Nanoformulations for co-delivery of multiple therapeutics and combinational therapies are expected to provide improved pharmacokinetics, reduced drug toxicities, and avert drug resistance. Methotrexate (MTX) and curcumin (CUR) were chosen as model drugs for this study primarily due to their anticancer activities in different phases of the cell cycle.
The nanoprecipitation technique was used to prepare CUR-, MTX-, and CUR/MTX-co-loaded poly(d,l-lactide-co-glycolide) (PLGA) nanoparticles.
Encapsulation and drug loading efficiencies were 87.9% and 8.0% for CUR-loaded PLGA nanoparticles and 81.4% and 13.5% for MTX-loaded PLGA nanoparticles, respectively. In addition, drug co-encapsulation and co-loading efficiencies were 77.2% and 6.5% for CUR and 86.1% and 15.5% for MTX, respectively. The results of cell cycle analysis indicated that CUR- and MTX-loaded PLGA nanoparticles additively complemented each other in the S and sub-G1 phases. The obtained combinational index (CI) also exhibited that CUR- and MTX-loaded PLGA nanoparticles produce additive cytotoxicity at Fa = 0.5 on U87MG glioma cells. In addition, necrosis dominates cell death in nanomediated monotherapy whereas apoptosis dominates co-delivery treatment. The highest amount of LDH released was related to co-delivery as well.
Nano-assisted combinational therapy and co-delivery can be used to obtain efficacies of drugs at lower concentrations, thus potentially increasing the therapeutic window of the drugs.
This is a preview of subscription content, log in to check access.
Buy single article
Instant access to the full article PDF.
Tax calculation will be finalised during checkout.
Subscribe to journal
Immediate online access to all issues from 2019. Subscription will auto renew annually.
Tax calculation will be finalised during checkout.
Cohen MH, Johnson JR, Pazdur R. Food and Drug Administration drug approval summary: temozolomide plus radiation therapy for the treatment of newly diagnosed glioblastoma multiforme. Clin Cancer Res. 2005;11(19):6767–71.
Cohen MH, Shen YL, Keegan P, Pazdur R. FDA drug approval summary: bevacizumab (Avastin®) as treatment of recurrent glioblastoma multiforme. Oncologist. 2009;14(11):1131–8.
Hryniuk WM. The mechanism of action of methotrexate in cultured L5178Y leukemia cells. Cancer Res. 1975;35(4):1085–92.
Shaikh N, Sardar M, Raj R, Jariwala P. A rapidly fatal case of low-dose methotrexate toxicity. Case Rep Med. 2018;2018:1–4.
Bertino JR, Göker E, Gorlick R, Li WW, Banerjee D. Resistance mechanisms to methotrexate in tumors. Oncologist. 1996;1(4):223–6.
Hanif R, Qiao L, Shiff SJ, Rigas B. Curcumin, a natural plant phenolic food additive, inhibits cell proliferation and induces cell cycle changes in colon adenocarcinoma cell lines by a prostaglandin-independent pathway. J Lab Clin Med. 1997;130(6):576–84.
Simon A, Allais DP, Duroux JL, Basly JP, Durand-Fontanier S, Delage C. Inhibitory effect of curcuminoids on MCF-7 cell proliferation and structure–activity relationships. Cancer Lett. 1998;129(1):111–6.
Yang K-Y, Lin L-C, Tseng T-Y, Wang S-C, Tsai T-H. Oral bioavailability of curcumin in rat and the herbal analysis from Curcuma longa by LC–MS/MS. J Chromatogr B. 2007;853(1):183–9.
Pignatello R, Puleo A, Puglisi G, Vicari L, Messina A. Effect of liposomal delivery on in vitro antitumor activity of lipophilic conjugates of methotrexate with lipoamino acids. Drug Deliv. 2003;10(2):95–100.
Wosikowski K, Biedermann E, Rattel B, Breiter N, Jank P, Löser R, et al. In vitro and in vivo antitumor activity of methotrexate conjugated to human serum albumin in human cancer cells. Clin Cancer Res. 2003;9(5):1917–26.
Nogueira DR, Tavano L, Mitjans M, Pérez L, Infante MR, Vinardell MP. In vitro antitumor activity of methotrexate via pH-sensitive chitosan nanoparticles. Biomaterials. 2013;34(11):2758–72.
Zhao Y, Guo Y, Li R, Wang T, Han M, Zhu C, et al. Methotrexate nanoparticles prepared with codendrimer from polyamidoamine (PAMAM) and oligoethylene glycols (OEG) dendrons: antitumor efficacy in vitro and in vivo. Sci Rep. 2016;6:28983.
