 6.24 (s, 1H), 4.03-3.98 (m, 1H), 3.76-3.69 (m, 1H), 3.51-3.35 (m, 1H), 2.73-2.68 (m, 1H), 2.60-2.54 (m, 1H), 1.72-1.69 (m, 2H), 1.49 (d, \u003cem\u003eJ\u003c/em\u003e = 6 Hz, 3H); LC/MS RT = 2.95 (M+H\u003csup\u003e+\u003c/sup\u003e: 424/426/428).\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eN-(3-((1-(5-bromo-1\u003cem\u003eH\u003c/em\u003e-indol-2-yl)ethyl)amino)propyl)-5-chlorothiophene-2-carboxamide (30).\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e5-Chlorothiophene-2-carboxylic acid (249 mg, 1.53 mmol), EDC hydrochloric acid (196 mg, 1.02 mmol), HOBt (69 mg, 0.51 mmol) and DIPEA (0.27 mL, 1.53 mmol) were dissolved in DMF (2 mL). After 5 minutes of stirring, N\u003csup\u003e1\u003c/sup\u003e-(1-(5-bromo-1\u003cem\u003eH\u003c/em\u003e-indol-2-yl)ethyl)propane-1,3-diamine (150 mg, 0.51 mmol) was added, and it was stirred overnight at room temperature. The reaction mixture was diluted with ethyl acetate, and it was washed with 1N hydrochloric acid, 10% sodium hydroxide and brine. The organic layer was dried over sodium sulfate. The organic layer was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-10% methanol/dichloromethane + 0.1% NH\u003csub\u003e4\u003c/sub\u003eOH) to give the product as a white solid (29 mg, 13%); \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (CDCl\u003csub\u003e3\u003c/sub\u003e) \u0026delta; 9.07 (bs, 1H), 7.64 (s, 1H), 7.26-7.06 (m, 2H), 6.79-6.78 (m, 2H), 6.26 (s, 1H), 4.05-4.00 (m, 1H), 3.66-3.60 (m, 1H), 3.51-3.39 (m, 1H), 2.75-2.71 (m, 1H), 2.62-2.54 (m, 1H) 1.73-1.70 (m, 2H), 1.51 (d, \u003cem\u003eJ\u003c/em\u003e = 7 Hz, 3H); LC/MS RT = 3.05 (M+H\u003csup\u003e+\u003c/sup\u003e: 440/442/444).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eN-(3-((1-(5-bromo-1\u003cem\u003eH\u003c/em\u003e-indol-2-yl)ethyl)amino)propyl)-4,5-dichlorothiophene-2-carboxamide (31).\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e4,5-Dichlorothiophene-2-carboxylic acid (162 mg, 0.82 mmol), EDC hydrochloric acid (157 mg, 0.82 mmol), HOBt (55 mg, 0.41 mmol) and DIPEA (0.23 mL, 1.26 mmol) were dissolved in DMF (5 mL). After 5 minutes of stirring, N\u003csup\u003e1\u003c/sup\u003e-(1-(5-bromo-1\u003cem\u003eH\u003c/em\u003e-indol-2-yl)ethyl)propane-1,3-diamine (120 mg, 0.41 mmol) was added, and it was stirred overnight at room temperature. The reaction mixture was diluted with ethyl acetate, and it was washed with 1N hydrochloric acid, 10% sodium hydroxide and brine. The organic layer was dried over sodium sulfate. The organic layer was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-10% methanol/dichloromethane + 0.1% NH\u003csub\u003e4\u003c/sub\u003eOH) to give the product as a white solid (63 mg, 32%); \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (CDCl\u003csub\u003e3\u003c/sub\u003e) \u0026delta; 8.88 (bs, 1H), 7.64 (s, 1H), 7.26-7.12 (m, 3H), 7.06 (bs, 1H), 6.26 (s, 1H), 4.04-3.98 (m, 1H), 3.65-3.58 (m, 1H), 3.49-3.40 (m, 1H), 2.75-2.69 (m, 1H), 2.63-2.57 (m, 1H) 1.74-1.68 (m, 2H), 1.49 (d, \u003cem\u003eJ\u003c/em\u003e = 7 Hz, 3H); LC/MS RT = 3.05 (M+H\u003csup\u003e+\u003c/sup\u003e: 474/476/478).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eN-(3-((1-(5-bromo-1\u003cem\u003eH\u003c/em\u003e-indol-2-yl)ethyl)amino)propyl)-3,4-dichlorobenzamide (32).