Stretching the structural envelope of isomeric imatinib analogs that reduce β-amyloid production by modulating both β- and γ-secretase cleavages of APP

Stretching the structural envelope of isomeric imatinib analogs that reduce β-amyloid production by modulating both β- and γ-secretase cleavages of APP | bioRxiv /* */ /* */ <!-- <!-- /*! * yepnope1.5.4 * (c) WTFPL, GPLv2 */ (function(a,b,c){function d(a){return"[object Function]"==o.call(a)}function e(a){return"string"==typeof a}function f(){}function g(a){return!a||"loaded"==a||"complete"==a||"uninitialized"==a}function h(){var a=p.shift();q=1,a?a.t?m(function(){("c"==a.t?B.injectCss:B.injectJs)(a.s,0,a.a,a.x,a.e,1)},0):(a(),h()):q=0}function i(a,c,d,e,f,i,j){function k(b){if(!o&&g(l.readyState)&&(u.r=o=1,!q&&h(),l.onload=l.onreadystatechange=null,b)){"img"!=a&&m(function(){t.removeChild(l)},50);for(var d in y[c])y[c].hasOwnProperty(d)&&y[c][d].onload()}}var j=j||B.errorTimeout,l=b.createElement(a),o=0,r=0,u={t:d,s:c,e:f,a:i,x:j};1===y[c]&&(r=1,y[c]=[]),"object"==a?l.data=c:(l.src=c,l.type=a),l.width=l.height="0",l.onerror=l.onload=l.onreadystatechange=function(){k.call(this,r)},p.splice(e,0,u),"img"!=a&&(r||2===y[c]?(t.insertBefore(l,s?null:n),m(k,j)):y[c].push(l))}function j(a,b,c,d,f){return q=0,b=b||"j",e(a)?i("c"==b?v:u,a,b,this.i++,c,d,f):(p.splice(this.i++,0,a),1==p.length&&h()),this}function k(){var a=B;return a.loader={load:j,i:0},a}var l=b.documentElement,m=a.setTimeout,n=b.getElementsByTagName("script")[0],o={}.toString,p=[],q=0,r="MozAppearance"in l.style,s=r&&!!b.createRange().compareNode,t=s?l:n.parentNode,l=a.opera&&"[object Opera]"==o.call(a.opera),l=!!b.attachEvent&&!l,u=r?"object":l?"script":"img",v=l?"script":u,w=Array.isArray||function(a){return"[object Array]"==o.call(a)},x=[],y={},z={timeout:function(a,b){return b.length&&(a.timeout=b[0]),a}},A,B;B=function(a){function b(a){var a=a.split("!"),b=x.length,c=a.pop(),d=a.length,c={url:c,origUrl:c,prefixes:a},e,f,g;for(f=0;f<d;f++)g=a[f].split("="),(e=z[g.shift()])&&(c=e(c,g));for(f=0;f<b;f++)c=x[f](c);return c}function g(a,e,f,g,h){var i=b(a),j=i.autoCallback;i.url.split(".").pop().split("?").shift(),i.bypass||(e&&(e=d(e)?e:e[a]||e[g]||e[a.split("/").pop().split("?")[0]]),i.instead?i.instead(a,e,f,g,h):(y[i.url]?i.noexec=!0:y[i.url]=1,f.load(i.url,i.forceCSS||!i.forceJS&&"css"==i.url.split(".").pop().split("?").shift()?"c":c,i.noexec,i.attrs,i.timeout),(d(e)||d(j))&&f.load(function(){k(),e&&e(i.origUrl,h,g),j&&j(i.origUrl,h,g),y[i.url]=2})))}function h(a,b){function c(a,c){if(a){if(e(a))c||(j=function(){var a=[].slice.call(arguments);k.apply(this,a),l()}),g(a,j,b,0,h);else if(Object(a)===a)for(n in m=function(){var b=0,c;for(c in a)a.hasOwnProperty(c)&&b++;return b}(),a)a.hasOwnProperty(n)&&(!c&&!--m&&(d(j)?j=function(){var a=[].slice.call(arguments);k.apply(this,a),l()}:j[n]=function(a){return function(){var b=[].slice.call(arguments);a&&a.apply(this,b),l()}}(k[n])),g(a[n],j,b,n,h))}else!c&&l()}var h=!!a.test,i=a.load||a.both,j=a.callback||f,k=j,l=a.complete||f,m,n;c(h?a.yep:a.nope,!!i),i&&c(i)}var i,j,l=this.yepnope.loader;if(e(a))g(a,0,l,0);else if(w(a))for(i=0;i (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];var j=d.createElement(s);var dl=l!='dataLayer'?'&l='+l:'';j.src='//www.googletagmanager.com/gtm.js?id='+i+dl;j.type='text/javascript';j.async=true;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-M677548'); Skip to main content Home About Submit ALERTS / RSS Search for this keyword Advanced Search New Results Stretching the structural envelope of isomeric imatinib analogs that reduce β-amyloid production by modulating both β- and γ-secretase cleavages of APP William J. Netzer , Anjana Sinha , Mondana Ghias , Emily Chang , Katherina Gindinova , Emily Mui , Ji-Seon Seo , View ORCID Profile Subhash C. Sinha doi: https://doi.org/10.1101/2024.07.14.602669 William J. Netzer 1 Laboratory of Molecular and Cellular Neuroscience, The Rockefeller University , New York, NY 10065 Find this author on Google Scholar Find this author on PubMed Search for this author on this site For correspondence: billnetzer{at}gmail.com sinhaanjana5819{at}gmail.com sus2044{at}med.cornell.edu Anjana Sinha 1 Laboratory of Molecular and Cellular Neuroscience, The Rockefeller University , New York, NY 10065 Find this author on Google Scholar Find this author on PubMed Search for this author on this site For correspondence: billnetzer{at}gmail.com sinhaanjana5819{at}gmail.com sus2044{at}med.cornell.edu Mondana Ghias 1 Laboratory of Molecular and Cellular Neuroscience, The Rockefeller University , New York, NY 10065 Find this author on Google Scholar Find this author on PubMed Search for this author on this site Emily Chang 1 Laboratory of Molecular and Cellular Neuroscience, The Rockefeller University , New York, NY 10065 Find this author on Google Scholar Find this author on PubMed Search for this author on this site Katherina Gindinova 1 Laboratory of Molecular and Cellular Neuroscience, The Rockefeller University , New York, NY 10065 Find this author on Google Scholar Find this author on PubMed Search for this author on this site Emily Mui 1 Laboratory of Molecular and Cellular Neuroscience, The Rockefeller University , New York, NY 10065 Find this author on Google Scholar Find this author on PubMed Search for this author on this site Ji-Seon Seo 1 Laboratory of Molecular and Cellular Neuroscience, The Rockefeller University , New York, NY 10065 Find this author on Google Scholar Find this author on PubMed Search for this author on this site Subhash C. Sinha 1 Laboratory of Molecular and Cellular Neuroscience, The Rockefeller University , New York, NY 10065 2 Appel Alzheimer’s Disease Research Institute, Feil Family Brain and Mind Research Institute , Weill Cornell Medicine, New York, NY 10021 Find this author on Google Scholar Find this author on PubMed Search for this author on this site ORCID record for Subhash C. Sinha For correspondence: billnetzer{at}gmail.com sinhaanjana5819{at}gmail.com sus2044{at}med.cornell.edu Abstract Full Text Info/History Metrics Supplementary material Preview PDF Abstract We previously showed that the anticancer drug imatinib mesylate (IMT, trade name: Gleevec) and a chemically distinct compound, DV2-103 (a kinase-inactive derivative of the potent Abl and Src kinase inhibitor, PD173955) lower Aβ levels at low micromolar concentrations primarily through a lysosome-dependent mechanism that renders APP less susceptible to proteolysis by BACE1 without directly inhibiting BACE1 enzymatic activity, or broadly inhibiting the processing of other BACE1 substrates. Additionally, IMT indirectly inhibits γ-secretase and stimulates autophagy, and thus may decrease Aβ levels through multiple pathways. In two recent studies we demonstrated similar effects on APP metabolism caused by derivatives of IMT and DV2-103. In the present study we investigated how so many structurally diverse compounds affect APP metabolism in the same way, with similar potencies and production of APP metabolites. To this end, we synthesized and tested radically altered IMT regioisomers that possess medium structural similarity to IMT. Independent of structural similarity, these isomers manifest widely differing potencies in altering APP metabolism. These will enable us to choose the most potent isomers for further derivatization. 