Camptothecin-Carrying Cuco2s4 Nanoparticles: Sustained Drug Release And Anticancer Activity

Camptothecin-Carrying Cuco2s4 Nanoparticles: Sustained Drug Release And Anticancer Activity | 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 Camptothecin-Carrying Cuco 2 s 4 Nanoparticles: Sustained Drug Release And Anticancer Activity Archana Sumohan Pillai, Aleyamma Alexander, Varnitha Manikantan, and 3 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-1190820/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract Nanocarriers of anticancer drugs are delicately designed with precision addition at every attempt. In this paper, we report CuCo 2 S 4 nanoparticles that show light-absorption in the NIR-II wavelength range and possess magnetic characteristics. The synthesized nanoparticles are characterized employing XRD, SEM, DLS, TGA, and XPS methods. The nanoparticles form a composite with biocompatible polymeric β-cyclodextrin. The nanoparticles possess a band gap energy of 2.25 eV. The magnetic property arises due to the cobalt-incorporation in the nanoparticles. The anticancer drug, camptothecin, is loaded in the nanocarrier with an 89% adsorption efficiency. The in vitro release of the drug occurs in a sustained fashion. Further, the in vitro anticancer potential of the nanocarrier is examined on breast cancer (MDA=MB-231) cell lines and the activities of the free- and the drug-loaded nanocarrier are compared. The cobalt-containing copper sulfide nanoparticle-poly-β-cyclodextrin composite works as a promising nanocarrier of camptothecin. Materials Chemistry Polymer Science Sulfide nanoparticles superparamagnetism NIR absorption nanocarrier anticancer activity. Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 1 Introduction Nanoparticles (NPs) have been investigated and utilized, with an increasing interest, as targeted drug delivery systems and for other biomedical applications. 1 – 3 The nanoparticles employed as drug carriers are designed as possessing properties that render them respond to stimuli like magnetic field, light, and pH. 4 – 6 These properties enable them to find application in diagnosis and treatment of chronic diseases like cancer. For instance, magnetic resonance imaging (MRI), and magnetic hyperthermia, and magnetic field-assisted anticancer drug-delivery are possible when magnetic NPs are used. 7 – 9 Some engineered NPs absorb near-infrared (NIR) radiation and heat up, thus acting as photothermal heating agents which allows the selective ablation of cancer cells. 10 Moreover, magnetic-luminescent NPs also have been developed with a focus on multiple-modal imaging. 11 In a nutshell, delicate design and synthesis of NPs with tailor-made characteristics have occupied a place of paramount importance in the field of targeted drug delivery. Besides the elemental composition of inorganic NPs used for biomedical applications, their size, shape, and surface modification have to be customized to make them effective therapeutic carriers. 12 , 13 The size of the NPs are desired to be in the range of 50-150 nm to make them reside longer in the circulatory system. 14 In addition, NPs with large aspect ratios undergo longer plasma residence. 15 Another essential requirement is to modify the surface of the NPs to render them biocompatible, dispersible in aqueous medium, and to enable the loading of anticancer drugs. 16 , 17 Biocompatible polymers serve the above-mentioned purposes. The loading of anticancer drug molecules in the polymer sheath that covers the inorganic NPs occurs mostly through non-covalent interactions. 18 For instance, the drug molecule gets inserted between the coiled polymeric strands. Nevertheless, an intriguing method of loading the drug is through the formation of supramolecular host: guest complexes on the surfaces of NPs. 19 It enables the loading of a larger am’ount of drugs. The inorganic NP-biocompatible polymer combination acts as a suitable choice of nanocarrier with precision-added properties. Copper sulfide NPs have emerged as new generation materials for photothermal therapy (PTT) as they display a large NIR absorption. 20 Attempts have been made to dope elements into CuS NPs to further fine-tune their properties. 21 , 22 We hypothesize that the substitution of a magnetic element like cobalt can make them magnetic while remaining NIR- absorbing materials. In addition, a nanocomposite (NC) of Co-doped copper sulfide with a carbohydrate polymer possessing cavity-containing molecular component can aid achieving an enhanced drug-loading and biocompatibility. For this purpose, we chose a β-cyclodextrin-containing polysaccharide as the coating polymer (poly-CD). Herein, we report the synthesis and characterization of Co-doped copper sulphide, preparation of its NC with the poly-CD, of the anticancer drug, camptothecin (CPT), and the anticancer potential of the drug loaded NC on breast cancer cell lines. 2 Materials And Methods 2.1 Materials The material needed for this work, namely copper chloride and cobalt nitrate, was obtained from Fischer Scientific. Thiourea was procured from Sigma Aldrich and ethyl acetate from NICE chemicals. Triethylamine and Cetyltrimethylammoniumbromide (CTAB) was obtained from Scientific chemical distributors. Analytical grade chemicals with purity above 98% was used. 2.2 Methods 2.2.1 Synthesis of CuCo 2 S 4 nanoparticles The β-CD polymer (poly-CD) was synthesized via a reported procedure. 23 A hydrothermal synthesis method was employed to synthesize poly-CD-coated CuCo 2 S 4 NPs. Copper chloride and thiourea was mixed in the ratio 1:3 respectively in 20 ml deionized water and subjected to stirring for 15 minutes. To the above solution, 0.02 M cobalt nitrate dissolved in 10 ml of water was added followed by addition of 0.2 g of CTAB and 0.2 g poly-CD. The mixture was then added to 6 ml triethylamine dissolved in 10 ml of ethyl acetate. It was sonicated for 2 hours at room temperature and then transferred to a Teflon stainless steel autoclave. The temperature was maintained at 180 °C for 18 hours. The obtained poly-CD-coated-CuCo 2 S 4 nanoparticles were alternatively washed with distilled water and ethanol to remove unreacted chemicals. The NPs were dried at room temperature. 2.2.2 Instrumentation The synthesized NPs were characterised for their crystalline phase using X-ray powder diffraction (Bruker D8-Advance) using Cu kα (λ = 0.154 Å) radiation in the range of 10–80°. High-resolution transmission electron microscope (HR TEM–Jeol/JEM 2100) was used to determine the topography, morphology and size of the nanoparticles. The composition of the NPs was determined using Jeol/JEM 2100 Energy dispersive X-ray (EDX) instrument. ICP-MS-Thermofisher CAP RQ ICP-MS device was used to measure elemental composition. The amount and the rate of weight changes of the nanoparticles with respect to the temperature was determined using Thermogravimetric Analysis (TGA), TA instruments, USA Model SDT Q600. The elemental composition was measured using a Thermofisher Scientific Model Nexsa bas X-ray photoelectron spectroscopy (XPS). M-H curves to determine the magnetic property were obtained using a Lake-shore 7410 Vibrating Sample Magnetometer (VSM). 