\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e3,4-Dichlorobenzoic acid (32 mg, 0.17 mmol), EDC hydrochloric acid (65 mg, 0.34 mmol), HOBt (23 mg, 0.17 mmol) and DIPEA (0.09 mL, 0.51 mmol) were disolved in dichloromethane (5 mL). After 5 minutes of stirring, N\u003csup\u003e1\u003c/sup\u003e-(1-(5-bromo-1\u003cem\u003eH\u003c/em\u003e-indol-2-yl)ethyl)propane-1,3-diamine (50 mg, 0.17 mmol) was added, and it was stirred overnight at room temperature. The reaction mixture was diluted with ethyl acetate, and it was washed with 1N hydrochloric acid, 10% sodium hydroxide and brine. The organic layer was dried over sodium sulfate. The organic layer was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-10% methanol/dichloromethane + 0.1% NH\u003csub\u003e4\u003c/sub\u003eOH) to give the product as colorless oil (14 mg, 18%); \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (CDCl\u003csub\u003e3\u003c/sub\u003e) \u0026delta; 9.97 (bs, 1H), 7.88 (d, \u003cem\u003eJ\u003c/em\u003e = 2 Hz, 1H), 7.64 (s, 1H), 7.53 (dd, \u003cem\u003eJ\u003c/em\u003e = 8 Hz, \u003cem\u003eJ\u003c/em\u003e = 2Hz, 2H), 7.42 (d, \u003cem\u003eJ\u003c/em\u003e = 8 Hz, 1H), 7.25-7.18 (m, 2H), 4.35-4.28 (m, 1H), 3.63-3.48 (m, 2H), 2.81-2.69 (m, 2H), 1.94-1.90 (m, 2H), 1.72 (d, J = 7 Hz, 3H); LC/MS RT = 3.12 (M+H\u003csup\u003e+\u003c/sup\u003e: 468/470/472).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eN-(3-((1-(5-bromo-1\u003cem\u003eH\u003c/em\u003e-indol-2-yl)ethyl)amino)propyl)-cyclohexanecarboxamide (33)\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eCyclohexane carboxylic acid (22 mg, 0.17 mmol), EDC hydrochloric acid (65 mg, 0.34 mmol), HOBt (23 mg, 0.17 mmol) and DIPEA (0.09 mL, 0.51 mmol) were dissolved in dichloromethane (5 mL). After 5 minutes of stirring, N\u003csup\u003e1\u003c/sup\u003e-(1-(5-bromo-1\u003cem\u003eH\u003c/em\u003e-indol-2-yl)ethyl)propane-1,3-diamine (50 mg, 0.17 mmol) was added, and it was stirred overnight at room temperature. The reaction mixture was diluted with ethyl acetate, and it was washed with 1N hydrochloric acid, 10% sodium hydroxide and brine. The organic layer was dried over sodium sulfate. The organic layer was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-10% methanol/dichloromethane + 0.1% NH\u003csub\u003e4\u003c/sub\u003eOH) to give the product as colorless oil (55 mg, 80%); \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz) (CDCl\u003csub\u003e3\u003c/sub\u003e) \u0026delta; 9.28 (bs, 1H), 7.64 (d, \u003cem\u003eJ\u003c/em\u003e = 2 Hz, 1H), 7.26 (d, \u003cem\u003eJ\u003c/em\u003e = 9 Hz, 1H), 7.19 (dd, \u003cem\u003eJ\u003c/em\u003e = 8 Hz, \u003cem\u003eJ\u003c/em\u003e = 2 Hz, 1H), 6.23 (d, \u003cem\u003eJ\u003c/em\u003e = 1 Hz, 1H), 5.68 (bs, 1H), 4.00-3.96 (m, 1H), 3.54-3.47 (m, 1H), 3.27-3.20 (m, 1H), 2.62-2.54 (m, 1H), 2.49-2.41 (m, 1H), 2.04-2.00 (m, 1H), 1.79-1.57 (m, 6H), 1.47 (d, \u003cem\u003eJ\u003c/em\u003e = 7 Hz, 3H), 1.38-1.14 (m, 6H); LC/MS RT = 2.88 (M+H\u003csup\u003e+\u003c/sup\u003e: 406/408).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e4-((1-(5-Bromo-1\u003cem\u003eH\u003c/em\u003e-indol-2-yl)ethyl)amino)-N-(3,4-dichlorophenyl)butanamide (34).\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eTo a solution of 1-(5-bromo-1\u003cem\u003eH\u003c/em\u003e-indol-2-yl)ethan-1-one (243 mg, 1.02 mmol) in ethanol (10 mL), 4-amino-\u003cem\u003eN\u003c/em\u003e-(3,4-dichlorophenyl)butanamide (253 mg, 1.02 mmol) and sodium cyanoborohydride (320 mg, 5.10 mmol) were added. The reaction mixture was stirred for 60 \u003csup\u003eo\u003c/sup\u003eC for 24 hours. The mixture was diluted with ethyl acetate, and the organic layer was washed with 10% sodium hydroxide, saturated ammonium chloride, and brine. The organic layer was then dried over sodium sulfate, and it was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (10% methanol/dichloromethane + 1% NH\u003csub\u003e4\u003c/sub\u003eOH) to give product as colorless oil (73 mg, 15%); \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz, DMSO-d\u003csub\u003e6\u003c/sub\u003e) d 11.30 (bs, 1H), 10.25 (bs, 1H), 8.00-7.99 (m, 1H), 7.72 (s, 1H), 7.60-7.55 (m, 1H), 7.49-7.44 (m, 1H), 7.38-7.36 (m, 1H), 7.23-7.21 (m, 1H), 6.48 (s, 1H), 4.33 (m, 1H), 2.76-2.60 (m, 2H), 2.42 (m, 2H), 1.84 (m, 2H), 1.57 (m, 3H).