1 Introduction Neurotoxic β-amyloid peptides (Aβ) are major drivers of Alzheimer’s disease (AD) and are formed by sequential cleavage of the amyloid precursor protein (APP) by β-secretase (BACE1/2) and γ-secretase, respectively. Both β- and γ-secretases can be pharmacologically inhibited to reduce production of Aβ peptides. Indeed, there has been great interest in the development of inhibitors and modulators of the secretases as potential AD therapeutics( Miranda et al. 2021 ; Kumar et al. 2018 ; Zhao et al. 2020 ; Portelius et al. 2010 ; Hur 2022 ; Panza et al. 2009 ; Golde et al. 2013 ; Rynearson et al. 2021 ) but at this time all clinical trials involving secretase inhibitors/modulators have failed. Reasons given have included timing of drug administration (too late in disease course for benefits to occur); non-specific inhibition of secretase substrates other than APP; lack of target engagement; toxicity; and even failure of the Amyloid hypothesis( Kim et al. 2022 ). In our previous study, we have shown that the anticancer drug IMT, which is a potent Abl kinase inhibitor( Druker et al. 1996 ), and PD173955( Nagar et al. 2002 ), an Abl/Src kinase inhibitor, reduce Aβ production in cultured N2a695 cells, rat embryonic neurons, and in guinea pig brain in vivo by indirectly inhibiting γ-secretase processing of APP, while sparing γ-secretase processing of Notch1 in cellular assays( Netzer et al. 2003 ). In a recent study we further showed that a kinase inactive derivative of PD173955, DV2-103, as well as IMT, reduced Aβ levels in cells mainly by indirectly inhibiting BACE cleavage of APP( Netzer et al. 2017 ), adding to our earlier study suggesting that the Aβ-lowering effect of IMT and DV2-103 are not only Abl kinase-independent but also broadly kinase-independent and affect both γ-secretase and BACE processing of APP. Additionally, IMT and DV2-103 decrease levels of APP-βCTF and sAPPβ, and raise levels of APP- αCTF, as well as a 141 amino acid APP-CTF (C141), and a 9 kDa APP-CTF (all consistent with reduced BACE processing of APP) in N2a695 cells( Netzer et al. 2017 ). Remarkably, we showed that this pattern of APP metabolites induced by IMT and DV2-103, and some of their analogs is observed when N2a695 cells are treated with a general, active-site-directed BACE inhibitor( Netzer et al. 2017 ; Sun et al. 2019 ; Sinha et al. 2019 ). We also demonstrated that IMT does not inhibit BACE1 enzymatic activity in two in vitro BACE1 assays at concentrations up to 100μM or inhibit processing of several non-APP BACE substrates in cells( Netzer et al. 2017 ). Additionally, we demonstrated that these inhibitory activities of IMT and DV2-103 require acidified lysosomes and we provided a model suggesting that the effects of these drugs on APP metabolism were a result of their effects on lysosomes, which caused APP to undergo increased trafficking to lysosomes and spend less time in the amyloidogenic pathway where Aβ and its direct precursor, the APP-βCTF, are formed( Netzer et al. 2017 ). To understand how so many structurally different compounds reduce levels of secreted Aβ in cells and have a characteristic effect on APP metabolite levels, we designed and synthesized a derivative of IMT, referred to here as IMT isomer 1 ( Fig. 1A ), as well as additional IMT isomers 2 - 3 ( Fig. 1E ) and tested their effects on APP metabolism by measuring the Aβ40 levels in cell supernatants. We found that isomer 2 , not 3 , showed effects comparable to IMT and isomer 1 (See Fig. 1 for the activity data of IMT and isomer 1 and Table 1 isomers 2 - 3 ). Subsequently, we focused on derivatives of isomer 1 and 2 and tested their effects on APP metabolism. The results are shown in Table 1 . Overall, our goal was to make a large change in the structure of IMT that would greatly alter the pharmacophore structurally but still maintain IMT’s physical properties, in particular its property as a weak base, which is necessary for its sequestration in lysosomes through ion trapping( Kazmi et al. 2013 ; Burger et al. 2015 ). Download figure Open in new tab Fig. 1. A Major Structural Change in IMT, results in IMT isomer 1, which retains IMT’s Aβ-reducing effect and its reduction of BACE processing of APP in N2a 695 cells. (A, E) Structures and properties of IMT and isomers 1 , 2 and 3 . Fingerprint (FP) similarity of iso -IMT 1 - 3 to parent IMT and their properties, including most basic PK, were calculated in silico. B) Western blots of N2a cell lysates (upper) and cell media (lower) from experiments using IMT and isomer 1 probed with antibody RU369 (anti-C terminal APP) and RU anti-C-terminal sAPPβ (bottom), respectively. Each western blot panel shows lanes from a single gel. However, the three lanes at the right of each, which refer to incubation with isomer 1 , are from a different part of the same gel. C) Quantification of secreted Aβ40 in N2a cells incubated with IMT or isomer 1 , 1-way Anova, p < 0.001. D) Quantification of sAPPβ levels. 1-way Anova, p < 0.001. View this table: View inline View popup Download powerpoint Table 1. Intermediates (5a-5b, 6a-6i), step(s), and methods used to prepare IMT isomer 1 and 1a-1n. 2 Material and Methods All commercial chemicals and solvents were reagent grade and used without further purification. All air-sensitive reactions were performed under argon protection. Column chromatography was performed using 230-400 mesh silica gel. Analytical thin layer chromatography was performed on 250 μM silica gel F 254 plates. Preparative thin layer chromatography was performed on 1000 μM silica gel F 254 plates. All final compounds were purified using HPLC. The identity and purity of each product was determined using MS, HPLC, TLC, and NMR analyses. 1 H NMR spectra were recorded on either a Bruker 400 or 600 MHz instrument. Chemical shifts are reported in δ values in ppm downfield from TMS as the internal standard. 1 H data are reported as follows: chemical shift, multiplicity (s = singlet, d = doublet, t = triplet, q = quartet, br = broad, m = multiplet), coupling constant (Hz), integration. Purity of target compounds has been determined to be >95% by LC/MS on a Waters autopurification system with PDA, MicroMass ZQ and ELSD detector and a reverse phase column (Waters X-Bridge C18, 4.6 x 150 mm, 5 µm) eluted with water/acetonitrile gradients, containing 0.1% TFA. All compounds tested in this study were prepared in house and their structures were confirmed using 1 H NMR and MS analyses (Spectral data provided for new compounds only). Yields were not optimized. All final compounds were obtained in >95% purity as judged by LCMS analysis. N2a695 were cultured in 1:1 OptiMem Reduced Serum Media (Life Technologies): Dulbecco’s Modified Eagle Medium ([+] 4.5 g/L D-glucose; [+] L-Glutamine; [-] Sodium pyruvate (Life Technologies) supplemented with 5% fetal bovine serum, 0.4% Penstrep and 0.4% Geneticin and incubated at 37 °C in 5% CO 2 . Antibodies were obtained from The Laboratory of Molecular and Cellular Neuroscience at The Rockefeller University. Human Aβ40 and Aβ42 ELISA plates (Life Technologies) and Plus MSD (Mesoscale Discovery) plates for Aβ Peptide (Aβ38, Aβ40 and Aβ42) Panel 1 (6E10) Kit (Catalog number K15200G) were obtained from Thermo Fisher, Life Technologies and Meso Scale Discovery. 2.1. Synthesis of IMT isomer 1 and analogs 1a-t Prepared using intermediates 5a - c ( Scheme 2 ). I) Intermediates 5a-c To prepare intermediate 5a , a solution of 4a (800 mg, 2.96 mmol) and o- toluidine (1 ml, 9.29 mmol) in i -PrOH (6 mL) was added 1N HCl (3 mL), and the mixture was heated at 125 °C using Microwave for 2 h. Solvents were removed under reduced pressure, residues treated with aqueous NaHCO 3 to neutralize, and the resulting mixture extracted with EtOAc. The combined organic layers were washed with brine, dried over anhydrous MgSO 4 , concentrated, and purified by Combi-Flash over Silica gel column using hexanes-EtOAc as eluants to afford intermediate 5a . 1 H NMR (600 MHz, CDCl 3 ) of 5a : δ 9.16 (s, 1H), 8.85 (s, 1H), 8.51 (s, 2H), 8.03 (d, J = 7.92 Hz, 1H), 7.35-7.30 (m, 2H), 7.14-7.07 (m, 2H), 2.38 (s, 3H); HRMS: m/z 341.0357 and 343.0337 [M+H] + , calcd. for C 16 H 14 BrN 4 ; found 341.0391 and 343.0370. Similarly, intermediate 4b was reacted with o-toluidine using the method described for 5a to afford intermediate 5b , and 4c reacted with 3-aminopyridine to afford 5c . 