2.2.3 In vitro drug release In vitro drug release was studied for NPs loaded with a chemotherapeutic drug camptothecin (CPT) and packed in a dialysis bag of MWCO 12-14 kDa. The bag was suspended in PBS solutions of pH 6.5 and 7.4. The bags were continuously stirred at room temperature. Samples were collected at regular time intervals. After sample collection, fresh PBS solution was added to maintain the pH throughout the release study. UV-vis absorption at 430 nm was recorded for each sample at each time point. The concentration of the released drug was determined by the standard calibration curve equation. The mean values for three independent time estimates were calculated. The percentage of drug loading and release was calculated using following equations: 2.2.4 Cell line culture and condition Triple negative breast cancer cell line MDA-MB-231, was obtained from NCCS, Pune and cultured in liquid medium i.e., Dulbecco’s Modified Eagle High Glucose (DMEM-HG) that was supplemented with 10% Fetal Bovine Serum Albumin (FBS), 1% sodium bicarbonate and 1% sodium pyruvate at 37 °C in a 5% CO 2 incubator. The cells were trypsinized using trypsin EDTA, and seeded in a 96 well plate for MTT and 6 well plate for flow cytometry analysis. 2.2.5 In-vitro MTT assay MDA-MB-231 cells were trypsinized and grown in a 96-well plate at a density of 1 × 10 6 cells/ml and incubated for 24 h. The cells were treated with free nanocarrier (poly-CD-coated NPs) and drug (CPT)-loaded nanocarriers. The concentrations of treatments were in the range of 0.25 μg/mL to 15 μg/mL. The sample was replicated three times It was then incubated for 24 h at 37°C. After incubation, the culture medium was removed from the treated plate, and 100 μL of MTT (0.5 mg/mL) was added to each well and incubated at 37 °C for 4 h. The resulting formazan crystals were dissolved in 100 μL of DMSO. The absorbance intensity was measured using a microplate reader (RT-2100 C) at 570 nm. The percentage o viable cells was plotted and the IC 50 value was calculated using GraphPad Prism 8.0 software. 3 Results And Discussion 3.1 Characterization of the NPs The Cu and Co-containing sulfide NPs were synthesized using a hydrothermal method. The synthesized NPs were coated with poly-CD. The polymer-coated NPs display an X-ray diffraction pattern as in Fig. 1 A. As the poly-CD possesses a crystalline nature, broad peaks are not obvious in the XRD. The sharp peaks correspond to the sulfide NPs. The prominent peaks are observed at 2θ 27.12° (2 2 0), 31.3° (3 1 1), 32.4° (2 2 2), 47.3° (4 2 2), 52.3° (3 3 3), 58.8° (5 3 1), and 68.9° (4 4 4). These reflections correspond to the CuCo 2 S 4 crystals of the face-centered cubic system. It is confirmed by their congruence with the JCPDS card number: 75-1570. The average grain size of the CuCo 2 S 4 crystals was determined employing the Debye-Scherrer expression: $$\text{D} =\frac{0.94{\lambda }}{{\beta } \text{c}\text{o}\text{s}{\theta }}$$ 3 where \({\lambda }, {\beta }, \text{a}\text{n}\text{d} {\theta }\) represent the wavelength of Cu (Kα), full-width-half-maximum in radians, and the angle of the most intense peak respectively. The crystalline size (D) is determined as 13.36 nm. Further, the dislocation density of the crystals was determined employing the following relation: $$\text{D}\text{i}\text{s}\text{l}\text{o}\text{c}\text{a}\text{t}\text{i}\text{o}\text{n} \text{d}\text{e}\text{n}\text{s}\text{i}\text{t}\text{y}, \delta =\frac{1}{{D}^{2}}$$ 4 where D represents the average crystallite size. The \(\delta\) value is 6.5 x 10 -3 nm - 1 . As the polymer contributes an extent of non-crystallinity to the sample of NPs, crystallinity index should be calculated. In a method of calculating the crystallinity, we utilized the following relation: $$\text{C}\text{r}\text{y}\text{s}\text{t}\text{a}\text{l}\text{l}\text{i}\text{n}\text{i}\text{t}\text{y} \text{I}\text{n}\text{d}\text{e}\text{x}, CI=\frac{Area of all the crystalline peaks}{Area of all the crystalline and amorphous peaks}$$ 5 The crystallinity thus determined is 46%. Figure 1 B shows the TEM images of the poly-CD-coated CuCo 2 S 4 NPs. Nucleated nano-structures are observed in hexagonal shapes. Some of them appear sideways in orientation revealing that the formed NPs are hexagonal plate-shaped NPs. These nanostructures are consistent with the observation in crystal data revealed by the XRD pattern. The mean length and width of the NPs are 120.71 and 42.79 nm respectively. The poly-CD has coated the NPs as a thin layer. In addition, such polymers are transparent in TEM and do not appear vivid on the dark contrast-sulfide NPs. 24 The size of the polymer-coated NPs was further confirmed by carrying out a particle-size analysis by dynamic light scattering experiment. The DLS shows a histogram with the mean size of the NPs as 122 nm. (Fig. 1 C). The size range matches with that observed in the TEM images. The elemental composition of the as-synthesized CuCo 2 S 4 NPs was determined qualitatively, employing EDX spectroscopy. The EDX of the NPs is displayed in Fig. 2 A. The peaks in the figure correspond to the elements viz., Cu, Co, and S. additional peaks pertaining to any other impurity is not observed. Thus, the EDX reveals the formation of pure CuCo 2 S 4 NPs. To add further evidence for such composition of the NPs and to determine the presence of various elements in specific electronic states, X-ray photoelectron spectra were recorded. For XPS measurements, the poly-CD-coated NPs were used. Figs. 2 B–F depict the Cu, Co, S, C, and O peaks of the XPS pattern. The 2p 3/2 and 2p 1/2 states of Cu appear at 932.25 and 952.20 eV respectively. (Fig. 2 B). the deconvoluted XPS of Co shows peaks at 779.64 and 796.33 eV due to the 2p 3/2 and 2p 1/2 electronic states. (Fig. 2 C). Fig. 2 D depicts the 2p 3/2 and 2p 1/2 peaks at 161.68 and 162.81 eV respectively. These spectra arise from the cobalt-substituted copper sulfide. The spectra C and O are contributed by the polymer that resides on the surface of the CuCo 2 S 4 NPs. The carbons of the C-O-C, C-C, and O-C=O functional groups display peaks at 284.04, 284.85, and 285.92 eV respectively, arising from the C 1s state. (Fig. 2 E). the O 1s peaks of the C-O and C=O groups show peaks at the binding energy values viz., 532.38 and 531.63 eV. The XPS peaks and their positions revel the composition of the poly-CD-coated CuCo 2 S 4 NPs. Additional peaks due to any impurity is not observed in the XPS data. It is apparent from the results discussed above that the synthesized material carries CuCo 2 S 4 NPs with the poly-CD immobilized on their surface. To further evaluate the amount of poly-CD on the surface of the NPs, thermogravimetric profiles of the coated CuCo 2 S 4 NPs were measured. Fig. 3 shows an initial weight loss of 4.42% and then plummeting down of the weight up to 12.46%. these decay profiles imply that the polymeric structure on the surface changes conformation and then starts melting. The melting is sharp at about 440°C. above this temperature, there is complete degradation of the coated polymer and it extends up to above 750°C. the residue remains with a weight percentage of 79.86 with an implication that the coated polymer offers about 20% of weight to the composite nanostructure. 3.2 Optical and magnetic characteristics of the NPs The UV-Vis absorption spectrum of the poly-CD-coated CuCo 2 S 4 NPs is shown in Fig. 4 A. the band at the visible spectral region is shorter (lower absorbance) than the NIR band, which is vividly due to the 3d electrons of the transition metals in the NPs. 25 The stronger absorption is further contributed by the Co incorporated into the crystals, which makes the normally observed CuS absorption spectrum enhance in absorbance at the NIR wavelength. 26 The aqueous dispersion of the poly-CD coated NPs, displaying such an NIR absorption, presents them suitable for photothermal heating. Further, the absorption wavelength was determined by an extrapolation of the line to the linear portion of the absorption spectrum. The optical band gap was determined employing the relation: $$Eg=\frac{hc}{\lambda }$$ 6 where h represents the Planck’s constant, c, the velocity of light, and λ is the wavelength derived from the tail of absorption. In the present work, the energy band gap calculated is Eg = 2.25 eV (Fig. 4 B). this value is slightly different from the reported secondary band gap of 2.12 eV for a smaller size CuCo 2 S 4 NPs. 27 Furthermore, it is larger than the band gap reported for CuS NPs (1.80 eV). 