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u003cem\u003etert\u003c/em\u003e\u003c/strong\u003e\u003cstrong\u003e-Butyl (4-((3,4-dichlorophenyl)amino)-4-oxobutyl)carbamate\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eTo a solution of 4-((\u003cem\u003etert\u003c/em\u003e-butoxy carbonyl)amino)butanoic acid (1.0 g, 4.9 mmol) in dichloromethane (50 mL), EDC hydrochloric acid (1.98 g, 10.3 mmol), HOBt (665 mg, 4.9 mmol) and DIPEA (1.80 mL, 10.33 mmol) were added. After the mixture stirred for 5 minutes, 3,4-dichloroaniline (797 mg, 4.92 mmol) was added. The mixture was then stirred for overnight at room temperature. The mixture was then diluted with dichloromethane, and the organic layer was washed with 1N hydrochloric acid, 10% sodium hydroxide, saturated ammonium chloride and brine. The organic layer was then dried over sodium sulfate, and it was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-30% ethyl acetate/hexane) to give product as a white solid (757 mg, 44%); \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz, CDCl\u003csub\u003e3\u003c/sub\u003e) d 9.47 (bs, 1H), 7.92 (s, 1H), 7.48 (d, \u003cem\u003eJ\u003c/em\u003e = 9 Hz, 1H), 7.37 (d, \u003cem\u003eJ\u003c/em\u003e = 9 Hz, 1H), 4.87 (m, 1H), 3.27-3.24 (m, 2H), 2.41-2.37 (m, 2H), 1.89-1.87 (m, 2H), 1.49 (s, 9H).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e4-Amino-\u003cem\u003eN\u003c/em\u003e-(3,4-dichlorophenyl)butanamide\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eTo a solution of \u003cem\u003etert\u003c/em\u003e-butyl (4-((3,4-dichlorophenyl)amino)-4-oxobutyl)carbamate (500 mg, 1.44 mmol) in methanol (0.36 mL), 4N hydrochloric acid (3.60 mL, 14.40 mmol) was added. The reaction mixture was stirred for 2 hours at room temperature. The mixture was diluted with ethyl acetate, and the organic layer was washed with 10% sodium hydroxide saturated ammonium chloride and brine. The organic layer was then dried over sodium sulfate, and it was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-100% ethyl acetate/hexane followed by 10% methanol/dichloromethane + 1% NH\u003csub\u003e4\u003c/sub\u003eOH) to give product as colorless oil (253 mg, 71%); \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz, CDCl\u003csub\u003e3\u003c/sub\u003e) d 9.85 (bs, 1H),7.78 (d, \u003cem\u003eJ\u003c/em\u003e = 2Hz, 1H), 7.38-7.32 (m, 2H), 2.90-2.86 (m, 2H), 2.54-2.49 (m, 2H), 1.80-1.84 (m, 2H), 1.64 (bs, 2H).\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eN-(1-(5-Bromo-1\u003cem\u003eH\u003c/em\u003e-indol-2-yl)ethyl)-4-(3,4-dichlorophenoxy)butan-1-amine (35).\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eTo a solution of 1-(5-bromo-1\u003cem\u003eH\u003c/em\u003e-indol-2-yl)ethan-1-one (190 mg, 0.80 mmol) in ethanol (10 mL), 4-(3,4-dichlorophenoxy)butan-1-amine (187 mg, 0.80 mmol) and sodium cyanoborohydride (251 mg, 4.00 mmol) were added. The reaction mixture was stirred for 60 \u003csup\u003eo\u003c/sup\u003eC for 24 hours. The mixture was diluted with ethyl acetate, and the organic layer was washed with 10% sodium hydroxide, saturated ammonium chloride and brine. The organic layer was then dried