1 H NMR: (600 MHz, CDCl 3 ) of 5b : δ 8.47 (d, J = 6.0 Hz, 1H), 8.22 (s, 1H), 8.09 (d, J = 6.0 Hz, 1H), 7.96 (d, J = 6.0 Hz, 1H), 7.62 (d, J = 6.0 Hz), 7.36 (t, J = 6.0 Hz, 1H), 7.29 (t, J = 6.0 Hz, 2H), 7.24 (d, J = 12 Hz, 1H), 7.11 (d, J = 6.0 Hz, 1H), 7.06 (t, J = 6.0 Hz, 1H), 6.95 (s, 1H), 2.37 (s, 3H); HRMS: m/z 340.0439 and 342.0418, Calcd. for C 17 H 14 BrN 3 . MS of 5c : m/z 264.12 [M+H] + . II) Compounds 1 and 1a-n To a degassed mixture of intermediates 5a (38 mg) and 6a (21 mg) in dioxane was added Pd 2 (dba) 3 (4 mg), XanthPhos (7 mg), and Cs 2 CO 3 (55 mg) and the resulting mixture was heated at 100°C temperature for 16 hours. Solvents were removed and worked up using EtOAc and water. Combined organic layers were dried over anhydrous MgSO 4 and concentrated under reduced pressure. The resulting residues were purified by preparative TLC (Silica gel, 1 mm plate; CH 2 Cl 2 :MeOH:Aq. NH 3 (90:10:1)) to afford the target product 1 . 1 H NMR (400 MHz, CDCl 3 ): d 9.01 (d, J = 1.5 Hz, 1H), 8.92 and 8.88 (s, 1H each), 8.47 (d, J = 5.12 Hz, 1H), 8.05 (d, J = 7.96 Hz, 1H), 7.88 (d, J = 7.96 Hz, 2H), 7.45 (d, J = 8.40 Hz, 1H), 7.28 (t, J = 4.28 Hz, 1H), 7.23 (d, J = 7.40 Hz, 1H), 7.15 (d, J = 5.16 Hz, 1H), 7.07-7.03 (m, 2H), 3.57 (s, 2H), 2.49 (br s, 8H), 2.35 and 2.30 (s, 3H each); HRMS: m/z 494.2624 [M+H] + , calcd. for C 29 H 32 N 7 O; found: 494.2653. Purity (HPLC): >98%. Compound 1a Buchwald coupling of 5b with 6a was performed as described for compound 1 to afford 1a . 1 H NMR (400 MHz, CDCl 3 ) of 1a : δ 8.46 (d, J = 5.20 Hz, 1H), 8.36 (s, 1H), 8.12 (d, 8.04 Hz, 1H), 8.02 (s, 1H), 7.87-7.82 (m, 4H), 7.52-7.46 (m, 3H), 7.31-7.23 (m, 1H), 7.17 (d, J = 5.16 Hz, 1H), 7.06 (t, J = 3.6 Hz, 1H), 6.96 (s, 1H), 3.56 (s, 2H), 2.52 (br s, 8H), 2.37 and 2.33 (s, 3H each); HRMS: m/z 493.2671 [M+H] + , calcd. for C 30 H 33 N 6 O; found: 493.2716. Compound 1b Intermediate 5a reacted with 6b under Buchwald conditions as described above for 1 to give the Boc protected compound 1b - Boc . MS: m/z 579.30. To a solution of 1b-Boc in EtOAc (3 mL) was added 4M HCl in dioxane (1 mL) at room temperature (RT) and the mixture was stirred for 2 hours or until the reaction was complete (monitored by TLC or LCMS). Solvents were removed under reduced pressure and purified over Silica gel column using CH 2 Cl 2 - MeOH/aq. NH 3 to give the Boc-deprotected compound 1b . HRMS: m/z 480.2467 [M+H] + , calcd. for C 28 H 30 N 7 O; found: 480.2506. Compound 1c Intermediate 5a underwent Buchwald coupling with 6c as described above for 1 to give compound 1c . 1 H NMR (600 MHz, CDCl 3 ): δ 9.03, 8.95 and 8.90 (s, 1H each), 8.50 (d, J = 6.0 Hz, 1H), 8.09 (d, J = 6.0 Hz, 1H), 7.87 (d, J = 6.0 Hz, 2H), 7.35 (d, J = 6.0 Hz, 2H), 7.32-7.29 (m, 1H), 7.25 (d, J = 6.0 Hz, 1H), 7.20 (d, J = 6.0 Hz, 1H), 7.08 (d, J = 6.0 Hz, 2H), 2.89 (t, J = 6.0 Hz, 2H), 2.59 (d, J = 6.0 Hz, 2H), 2.37 (s, 3H), 2.33 (s, 6H); HRMS: m/z 453.2358 [M+H] + , calcd. for C 27 H 29 N 6 O; found: 453.2403. Compound 1d Intermediate 5b underwent Buchwald coupling with 6c as described for 1 to give compound 1d . 1 H NMR (600 MHz, CDCl 3 ): δ 8.49 (d, J = 6.0 Hz, 1H), 8.38 (s, 1H), 8.15 (d, J = 6.0 Hz, 1H), 7.99 (br s, 1H), 7.88-7.83 (m, 4H), 7.68 (d, J = 6.0 Hz, 1H), 7.53 (t, J = 6.0 Hz, 1H), 7.42-7.38 (m, 2H), 7.33 (d, J = 12.0 Hz, 1H), 7.26 (d, J = 6.0 Hz, 1H), 7.21 (d, J = 6.0 Hz, 1H), 7.08 (t, J = 6.0 Hz, 1H), 6.97 (s, 1H), 2.29 (q, J = 6.0 Hz, 2H), 2.61 (q, J = 6.0 Hz, 2H), 2.40 (s, 3H), 2.38 (s, 6H); HRMS: m/z 452.2406 [M+H] + , calcd. for C 28 H 30 N 5 O; found 452.2450. Purity (HPLC): >98%. Compound 1e Intermediate 5a underwent Buchwald coupling with 6d as described for 1 to give compound 1e . HRMS: m/z 439.2202 [M+H] + , calcd. for C 26 H 27 N 6 O; found: 439.2243. Compound 1f Intermediate 5a underwent Buchwald coupling with 6e as described for 1 to give the Boc-protected derivative, 1f-Boc , and the latter underwent Boc-deprotection to give compound 1f . 1 H NMR (400 MHz, CDCl 3 ) of 1f-Boc : δ 9.04 (s, 1H), 8.43 (s, 1H), 8.85 (s, 1H), 8.49 (d, J = 5.04 Hz, 1H), 8.07 (d, J = 8.0 Hz, 1H), 7.89-7.87 (br, 2H), 7.27-7.17 (m, 6H), 7.07-7.03 (m, 2H), 4.58 (br, 2H), 3.16(br s, 1H), 3.07 (br s, 1H), 2.36 (s, 3H), 1.50 and 1.34 (s, 6H and 3H); 0.97 (s, 9H); MS: m/z 581.32 [M+H] + . 1 H NMR (400 MHz, CDCl 3 ) of 1f : δ 9.06, 8.94 and 8.85 (s, 1H each), 8.50 (d, J = 5.04 Hz, 1H), 8.10 (d, J = 8.0 Hz, 1H), 8.0 (s, 1H), 8.07 (d, J = 8.0 Hz, 1H), 7.89 (d, J = 7.92 Hz, 2H), 7.52 (d, J = 7.80 Hz, 2H), 7.24-7.21 (m, 4H), 7.08 (t, J = 4.00 Hz, 1H), 7.00 (s, 1H), 3.92 (s, 2H), 2.38 (s, 2H), 2.37 (s, 3H), 0.95 (s, 9H); HRMS: m/z 481.2671 [M+H] + , calcd.; for: C 29 H 33 N 6 O; found 481.2716. Compound 1g Intermediate 5b underwent Buchwald coupling with 6e as described for 1 to give the Boc-protected derivative, 1g-Boc , and the latter underwent Boc-deprotection to give compound 1g . MS of 1g-Boc : m/z 580.33 [M+H] + . 1 H NMR (600 MHz, CDCl 3 ) of 1g : δ 8.47, 8.36 and 8.14 (s, 1H each), 7.88-7.83 (m, 4H), 7.50 (d, J = 7.40 Hz, 4H), 7.24-7.19 (m, 3H), 7.08--7.01 (m, 2H), 3.90 (s, 2H), 3.49 (s, 2H), 2.34 (s, 3H), 0.95 (m, 9H); HRMS: m/z 480.2719 [M+H] + , calcd. for C 30 H 34 N 5 O; found: 480.2776. Compound 1h Intermediate 5a underwent Buchwald coupling with 6f as described for 1 to give the Boc-protected derivative, 1h-Boc , and the latter underwent Boc-deprotection to give compound 1h . 1 H NMR (400 MHz, CDCl 3 ) of compound 1h-Boc : δ 9.04 (br s, 1H), 8.97 and 8.88 (s, 1H each), 8.72 (br, 1H), 8.47 (d, J = 4.28 Hz, 1H), 8.06 (d, J = 7.92 Hz, 4H), 7.94 (m, 2H), 7.44 7.36-7.17 (m, 5H), 7.06 (m, 2H), 4.42 (br s, 2H), 4.08 (m, 1H), 2.36 (s, 3H), 1.70-1.4 (m, 8H), 1.35-1.26 (m, (9H+2H); MS: m/z 593.32 [M+H] + . 1 H NMR (400 MHz, CDCl 3 ) of compound 1h : δ 9.03 (br s, 1H), 8.94 and 8.85 (s, each 1H), 8.48 (d, J = 4.84 Hz, 1H), 8.34 9 (br, 1H), 8.08 (d, J = 7.96 Hz, 1H), 7.88 (d, J = 7.80 Hz, 2H), 7.49 (d, J = 7.64 Hz, 2H), 7.30 (d, J = 7.56 Hz, 2H), 7.25-7.23 (m, 3H), 7.18 (d, J = 4.72 Hz, 1H), 7.09-7.05 (m, 2H), 3.94 (s, 2H), 2.57 (m, 1H), 2.37 (s, 3H), 1.98-1.45 (m, 8H), 1.29-1.18 (m, 2H); HRMS: m/z 493.2671 [M+H] + , calcd. for: C 30 H 33 N 6 O; found: 493.2718. Compound 1i Intermediate 5b underwent Buchwald coupling with 6f as described for 1 to give the Boc-protected derivative, 1i-Boc , and the latter underwent Boc-deprotection to give compound 1i . MS of 1i-Boc : m/z 592.33 [M+H] + . 1 H NMR (600 MHz, CDCl 3 ) of 1i : δ 8.41 (d, J = 5.04 Hz, 1H), 8.31 (s, 1H), 8.04 (d, J = 7.96 Hz, 1H), 7.88 (d, J = 7.72 Hz, 3H), 7.78 (d, J = 7.64 Hz, 1H), 7.49-7.42 (m, 4H), 7.27-7.24 (m, 3H), 7.15 (d, J = 4.96 Hz, 1H), 7.03 (t, J = 7.24 Hz, 1H), 3.90 (s, 2H), 3.38 (m, 1H), 2.34 (s, 3H), 1-97-1.62 (m, 6H), 1.26-1.16 (m, 4H); HRMS: m/z 492.2719 [M+H] + , calcd. for C 31 H 34 N 5 O; found 492.2772. Compound 1j Intermediate 5a underwent Buchwald coupling with 6g as described for 1 to give compound 1j . 1 H NMR (600 MHz, CDCl 3 ): δ 9.04 (s, 1H), 8.94 (s, 1H), 8.86 (d, J = 6.0 Hz, 1H), 8.13 (dd, J = 12.0, 6.0 Hz, 1H), 7.87 (d, J = 6.0 Hz, 2H), 7.32 (t, J = 6.0 Hz, 2H), 7.265 (d, J = 6.0 Hz, 1H), 7.215 (d, J = 6.0 Hz, 1H), 7.08 (t, J = 6.0 Hz, 1H), 6.975 (d, J = 6.0 Hz, 2H), 3.40 (t, J = 6.0 Hz, 4H), 2.62 (t, J = 6.0 Hz, 4H), 2.40 and 2.39 (s, 3H each); MS: m/z 480.25 [M+H] + , calcd. for C 28 H 30 N 7 O; found: 480.25. Compound 1k Intermediate 5b underwent Buchwald coupling with 6g as described for 1 to give compound 1k . 1 H NMR (600 MHz, CDCl 3 ) of 1k : δ 8.49 (d, J = 6.0 Hz, 1H), 8.36 (s, 1H), 8.17 (dd, J = 6.0, 12.0 Hz, 1H), 7.88-7.83 (m, 3H), 7.51 (t, J = 6.0 Hz, 1H), 7.33-7.29 (m, 2H), 7.27 (d, J = 6.0 Hz), 7.22 (d, J = 6.0 Hz, 1H), 7.09 (t, J = 6.0 Hz, 1H), 6.99 (d, J = 6.0 Hz, 2H), 3.41 (t, J = 6.0 Hz, 4H), 2.63 (br t, 4H), 2.41 and 2.40 (s, 3H each); MS: m/z 479.25 [M+H] + , calcd. for C 28 H 31 N 6 O; found: 479.25. Compound 1l Intermediate 5a underwent Buchwald coupling with 6h as described for 1 to give the Boc-protected derivative, 1l-Boc , and the latter underwent Boc-deprotection to give compound 1l . 