Naksuriya O, Okonogi S, Schiffelers RM, Hennink WE. Curcumin nanoformulations: a review of pharmaceutical properties and preclinical studies and clinical data related to cancer treatment. Biomaterials. 2014;35(10):3365–83.
Yallapu MM, Jaggi M, Chauhan SC. Curcumin nanoformulations: a future nanomedicine for cancer. Drug Discov Today. 2012;17(1):71–80.
Zanotto-Filho A, Coradini K, Braganhol E, Schröder R, De Oliveira CM, Simões-Pires A, et al. Curcumin-loaded lipid-core nanocapsules as a strategy to improve pharmacological efficacy of curcumin in glioma treatment. Eur J Pharma Biopharm. 2013;83(2):156–67.
Schiborr C, Kocher A, Behnam D, Jandasek J, Toelstede S, Frank J. The oral bioavailability of curcumin from micronized powder and liquid micelles is significantly increased in healthy humans and differs between sexes. Mol Nutr Food Res. 2014;58(3):516–27.
Gerards AH, Landewé RBM, Prins APA, Bruijn GAW, Laan R, Dijkmans BAC. Cyclosporin A monotherapy versus cyclosporin A and methotrexate combination therapy in patients with early rheumatoid arthritis: a double blind randomised placebo controlled trial. Ann Rheum. 2003;62(4):291–6.
Mulder JH, Smink T, Van Putten LM. 5-Fluorouracil and methotrexate combination chemotherapy: the effect of drug scheduling. Eur J Canc Clin Oncol. 1981;17(7):831–7.
Murad AM, Santiago FF, Petroianu A, Rocha PRS, Rodrigues MAG, Rausch M. Modified therapy with 5-fluorouracil, doxorubicin, and methotrexate in advanced gastric cancer. Cancer. 1993;72(1):37–41.
Hossain MM, Banik NL, Ray SK. Synergistic anti-cancer mechanisms of curcumin and paclitaxel for growth inhibition of human brain tumor stem cells and LN18 and U138MG cells. Neurochem Int. 2012;61(7):1102–13.
Zanotto-Filho A, Braganhol E, Klafke K, Figueiró F, Terra SR, Paludo FJ, et al. Autophagy inhibition improves the efficacy of curcumin/temozolomide combination therapy in glioblastomas. Cancer Lett. 2015;358(2):220–31.
Dilnawaz F, Sahoo SK. Enhanced accumulation of curcumin and temozolomide loaded magnetic nanoparticles executes profound cytotoxic effect in glioblastoma spheroid model. Eur J Pharm Biopharm. 2013;85(3):452–62.
Qi S-S, Sun J-H, Yu H-H, Yu S-Q. Co-delivery nanoparticles of anti-cancer drugs for improving chemotherapy efficacy. Drug Deliv. 2017;24(1):1909–26.
Banji D, Pinnapureddy J, Banji OJ, Saidulu A, Hayath MS. Synergistic activity of curcumin with methotrexate in ameliorating Freund’s complete adjuvant induced arthritis with reduced hepatotoxicity in experimental animals. Eur J Pharmacol. 2011;668(1–2):293–8.
Dhanasekaran S, Biswal BK, Sumantran VN, Verma RS. Augmented sensitivity to methotrexate by curcumin induced overexpression of folate receptor in KG-1 cells. Biochimie. 2013;95(8):1567–73.
Fang JH, Lai YH, Chiu TL, Chen YY, Hu SH, Chen SY. Magnetic core–shell nanocapsules with dual-targeting capabilities and co-delivery of multiple drugs to treat brain gliomas. Adv Healthc Mater. 2014;3(8):1250–60.
Madani F, Esnaashari SS, Mujokoro B, Dorkoosh F, Khosravani M, Adabi M. Investigation of effective parameters on size of paclitaxel loaded PLGA nanoparticles. Adv Pharm Bull. 2018;8(1):77–84.
Malekpour MR, Naghibzadeh M, Najafabadi MRH, Esnaashari SS, Adabi M. Effect of various parameters on encapsulation efficiency of mPEG-PLGA nanoparticles: artificial neural network. Biointerface Res Appl Chem. 2018;8(3):3267–72.
Esnaashari SS, Amani A. Optimization of noscapine-loaded mPEG-PLGA nanoparticles and release study: a response surface methodology approach. J Pharm Innov. 2018;13(3):237–46.
Tsai Y-M, Chien C-F, Lin L-C, Tsai T-H. Curcumin and its nano-formulation: the kinetics of tissue distribution and blood–brain barrier penetration. Int J Pharm. 2011;416(1):331–8.