over sodium sulfate, and it was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (10% methanol/dichloromethane + 1% NH\u003csub\u003e4\u003c/sub\u003eOH) to give product as colorless oil (47 mg, 13%); \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz, CDCl\u003csub\u003e3\u003c/sub\u003e) d 9.11 (bs, 1H), 7.71 (s, 1H), 7.31-7.28 (m, 3H), 6.91 (s, 1H), 6.62 (d, \u003cem\u003eJ\u0026nbsp;\u003c/em\u003e= 9 Hz, 1H), 6.44 (s, 1H), 4.11-4.39 (m, 1H), 3.87 (m, 2H), 2.78 (m, 2H), 1.81 (m, 4H), 1.72 (d, \u003cem\u003eJ\u003c/em\u003e = 7 Hz, 3H).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e2-(4-(3,4-Dichlorophenoxy)butyl)isoindoline-1,3-dione\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eTo a solution of 2-(4-bromobutyl)isoindoline-1,3-dione (500 mg, 1.77 mmol) in DMF (5 mL), 3,4-dichlorophenol (289 mg, 1.77 mmol) and potassium carbonate (245 mg, 1.77 mmol) were added. The reaction mixture was stirred for 5 hours at room temperature. The mixture was diluted with ethyl acetate, the organic layer was washed with water and brine. The organic layer was then dried over sodium sulfate, and it was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-30% ethyl acetate/hexane) to give product as colorless oil (545 mg, 84%); \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz, CDCl\u003csub\u003e3\u003c/sub\u003e) d 7.89-7.86 (m, 2H), 7.76-7.73 (m, 2H), 7.29 (d, \u003cem\u003eJ\u003c/em\u003e = 9 Hz, 1H), 6.98 (d, \u003cem\u003eJ\u003c/em\u003e = 3 Hz, 1H), 6.75 (dd, \u003cem\u003eJ\u003c/em\u003e = 9 Hz, \u003cem\u003eJ\u003c/em\u003e = 3 Hz, 1H), 4.00-3.96 (m, 2H), 3.81-3.77 (m, 2H), 1.89-1.87 (m, 4H).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e4-(3,4-Dichlorophenoxy)butan-1-amine\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eTo a solution of 2-(4-(3,4-dichlorophenoxy)butyl)isoindoline-1,3-dione (545 mg, 1.50 mmol) in methanol (15 mL), hydrazine monohydrate (0.15 mL, 3.00 mmol) was added at room temperature. The reaction mixture was stirred for 2 hours at 60 \u003csup\u003eo\u003c/sup\u003eC. The white suspension was formed, and the suspension was filtered. The suspension was concentrated under reduced pressure, and the residue was diluted with ethyl acetate. The organic layer was washed with 10% sodium hydroxide and brine. The organic layer was then dried over sodium sulfate, and it was concentrated under reduced pressure. The residue was purified on an ISCO chromatograph (0-100% ethyl acetate/hexane followed by 10% methanol/dichloromethane + 1% NH\u003csub\u003e4\u003c/sub\u003eOH) to give product as colorless oil (187 mg, 53%); \u003csup\u003e1\u003c/sup\u003eH NMR (300 MHz, CDCl\u003csub\u003e3\u003c/sub\u003e) d 7.31 (d, \u003cem\u003eJ\u003c/em\u003e = 9 Hz, 1H), 6.99 (d, \u003cem\u003eJ\u003c/em\u003e = 3 Hz, 1H), 6.75 (dd, \u003cem\u003eJ\u003c/em\u003e = 9 Hz, \u003cem\u003eJ\u003c/em\u003e = 3 Hz, 1H), 3.97-3.92 (m, 2H), 2.80-2.75 (m, 2H), 1.85-1.80 (m, 2H), 1.66-1.58 (m, 2H), 1.23 (bs, 2H).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eIntrinsic MIC Assays\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eMIC assays were conducted in accordance with Clinical and Laboratory Standards Institute (CLSI) guidelines for broth microdilution. A 96-well plate containing cation-adjusted Mueller-Hinton (CAMH) broth with 2-fold serial dilution of compounds was inoculated with log-phase bacteria at 5x10\u003csup\u003e5\u003c/sup\u003e CFU/mL. The final volume in each well was 100 \u0026micro;L. Each compound was tested in duplicate. The microtiter plates were incubated in an aerobic environment for 18 hours at 37 \u0026deg;C. Then the bacterial growth was tested by reading the plate with a VersaMax plate reader (Molecular Devices, Inc.) at 600 nm. The MIC was defined as the lowest compound concentration that inhibited 90% of bacterial growth.