1 HNMR (600 MHz, CDCl 3 ) of 1l-Boc : δ 9.04 (s, 1H), 8.95 (s, 1H), 8.88 (s, 1H), 8.50 (s, 2H), 8.10 (d, J = 6.0 Hz, 1H), 7.91 (d, J = 6.0 Hz, 2H), 7.37 (br t, J =, 2H), 7.30 (d, J = 6.0 Hz, 1H), 7.26 (d, J = 6.0 Hz, 1H), 7.20 (m, 2H), 4.18 (m, 1H), 2.82 (br, 2H), 2.40 (s, 3H), 2.08 (m, 1H), 1.82 (m, 1H), 1.69 (m, 2H), 1.61 (s, 3H), 1.49 (s, 6H), 1.48 (m, 2H); MS (ESI) m/z 564.28 [M] + , calcd. for C 33 H 36 N 6 O 3 ; found: 565.29 [M+H] + . HRMS of 1l : m/z 465.2358 [M+H] + , calcd. for C 28 H 29 N 6 O; found 465.2401. Compound 1m Intermediate 5b underwent Buchwald coupling with 6h as described for 1 to give the Boc-protected derivative, 1m-Boc , and the latter underwent Boc-deprotection to give compound 1m . 1 HNMR (600 MHz, CDCl 3 ) of 1m-Boc : δ 8.50 (d, J = 6.0 Hz, 1H), 8.37 (s, 1H), 8.16 (d, J = 12.0 Hz, 1H), 7.93 (s, 1H), 7.89-7.86 (m, 3H), 7.545 (t, J = 6.0 Hz, 1H), 7.42 (d, J = 6.0 Hz, 1H), 7.32 (t, J = 12.0 Hz, 2H), 7.27 (d, J = 6.0 Hz, 1H), 7.22 (d, J = 6.0 Hz, 1H), 7.08 (t, J = 6.0 Hz, 1H), 6.96 (s, 1H), 4.18 (m, 1H), 2.81 (br, 2H), 2.40 (s, 3H), 2.08 (m, 1H), 1.81 (m, 1H), 1.68 (m, 2H), 1.59 (s, 3H), 1.51 (s, 6H), 1.48 (m, 2H); MS: m/z 564.29 [M+H] + . HRMS of 1m : m/z 464.2406 [M+H] + , calcd. for C 29 H 29 N 5 O; found 464.2459. Purity (HPLC): >98%. Compound 1n Intermediate 5b underwent Buchwald coupling with 6i as described for 1 to give the Boc-protected derivative, 1n-Boc , and the latter underwent Boc-deprotection to give compound 1n . MS of 1n-Boc : m/z 564.29 [M+H] + . MS of 1n : m/z 463.24 [M+H] + , calcd. for C 29 H 30 N 5 O; found 464.24. Compound 1o NaCNBH 3 (3 equiv.) and AcOH (1 equiv.) were added sequentially to a solution of amine 1l (1 equiv.) and paraformaldehyde (3 equiv.) in MeOH (3-5 mL/mmol) at ice-water temperature and the reaction mixture was stirred at RT for another 2-8 hrs. Solvents were removed under reduced pressure, and the residue was suspended in CH 2 Cl 2 and washed using water. Combined organic layers were dried using Na 2 SO 4 , filtered, and concentrated under reduced pressure. The residue was purified by Silica gel column to afford 1o . 1 H NMR (600 MHz, CD 3 OD+CDCl 3 ): δ 8.99 (br s, 1H), 8.94 (s, 1H), 8.45 (s, 1H), 8.02 (d, J = 6.0 Hz, 2H), 7.74 (d, J = 6.0 Hz, 1H), 7.49 (d, J = 6.0 Hz, 2H), 7.30-7.24 (m, 3H), 7.10 (t, J = 6.0 Hz, 1H), 3.49 (m, 2H), 3.10 (t, J = 12.0 Hz, 1H), 2.95 (t, J = 12.0 Hz, 1H), 2.87 (m, 1H), 2.81 (s, 3H), 2.34 (s, 3H), 2.10-1.64 (m, 4H); HRMS: m/z 479.2515 [M+H] + , calcd. for C 29 H 31 N 6 O; found: 479.2560. Compound 1p Prepared by reductive amination of amine 1m with paraformaldehyde and NaCNBH 3 as described for 1o . 1 HNMR (600 MHz, CDCl 3 ) of 1p : δ 8.42 (d J = 6.0 Hz, 1H), 8.36 (s, 1H), 8.05 (d, J = 6.0 Hz, 1H), 7.87 (d, J = 6.0 Hz, 2H), 7.80 (d, J = 6.0 Hz, 1H), 7.47 (t, J = 6.0 Hz, 1H), 7.34 (d, J = 12.0 Hz, 2H), 7.265 (d, J = 6.0 Hz, 1H), 7.23 (d, J = 6.0 Hz, 1H), 7.16 (q, J = 6.0 Hz, 1H), 7.05 (t, J = 6.0 Hz, 1H), 3.29 (d, J = 12.0 Hz, 2H), 3.10 (t, J = 6.0 Hz, 1H), 2.66 (s, 3H), 2-63-2.54 (m, 2H), 2.34 (s, 3H), 2.03-1.92 (m, 3H), 1.65-1.61 (m, 1H); HRMS: m/z 478.2562 [M+H] + , calcd. for C 30 H 32 N 5 O; Found 478.2602. Compound 1q To a degassed solution of intermediate 5a (1 equiv.) and 4 - formylbenzene boronic acid (1 equiv.), and 2M aq. K 2 CO 3 solution (1.5 mL/mmol) in 1,4-dioxane (4.5 mL/mmol) in a microwave vial was added Pd(PPh 3 ) 4 (0.05 equiv.) and the mixture was heated at 100°C for 2 min. using microwave. The reaction mixture was diluted using water, extracted using CH 2 Cl 2 , and the combined organic layers concentrated under reduced pressure and purified over Silica gel to afford 1q . 1 H NMR (400 MHz, CDCl 3 ): δ 9.18 (s, 1H), 8.92 (s, 1H), 8.56 (s, 1H), 8.51 (d, J = 5.04 Hz, 1H), 8.08 (d, J = 8.00 Hz, 1H), 7.63 (d, J = 7.88 Hz, 2H), 7.48 (d, J = 7.88 Hz, 2H), 7.25-7.21 (m, 4H), 7.09-7.05 (m, 1H), 3.49 (s, 2H), 2.37 (s, 3H), 2.32 (s, 6H); HRMS: m/z 396.2144 [M+H] + , calcd. for C 25 H 26 N 5 ; found 396.2197. Compound 1r Prepared by Suzuki reaction of 5a with 7 as described above for 1q . 1 H NMR (400 MHz, CDCl 3 ) of 1r : d 9.14 (s, 1H), 8.92 (s, 1H), 8.52 (d, J = 4.52 Hz, 2H), 8.13 (d, J = 6.0 Hz, 1H), 7.60 (d, J = 8.56 Hz, 2H), 7.48 (d, J = 8.56 Hz, 1H), 7.25-7.21 (m, 2H), 7.10-6.96 (m, 3H), 3.34-3.23 (m, 8H), 1H), 2.81 (s, 3H), 2.34 (s, 3H), 2.10-1.64 (m, 4H); HRMS: m/z 437.2409 [M+H] + , calcd. for C 27 H 29 N 6 ; found 437.2449. Compound 1s Prepared by amide formation between 5c and 6j using EDC/HOBt coupling. 1 H NMR (600 MHz, CDCl 3 ): d 8.49 (d, J = 6.0 Hz, 1H), 8.38 (s, 1H), 8.15 (d, J = 6.0 Hz, 1H), 7.99 (br s, 1H), 7.88-7.83 (m, 4H), 7.68 (d, J = 6.0 Hz, 1H), 7.53 (t, J = 6.0 Hz, 1H), 7.42-7.38 (m, 2H), 7.33 (d, J = 12.0 Hz, 1H), 7.26 (d, J = 6.0 Hz, 1H), 7.21 (d, J = 6.0 Hz, 1H), 7.08 (t, J = 6.0 Hz, 1H), 6.97 (s, 1H), 2.29 (q, J = 6.0 Hz, 2H), 2.61 (q, J = 6.0 Hz, 2H), 2.40 (s, 3H), 2.38 (s, 6H); HRMS: m/z 439.2202 [M+H] + , calcd. for C 28 H 30 N 5 O; found 439.2256. Purity (HPLC): >98%. Compound 1t Prepared by amide formation between 5c and 6k using EDC/HOBt coupling. MS of 1t : m/z 464.23 [M+H] + , calcd. for C 28 H 29 N 6 O; found 465.24. 2.2. Synthesis of IMT isomer 2 and analogs 2a-b Prepared using intermediates 5a - c ( Scheme 3A ). IMT isomer 2 Buchwald coupling of intermediate 8 with amine 9 as described above for 1 afforded intermediate 10 . MS of 10 : m/z 409.15 [M] + , calcd. for C 24 H 19 N 5 O 2 ; found 410.16 [M+H] + . Intermediate 10 underwent reductive amination with 4-methylpiperazine using NaCNBH 3 as described for 1o to give IMT isomer 2 . 1 H NMR (400 MHz, CDCl 3 + CD 3 OD) of 2 : δ 9.15 (s, 1H), 8.81 (s, 1H), 8.46 (d, J = 4.88 Hz, 1H), 8.35 (s, 1H), 8.27 (d, J = 6.88 Hz, 1H), 8.07 (d, J = 7.64 Hz, 2H), 7.85 (m, 1H), 7.57 (d, J = 8.0 Hz, 1H), 7.47 (d, J = 8.8 Hz, 1H), 7.45 (d, J = 8.0 Hz, 2H), 7.37 (m, 1H), 7.24 (d, J = 8.0 Hz, 1H), 7.15 (d, J = 4.96 Hz, 1H), 3.58 (s, 2H), 2.52 (br s, 8H), 2.33 (s, 6H); MS: m/z 494.2624 [M+H] + , calcd. for C 29 H 32 N 7 O; found: 494.2678. Compounds 2a and 2b Aldehyde 8 underwent reductive amination with neopentyl and cyclohexyl amine, NaCNBH 3 , and AcOH, as described above for intermediate 10 , and the resulting products were reacted with Boc-anhydride to afford intermediates 11a and 11b . Next, Buchwald coupling of 11a and 11b with amine 9 as described above for 1 afforded the Boc-protected derivatives of compounds 2a and 2b . Finally, Boc-deprotection in the latter products using 4M HCl in dioxane gave the title products 2a and 2b . MS of 2a : m/z 481.27 [M+H] + . MS of 2b : m/z 493.27 [M+H] + . 2.2. Synthesis of IMT isomer 3 Prepared using intermediates 12 and 14 as outlined in Scheme 3B . Intermediate 12 2-Aminopyrimidine underwent Buchwald coupling with 3-bromo-4-methylbenzaldehyde to afford intermediate 12 . MS of 12 : m/z 213.09 [M] + , calcd. for C 12 H 11 N 3 O; found 214.09 [M+H] + . Intermediate 14 To a solution of 3-aminopyridine (1 equiv.) and DIEA (3 equiv.) in dry THF (5 mL/mmol) was added 4-chloromethylbenzoyl chloride (1.2 equiv.) at room temperature and the resulting mixture was stirred for 16 hours to afford intermediate 13 after purification. MS of 13 : m/z 246.06/248.05 [M] + , calcd. for C 12 H 11 ClN 2 O; found 247.06/249.06 [M+H] + . A solution of intermediate 13 (1 equiv.), N-Boc-piperidine (1 equiv.), and DIEA (3 equiv.) in dry THF (5 mL/mmol) was heated at 90 °C for 16 h. Reaction mixture was worked-up using water and CH 2 Cl 2 , and the combined organic layers concentrated under reduced pressure and chromatographed over Silica gel using CH 2 Cl 2 -MeOH-aq. NH 3 to afford Boc-protected- 14 . The latter underwent Boc deprotection to afford 14 . MS: m/z 296.16 [M] + , calcd. for C 17 H 20 N 4 O; found 297.17 [M+H] + . IMT isomer 3 A solution of 12 and 14 in methanol underwent reductive amination using NaCNBH 3 and AcOH to give IMT isomer 3 . 