28 The photoluminescence spectrum of the poly-CD-coated CuCo 2 S 4 NPs is shown in Fig. 4 C. The luminescence spectral band is broad and intense, showing wavelength of emission at 510 nm. This property presents the NPs to be followed employing luminescence imaging. The magnetic properties of the poly-CD-coated NPs were investigated by employing VSM. The room-temperature magnetization property, with n applied sweeping magnetic field, of the NPs is displayed in Fig. 4 D. the magnetization curve shows a small coercivity value of 78.09 Oe which is close to superparamagnetic limit. 29 The saturation magnetization (Ms) value is 0.024 emu g - 1 . Such magnetic NPs attain saturation quickly at the application of an external magnetic field. In addition, they relax quickly due to the absence of broad hysteresis loops. Therefore, the synthesized NPs are suitable for application in bio-imaging using MRI. 30 3.3 Drug-loading and release The polymer-coating on the NPs poses the advantages viz., (i) it makes the NPs dispersible in aqueous medium, (ii) renders them biocompatible, and (iii) enables the loading of organic therapeutic molecules. A drug carrier should accommodate an appreciable amount of the therapeutics. In addition, the release of the drug is preferred to be sustained and slow. 31 Therefore, the drug loading and the drug-release kinetics were investigated in vitro. The experimental procedures have been elaborated in the methods section. The percentage of the model drug CPT, loaded in the poly-CD-coated CuCo 2 S 4 NPs (nanocarrier) is estimated employing the equation ( 1 ). The percentage of adsorbed CPT on the nanocarrier is 89.04%. It implies a large amount of drug is loaded on the nanocarrier. CPT forms an inclusion complex with the β-CD molecule. 32 Therefore, the poly-CD is a suitable choice to be employed as the polymeric host structure that is immobilized on the surface of the NPs. The CPT release profile from the nanocarrier (Fig. 5 ) reveals that it is disseminated in a sustained fashion, well over a period of 360 hours at the physiological pH of 7.4. The cumulative release of CPT at 24 hours is 25%. In addition, the release rate is accelerated when the pH is made slightly acidic (6.5). In this lower pH, release is enhanced to 40% at 24 hours. Therefore, it is obviously seen that the drug release is pH dependent and it is tunable with the application of a stimulus. It is worth mentioning that the tumor microenvironment is acidic in nature. 33 Therefore, the designed nanocarrier is suitable for selective release of CPT at the tumor site. 3.4 In vitro anticancer activity The in vitro anticancer activities of the free- and the nanocarrier were carried out on human breast cancer cell lines (MDA-MB-231). After an incubation period of 24 hours as described in the experimental section, the cytotoxicity was analyzed employing an MTT assay. Figs. 6 A and B represent the percentage cell viability as a function of concentration of the free- and CPT-loaded nanocarriers respectively. The cytotoxicity is observed to be dose-dependent in both the cases. Nevertheless, it is more in the case of CPT-loaded nanocarrier than the free-nanocarrier. The half-maximal inhibitory concentration (IC 50 ) of the free nanocarrier is 161.5 µg mL - 1 , indicating that the cells are considerably viable at the addition of the nanocarrier alone. Contrary to that, the loading of CPT in the nanocarrier leads to an enhancement of the anticancer activity. The IC 50 value of the CPT-loaded nanocarrier is 68.93 µg mL - 1 . Therefore, it can be concluded that the loaded drug gets slowly released from the nanocarrier and destroys the cancer cells. Further, all of the amount of loaded CPT is not released at the time period of 24 hours. The cumulative release as discussed in the previous section suggests a 40% of CPT disseminated over a period of 24 hours. Therefore, it follows that the anticancer activity occurs due to the portion of the CPT that is released from the nanocarrier. The reported IC 50 of the free-CPT is indeed more than the CPT-loaded nanocarrier reported herein. Nevertheless, it should be noted that such direct application of CPT to the culture of cells renders the former rapidly and unrestrictedly diffusing into mammalian cells and hence make them more cytotoxic. 34 This is contrary to the slow internalization of the drug-loaded nanocarrier through the endocytosis pathway. Therefore, the poly-CD-coated CuCo 2 S 4 nanocarrier is a promising nanocarrier of CPT, that possesses additional features of magnetic, NIR absorbing, and photoluminescence characteristics. 4 Conclusions We synthesized CuCo 2 S 4 nanoparticles and coated them with poly-CD. The nanoparticles show a hexagonal crystal structure with a mean size of 122 nm which is a size suitable for employing as a nanocarrier. The NIR-II absorption of the NPs is broad and more intense than the visible spectral band. The optical band gap is 2.25 eV. In addition, the nanocomposite reveals a superparamagnetic behavior due to the presence of incorporated cobalt. It makes them suitable for control by external magnetic field. The poly-CD coating renders the material biocompatible and enables the loading of CPT by means of a non-covalent host : guest association. The drug-release from the nanocarrier takes place over a period of about 360 hours. Further, the drug-release is pH-dependent and it is accelerated at pH 6.0 compared to the normal physiological pH (7.4). the drug-loaded nanocarrier reveals an anticancer efficacy (IC 50 = 68.93 µg mL - 1 ) that presents it as a suitable choice as a CPT-transporter. The cobalt-addition and the poly-CD coating thus add precision to the promising sulfide NPs. Declarations Acknowledgement The research work was carried out without any funding by external agencies. The authors thank the management of KITS for providing necessary facilities. 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Kaliyamoorthi, S. Ramasamy, A. S. Pillai, A. Alexander, A. Arivarasu, I. V. M. V. Enoch, Camptothecin-loaded holmium ferrite nanocarrier. Expanded activity on breast cancer cells. Mater. Lett. 285 , 129164 (2021). https://doi.org/10.1016/j.matlet.2020.129164. K. Kaliyamoorthy, A. S. Pillai, A. Alexander, A. Arivarasu, I. V. M. V Enoch, S. Ramasamy, β-Cyclodextrin-folate functionalized poly (lactic-co-glycolide)–superparamagnetic ytterbium ferrite hybrid nanocarrier for targeted delivery of camptothecin. Mater. Sci. Eng. C 122 , 111796 (2021). DOI: 10.1016/j.msec.2020.111796. Y. Kato, S. Ozawa, C. Miyamoto, Y. Maehata, A. Suzuki, T. Maeda, Y. Baba, Acidic extracellular microenvironment and cancer. Cancer Cell Int. 13 , 89-96 (2013). https://doi.org/10.1186/1475-2867-13-89. L. Kou, J. Sun, Y. Zhai, The endocytosis and intracellular fate of nanomedicine: implication for rational design. Asian J. Pharm. Sci. 8 , 1-10 (2013). https://doi.org/10.1016/j.ajps.2013.07.001. Supplementary Files Graphicalabstract.jpeg Cite Share Download PDF Status: Posted Version 1 posted 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-1190820","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":71832134,"identity":"313de9c6-7aab-488b-81a2-45a10dc1e1d9","order_by":0,"name":"Archana Sumohan Pillai","email":"","orcid":"","institution":"Centre for Nanoscience and Genomics","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Archana","middleName":"Sumohan","lastName":"Pillai","suffix":""},{"id":71832135,"identity":"6f351fe5-c2ff-4e27-b404-be0dd20b6d5a","order_by":1,"name":"Aleyamma Alexander","email":"","orcid":"","institution":"Centre