Synergism of Curcumin and Doxorubicin Proves an Effective Anticancer Therapeutics Against Breast Cancer: An in vitro Study

Abstract

Background: Curcumin is a polyphenol phyto-compound found in turmeric (Curcuma longa), which inhibits tumorigenesis by introducing apoptosis as well as by restricting cell survival and proliferation. This in vitro research article focuses on the pharmacodynamics interactions of Curcumin (Cur.) combined with the commercial drug Doxorubicin (Doxo.) to enhance the cytotoxicity of doxo. at lower doses against breast cancer cells MDA-MB-231 and MCF-7 with the chemo-protective effect against normal HEK-293. The synergism of two drugs is calculated based on the combination index (CI) and median-effect equation, calculated by the software Compusyn. In this study, we observed the dose-dependent cytotoxicity, increased ROS generation, down-regulation of mitochondrial membrane potential (MMP), and increased chromatin condensation in combination doses, compared to the single drugs. Moreover, the cell cycle arrest and overexpression of checkpoints regulatory genes ATM, CHEK2, BRCA1, BRCA2, and TP53 were observed for preventing cell proliferation. Methodology: MTT analysis is performed to determine cell viability at different doses, ROS-generation is observed using DCFH-DA stained fluorescence images, reducing MMP is detected by Rhodamine123 staining method, condensation of DNA is detected by Hoechst33342 stained photomicrographs, apoptosis analysis is performed by both AO/EtBr staining and Annexin-V/FITC & PI flow cytometry. To validate the findings, mRNA expression of cell-cycle check-point markers is quantified by rt-qPCR. Result & Conclusion: The calculated combination dose showing maximum growth inhibition is 33.117µM Cur. + 0.331µM Doxo. against MDA-MB-231 and 14.361µM Cur. + 0.14361µM Doxo. against MCF-7 with negligible toxicity against normal HEK-293 cells. Significant increase in mRNA expressions of TP53, BRCA1, BRCA2, ATM, and CHEK2 genes (Ct-value) were evident with G0/G1 and S-phase cell cycle arrest. Thus, Curcumin synergistically chemo-sensitizes the anticancer activity of Doxorubicin and enhances the responses towards conventional chemo-therapy attenuating breast cancer