1 H NMR (400 MHz, CDCl 3 + CD 3 OD) of 3 : d 8.68 (s, 1H), 8.37 (s, 1H, and d, J = 4.6 Hz, 2H), 8.30 (s, 2H), 7.83 (d, J = 7.13 Hz, 2H), 7.82 (s, 1H), 7.43 (d, J = 7.88 Hz, 2H), 7.31 (dd, J = 7.96, 4.6 Hz, 1H), 7.17 (d, J = 7.64 Hz, 1H), 7.01 (d, J = 7.48 Hz, 1H), 6.89 (s, 1H), 6.69 (t, J = 4.72 Hz, 1H), 3.57 (s, 4H), 3.53 (s, 4H), 2.29 (s, 3H); HRMS: m/z 494.2624 [M+H] + , calcd. for C 29 H 32 N 7 O; found: 494.2672. 2.2. Screening and evaluation of IMT isomers and analogs( Sun et al. 2019 ; Sinha et al. 2019 ) N2a695 cells were used to screen all new compounds and in the follow-up studies with compounds found active in the preliminary screen. In a typical experiment, 6-well tissue culture plates (Corning) were seeded at 4.0×10 5 – 4.5×10 5 N2a695 cells/mL, 2 mL/well for overnight incubation. When cells were >95% confluent, media were exchanged with fresh media containing 10 µM solutions of compounds and cells were incubated at 37 °C in 5% CO 2 for 5 hours. Culture media were collected and soluble Aβ concentrations in the media were determined by ELISA or MSD plates for human Aβ Peptides as per manufacturer instructions. Signals for Aβ were measured using Perkin Elmer Envision and SQ120 MSD ELISA reader. Follow-up studies with N2a695 cells were performed similarly. 2.3. Effects of IMT and isomers on APP metabolism( Netzer et al. 2017 ; Sun et al. 2019 ; Sinha et al. 2019 ) N2a695 cells were treated with compounds for 5 hours as described above, and media were aspirated out (or collected for determination of Aβ levels). Cells were scraped in cold Dulbecco’s PBS buffer (1 mL) containing mini EDTA-free protease inhibitor (Roche) and centrifuged for 1 minute at 13,000 rpm at 4 °C to form a cell pellet. The buffer was aspirated and the cell pellets were lysed in 3% SDS plus protease inhibitor cocktail by sonication on ice for two rounds of 20 seconds on a low setting. Protein concentrations were measured using the Pierce BCA Protein Assay (Thermo Fisher) kit in accordance with the manufacturer’s instructions. To perform WBs, N2a695 cell lysates from 1a and analogs-treated samples were run on a 10-20% or a 16.5 % Tris-Tricine gel (Criterion) and electro transferred to PVDF membranes (EMD Millipore) overnight at 30V. PVDF membranes were incubated in PBS containing 0.25% glutaraldehyde (Sigma) for 30 min after electro transference, blocked for 30 minutes in milk PBST, incubated with primary antibody RU369 for 1 hour at room temperature followed by washing and incubation with an HRP-linked secondary antibody and detected with enhanced chemiluminescence ECL reagents. WB images were analyzed using ImageJ to quantify the prominent bands. To determine effects of compounds on BACE1 vs. GS inhibition, we used N2a cells transiently transfected with full length APP (APP-FL) or with APP99 (APP-βCTF) as described previously( Netzer et al. 2017 ; Sun et al. 2019 ). After 48 hours, media were removed and fresh media containing compound 1a and analogs were added. Following 5 hours of incubation, cell supernatants were collected, and analyzed using MSD-ELISA for Aβ and for sAPPα and β. 2.4. In vivo brain permeability and retention of IMT analogs( Sun et al. 2019 ) All procedures involving animals were approved by The Rockefeller University Institutional Animal Care and Use Committee and were in accordance with the National Institutes of Health guidelines. Mesylate salts of the isomeric IMT analogs (1 or 3 mg/mL in water, 125 µL, 50 mg/kg) were administered intraperitoneally (i.p.) or through oral gavage to 8 weeks old C57BL/6J WT mice. Mice were euthanized 4 hours post drug administration and brain hemispheres and plasma were harvested and collected in pre-weighted tubes and snap-frozen in liquid nitrogen. To measure brain and plasma concentrations of the specific compounds, mouse brain tissue was homogenized and extracted using Ethanol, and plasma samples were extracted using Acetonitrile. Concentration of the drug and metabolites in brain and in plasma was determined by LC-MS/MS analysis. 2.5. Drug Extraction from Brain( Sun et al. 2019 ) After tubes were weighed to calculate brain weight and thawed to room temperature, 1 mL of EtOH (200 Proof) was added to the microcentrifuge tubes containing the harvested right brain hemispheres. 10 µL of 1 µM internal standard (ABG190, a synthetic analog of 1a) was added to each tube and samples were sonicated to homogeneity (∼2 min). Tubes were shaken at 40 min at room temperature (1K RPM) and centrifuged for 8 minutes at 13K RPM. The supernatant (0.9 mL) was transferred to a new collection tube and 0.5 mL EtOH was added to the pellet for a second round of extraction as described above. 600 µL of the supernatant was combined with the first collection before samples were submitted for LCMS-MS analysis. 2.6. Drug Extraction from Blood( Sun et al. 2019 ) 300 µL of acetonitrile was added to collected blood samples. 10 µL of 1 µM internal standard (ABG190) was added to each tube and samples were sonicated to homogeneity (∼2 min). Tubes were contributed at 13K RPM for 9 minutes. 300 µL of the supernatant was collected and combined with 500 µL of 5 mM ammonium formate before samples were submitted for LCMS-MS analysis. 2.7. In vitro kinase activity assay( Jester et al. 2010 ) The assay was performed by Luceome Biotechnologies, LLC. Typically, 10 mM stock solutions of the compounds were diluted in DMSO to a concentration of 250 μM. Prior to initiating the assay, all test compounds were evaluated for false positive against split-luciferase( Jester et al. 2010 ). For kinase assays, each Cfluc-Kinase was translated along with Fos-Nfluc using a cell-free system (rabbit reticulocyte lysate) at 30 °C for 90 min. 24 μL aliquot of this lysate containing either 1 μL of DMSO (for no-inhibitor control) or compound solution in DMSO (10 μM final concentration) was incubated for 30 minutes at room temperature followed by 1 hour in presence of a kinase specific probe. 80 μL of luciferin assay reagent was added to each solution and luminescence was immediately measured on a luminometer. The percent Inhibition was calculated using the following equation: % Inhibition = (ALUcontrol– ALUsamplex 100)/ALUcontrol. 3 Results 3.1 Chemistry IMT isomers 1 - 3 possess all five rings and the chemical functions that broadly match the parent compound ( Fig. 1 ). We designed these isomers by making one or two hypothetical fragmentations across C-N and C-C bonds and re-joining the resulting fragments through other ring(s) and keeping the functionalities similar to IMT, as outlined in Scheme 1 . Arrow ‘a-c’ shown in RED and the double arrow shown in BLUE denote the site(s) of fragmentation and re-attachment of various bonds, respectively. Thus, the left part (three ‘ABC’ rings) of IMT will separate from the right two ‘DE’ rings involving a C-N bond fragmentation (designated by ‘a’, Eqn. 1) next to ring ‘C’, and the resulting left fragment, I , will connect through ring ‘A’ (pyridine ring) to amide ‘N’ of the right fragment, II , giving isomer 1 . Alternatively, IMT will undergo two cleavages across the C-C bond between ring A and B (step ‘b’) for both isomers 2 and 3 , and another (1) C-C bond cleavage between -C(O)- and ring ‘D’ (step ‘c’, Eqn. 2) for isomer 2 and (2) C-N bond cleavage next to ring ‘C’ (step ‘a’, Eqn. 3) for isomer 3 . These cleavages would give three fragments each, III , IV and V for isomer 2 , and III , VI and II for isomer 3 . Note that II is a common fragment for both the isomers 1 and 3 , and III is common for isomers 2 and 3 . To generate isomer 2 , fragment III will combine with the amide carbon in IV and the ring ‘B’ of fragment IV with ring ‘D’ of V , both involving the C-C bond formation (Eqn. 2). Finally, fragment III will combine (C-N bond formation) with the amide nitrogen in II , and the ring ‘C’ of fragment VI with ‘N-Me’ ring ‘E’ of II (C-C bond formation) giving isomer 3 (Eqn. 3). Download figure Open in new tab Scheme 1. Design of IMT isomers 1 , 2 and 3 . Shown are hypothetical fragmentation of IMT involving (a) C-N bond or (b, c) C-C bond cleavage giving fragments I - VI , and re-assembly of these fragments to afford IMT isomers 1 , 2 and 3 . Note: fragment II is common for both IMT isomers 1 and 3 , and III for isomers 2 and 3 . Key: Red arrow, site of C-C or C-N bond cleavage for fragmentation; Blue arrow, C-C or C-N bond connection for re-assembly of the molecules. Synthesis of IMT isomers 1 - 3 and analogs. We prepared IMT isomer 1 and its analogs 1a - 1t using the readily available intermediates, as outlined in Schemes 2A-D. First, to prepare IMT isomer 1 , intermediate 4a was reacted with o-toluidine and the resulting product 5a underwent Buchwald coupling( Ruiz-Castillo and Buchwald 2016 ) with amide 6 to give isomer 1 ( Scheme 1A ). Similarly, intermediate 4b reacted with o-toluidine to give 5b , and both 5a and 5b underwent Buchwald coupling( Ruiz-Castillo and Buchwald 2016 ) with various amides 6a - i giving products 1a - n , several after Boc deprotection as needed ( Scheme 2A and Table 1 ). Analogs 1o and 1p were prepared by reaction of 1l and 1m with formaldehyde under the reductive amination conditions using NaCNBH 3 ( Scheme 2B ). Analog 1q was prepared by Suzuki coupling of 5a with 4-formylphenylboronic acid, 7a , followed by reductive amination of the resulting product with dimethylamine, and 1r by Suzuki coupling of 5a with boronic acid 7b ( Scheme 2C )( Miyaura and Suzuki 1995 ). The remaining analogs of 1 , i.e., 1s and 1t were obtained by reacting 4c with 3-aminopyridine, followed by Boc-deprotection giving amine 5c and reacting the latter with acids 6j and 6k ( Scheme 2D ). Download figure Open in new tab Scheme 2. Synthesis of IMT isomer 1 and analogs 1a-t. Key: a) 3N HCl, Dioxane. microwave, 100°C, 2 h. b) Pd 2 (dba) 3 , XanthPhos, Cs 2 CO 3 , 1,4-Dioxane, microwave, 100°C. c) 4M HCl in dioxane, EtOAc, RT, 2 h. d) CH 2 O, NaCNBH 3 , DCE, 0 °C - RT, 16 h. e) Pd(PPh 3 ) 4 , aq. K 2 CO 3 , 1,4-Dioxane, microwave, 100°C, 2 h. f) Me 2 NH, Na(OAc) 2 BH, AcOH, DCE. g) EDC, HOBt, CH 2 Cl 2 , RT, 16 h. All compounds possess original A-E lettering for IMT shown in Fig. 1 and Scheme 1 . Next, we prepared IMT isomer 2 and its analogs 2a - b using intermediates 8 and 9 , as described in Scheme 3A . Intermediates 8 and 9 reacted together under the Buchwald coupling conditions affording 10 , which underwent reductive amination with N-methylpiperazine to give IMT isomer 2 . Alternatively, intermediate 8 underwent reductive amination with cyclohexyl amine and neopentyl amine and Boc-protection of the resulting amines to give intermediates 11a and 11b , which reacted with intermediate 9 under the Buchwald coupling conditions, followed by Boc-deprotection to give analogs 2a - b . Finally, to prepare IMT isomer 3 , we prepared intermediate 12 by reacting 3-bromo-4-methylbenzaldehyde with 2-amino-pyrimidine under Buchwald conditions, and intermediate 14 by reacting 4-chloro-mthylbenzoyl chloride with 3-aminopyridine 3-amino-pyridine and then with N-Boc-piperazine, followed by N-deprotection. Subsequently, we coupled intermediates 12 and 14 together under the reductive amination conditions using NaCNBH 4 to give the title product 3 ( Afanasyev et al. 2019 ). Download figure Open in new tab Scheme 3. Synthesis of IMT isomers 2 and 3 . Key: a) Pd(dba) 3 , XanthPhos, Cs 2 CO 3 , 1,4-Dioxane, microwave, 100°C, 2 h. b) Na(OAc) 3 BH, DCE, AcOH. c) Boc 2 O, ACN. d) 4M HCl in dioxane, EtOAc, RT, 2 h. d) Methanolic HCl, EtOAc. e) DIEA, THF, RT, 3 h. f) DIEA, THF, 90°C, 2 h. 3.2 Structural diversity The majority of IMT isomer 1 analogs, including 1a - 1n and both analogs of isomer 2 , i.e., 2a and 2b , differ from one-another in ring ‘A’ and/or in ‘E’ (See: Fig. 1 for lettering of the rings, which are based on the original A-E lettering for IMT shown in Scheme 1 ). New IMT isomer 1 analogs contain piperazine ring, a cyclic amine or piperidine ring connected through C-C or C-N bond to ring D, while all other isomer 1 and both isomer 2 analogs possess a substituted alkylamine instead of the ring E. These modifications were made to compare the similar changes made in IMT analogs. There was no additional difference between two analogs, 2a and 2b , of isomer 2 . All 20 analogs of isomer 1 also have rings ‘A-D’ and their arrangement is similar with three exceptions. 1) Nine compounds possess 1,3-substituted benzene and the remaining 11 analogs contain 3,5-substituted pyridine (Py) as the middle ring ‘A’, 2) The first ring from the left (ring ‘C’) in 2 analogs, 1s and 1t , is 3-aminopyridine instead of o-toluidine in all remaining 18 compounds. 3) Analogs 1q and 1r do not possess the ‘amide group’ that connects the middle ring ‘A’ to the 4th ring ‘D’. 3.3 Evaluation Previously, we showed that two chemically distinct compounds, IMT and DV2-103 lower Aβ production primarily by reducing BACE processing of APP( Netzer et al. 2017 ). Similarly, numerous analogs of IMT also lowered Aβ production by reducing BACE processing of APP( Sun et al. 2019 ). In the present study we further examined the effects of these compounds on γ-secretase catalyzed Aβ formation and compared these to the more radically isomeric analogs of IMT. Our results show that IMT, DV2-103, and IMT isomer 1 are γ-secretase modulators; i.e. These compounds favor production or inhibition of different lengths of Aβ peptides (differing in their C-termini). Specifically, we exposed N2a695 cells to increasing concentrations of each compound and measured the production of Aβ38, 40, and 42. IMT, DV2-103 and IMT isomer 1 ( Fig. 2A ) inhibit the formation of Aβ38 least, compared to Aβ40 and 42, and even boost levels of Aβ38 above controls at a drug concentration of 5μM. Remarkably, this occurs for all Aβ peptides tested shorter than 40 amino acids ( Fig. 2B,C )). Thus, both IMT, DV2-103, and IMT isomer 1 are γ-secretase modulators. Remarkably, while each compound modulates γ-secretase activity, most conspicuously at 5μM, this effect vanishes at 10μM, relative to controls. Download figure Open in new tab Fig. 2. IMT, DV2-103, and IMT isomer 1 are γ-secretase modulators. (A) N2a695 cells incubated with IMT, DV2-103 and IMT isomer 1 lower levels of Aβ40 and 42 more than Aβ38, especially at a drug concentration of 5μM, as measured by ELISA. Means differ significantly for IMT and DV2-103 compared to DMSO controls, N = 3 x 3. Data for Isomer 1 are from a representative sample. Differences between means for (A)are analyzed by One-way Anova for IMT and DV2-103 treated cells. Differences among means comparing 5μM IMT and DMSO controls (B,C)) are analyzed by Student’s T test (S.E.M.). Subsequently, we screened all IMT derivatives shown in Table 1 for levels of Aβ40, Aβ38, and Aβ42 peptides. We found that the majority of compounds reduced production of Aβ40 and Aβ42 more than Aβ38 peptide ( Table 2 and Supporting Information (SI) Fig. S-1). This indicates that isomer 1 and analogs modulate γ-secretase cleavage of C-terminal APP since the differences in lengths of these peptides is determined by γ-secretase according to differences in utilization of the APP γ-secretase cleavage sites. View this table: View inline View popup Download powerpoint Table 2. Isomeric IMT analogs are γ-secretase modulators a . To test whether these compounds lower Aβ levels by affecting the BACE and/or γ-secretase cleavages, we examined their effects using wild-type (wt) cells transfected with APP-FL and APP βCTF (C99), respectively. BACE inhibitor, MK8931, and γ-secretase inhibitor, DAPT, were used as controls and the experiment was performed and processed as described previously 13 . The results shown in Fig. 3A and 3B revealed that all 4 compounds reduced β- and γ-cleavages of APP similarly to IMT. There were reductions in Aβ production in both cases, but more so in cells transfected with full-length APP indicating that these compounds, like IMT ( Netzer et al. 2017 ), reduce both BACE and γ-secretase cleavages of APP but that attenuation of BACE processing accounted for the greater part of Aβ reduction ( Fig. 3A, B ). None of these compounds showed any toxicity to N2a695 cells at 10µM concentration ( Fig. 3C ). This was assessed by measuring the percentage of viable cells in the drug treated groups compared to the DMSO control after 5 hours incubation under the conditions used for the Aβ assay. Download figure Open in new tab Fig. 3. IMT isomer 1 and its analogs 1d , 1k and 1s lower BACE and γ-secretase cleavage of APP and lower levels of Aβ in N2a cells transiently transfected with APP 695 (left graph) or APP C99 (right graph): (A) full length APP695 (APP-FL) or (B) APP C99 (β-CTF). (C) Percentage of viable wild-type (WT) N2a cells upon treatment with IMT isomers 1 , cpd. 1d , cpd. 1k and cpd . 