\u003c/p\u003e\n\u003cp\u003eThe intrinsic MIC of the experimental EPIs was tested with the method described above. The 2-fold serial dilution begins with 100 \u0026micro;g/mL of tested compound in the first column of the 96-well plates. The following Gram-negative bacterial strains were included in these assays:\u003c/p\u003e\n\u003cp\u003e\u003cem\u003eEscherichia coli\u003c/em\u003e ATCC 25922\u003c/p\u003e\n\u003cp\u003e\u003cem\u003eKlebsiella pneumoniae\u003c/em\u003e ATCC 13883 and ATCC 10031\u003c/p\u003e\n\u003cp\u003e\u003cem\u003ePseudomonas aeruginosa\u003c/em\u003e PAO1\u003c/p\u003e\n\u003cp\u003e\u003cem\u003eAcinetobacter baumannii\u003c/em\u003e ATCC 19606\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eMIC Assays in the Presence of a Bacterial Efflux Inhibitor\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe EPI assay for the purposes of these studies represents a MIC assay in which the MIC of the antibiotic against the bacteria is tested in the presence of an experimental efflux pump inhibitor (EPI), N-(((2S,4R)-4-(aminomethyl)pyrrolidin-2-yl)methyl)-6-(4-fluorophenyl)-1H-indole-2-carboxamide. The highest concentration of the EPI present in the assay typically is \u0026frac12; of the intrinsic MIC of the compound. If the intrinsic MIC of the EPI is greater than 100 \u0026micro;g/mL, the EPI assay was tested with 50 \u0026micro;g/mL. Using serial dilutions of the EPI, its enhancement of antibiotic activity was then evaluated. The relative EPI activity was decided by comparing the MIC of the antibiotic in the presence of the EPI compound with the intrinsic MIC of the antibiotic alone. For comparative purposes, we used this EPI at concentration of 12.5 \u0026micro;g/mL against varying concentration of our test compounds.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eATPase Assays with \u003cem\u003eE. coli\u003c/em\u003e MreB (EcMreB)\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe cloning and expression of\u003cem\u003e\u0026nbsp;E. coli\u003c/em\u003e MreB as well as all ATPase assays were conducted as described previously (23).\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eNotes\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors declare the following competing financial interest(s): \u0026nbsp;Dr. LaVoie and Dr. Pilch are co-founders of TAXIS Pharmaceuticals and Dr. Parhi is a shareholder and therefore have a financial interest in the company.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAcknowledgement\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThis research was supported in part by Research Agreement between Rutgers University and TAXIS Pharmaceuticals, Inc.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\n \u003cli\u003eFigge, RM, Divakaruni, AV, Gober, JW. MreB, the cell shape-determining bacterial actin homologue, co-ordinates cell wall morphogenesis in \u003cem\u003eCaulobacter crescentus\u003c/em\u003e. Mol. Microbiol. 2004;\u003cem\u003e51\u003c/em\u003e(5): 1321-1332.\u003c/li\u003e\n \u003cli\u003eStrahl, H, B\u0026uuml;rmann, F, Hamoen. LW. The actin homologue MreB organizes the bacterial cell membrane. Nat. Commun.\u003cem\u003e\u0026nbsp;\u003c/em\u003e2013;\u003cem\u003e5\u003c/em\u003e(3442): 1-11.