Screening the Combination of Gemcitabine, Clomipramine, and Resveratrol in HL-60 Leukemia Cells

Background and Objectives: Potential anti-neoplastic effects of resveratrol, which has antioxidant features combined with clomipramine, which has antineoplastic features, or with gemcitabine, used as a nucleoside analog widely used in chemotherapy, were evaluated together and individually on the HL-60 leukemia cells in this in vitro screening study. Materials and Methods: HL-60 cells were treated with gemcitabine, clomipramine, resveratrol, or their combinations at concentrations ranging from 1 to 200 µM. Cell viability was assessed at 24, 48, and 72 h using the trypan blue exclusion method, and results are expressed as a percentage of time-matched untreated controls. Cell proliferation was further evaluated by bromodeoxyuridine (BrdU) immunohistochemical labeling. All experiments were performed in triplicate, and statistical analyses were conducted using one-way analysis of variance (ANOVA) with post hoc comparisons. Results: Gemcitabine markedly reduced HL-60 cell viability at all concentrations and time points (p < 0.001), indicating strong time-dependent cytotoxicity, with a significant drop in BrdU proliferation index at 48 h (p < 0.001). Clomipramine exhibited a biphasic response: high concentrations decreased viability (p < 0.05), while low concentrations allowed partial recovery by 72 h. Resveratrol showed concentration-dependent cytotoxicity, with reduced viability at high concentration and near-control levels at low concentration by 72 h; BrdU indices remained significantly lower than control (p < 0.001). Combination treatments with gemcitabine showed no additive cytotoxic or antiproliferative effects (p > 0.05). A transient enhanced effect was observed in the clomipramine + resveratrol group at 24 h (p < 0.01 vs. clomipramine; p < 0.05 vs. gemcitabine). Conclusions: Gemcitabine, clomipramine, and resveratrol all exhibited inhibitory effects on cell proliferation in HL-60 cell cultures. However, the combination treatments did not show additional cytotoxicity or additive effects. These findings suggest that while each of these compounds individually has the potential to inhibit cell growth, their combined application does not enhance the cytotoxic effects beyond those observed with single treatments. These findings highlight the necessity of a rational approach when considering novel drug combinations.