for Nanoscience and Genomics","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Aleyamma","middleName":"","lastName":"Alexander","suffix":""},{"id":71832136,"identity":"51ade55a-2c3a-4a98-aa0b-bd52e5007977","order_by":2,"name":"Varnitha Manikantan","email":"","orcid":"","institution":"Centre for Nanoscience and Genomics","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Varnitha","middleName":"","lastName":"Manikantan","suffix":""},{"id":71832137,"identity":"5d77a2ec-5d0d-4a30-aeea-43042c15ee27","order_by":3,"name":"Govindaraj Sri Varalakshmi","email":"","orcid":"","institution":"Karunya Institute of Technology and Sciences","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Govindaraj","middleName":"Sri","lastName":"Varalakshmi","suffix":""},{"id":71832138,"identity":"7d9619a9-05dc-4f6b-b500-725ae2b699fe","order_by":4,"name":"Bose Allben Akash","email":"","orcid":"","institution":"Karunya Institute of Technology and Sciences","correspondingAuthor":false,"submittingAuthor":false,"prefix":"","firstName":"Bose","middleName":"Allben","lastName":"Akash","suffix":""},{"id":71832139,"identity":"df307e21-ecda-4081-adb4-59dc47663eb4","order_by":5,"name":"Israel V M V Enoch","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAA8klEQVRIiWNgGAWjYDCCA4wNIEoGRDAzVIBI5gaitPBAtJwBkYyEtEAoiBbGNhBFQAvf7cPNnytqbHj4pZsPfi6cVxvN3w7U8qNiG04tkucS2yTPHEvjkZxzLFl65rbjuTMOMzYw9py5jVOLwRnGNsYGtsM8BjdyzJh5tx3LbQBqAboQr5bmjw3//vPYg7XMOZY7nwgtDZKNbQd4DCRAWhpqcjcQ0iIJdJhkY18yj8SNtGRpnmMHcjcCtRzE5xe+M+yPPzZ8s5Pjn5F88DNPTV3uvPOHDz74UYFbCzo4DCYPEK0eCOpIUTwKRsEoGAUjBAAABSRaaFYTKaMAAAAASUVORK5CYII=","orcid":"https://orcid.org/0000-0002-8489-531X","institution":"Centre for Nanoscience and Genomics","correspondingAuthor":true,"submittingAuthor":false,"prefix":"","firstName":"Israel","middleName":"V M V","lastName":"Enoch","suffix":""}],"badges":[],"createdAt":"2021-12-21 06:32:41","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-1190820/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-1190820/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":16781927,"identity":"38739b0a-2620-43ea-8fbe-45a93d58f45f","added_by":"auto","created_at":"2021-12-28 00:37:55","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":249220,"visible":true,"origin":"","legend":"\u003cp\u003e(a) XRD, (B) TEM, and (C) DLS of CuCO\u003csub\u003e2\u003c/sub\u003eS\u003csub\u003e4\u003c/sub\u003e-poly-CD nanocomposites.\u003c/p\u003e","description":"","filename":"Figure1.png","url":"https://assets-eu.researchsquare.com/files/rs-1190820/v1/c4a6575235391082ee35e572.png"},{"id":16781930,"identity":"31edc6be-a3ed-4486-a5e2-409e5d41f17d","added_by":"auto","created_at":"2021-12-28 00:37:55","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":297639,"visible":true,"origin":"","legend":"\u003cp\u003e(a) EDX spectrum of CuCo\u003csub\u003e2\u003c/sub\u003eS\u003csub\u003e4\u003c/sub\u003e-poly-CD, X-ray photoelectron spectrum of elements present in CuCo\u003csub\u003e2\u003c/sub\u003eS\u003csub\u003e4\u003c/sub\u003e-poly-CD NPs (Cu, Co, S, C, O shown from B to F respectively).\u003c/p\u003e","description":"","filename":"Figure2.png","url":"https://assets-eu.researchsquare.com/files/rs-1190820/v1/5fe58090ea998df586569270.png"},{"id":16781932,"identity":"6a9aa935-13b3-4f04-acfd-a8a836963f2b","added_by":"auto","created_at":"2021-12-28 00:37:55","extension":"png","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":33350,"visible":true,"origin":"","legend":"\u003cp\u003eTGA profile of CuCo\u003csub\u003e2\u003c/sub\u003eS\u003csub\u003e4\u003c/sub\u003e-poly-CD NPs.\u003c/p\u003e","description":"","filename":"Figure3.png","url":"https://assets-eu.researchsquare.com/files/rs-1190820/v1/431786bfaeac4e326166e0f5.png"},{"id":16782015,"identity":"03c586ba-ce67-4784-a433-8fb1dd08c3c5","added_by":"auto","created_at":"2021-12-28 00:40:55","extension":"png","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":94630,"visible":true,"origin":"","legend":"\u003cp\u003eA. Absorbance graph of CuCo\u003csub\u003e2\u003c/sub\u003eS\u003csub\u003e4\u003c/sub\u003e-poly-CD NPs, B shows the band gap energy of CuCo\u003csub\u003e2\u003c/sub\u003eS\u003csub\u003e4\u003c/sub\u003e-poly-CD NPs calculated using Tauc’s plot, C and D show the magnetization profile and photoluminescence spectra of CuCo\u003csub\u003e2\u003c/sub\u003eS\u003csub\u003e4\u003c/sub\u003e-poly-CD NPs respectively.\u003c/p\u003e","description":"","filename":"Figure4.png","url":"https://assets-eu.researchsquare.com/files/rs-1190820/v1/0fc8f1cc13194e622115d8c3.png"},{"id":16781926,"identity":"3925a27b-4d25-42b3-9c96-817d09d2fe18","added_by":"auto","created_at":"2021-12-28 00:37:55","extension":"png","order_by":5,"title":"Figure 5","display":"","copyAsset":false,"role":"figure","size":71562,"visible":true,"origin":"","legend":"\u003cp\u003eDrug release profile of CuCo\u003csub\u003e2\u003c/sub\u003eS\u003csub\u003e4\u003c/sub\u003e-poly-CD NPs at different pH’s.\u003c/p\u003e","description":"","filename":"Figure5.png","url":"https://assets-eu.researchsquare.com/files/rs-1190820/v1/40771e85d10ab65d642152df.png"},{"id":16781931,"identity":"ad2c2e86-c239-49db-9777-2cedb96bc98f","added_by":"auto","created_at":"2021-12-28 00:37:55","extension":"png","order_by":6,"title":"Figure 6","display":"","copyAsset":false,"role":"figure","size":168830,"visible":true,"origin":"","legend":"\u003cp\u003eA and B show the in-vitro cytotoxicity assay on MDA-MB-231 cell lines of CuCo\u003csub\u003e2\u003c/sub\u003eS\u003csub\u003e4\u003c/sub\u003e and CPT loaded CuCo\u003csub\u003e2\u003c/sub\u003eS\u003csub\u003e4\u003c/sub\u003e-poly-CD respectively.\u003c/p\u003e","description":"","filename":"Figure6.png","url":"https://assets-eu.researchsquare.com/files/rs-1190820/v1/963e01ba2af40b1098c103ec.png"},{"id":17178372,"identity":"d304da12-9a3a-4aa8-a3a5-085ffd59a19d","added_by":"auto","created_at":"2022-01-10 19:57:35","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":1244505,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-1190820/v1/46c67d8d-6d43-4fbe-ac08-1c874b9886b7.pdf"},{"id":16782016,"identity":"ef1909a7-6c86-4e28-8f8f-30860d52a230","added_by":"auto","created_at":"2021-12-28 00:40:55","extension":"jpeg","order_by":1,"title":"","display":"","copyAsset":false,"role":"supplement","size":143548,"visible":true,"origin":"","legend":"","description":"","filename":"Graphicalabstract.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-1190820/v1/d5bca5f5cdf7460fbbb31b55.jpeg"}],"financialInterests":"","formattedTitle":"\u003cp\u003eCamptothecin-Carrying Cuco\u003csub\u003e2\u003c/sub\u003es\u003csub\u003e4\u003c/sub\u003e Nanoparticles: Sustained Drug Release And Anticancer Activity\u003c/p\u003e","fulltext":[{"header":"1 Introduction","content":"\u003cp\u003eNanoparticles (NPs) have been investigated and utilized, with an increasing interest, as targeted drug delivery systems and for other biomedical applications.\u003csup\u003e\u003cspan additionalcitationids=\"CR2\" citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e\u003c/sup\u003e The nanoparticles employed as drug carriers are designed as possessing properties that render them respond to stimuli like magnetic field, light, and pH.\u003csup\u003e\u003cspan additionalcitationids=\"CR5\" citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e\u003c/sup\u003e These properties enable them to find application in diagnosis and treatment of chronic diseases like cancer. For instance, magnetic resonance imaging (MRI), and magnetic hyperthermia, and magnetic field-assisted anticancer drug-delivery are possible when magnetic NPs are used.\u003csup\u003e\u003cspan additionalcitationids=\"CR8\" citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e\u003c/sup\u003e Some engineered NPs absorb near-infrared (NIR) radiation and heat up, thus acting as photothermal heating agents which allows the selective ablation of cancer cells.