Regulatory Effects of Apatinib in Combination with Piperine on MDM-2 Gene Expression, Glutathione Peroxidase Activity and Nitric Oxide level as Mechanisms of Cytotoxicity in Colorectal Cancer Cells

Purpose: Apatinib has been utilized in colon cancer therapies but its efficiency and molecularmechanism are not fully understood. Chemotherapy in combination with non-toxic compoundscan be an effective treatment strategy for cancer. Consequently, this study was carried out toevaluate the effects of apatinib and piperine on colorectal cancer (CRC) cell line and theirpotential anti-cancerous mechanisms in vitro. Methods: The effects of apatinib and piperine on HCT-116 CRC cells were detected byassessing cell viability using MTT assay. The potential cytotoxic mechanisms of apatinib andpiperine were investigated by evaluating MDM-2 gene expression ratio using real-time PCRassay. Moreover, the glutathione peroxidase (GPX) activity and nitric oxide (NO) levels wereassessed by colorimetric assays. Results: The proliferation rate of CRC cells decreased by increasing the concentrations ofpiperine or apatinib. When HCT-116 cells were treated with different concentrations of apatinibin combination with piperine, the synergistic effects were observed (combination index < 1).In HCT-116 cells treated with apatinib and piperine at the concentrations of 0.5×IC50 and0.2×IC50, the MDM-2 gene expression was downregulated and NO levels increased comparedto the untreated control cells and related single treatments. In addition, GPX activity significantlydecreased in combination treatment at 0.5×IC50 concentration of both agents versus singletreatments. Conclusion: Apatinib in combination with piperine could significantly inhibit the growth ofCRC cells. These cytotoxic effects were induced by regulation of MDM-2 gene expression andinhibition of antioxidant marker.