1s compared to DMSO control under the same conditions used to test Aβ production. (D) Effects of isomer 1 on Abl1 kinase in vitro. (E) Brain and plasma concentrations of isomer 1 and cpd. 1d in 2 months old WT mice 4 hours after i.p. injection of 50 mg/kg of each drug. Data for A-C are from representative samples. IMT inhibits Abl1 kinase with low nanomolar affinity. Earlier, we prepared and evaluated numerous IMT analogs to find that many of these analogs reduced Aβ levels in cells similarly to IMT, while inhibiting Abl kinase less potently, compared to IMT. In other words, there isn’t a good correlation between the Abl kinase inhibitory activity vs. the Aβ lowering effects in cells contacted with the IMT analogs. We have evaluated IMT isomer 1 to find that it inhibits Abl kinase less potently (IC 50 : 1.172 µM) ( Fig. 3D ) than IMT IC 50 : 0.038 µM)( Buchdunger et al. 1996 ), while it reduced Aβ levels more potently than IMT ( Fig. 1C ). This result further reinforces our prior observation that there is no or little connection between the Aβ-lowering activity of IMT and its inhibition of Abl1 kinase( Netzer et al. 2003 ). Finally, we tested the brain permeability of compound 1d by administering it to 2 months old mice. Plasma and brain tissue were collected 4 hours post drug administration, and LC-MS/MS analysis of the acetonitrile and ethanol extracts was used to measure drug concentration. Compound 1d possesses similarity to IMT isomer- 1a and is isomeric to an ABG-179( Sun et al. 2019 ) analog that possessed a benzene instead of the pyridine (A) ring (see: Fig. 1 for the ring numbering). Earlier, we have shown that ABG-179 possesses superior brain exposure compared to IMT and reduced both Aβ40 and 42 levels significantly in AD mice when delivered acutely for 5 days, and now found that compounds 1d and ABG-179 possess comparable brain exposure but the latter possess superior plasma half-life ( Fig. 3E )( Sun et al. 2019 ). 4 Discussion Based on the number and variety of chemically distinct compounds that produce the same biochemical effects on APP metabolism( Druker et al. 1996 ; Nagar et al. 2002 ; Netzer et al. 2017 ) and that all active compounds are active at low micromolar concentration, we postulate that IMT, DV2-103 and their analogs are likely to produce their effects on APP metabolism by virtue of their physical rather than stereological properties. For example, physical properties would include acting as a weak base that would cause these molecules to be lysosomotropic. We came to this conclusion by showing that the effects of IMT and DV2-103 on APP metabolism are dependent on acidified lysosomes( Netzer et al. 2017 ). Reduced dependence on stereological factors would suggest that IMT, DV2-103 and their active derivatives might bind to a polyspecific receptor where binding is less dependent on structural and electrostatic complementarity. As a first step to test this, we designed derivatives of IMT, referred to here as IMT isomers, and then synthesized derivatives of these compounds and tested their effects on APP metabolism. Our goal was to make a large change in the structure of IMT that would destroy a structural/stereological pharmacophore but still maintain IMT’s physical properties, in particular its property as a weak base, which is necessary for its sequestration in lysosomes through ion trapping( Burger et al. 2015 ). These properties are preserved in IMT isomer 1 , and we demonstrated that isomer 1 not only lowers levels of Aβ peptide in N2a 695 cells but also lowers BACE processing of APP, and in each case with equal activity or more potently than IMT, while producing the same metabolites of APP in cells exposed to IMT. Other IMT isomers were synthesized and a subset of these recapitulated IMT’s APP phenotype. Isomers 1 and 1d also showed increased brain accumulation compared to IMT administered in mice. Also, isomer 1 inhibited Abl kinase activity with over a 100 fold reduction in potency compared to previously published reports of IMT ( Buchdunger et al. 1996 ). Although we had compared the relative effects of γ-secretase and BACE modulation of Aβ generation in cells, we could not rule out that the lowering of Aβ and sAPPβ was not a result of IMT’s effect of stimulating autophagy, since autophagy was previously shown to accelerate lysosomal degradation of APP-βCTF and Aβ( Tian et al. 2011 ). Nevertheless, we show that IMT, DV2-103, and the IMT isomers tested in this study are modulators of γ-secretase by virtue of the observation that their Aβ-lowering potency differentially affects Aβ peptide lengths depending on drug concentration. Remarkably, Aβ1-42 production is lowered at 5μM drug concentrations, while Aβ1-38 production is inhibited least and in some cases raised. This is important because relative heightened production of Aβ38 has been considered benign, and more recently therapeutic( Cullen et al. 2022 ), while lowered production of Aβ42 is considered therapeutic; in either case, a decrease in Aβ peptide aggregation may occur. 5 Conclusion In summary, we suggest that IMT and a subset of its isomers, and related lysosomotropic drugs affect APP metabolism through a lysosomal mechanism that increases trafficking of full-length APP directly to lysosomes where it is degraded, thus causing APP to evade processing by BACE and γ-secretase in earlier secretory compartments along the amyloidogenic pathway. The fact that many of these compounds (structurally related or not) are γ-secretase modulators is consistent with a mechanism involving altered trafficking of APP that affects the specificity of γ-secretase cleavage sites in the formation of Aβ peptides. 6 Data availability statement Synthetic methods and analytical data for new compounds and their effects on Abeta levels depicted as a bar graph. AUTHOR INFORMATION Corresponding Authors. William J. Netzer, E-mail: billnetzer{at}gmail.com , Anjana Sinha, Email: sinhaanjana5819{at}gmail.com , and Subhash C. Sinha, E-mail: sus2044{at}med.cornell.edu . ORCID 0000-0001-8916-5677 Notes: The authors declare no competing financial interest. ACKNOWLEDGEMENTS We are thankful to Dr. Paul Greengard (Deceased) of the Rockefeller University for his enthusiastic support to this work and Dr. Victor H. Bustos for helpful discussion. Funding support from JPB (#322 and #839 to SCS) and Fisher Center for Alzheimer’s Research Foundation (PG) is duly acknowledged. References ↵ Afanasyev , Oleg I. , Ekaterina Kuchuk , Dmitry L. Usanov , and Denis Chusov . 2019 . ‘ Reductive Amination in the Synthesis of Pharmaceuticals ’, Chemical reviews , 119 : 11857 – 911 . OpenUrl ↵ Buchdunger , E. , J. Zimmermann , H. Mett , T. Meyer , M. Müller , B. J. Druker , and N. B. Lydon . 1996 . ‘ Inhibition of the Abl protein-tyrosine kinase in vitro and in vivo by a 2-phenylaminopyrimidine derivative ’, Cancer Res , 56 : 100 – 4 . OpenUrl Abstract / FREE Full Text ↵ Burger , H. , A. T. den Dekker , S. Segeletz , A. W. Boersma , P. de Bruijn , M. Debiec-Rychter , T. Taguchi , S. Sleijfer , A. Sparreboom , R. H. Mathijssen , and E. A. Wiemer . 2015 . ‘ Lysosomal Sequestration Determines Intracellular Imatinib Levels ’, Mol Pharmacol , 88 : 477 – 87 . OpenUrl Abstract / FREE Full Text ↵ Cullen , N. , S. Janelidze , S. Palmqvist , E. Stomrud , N. Mattsson-Carlgren , and O. Hansson . 