\u003csup\u003e\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e\u003c/sup\u003e Moreover, magnetic-luminescent NPs also have been developed with a focus on multiple-modal imaging.\u003csup\u003e\u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e\u003c/sup\u003e In a nutshell, delicate design and synthesis of NPs with tailor-made characteristics have occupied a place of paramount importance in the field of targeted drug delivery.\u003c/p\u003e \u003cp\u003eBesides the elemental composition of inorganic NPs used for biomedical applications, their size, shape, and surface modification have to be customized to make them effective therapeutic carriers.\u003csup\u003e\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e,\u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e\u003c/sup\u003e The size of the NPs are desired to be in the range of 50-150 nm to make them reside longer in the circulatory system.\u003csup\u003e\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e\u003c/sup\u003e In addition, NPs with large aspect ratios undergo longer plasma residence.\u003csup\u003e\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e\u003c/sup\u003e Another essential requirement is to modify the surface of the NPs to render them biocompatible, dispersible in aqueous medium, and to enable the loading of anticancer drugs.\u003csup\u003e\u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e,\u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e\u003c/sup\u003e Biocompatible polymers serve the above-mentioned purposes. The loading of anticancer drug molecules in the polymer sheath that covers the inorganic NPs occurs mostly through non-covalent interactions.\u003csup\u003e\u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e18\u003c/span\u003e\u003c/sup\u003e For instance, the drug molecule gets inserted between the coiled polymeric strands. Nevertheless, an intriguing method of loading the drug is through the formation of supramolecular host: guest complexes on the surfaces of NPs.\u003csup\u003e\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e\u003c/sup\u003e It enables the loading of a larger am\u0026rsquo;ount of drugs. The inorganic NP-biocompatible polymer combination acts as a suitable choice of nanocarrier with precision-added properties.\u003c/p\u003e \u003cp\u003eCopper sulfide NPs have emerged as new generation materials for photothermal therapy (PTT) as they display a large NIR absorption.\u003csup\u003e\u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e20\u003c/span\u003e\u003c/sup\u003e Attempts have been made to dope elements into CuS NPs to further fine-tune their properties.\u003csup\u003e\u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e,\u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e22\u003c/span\u003e\u003c/sup\u003e We hypothesize that the substitution of a magnetic element like cobalt can make them magnetic while remaining NIR- absorbing materials. In addition, a nanocomposite (NC) of Co-doped copper sulfide with a carbohydrate polymer possessing cavity-containing molecular component can aid achieving an enhanced drug-loading and biocompatibility. For this purpose, we chose a β-cyclodextrin-containing polysaccharide as the coating polymer (poly-CD). Herein, we report the synthesis and characterization of Co-doped copper sulphide, preparation of its NC with the poly-CD, of the anticancer drug, camptothecin (CPT), and the anticancer potential of the drug loaded NC on breast cancer cell lines.\u003c/p\u003e"},{"header":"2 Materials And Methods","content":"\u003cp\u003e\u003cstrong\u003e2.1 Materials\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe material needed for this work, namely copper chloride and cobalt nitrate, was obtained from Fischer Scientific. Thiourea was procured from Sigma Aldrich and ethyl acetate from NICE chemicals. Triethylamine and Cetyltrimethylammoniumbromide (CTAB) was obtained from Scientific chemical distributors. Analytical grade chemicals with purity above 98% was used.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e2.2 Methods\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e2.2.1 Synthesis of CuCo\u003csub\u003e2\u003c/sub\u003eS\u003csub\u003e4\u003c/sub\u003e nanoparticles\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe \u0026beta;-CD polymer (poly-CD) was synthesized via a reported procedure.\u003csup\u003e23\u003c/sup\u003e A hydrothermal synthesis method was employed to synthesize poly-CD-coated CuCo\u003csub\u003e2\u003c/sub\u003eS\u003csub\u003e4\u003c/sub\u003e NPs. Copper chloride and thiourea was mixed in the ratio 1:3 respectively in 20 ml deionized water and subjected to stirring for 15 minutes. To the above solution, 0.02 M cobalt nitrate dissolved in 10 ml of water was added followed by addition of 0.2 g of CTAB and 0.2 g poly-CD. The mixture was then added to 6 ml triethylamine dissolved in 10 ml of ethyl acetate. It was sonicated for 2 hours at room temperature and then transferred to a Teflon stainless steel autoclave. The temperature was maintained at 180 \u0026deg;C for 18 hours. The obtained poly-CD-coated-CuCo\u003csub\u003e2\u003c/sub\u003eS\u003csub\u003e4\u003c/sub\u003e nanoparticles were alternatively washed with distilled water and ethanol to remove unreacted chemicals. The NPs were dried at room temperature.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e2.2.2 Instrumentation\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe synthesized NPs were characterised for their crystalline phase using X-ray powder diffraction (Bruker D8-Advance) using Cu k\u0026alpha; (\u0026lambda; = 0.154 \u0026Aring;) radiation in the range of 10\u0026ndash;80\u0026deg;. High-resolution transmission electron microscope (HR TEM\u0026ndash;Jeol/JEM 2100) was used to determine the topography, morphology and size of the nanoparticles. The composition of the NPs was determined using Jeol/JEM 2100 Energy dispersive X-ray (EDX) instrument. ICP-MS-Thermofisher CAP RQ ICP-MS device was used to measure elemental composition. The amount and the rate of weight changes of the nanoparticles with respect to the temperature was determined using Thermogravimetric Analysis (TGA), TA instruments, USA Model SDT Q600. The elemental composition was measured using a Thermofisher Scientific Model Nexsa bas X-ray photoelectron spectroscopy (XPS). M-H curves to determine the magnetic property were obtained using a Lake-shore 7410 Vibrating Sample Magnetometer (VSM). \u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e2.2.3 In vitro drug release\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eIn vitro drug release was studied for NPs loaded with a chemotherapeutic drug camptothecin (CPT) and packed in a dialysis bag of MWCO 12-14 kDa. The bag was suspended in PBS solutions of pH 6.5 and 7.4. The bags were continuously stirred at room temperature. Samples were collected at regular time intervals. After sample collection, fresh PBS solution was added to maintain the pH throughout the release study. UV-vis absorption at 430 nm was recorded for each sample at each time point. The concentration of the released drug was determined by the standard calibration curve equation. The mean values for three independent time estimates were calculated. The percentage of drug loading and release was calculated using following equations:\u003c/p\u003e\n\u003cp\u003e\u003cimg 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\"\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e2.2.4 Cell line culture and condition\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eTriple negative breast cancer cell line MDA-MB-231, was obtained from NCCS, Pune and cultured in liquid medium i.e., Dulbecco\u0026rsquo;s Modified Eagle High Glucose (DMEM-HG) that was supplemented with 10% Fetal Bovine Serum Albumin (FBS), 1% sodium bicarbonate and 1% sodium pyruvate at 37 \u0026deg;C in a 5% CO\u003csub\u003e2\u003c/sub\u003e incubator. The cells were trypsinized using trypsin EDTA, and seeded in a 96 well plate for MTT and 6 well plate for flow cytometry analysis.