\u003c/li\u003e\n \u003cli\u003eWang, H, Xie, L, Luo, H, Xie, J. Bacterial cytoskeleton and implications for new antibiotic targets. J. Drug Target. 2016;2\u003cem\u003e4\u003c/em\u003e(5): 392-398.\u003cstrong\u003e\u003cem\u003e\u003c/em\u003e\u003c/strong\u003e\u003c/li\u003e\n \u003cli\u003eBusiek, K, Margolin, W. Bacterial Actin and Tubulin Homologs in Cell Growth and Division. Curr. Biol\u003cem\u003e.\u003c/em\u003e 2015;\u003cem\u003e25\u003c/em\u003e(6); R243-R254.\u003c/li\u003e\n \u003cli\u003eFoss, MH, Eun, Y-J, Weibel, DB. Chemical-biological studies of subcellular organization in bacteria. Biochemistry 2011;\u003cem\u003e50\u003c/em\u003e:7719-7734.\u003c/li\u003e\n \u003cli\u003eWhite, CL, Gober, JW. MreB: pilot or passenger of cell wall synthesis? Trends Microbiol\u003cem\u003e.\u003c/em\u003e 2012; \u003cem\u003e20\u003c/em\u003e(2), 74-79.\u003c/li\u003e\n \u003cli\u003eFenton, AK, Gerdes, K. Direct Interaction of FtsZ and MreB is required for septum synthesis and cell division in \u003cem\u003eEscherichia coli\u003c/em\u003e.\u003cem\u003e\u0026nbsp;\u003c/em\u003eEMBO J.\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e2013;\u003cem\u003e32\u003c/em\u003e(13): 1953-1965.\u003c/li\u003e\n \u003cli\u003eVollmer, W. The prokaryotic cytoskeleton: a putative target for inhibitors and antibiotics? Appl. Microbiol. Biotechnol. 2006;73: 37-47.\u003c/li\u003e\n \u003cli\u003eNoguchi, N, Yanagimoto, K, Nakaminami, H, Wakabayashi, M, Iwai, N, Wachi, M, Sasatsu, M. Anti-infectious effect of S-benzylisothiourea compound A22, which inhibits the actin-like protein, MreB, in \u003cem\u003eShigella flexneri\u003c/em\u003e. Biol. Pharm. Bull. 2008;\u003cem\u003e31\u003c/em\u003e(7): 1327-1332.\u003c/li\u003e\n \u003cli\u003eBarker, CA, Allison, SE, Zlitni, S, Nguyen, ND, Das, R, Melacini, G, Capretta, AA, Brown, ED. Degradation of MAC13243 and studies of the interaction of resulting thiourea compounds with the lipoprotein targeting chaperone LolA. Bioorg. Med. Chem. Lett. 2013;\u003cem\u003e\u0026nbsp;\u003c/em\u003e23: 2426-2431.\u003c/li\u003e\n \u003cli\u003eBonez, PC, Ramos, AP, Nascimento, K, Copetti, PM, Souza, ME, Rossi, GG, Agertt, VA, Sagrillo, MR, Santos, RCV, Campos, MMA. Antibacterial, cyto and genotoxic activities of A22 compound ((S-3, 4-dichlorobenzyl) isothiourea hydrochloride). Microb. Pathog. 2016;99: 14-18.\u003c/li\u003e\n \u003cli\u003eIwai, N, Fujii, T, Nagura, H, Wachi, M, Kitazume, T. Structure-activity relationship study of the bacterial actin-like protein MreB inhibitors: Effects of substitution of benzyl group in S-benzylisothiourea. Biosci. Biotechnol. Biochem. 2007;\u003cem\u003e71\u003c/em\u003e(1): 246-248.\u003c/li\u003e\n \u003cli\u003eIwai, N, Ebata, T, Nagura, H, Kitazume, T, Nagai, K, Wachi, M. Structure-activity relationship of S-benzylisothiourea derivatives to induce spherical cells in \u003cem\u003eEscherichia coli.\u003c/em\u003e Biosci. Biotechnol. Biochem\u003cem\u003e.\u003c/em\u003e, 2004; \u003cem\u003e68\u003c/em\u003e(11): 2265-2269.\u003c/li\u003e\n \u003cli\u003eNicholson, A, Perry, JD, James, AL, Stanforth, SP, Carnell, S, Wilkinson, K, Kahn, CMA, De Soyza, A, Gould, FK. In vitro activity of S-(3,4-dichlorobenzyl)isothiourea hydrochloride and novel structurally related compounds against multidrug-resistant bacteria, including \u003cem\u003ePseudomonas aeruginosa\u003c/em\u003e and \u003cem\u003eBurkholderia cepacia\u003c/em\u003e complex. \u003cem\u003eInt. J. Antimicrob. Agents\u003c/em\u003e\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e2012;\u003cem\u003e39\u003c/em\u003e: 27-32.