2022 . ‘ Association of CSF Aβ(38) Levels With Risk of Alzheimer Disease-Related Decline ’, Neurology , 98 : e958 – e67 . OpenUrl CrossRef ↵ Druker , Brian J. , Shu Tamura , Elisabeth Buchdunger , Sayuri Ohno , Gerald M. Segal , Shane Fanning , Jürg Zimmermann , and Nicholas B. Lydon . 1996 . ‘ Effects of a selective inhibitor of the Abl tyrosine kinase on the growth of Bcr–Abl positive cells ’, Nature Medicine , 2 : 561 – 66 . OpenUrl CrossRef PubMed Web of Science ↵ Golde , T. E. , E. H. Koo , K. M. Felsenstein , B. A. Osborne , and L. Miele . 2013 . ‘ γ-Secretase inhibitors and modulators ’, Biochim Biophys Acta , 1828 : 2898 – 907 . OpenUrl CrossRef PubMed Web of Science ↵ Hur , Ji-Yeun . 2022 . ‘ γ-Secretase in Alzheimer’s disease ’, Experimental & Molecular Medicine , 54 : 433 – 46 . OpenUrl ↵ Jester , Benjamin W. , Kurt J. Cox , Alicia Gaj , Carolyn D. Shomin , Jason R. Porter , and Indraneel Ghosh . 2010 . ‘ A Coiled-Coil Enabled Split-Luciferase Three-Hybrid System: Applied Toward Profiling Inhibitors of Protein Kinases ’, Journal of the American Chemical Society , 132 : 11727 – 35 . OpenUrl CrossRef PubMed ↵ Kazmi , F. , T. Hensley , C. Pope , R. S. Funk , G. J. Loewen , D. B. Buckley , and A. Parkinson . 2013 . ‘ Lysosomal sequestration (trapping) of lipophilic amine (cationic amphiphilic) drugs in immortalized human hepatocytes (Fa2N-4 cells) ’, Drug Metab Dispos , 41 : 897 – 905 . OpenUrl Abstract / FREE Full Text ↵ Kim , C. K. , Y. R. Lee , L. Ong , M. Gold , A. Kalali , and J. Sarkar . 2022 . ’Alzheimer’s Disease: Key Insights from Two Decades of Clinical Trial Failures’ , J Alzheimers Dis , 87 : 83 – 100 . OpenUrl ↵ Kumar , Devendra , Ankit Ganeshpurkar , Dileep Kumar , Gyan Modi , Sanjeev Kumar Gupta , and Sushil Kumar Singh . 2018 . ’Secretase inhibitors for the treatment of Alzheimer’s disease: Long road ahead’ , European Journal of Medicinal Chemistry , 148 : 436 – 52 . OpenUrl CrossRef ↵ Miranda , Alvaro , Enrique Montiel , Henning Ulrich , and Cristian Paz . 2021 . ‘ Selective Secretase Targeting for Alzheimer’s Disease Therapy ’, Journal of Alzheimer’s Disease , 81 : 1 – 17 . OpenUrl ↵ Miyaura , Norio , and Akira Suzuki . 1995 . ‘ Palladium-Catalyzed Cross-Coupling Reactions of Organoboron Compounds ’, Chemical reviews , 95 : 2457 – 83 . OpenUrl CrossRef Web of Science ↵ Nagar , B. , W. G. Bornmann , P. Pellicena , T. Schindler , D. R. Veach , W. T. Miller , B. Clarkson , and J. Kuriyan . 2002 . ‘ Crystal structures of the kinase domain of c-Abl in complex with the small molecule inhibitors PD173955 and imatinib (STI-571) ’, Cancer Res , 62 : 4236 – 43 . OpenUrl Abstract / FREE Full Text ↵ Netzer , W. J. , K. Bettayeb , S. C. Sinha , M. Flajolet , P. Greengard , and V. Bustos . 2017 . ‘ Gleevec shifts APP processing from a β-cleavage to a nonamyloidogenic cleavage ’, Proc Natl Acad Sci U S A , 114 : 1389 – 94 . OpenUrl Abstract / FREE Full Text ↵ Netzer , W. J. , F. Dou , D. Cai , D. Veach , S. Jean , Y. Li , W. G. Bornmann , B. Clarkson , H. Xu , and P. Greengard . 2003 . ‘ Gleevec inhibits beta-amyloid production but not Notch cleavage ’, Proc Natl Acad Sci U S A , 100 : 12444 – 9 . OpenUrl Abstract / FREE Full Text ↵ Panza , Francesco , Vincenzo Solfrizzi , Vincenza Frisardi , Cristiano Capurso , Alessia D’Introno , Anna M. Colacicco , Gianluigi Vendemiale , Antonio Capurso , and Bruno P. Imbimbo . 2009 . ‘ Disease-Modifying Approach to the Treatment of Alzheimer’s Disease ’, Drugs & Aging , 26 : 537 – 55 . OpenUrl CrossRef PubMed ↵ Portelius , E. , B. Van Broeck , U. Andreasson , M. K. Gustavsson , M. Mercken , H. Zetterberg , H. Borghys , and K. Blennow . 2010 . ‘ Acute effect on the Aβ isoform pattern in CSF in response to γ-secretase modulator and inhibitor treatment in dogs ’, J Alzheimers Dis , 21 : 1005 – 12 . OpenUrl ↵ Ruiz-Castillo , Paula , and Stephen L. Buchwald . 2016 . ‘ Applications of Palladium-Catalyzed C–N Cross-Coupling Reactions ’, Chemical reviews , 116 : 12564 – 649 . OpenUrl CrossRef PubMed ↵ Rynearson , Kevin D. , Moorthi Ponnusamy , Olga Prikhodko , Yuhuan Xie , Can Zhang , Phuong Nguyen , Brenda Hug , Mariko Sawa , Ann Becker , Brian Spencer , Jazmin Florio , Michael Mante , Bahar Salehi , Carlos Arias , Douglas Galasko , Brian P. Head , Graham Johnson , Jiunn H. Lin , Steven K. Duddy , Robert A. Rissman , William C. Mobley , Gopal Thinakaran , Rudolph E. Tanzi , and Steven L. Wagner . 2021 . ‘ Preclinical validation of a potent γ-secretase modulator for Alzheimer’s disease prevention ’, Journal of Experimental Medicine , 218 . ↵ Sinha , A. , K. Gindinova , E. Mui , W. J. Netzer , and S. C. Sinha . 2019 . ‘ Development of Kinase Inactive PD173955 Analogues for Reducing Production of Aβ Peptides ’, ACS Med Chem Lett , 10 : 1430 – 35 . OpenUrl ↵ Sun , W. , W. J. Netzer , A. Sinha , K. Gindinova , E. Chang , and S. C. Sinha . 2019 . ‘ Development of Gleevec Analogues for Reducing Production of β-Amyloid Peptides through Shifting β-Cleavage of Amyloid Precursor Proteins ’, J Med Chem , 62 : 3122 – 34 . OpenUrl ↵ Tian , Y. , V. Bustos , M. Flajolet , and P. Greengard . 2011 . ‘ A small-molecule enhancer of autophagy decreases levels of Abeta and APP-CTF via Atg5-dependent autophagy pathway ’, Faseb j , 25 : 1934 – 42 . OpenUrl CrossRef PubMed Web of Science ↵ Zhao , J. , X. Liu , W. Xia , Y. Zhang , and C. Wang . 2020 . ’Targeting Amyloidogenic Processing of APP in Alzheimer’s Disease’ , Front Mol Neurosci , 13 : 137 . OpenUrl View the discussion thread. Back to top Previous Next Posted July 17, 2024. Download PDF Supplementary Material Email Thank you for your interest in spreading the word about bioRxiv. NOTE: Your email address is requested solely to identify you as the sender of this article. Your Email * Your Name * Send To * Enter multiple addresses on separate lines or separate them with commas. You are going to email the following Stretching the structural envelope of isomeric imatinib analogs that reduce β-amyloid production by modulating both β- and γ-secretase cleavages of APP Message Subject (Your Name) has forwarded a page to you from bioRxiv Message Body (Your Name) thought you would like to see this page from the bioRxiv website. Your Personal Message CAPTCHA This question is for testing whether or not you are a human visitor and to prevent automated spam submissions. Share Stretching the structural envelope of isomeric imatinib analogs that reduce β-amyloid production by modulating both β- and γ-secretase cleavages of APP William J. Netzer , Anjana Sinha , Mondana Ghias , Emily Chang , Katherina Gindinova , Emily Mui , Ji-Seon Seo , Subhash C. Sinha bioRxiv 2024.07.14.602669; doi: https://doi.org/10.1101/2024.07.14.602669 Share This Article: Copy Citation Tools Stretching the structural envelope of isomeric imatinib analogs that reduce β-amyloid production by modulating both β- and γ-secretase cleavages of APP William J. Netzer , Anjana Sinha , Mondana Ghias , Emily Chang , Katherina Gindinova , Emily Mui , Ji-Seon Seo , Subhash C. Sinha bioRxiv 2024.07.14.602669; doi: https://doi.org/10.1101/2024.07.14.602669 Citation Manager Formats BibTeX Bookends EasyBib EndNote (tagged) EndNote 8 (xml) Medlars Mendeley Papers RefWorks Tagged Ref Manager RIS Zotero Tweet Widget Facebook Like Google Plus One Subject Area Pharmacology and Toxicology Subject Areas All Articles Animal Behavior and Cognition (7646) Biochemistry (17728) Bioengineering (13917) Bioinformatics (42038) Biophysics (21489) Cancer Biology (18637) Cell Biology (25554) Clinical Trials (138) Developmental Biology (13403) Ecology (19941) Epidemiology (2067) Evolutionary Biology (24368) Genetics (15624) Genomics (22547) Immunology (17764) Microbiology (40475) Molecular Biology (17208) Neuroscience (88756) Paleontology (667) Pathology (2842) Pharmacology and Toxicology (4834) Physiology (7659) Plant Biology (15175) Scientific Communication and Education (2047) Synthetic Biology (4304) Systems Biology (9835) Zoology (2272)