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e2.2.5 In-vitro MTT assay\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eMDA-MB-231 cells were trypsinized and grown in a 96-well plate at a density of 1 \u0026times; 10\u003csup\u003e6\u003c/sup\u003e cells/ml and incubated for 24 h. The cells were treated with free nanocarrier (poly-CD-coated NPs) and drug (CPT)-loaded nanocarriers. The concentrations of treatments were in the range of 0.25 \u0026mu;g/mL to 15 \u0026mu;g/mL. The sample was replicated three times It was then incubated for 24 h at 37\u0026deg;C. After incubation, the culture medium was removed from the treated plate, and 100 \u0026mu;L of MTT (0.5 mg/mL) was added to each well and incubated at 37 \u0026deg;C for 4 h. The resulting formazan crystals were dissolved in 100 \u0026mu;L of DMSO. The absorbance intensity was measured using a microplate reader (RT-2100 C) at 570 nm. The percentage o viable cells was plotted and the IC\u003csub\u003e50\u003c/sub\u003e value was calculated using GraphPad Prism 8.0 software.\u003c/p\u003e"},{"header":"3 Results And Discussion","content":"\u003cdiv id=\"Sec11\" class=\"Section2\"\u003e \u003ch2\u003e3.1 Characterization of the NPs\u003c/h2\u003e \u003cp\u003eThe Cu and Co-containing sulfide NPs were synthesized using a hydrothermal method. The synthesized NPs were coated with poly-CD. The polymer-coated NPs display an X-ray diffraction pattern as in Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eA. As the poly-CD possesses a crystalline nature, broad peaks are not obvious in the XRD. The sharp peaks correspond to the sulfide NPs. The prominent peaks are observed at 2θ 27.12\u0026deg; (2 2 0), 31.3\u0026deg; (3 1 1), 32.4\u0026deg; (2 2 2), 47.3\u0026deg; (4 2 2), 52.3\u0026deg; (3 3 3), 58.8\u0026deg; (5 3 1), and 68.9\u0026deg; (4 4 4). These reflections correspond to the CuCo\u003csub\u003e2\u003c/sub\u003eS\u003csub\u003e4\u003c/sub\u003e crystals of the face-centered cubic system. It is confirmed by their congruence with the JCPDS card number: 75-1570. The average grain size of the CuCo\u003csub\u003e2\u003c/sub\u003eS\u003csub\u003e4\u003c/sub\u003e crystals was determined employing the Debye-Scherrer expression:\u003c/p\u003e \u003cp\u003e \u003cdiv id=\"Equ3\" class=\"Equation\"\u003e \u003cdiv format=\"TEX\" class=\"mathdisplay\" id=\"FileID_Equ3\" name=\"EquationSource\"\u003e\n$$\\text{D} =\\frac{0.94{\\lambda }}{{\\beta } \\text{c}\\text{o}\\text{s}{\\theta }}$$\u003c/div\u003e \u003cdiv class=\"EquationNumber\"\u003e3\u003c/div\u003e\u003c/div\u003e \u003c/p\u003e \u003cp\u003ewhere \u003cspan class=\"InlineEquation\"\u003e\u003cspan class=\"mathinline\"\u003e\\({\\lambda }, {\\beta }, \\text{a}\\text{n}\\text{d} {\\theta }\\)\u003c/span\u003e\u003c/span\u003e represent the wavelength of Cu (Kα), full-width-half-maximum in radians, and the angle of the most intense peak respectively. The crystalline size (D) is determined as 13.36 nm. Further, the dislocation density of the crystals was determined employing the following relation:\u003cdiv id=\"Equ4\" class=\"Equation\"\u003e\u003cdiv format=\"TEX\" class=\"mathdisplay\" id=\"FileID_Equ4\" name=\"EquationSource\"\u003e\n$$\\text{D}\\text{i}\\text{s}\\text{l}\\text{o}\\text{c}\\text{a}\\text{t}\\text{i}\\text{o}\\text{n} \\text{d}\\text{e}\\text{n}\\text{s}\\text{i}\\text{t}\\text{y}, \\delta =\\frac{1}{{D}^{2}}$$\u003c/div\u003e\u003cdiv class=\"EquationNumber\"\u003e4\u003c/div\u003e\u003c/div\u003e\u003c/p\u003e \u003cp\u003ewhere D represents the average crystallite size. The \u003cspan class=\"InlineEquation\"\u003e\u003cspan class=\"mathinline\"\u003e\\(\\delta\\)\u003c/span\u003e\u003c/span\u003e value is 6.5 x 10\u003csup\u003e-3\u003c/sup\u003e nm\u003csup\u003e-\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e\u003c/sup\u003e. As the polymer contributes an extent of non-crystallinity to the sample of NPs, crystallinity index should be calculated. In a method of calculating the crystallinity, we utilized the following relation:\u003cdiv id=\"Equ5\" class=\"Equation\"\u003e\u003cdiv format=\"TEX\" class=\"mathdisplay\" id=\"FileID_Equ5\" name=\"EquationSource\"\u003e\n$$\\text{C}\\text{r}\\text{y}\\text{s}\\text{t}\\text{a}\\text{l}\\text{l}\\text{i}\\text{n}\\text{i}\\text{t}\\text{y} \\text{I}\\text{n}\\text{d}\\text{e}\\text{x}, CI=\\frac{Area of all the crystalline peaks}{Area of all the crystalline and amorphous peaks}$$\u003c/div\u003e\u003cdiv class=\"EquationNumber\"\u003e5\u003c/div\u003e\u003c/div\u003e\u003c/p\u003e \u003cp\u003eThe crystallinity thus determined is 46%.\u003c/p\u003e \u003cp\u003eFigure\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eB shows the TEM images of the poly-CD-coated CuCo\u003csub\u003e2\u003c/sub\u003eS\u003csub\u003e4\u003c/sub\u003e NPs. Nucleated nano-structures are observed in hexagonal shapes. Some of them appear sideways in orientation revealing that the formed NPs are hexagonal plate-shaped NPs. These nanostructures are consistent with the observation in crystal data revealed by the XRD pattern. The mean length and width of the NPs are 120.71 and 42.79 nm respectively. The poly-CD has coated the NPs as a thin layer. In addition, such polymers are transparent in TEM and do not appear vivid on the dark contrast-sulfide NPs.\u003csup\u003e\u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e24\u003c/span\u003e\u003c/sup\u003e The size of the polymer-coated NPs was further confirmed by carrying out a particle-size analysis by dynamic light scattering experiment. The DLS shows a histogram with the mean size of the NPs as 122 nm. (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eC). The size range matches with that observed in the TEM images.\u003c/p\u003e \u003cp\u003eThe elemental composition of the as-synthesized CuCo\u003csub\u003e2\u003c/sub\u003eS\u003csub\u003e4\u003c/sub\u003e NPs was determined qualitatively, employing EDX spectroscopy. The EDX of the NPs is displayed in Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eA. The peaks in the figure correspond to the elements viz., Cu, Co, and S. additional peaks pertaining to any other impurity is not observed. Thus, the EDX reveals the formation of pure CuCo\u003csub\u003e2\u003c/sub\u003eS\u003csub\u003e4\u003c/sub\u003e NPs. To add further evidence for such composition of the NPs and to determine the presence of various elements in specific electronic states, X-ray photoelectron spectra were recorded. For XPS measurements, the poly-CD-coated NPs were used. Figs.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eB\u0026ndash;F depict the Cu, Co, S, C, and O peaks of the XPS pattern. The 2p\u003csub\u003e3/2\u003c/sub\u003e and 2p\u003csub\u003e1/2\u003c/sub\u003e states of Cu appear at 932.25 and 952.20 eV respectively. (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eB). the deconvoluted XPS of Co shows peaks at 779.64 and 796.33 eV due to the 2p\u003csub\u003e3/2\u003c/sub\u003e and 2p\u003csub\u003e1/2\u003c/sub\u003e electronic states. (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eC). Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eD depicts the 2p\u003csub\u003e3/2\u003c/sub\u003e and 2p\u003csub\u003e1/2\u003c/sub\u003e peaks at 161.68 and 162.81 eV respectively. These spectra arise from the cobalt-substituted copper sulfide. The spectra C and O are contributed by the polymer that resides on the surface of the CuCo\u003csub\u003e2\u003c/sub\u003eS\u003csub\u003e4\u003c/sub\u003e NPs. The carbons of the C-O-C, C-C, and O-C=O functional groups display peaks at 284.04, 284.85, and 285.92 eV respectively, arising from the C 1s state. (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eE). the O 1s peaks of the C-O and C=O groups show peaks at the binding energy values viz., 532.38 and 531.63 eV. The XPS peaks and their positions revel the composition of the poly-CD-coated CuCo\u003csub\u003e2\u003c/sub\u003eS\u003csub\u003e4\u003c/sub\u003e NPs. Additional peaks due to any impurity is not observed in the XPS data.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eIt is apparent from the results discussed above that the synthesized material carries CuCo\u003csub\u003e2\u003c/sub\u003eS\u003csub\u003e4\u003c/sub\u003e NPs with the poly-CD immobilized on their surface. To further evaluate the amount of poly-CD on the surface of the NPs, thermogravimetric profiles of the coated CuCo\u003csub\u003e2\u003c/sub\u003eS\u003csub\u003e4\u003c/sub\u003e NPs were measured. Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003e shows an initial weight loss of 4.42% and then plummeting down of the weight up to 12.46%. these decay profiles imply that the polymeric structure on the surface changes conformation and then starts melting. The melting is sharp at about 440\u0026deg;C. above this temperature, there is complete degradation of the coated polymer and it extends up to above 750\u0026deg;C. the residue remains with a weight percentage of 79.86 with an implication that the coated polymer offers about 20% of weight to the composite nanostructure.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec12\" class=\"Section2\"\u003e \u003ch2\u003e3.2 Optical and magnetic characteristics of the NPs\u003c/h2\u003e \u003cp\u003eThe UV-Vis absorption spectrum of the poly-CD-coated CuCo\u003csub\u003e2\u003c/sub\u003eS\u003csub\u003e4\u003c/sub\u003e NPs is shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003eA. the band at the visible spectral region is shorter (lower absorbance) than the NIR band, which is vividly due to the 3d electrons of the transition metals in the NPs.\u003csup\u003e\u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e25\u003c/span\u003e\u003c/sup\u003e The stronger absorption is further contributed by the Co incorporated into the crystals, which makes the normally observed CuS absorption spectrum enhance in absorbance at the NIR wavelength.\u003csup\u003e\u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e26\u003c/span\u003e\u003c/sup\u003e The aqueous dispersion of the poly-CD coated NPs, displaying such an NIR absorption, presents them suitable for photothermal heating. Further, the absorption wavelength was determined by an extrapolation of the line to the linear portion of the absorption spectrum. The optical band gap was determined employing the relation:\u003c/p\u003e \u003cp\u003e \u003cdiv id=\"Equ6\" class=\"Equation\"\u003e \u003cdiv format=\"TEX\" class=\"mathdisplay\" id=\"FileID_Equ6\" name=\"EquationSource\"\u003e\n$$Eg=\\frac{hc}{\\lambda }$$\u003c/div\u003e \u003cdiv class=\"EquationNumber\"\u003e6\u003c/div\u003e\u003c/div\u003e \u003c/p\u003e \u003cp\u003ewhere h represents the Planck\u0026rsquo;s constant, c, the velocity of light, and λ is the wavelength derived from the tail of absorption. In the present work, the energy band gap calculated is Eg = 2.25 eV (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003eB). this value is slightly different from the reported secondary band gap of 2.12 eV for a smaller size CuCo\u003csub\u003e2\u003c/sub\u003eS\u003csub\u003e4\u003c/sub\u003e NPs.\u003csup\u003e\u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e27\u003c/span\u003e\u003c/sup\u003e Furthermore, it is larger than the band gap reported for CuS NPs (1.80 eV).\u003csup\u003e\u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e28\u003c/span\u003e\u003c/sup\u003e The photoluminescence spectrum of the poly-CD-coated CuCo\u003csub\u003e2\u003c/sub\u003eS\u003csub\u003e4\u003c/sub\u003e NPs is shown in Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003eC. The luminescence spectral band is broad and intense, showing wavelength of emission at 510 nm. This property presents the NPs to be followed employing luminescence imaging.\u003c/p\u003e \u003cp\u003eThe magnetic properties of the poly-CD-coated NPs were investigated by employing VSM. The room-temperature magnetization property, with n applied sweeping magnetic field, of the NPs is displayed in Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003eD. the magnetization curve shows a small coercivity value of 78.09 Oe which is close to superparamagnetic limit.\u003csup\u003e\u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e29\u003c/span\u003e\u003c/sup\u003e The saturation magnetization (Ms) value is 0.024 emu g\u003csup\u003e-\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e\u003c/sup\u003e. Such magnetic NPs attain saturation quickly at the application of an external magnetic field. In addition, they relax quickly due to the absence of broad hysteresis loops. Therefore, the synthesized NPs are suitable for application in bio-imaging using MRI.\u003csup\u003e\u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e30\u003c/span\u003e\u003c/sup\u003e\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec13\" class=\"Section2\"\u003e \u003ch2\u003e3.3 Drug-loading and release\u003c/h2\u003e \u003cp\u003eThe polymer-coating on the NPs poses the advantages viz., (i) it makes the NPs dispersible in aqueous medium, (ii) renders them biocompatible, and (iii) enables the loading of organic therapeutic molecules. A drug carrier should accommodate an appreciable amount of the therapeutics. In addition, the release of the drug is preferred to be sustained and slow.\u003csup\u003e\u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e31\u003c/span\u003e\u003c/sup\u003e Therefore, the drug loading and the drug-release kinetics were investigated in vitro. The experimental procedures have been elaborated in the \u003cspan refid=\"Sec4\" class=\"InternalRef\"\u003emethods\u003c/span\u003e section. The percentage of the model drug CPT, loaded in the poly-CD-coated CuCo\u003csub\u003e2\u003c/sub\u003eS\u003csub\u003e4\u003c/sub\u003e NPs (nanocarrier) is estimated employing the equation (\u003cspan refid=\"Equ1\" class=\"InternalRef\"\u003e1\u003c/span\u003e). The percentage of adsorbed CPT on the nanocarrier is 89.04%. It implies a large amount of drug is loaded on the nanocarrier. CPT forms an inclusion complex with the β-CD molecule.\u003csup\u003e\u003cspan citationid=\"CR32\" class=\"CitationRef\"\u003e32\u003c/span\u003e\u003c/sup\u003e Therefore, the poly-CD is a suitable choice to be employed as the polymeric host structure that is immobilized on the surface of the NPs. The CPT release profile from the nanocarrier (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003e) reveals that it is disseminated in a sustained fashion, well over a period of 360 hours at the physiological pH of 7.4. The cumulative release of CPT at 24 hours is 25%. In addition, the release rate is accelerated when the pH is made slightly acidic (6.5). In this lower pH, release is enhanced to 40% at 24 hours. Therefore, it is obviously seen that the drug release is pH dependent and it is tunable with the application of a stimulus. It is worth mentioning that the tumor microenvironment is acidic in nature.\u003csup\u003e\u003cspan citationid=\"CR33\" class=\"CitationRef\"\u003e33\u003c/span\u003e\u003c/sup\u003e Therefore, the designed nanocarrier is suitable for selective release of CPT at the tumor site.