Synergistic anticancer potential of Andrographis paniculata and Datura metel combined with Doxorubicin in inhibiting breast cancer cells

Abstract

Breast cancer remains one of the prevalent malignancies affecting women worldwide. This study investigates the synergistic anticancer potential of triple-combinatorial phytotherapeutic agents, Andrographis paniculata and Datura metel, along with the chemotherapeutic drug doxorubicin (DOX), on estrogen-receptor-positive (MCF-7) and triple-negative (MDA-MB-231) breast carcinoma cells and normal breast epithelial cells (MCF-10A). The objective was to identify a synergistic drug–natural product combination that enhances cytotoxic efficacy while minimizing off-target toxicity. Sequential ultrasound-assisted extractions were performed using hexane, ethyl acetate, methanol, and aqueous ethanol. SRB-based cytotoxic screening revealed that ethyl acetate extract of A. paniculata (AP) and methanolic extract of D. metel (DM) were most potent, demonstrating IC₅₀ values at 168.2 µg/mL (MCF-7) and 90.26 µg/mL (MDA-MB-231) for A. paniculata, and 0.23 µg/mL (MCF-7) and 21.59 µg/mL (MDA-MB-231) for D. metel after 72 h, with relatively low cytotoxicity toward MCF 10A. Bioactivity-guided fractionation of A. paniculata ethyl acetate extract was performed using normal-phase column chromatography with a 10% gradient solvent system consisting of hexane and ethyl acetate. Methanolic extract of D. metel was subjected to Kupchan’s solvent-solvent partitioning to isolate active constituents. Fraction C8 of A. paniculata (AP-C8) exerted denotable cytotoxicity with IC₅₀ values at 32.99 ± 15.27 µg/mL on MCF-7 and 4.674 ± 0.73 µg/mL on MDA-MB-231 at 48 h post treatment. The chloroform fraction of D. metel (DM-F2) demonstrated the highest cytotoxicity with IC₅₀ at 16.04 ± 0.33 µg/mL and 2.535 ± 0.09 µg/mL on MCF-7 and MDA-MB-231, respectively. Triple combination treatments (DOX + AP-C8 + DM-F2) were assessed in various fixed dose ratios using the SRB assay. CompuSyn software analysis confirmed that the highest synergistic interactions were observed at a ratio of 1:64:256 on MCF-7 (combination index [CI]=0.71846) and at 1:1.25:10 on MDA MB-231 (CI=0.77291). Both combinations demonstrated higher dose reduction index (DRI) values in a favorable range, especially for DOX, indicating potential for therapeutic dose minimization. Mainly, the combinations exhibited minimal cytotoxic effects in MCF-10A cells, supporting their selective anticancer potential in the selected combination ratios.