Synergistic Anticancer Efficacy of Curcumin and Doxorubicin Combination Treatment Inducing S-phase Cell Cycle Arrest...

Curcumin (Cur) is a polyphenol phyto-compound found in turmeric (Curcuma longa) that inhibits tumorigenesis by introducing apoptosis and restricting cell survival and proliferation. This in vitro research article focuses on the pharmacodynamic interactions of Cur combined with the commercial drug doxorubicin (Doxo) to enhance the cytotoxicity of Doxo at lower doses against triple-negative breast cancer cells (MDA-MB-231) with the chemo-protective effect against normal HEK293 cells. In this study, we observed the dose-dependent cytotoxicity, increased reactive oxygen species (ROS) generation, and increased chromatin condensation in combination doses compared to single doses. Moreover, the cell cycle arrest and overexpression of checkpoint regulatory genes ATM, P53, CHEK2, BRCA-1, and BRCA-2 were observed to prevent cell proliferation.

3-(4,4-Dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) analysis is performed to determine cell viability at different doses. ROS generation is observed using DCFH-DA-stained fluorescence images. Hoechst33342-stained photomicrographs detect DNA condensation. Apoptosis analysis is performed using Annexin V/FITC and PI flow cytometry. To validate the findings, mRNA expression of cell-cycle checkpoint markers is quantified using reverse transcription quantitative polymerase chain reaction analysis.

The calculated combination dose showing maximum growth inhibition is 33.12 µM Cur + 0.33 µM Doxo against MDA-MB-231 cells with negligible cytotoxicity against normal HEK293 cells. There is a significant increase in mRNA expressions of P53 (4.43-fold), CHEK2 (2.58-fold),BRCA-1 (2.01-fold), BRCA-2 (1.60-fold),and ATM (0.91-fold) genes (2^{- ∆∆ Ct}) after treatment with the combination doses, evident with the major S-phase cell cycle arrest in MDA-MB-231 cells.

Cur synergistically chemo-sensitizes the anticancer activity of Doxo and enhances the responses toward conventional chemotherapy attenuating breast cancer.