Fessi H, Puisieux F, Devissaguet JP, Ammoury N, Benita S. Nanocapsule formation by interfacial polymer deposition following solvent displacement. Int J Pharm. 1989;55(1):R1–4.
Zhang F, Koh GY, Jeansonne DP, Hollingsworth J, Russo PS, Vicente G, et al. A novel solubility-enhanced curcumin formulation showing stability and maintenance of anticancer activity. J Pharm Sci. 2011;100(7):2778–89.
Chou TC, Martin N. CompuSyn for drug combinations and for general dose-effect analysis, software and user’s guide: a computer program for quantitation of synergism and antagonism in drug combinations, and the determination of IC50 and ED50 and LD50 values. Paramus: ComboSyn Inc.; 2005.
Blanco E, Shen H, Ferrari M. Principles of nanoparticle design for overcoming biological barriers to drug delivery. Nat Biotechnol. 2015;33(9):941–51.
Nicolete R, dos Santos DF, Faccioli LH. The uptake of PLGA micro or nanoparticles by macrophages provokes distinct in vitro inflammatory response. Int Immunopharmacol. 2011;11(10):1557–63.
Maeda H. Macromolecular therapeutics in cancer treatment: the EPR effect and beyond. J Control Release. 2012;164(2):138–44.
Jain A, Jain A, Garg NK, Tyagi RK, Singh B, Katare OP, et al. Surface engineered polymeric nanocarriers mediate the delivery of transferrin–methotrexate conjugates for an improved understanding of brain cancer. Acta Biomater. 2015;24:140–51.
Trapani A, Denora N, Iacobellis G, Sitterberg J, Bakowsky U, Kissel T. Methotrexate-loaded chitosan-and glycolchitosan-based nanoparticles: a promising strategy for the administration of the anticancer drug to brain tumors. AAPS PharmSciTech. 2011;12(4):1302–11.
Rescignano N, Tarpani L, Romani A, Bicchi I, Mattioli S, Emiliani C, et al. In-vitro degradation of PLGA nanoparticles in aqueous medium and in stem cell cultures by monitoring the cargo fluorescence spectrum. Polym Degrad Stabil. 2016;134:296–304.
Zolnik BS, Burgess DJ. Effect of acidic pH on PLGA microsphere degradation and release. J Control Release. 2007;122(3):338–44.
Qian Y, Ma J, Guo X, Sun J, Yu Y, Cao B, et al. Curcumin enhances the radiosensitivity of U87 cells by inducing DUSP-2 up-regulation. Cell Physiol Biochem. 2015;35(4):1381–93.
Palmer AC, Sorger PK. Combination cancer therapy can confer benefit via patient-to-patient variability without drug additivity or synergy. Cell. 2017;171(7):1678–91.
Su C-C, Wang M-J, Chiu T-L. The anti-cancer efficacy of curcumin scrutinized through core signaling pathways in glioblastoma. Int J Mol Med. 2010;26(2):217–24.
Liu E, Wu J, Cao W, Zhang J, Liu W, Jiang X, et al. Curcumin induces G2/M cell cycle arrest in a p53-dependent manner and upregulates ING4 expression in human glioma. J Neuro-Oncol. 2007;85(3):263–70.
Figueiró F, de Oliveira CP, Rockenbach L, Mendes FB, Bergamin LS, Jandrey EHF, et al. Pharmacological improvement and preclinical evaluation of methotrexate-loaded lipid-core nanocapsules in a glioblastoma model. J Biomed Nanotechnol. 2015;11(10):1808–18.
Harasym TO, Liboiron BD, Mayer LD. Drug ratio-dependent antagonism: a new category of multidrug resistance and strategies for its circumvention. Multi-Drug Resistance Cancer. Springer; 2010;291–323.
Mayer LD, Harasym TO, Tardi PG, Harasym NL, Shew CR, Johnstone SA, et al. Ratiometric dosing of anticancer drug combinations: controlling drug ratios after systemic administration regulates therapeutic activity in tumor-bearing mice. Mol Cancer Ther. 2006;5(7):1854–63.
This work was supported by the Tehran University of Medical Sciences (Grant No. 96-01-103-34141).
Conflict of Interest
The authors declare that they have no conflict of interest.
Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
About this article
Cite this article
Mujokoro, B., Madani, F., Esnaashari, S.S. et al. Combination and Co-delivery of Methotrexate and Curcumin: Preparation and In Vitro Cytotoxic Investigation on Glioma Cells. J Pharm Innov 15, 617–626 (2020). https://doi.org/10.1007/s12247-019-09406-3
- Combinational therapies