Implementation of the Chou-Talalay method for studying the in vitro pharmacodynamic interactions of binary and ternary drug combinations on MDA-MB-231 triple negative breast cancer cells

Triple Negative Breast Cancer (TNBC) treatment is more challenging than other subtypes of breast malignancy and due to the lack of markers for the molecularly targeted therapies (ER, PR, and HER-2/Neu), the conventional chemotherapeutic agents are still the mainstay of the therapeutic protocols of its patients. Unfortunately, the initial good response to the chemotherapy eventually turns into a refractory drug-resistance, therefore; more efficient therapeutic regimens are urgently required. Here, we examined the single and combined cytotoxicity of PU-H71, Dehydroepiandrosterone, Berberine, and Sorafenib on the MDA-MB-231 triple negative breast cancer cells after 48 h incubation period through the neutral red uptake assay. Based on Median Effect Equation (Chou), Combination Index Theorem and Dose Reduction Index Equation (Chou-Talalay), we tested six binary combinations and four ternary ones to define and quantify their pharmaco-dynamic interactions (synergism, antagonism or additivity). The highest-to-lowest order of potency of a single drug treatment was PU-H71 > Sorafenib > Berberine > Dehydroepiandrosterone. At fractional affected level (fa) ≥ 0.90, almost all the actual and computer-simulated combinations exerted synergistic effects, where (PU-H71 plus Sorafenib), (Berberine plus Sorafenib) and (PU-H71 plus Berberine plus Sorafenib) were the strongest synergistic combinations with CI value < 0.30. Based on our in vitro combination results, we suggest subsequent downstream investigations to understand the molecular mechanisms of such promising synergistic combinations. Additionally, we recommend the application of such combinations on TNBC-xenografted animal models to effectively establish the go\no-go decision of the further application in clinical settings.