\u003c/li\u003e\n \u003cli\u003eBean, GJ, Flickinger, ST, Westler, WM, McCully, ME, Sept, D, Weibel, DB, Amann, KJ. A22 disrupts the bacterial actin cytoskeleton by directly binding and inducing a low-affinity state in MreB. \u003cem\u003eBiochemistry\u003c/em\u003e, 2009;\u003cem\u003e48\u003c/em\u003e(22): 4852-4857.\u003c/li\u003e\n \u003cli\u003eTakacs, CN, Poggio, S, Charbon, G, Pucheault, M, Vollmer, W, Wagner, CJ. MreB drives de novo rod morphogenesis in \u003cem\u003eCaulobacter crescentus\u003c/em\u003e via remodeling of the cell wall.\u003cem\u003e\u0026nbsp;\u003c/em\u003eJ. Bacteriol. 2010: 1671-1684.\u003c/li\u003e\n \u003cli\u003eGerdes, K, M\u0026oslash;ller-Jensen, J, Ebersbach, G, Kruse, T, Nordstr\u0026ouml;m, K. Bacterial mitotic machineries. Cell 2004;116(3): 359-366.\u003c/li\u003e\n \u003cli\u003eKruse, T, Gerdes, K. Bacterial DNA segregation by the actin-like MreB protein. Trends Cell Biol. 2005;15(7): 343-345.\u003c/li\u003e\n \u003cli\u003eKruse, T, Blagoev, B, Lobner-Olesen, A, Wachi, M, Sasaki, K, Iwai, N, Mann, M, Gerdes, K. Actin homolog MreB and RNA polymerase interact and are both required for chromosome segregation in \u003cem\u003eEscherichia coli\u003c/em\u003e. Genes Dev.\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e2006;20: 113-124.\u003c/li\u003e\n \u003cli\u003eKruse, T, Moller-Jensen, J, Lobner-Olesen, A, Gerdes, K. Dysfunctional MreB inhibits chromosome segregation in \u003cem\u003eEscherichia coli\u003c/em\u003e. EMBO J. 2003;\u003cem\u003e22\u003c/em\u003e(19): 5283-5292.\u003c/li\u003e\n \u003cli\u003eGitai, Z, Dye, NA, Reisenauer, A, Wachi, M, Shapiro, L. MreB actin-mediated segregation of a specific region of a bacterial chromosome. Cell 2005;120: 329-341.\u003c/li\u003e\n \u003cli\u003eRobertson, GT, Doyle, TB, Du, Q, Duncan, L, Mdluli, KE, Lynch, AS. A novel indole compound that inhibits \u003cem\u003ePseudomonas aeruginosa\u0026nbsp;\u003c/em\u003egrowth by targeting MreB is a substrate for MexAB-OprM. J. Bacteriol\u003cem\u003e.\u003c/em\u003e 2007;189(19): 6870-6881.\u003c/li\u003e\n \u003cli\u003eBryan, EJ, Sagong HY, Parhi AK, Grier MC, Roberge JC, LaVoie EJ, Pilch, DS. TXH11106: A third-generation MreB inhibitor with enhanced activity against a broad range of Gram-negative bacterial pathogens. Antibiotics 2022;11: 693-709.\u003c/li\u003e\n \u003cli\u003eLaVoie, E, Parhi, A, Sagong, HY, inventors; Therapeutic compounds and methods to treat infection.US 0155507 A1, 2020 May 21.\u003c/li\u003e\n\u003c/ol\u003e"},{"header":"Tables","content":"\u003cp\u003eTables 1 to 3 are available in the Supplementary Files section\u003c/p\u003e"},{"header":"scheme","content":"\u003cp\u003eScheme 1 to 9 are available in the Supplementary Files section.\u003c/p\u003e\n"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":false,"highlight":"","institution":"","isAcceptedByJournal":true,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"medicinal-chemistry-research","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"mcre","sideBox":"Learn more about [Medicinal Chemistry Research](https://www.springer.com/journal/44)","snPcode":"44","submissionUrl":"https://submission.nature.com/new-submission/44/3","title":"Medicinal Chemistry Research","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"stoa","reportingPortfolio":"Springer Hybrid","inReviewEnabled":true,"inReviewRevisionsEnabled":false},"keywords":"MreB inhibitors, Gram (-) Antibiotics, E. coli MreB ATPase Inhibition, Structure Activity Relationships","lastPublishedDoi":"10.21203/rs.3.rs-1824695/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-1824695/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eMreB is a cytoskeleton protein present in rod-shaped bacteria that is both essential for bacterial cell division and highly conserved.