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec14\" class=\"Section2\"\u003e \u003ch2\u003e3.4 In vitro anticancer activity\u003c/h2\u003e \u003cp\u003eThe in vitro anticancer activities of the free- and the nanocarrier were carried out on human breast cancer cell lines (MDA-MB-231). After an incubation period of 24 hours as described in the experimental section, the cytotoxicity was analyzed employing an MTT assay. Figs.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003eA and B represent the percentage cell viability as a function of concentration of the free- and CPT-loaded nanocarriers respectively. The cytotoxicity is observed to be dose-dependent in both the cases. Nevertheless, it is more in the case of CPT-loaded nanocarrier than the free-nanocarrier. The half-maximal inhibitory concentration (IC\u003csub\u003e50\u003c/sub\u003e) of the free nanocarrier is 161.5 \u0026micro;g mL\u003csup\u003e-\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e\u003c/sup\u003e, indicating that the cells are considerably viable at the addition of the nanocarrier alone. Contrary to that, the loading of CPT in the nanocarrier leads to an enhancement of the anticancer activity. The IC\u003csub\u003e50\u003c/sub\u003e value of the CPT-loaded nanocarrier is 68.93 \u0026micro;g mL\u003csup\u003e-\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e\u003c/sup\u003e. Therefore, it can be concluded that the loaded drug gets slowly released from the nanocarrier and destroys the cancer cells. Further, all of the amount of loaded CPT is not released at the time period of 24 hours. The cumulative release as discussed in the previous section suggests a 40% of CPT disseminated over a period of 24 hours. Therefore, it follows that the anticancer activity occurs due to the portion of the CPT that is released from the nanocarrier. The reported IC\u003csub\u003e50\u003c/sub\u003e of the free-CPT is indeed more than the CPT-loaded nanocarrier reported herein. Nevertheless, it should be noted that such direct application of CPT to the culture of cells renders the former rapidly and unrestrictedly diffusing into mammalian cells and hence make them more cytotoxic.\u003csup\u003e\u003cspan citationid=\"CR34\" class=\"CitationRef\"\u003e34\u003c/span\u003e\u003c/sup\u003e This is contrary to the slow internalization of the drug-loaded nanocarrier through the endocytosis pathway. Therefore, the poly-CD-coated CuCo\u003csub\u003e2\u003c/sub\u003eS\u003csub\u003e4\u003c/sub\u003e nanocarrier is a promising nanocarrier of CPT, that possesses additional features of magnetic, NIR absorbing, and photoluminescence characteristics.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003c/div\u003e"},{"header":"4 Conclusions","content":"\u003cp\u003eWe synthesized CuCo\u003csub\u003e2\u003c/sub\u003eS\u003csub\u003e4\u003c/sub\u003e nanoparticles and coated them with poly-CD. The nanoparticles show a hexagonal crystal structure with a mean size of 122 nm which is a size suitable for employing as a nanocarrier. The NIR-II absorption of the NPs is broad and more intense than the visible spectral band. The optical band gap is 2.25 eV. In addition, the nanocomposite reveals a superparamagnetic behavior due to the presence of incorporated cobalt. It makes them suitable for control by external magnetic field. The poly-CD coating renders the material biocompatible and enables the loading of CPT by means of a non-covalent host : guest association. The drug-release from the nanocarrier takes place over a period of about 360 hours. Further, the drug-release is pH-dependent and it is accelerated at pH 6.0 compared to the normal physiological pH (7.4). the drug-loaded nanocarrier reveals an anticancer efficacy (IC\u003csub\u003e50\u003c/sub\u003e = 68.93 \u0026micro;g mL\u003csup\u003e-\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e\u003c/sup\u003e) that presents it as a suitable choice as a CPT-transporter. The cobalt-addition and the poly-CD coating thus add precision to the promising sulfide NPs.\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eAcknowledgement\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe research work was carried out without any funding by external agencies. The authors thank the management of KITS for providing necessary facilities.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eCompliance with ethical standards\u003c/strong\u003e\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConflict of interest\u0026nbsp;\u003c/strong\u003eNone declared.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\n \u003cli\u003eM. J. Mitchell, M. M. Billingsley, R. M. Haley, M. E. Wechsler, N. A. Peppas, R. Langer, Engineering precision nanoparticles for drug delivery. Nat. Rev. Drug Discov. \u003cstrong\u003e20\u003c/strong\u003e, 101-124 (2021). https://doi.org/10.1038/s41573-020-0090-8.\u003c/li\u003e\n \u003cli\u003eK. Kaliyamoorthi, A. S. Pillai, A. Alexander, S. Ramasamy, A. Arivarasu, I. V. M. V. Enoch, Designed poly (ethylene glycol) conjugate-erbium-doped magnetic nanoparticle hybrid carrier: enhanced activity of anticancer drug. J. Mater. 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Zhai, The endocytosis and intracellular fate of nanomedicine: implication for rational design. Asian J. Pharm. Sci. \u003cstrong\u003e8\u003c/strong\u003e, 1-10 (2013). https://doi.org/10.1016/j.ajps.2013.07.001.\u003c/li\u003e\n\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":true,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"Sulfide nanoparticles, superparamagnetism, NIR absorption, nanocarrier, anticancer activity.","lastPublishedDoi":"10.21203/rs.3.rs-1190820/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-1190820/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eNanocarriers of anticancer drugs are delicately designed with precision addition at every attempt. In this paper, we report CuCo\u003csub\u003e2\u003c/sub\u003eS\u003csub\u003e4\u003c/sub\u003e nanoparticles that show light-absorption in the NIR-II wavelength range and possess magnetic characteristics. The synthesized nanoparticles are characterized employing XRD, SEM, DLS, TGA, and XPS methods. The nanoparticles form a composite with biocompatible polymeric β-cyclodextrin. The nanoparticles possess a band gap energy of 2.25 eV. The magnetic property arises due to the cobalt-incorporation in the nanoparticles. The anticancer drug, camptothecin, is loaded in the nanocarrier with an 89% adsorption efficiency. The in vitro release of the drug occurs in a sustained fashion. Further, the in vitro anticancer potential of the nanocarrier is examined on breast cancer (MDA=MB-231) cell lines and the activities of the free- and the drug-loaded nanocarrier are compared. The cobalt-containing copper sulfide nanoparticle-poly-β-cyclodextrin composite works as a promising nanocarrier of camptothecin.\u003c/p\u003e","manuscriptTitle":"Camptothecin-Carrying Cuco2s4 Nanoparticles: Sustained Drug Release And Anticancer Activity","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2021-12-28 00:37:53","doi":"10.21203/rs.3.rs-1190820/v1","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"218eb700-47f3-4a17-9c85-ee5899ac5b85","owner":[],"postedDate":"December 28th, 2021","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[{"id":9344900,"name":"Materials Chemistry"},{"id":9344901,"name":"Polymer Science"}],"tags":[],"updatedAt":"2022-01-10T19:57:30+00:00","versionOfRecord":[],"versionCreatedAt":"2021-12-28 00:37:53","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-1190820","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-1190820","identity":"rs-1190820","version":["v1"]},"buildId":"WrCJVZZCHTDjtuVLN7oU0","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}