\u0026nbsp;Because most Gram (-) bacteria require MreB for cell division, chromosome segregation, cell wall morphogenesis, and cell polarity, it is an attractive target for antibacterial drug discovery.\u0026nbsp;As MreB modulation is not associated with the activity of antibiotics in clinical use, acquired resistance to MreB inhibitors is also unlikely.\u0026nbsp;Compounds, such as A22 and CBR-4830, are known to disrupt MreB function by inhibition of ATPase activity.\u0026nbsp;However, the toxicity of these compounds has hindered efforts to assess the in vivo efficacy of these MreB inhibitors.\u0026nbsp;The present study further examines the structure-activity of analogs related to CBR-4830 as it relates to relative antibiotic activity and improved drug properties.\u0026nbsp;These data reveal that certain analogs have enhanced antibiotic activity.\u0026nbsp;In addition, we evaluated several representative analogs (\u003cstrong\u003e9\u003c/strong\u003e, \u003cstrong\u003e10\u003c/strong\u003e, \u003cstrong\u003e14\u003c/strong\u003e, \u003cstrong\u003e26\u003c/strong\u003e, and \u003cstrong\u003e31\u003c/strong\u003e) for their abilities to target purified \u003cem\u003eE. coli\u003c/em\u003e MreB (EcMreB) and inhibit its ATPase activity.\u0026nbsp;Except for \u003cstrong\u003e14\u003c/strong\u003e, all these analogs were more potent than CBR-4830 as inhibitors of the ATPase activity of EcMreB with corresponding IC\u003csub\u003e50\u003c/sub\u003e values ranging from 6.3 ± 2.0 to 29.4 ± 8.8 uM.\u0026nbsp;\u0026nbsp;\u003c/p\u003e","manuscriptTitle":"Novel MreB Inhibitors with Antibacterial Activity Against Gram (-) Bacteria","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2022-07-20 14:57:26","doi":"10.21203/rs.3.rs-1824695/v1","editorialEvents":[{"type":"communityComments","content":0},{"type":"decision","content":"Minor Revisions","date":"2022-08-08T09:55:40+00:00","index":"","fulltext":""},{"type":"reviewerAgreed","content":"","date":"2022-07-28T22:07:48+00:00","index":0,"fulltext":""},{"type":"reviewersInvited","content":"","date":"2022-07-12T16:55:41+00:00","index":"","fulltext":""},{"type":"editorAssigned","content":"","date":"2022-07-07T20:29:32+00:00","index":"","fulltext":""},{"type":"submitted","content":"Medicinal Chemistry Research","date":"2022-07-04T13:10:14+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"medicinal-chemistry-research","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"mcre","sideBox":"Learn more about [Medicinal Chemistry Research](https://www.springer.com/journal/44)","snPcode":"44","submissionUrl":"https://submission.nature.com/new-submission/44/3","title":"Medicinal Chemistry Research","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"stoa","reportingPortfolio":"Springer Hybrid","inReviewEnabled":true,"inReviewRevisionsEnabled":false}}],"origin":"","ownerIdentity":"6cffcc1a-443a-4507-ba94-d24c118de5a9","owner":[],"postedDate":"July 20th, 2022","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"under-review","subjectAreas":[],"tags":[],"updatedAt":"2022-08-30T16:45:40+00:00","versionOfRecord":[],"versionCreatedAt":"2022-07-20 14:57:26","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-1824695","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-1824695","identity":"rs-1824695","version":["v1"]},"buildId":"_2-kVJe1T_tPrBINL-cwx","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}