The effects of papaverine on proliferation and cell death induction in cancer cells

Abstract

Cancer is a leading cause of mortality with close to 10 million deaths reported in 2020 with lung- and breast cancer being the most diagnosed types of cancer. Papaverine (PPV), a natural occurring, non-narcotic benzylisoquinoline alkaloid isolated from the Papaver somniferum plant, is currently approved by the Food and Drug Administration of the United States for vasodilation purposes. Previous studies have indicated that papaverine inhibits cell growth and potentially induces cell death in tumourigenic cell lines. However, specific effects on oxidative stress, cell migration, vascular endothelial growth factor (VEGF) and cell cycle progression in tumourigenic cell lines remains elusive. The influence of papaverine was evaluated in this study on cell proliferation by using crystal violet staining (spectrophotometry), morphology by means of light microscopy, hydrogen peroxide production by utilising 2,7-dichlorofluoresceindiacetate staining (fluorescent microscopy), cell cycle progression by means of propidium iodide staining (flow cytometry), cell migration utilising a scratch assay (light microscopy), focal adhesion tyrosine kinase (FAK) expression by using enzyme-linked immunosorbent assay (ELISA) and VEGF expression by the quantification of vascular endothelial growth factor receptor 1 (VEGF-R1), vascular endothelial growth factor receptor 2 (VEGF-R2) and vascular endothelial growth factor ligand B (VEGF-B) levels using ELISAs in a triple negative breast cancer cell line (MDA-MB-231), adenocarcinoma alveolar cancer cell line (A549) and a prostate cancer cell line (DU145). Results indicated that exposure to PPV resulted in time- and dose-dependent antiproliferative activity in all three tumourigenic cell lines. Exposure to 150 μM PPV for 48 h reduced cell growth to 56%, 53%, and 64% in the MDA-MB-231 cell line, A549 cell line and DU145 cell line, respectively. Light microscopy revealed that PPV exposure for 72 h increased cellular protrusions in MDA-MB-231- and A549 cells to 34% and 23%, respectively. PPV-treated cells also demonstrated an increase in hydrogen peroxide with a fold increase to 1.27, 1.31 and 1.44 in MDA-MB-231-, A549- and DU145 cells, respectively. Furthermore, exposure to PPV for 72 h resulted in an increase of cells in the sub-G1 phase to 46% and endoreduplication by 10%. The migration assay revealed that after 48 h, PPV (100 µM) reduces cell migration to 81%, 91% and 71% in MDA-MB-231-, A549- and DU145 cells, respectively. Furthermore, VEGF B expression was reduced to 0.79-, 0.71- and 0.73 fold when exposed to PPV in MDA-MB-231-, A549- and DU145 cells after 48 h treatment with PPV whilst exposure to PPV for 48 h increased VEGF R1 expression in MDA-MB-231- and DU145 cells to 1.38 and 1.46 whilst a fold decrease in VEGF R1 expression was observed in A549 cells to 0.90 after exposure to 150 µM. No significant effects were observed on VEGF R2- and FAK expression after exposure to PPV for 48 h. This study aided in the understanding regarding the influence of PPV on tumourigenic cell lines. Moreover, data obtained in this study may improve the understanding and use of phytomedicinal compounds in biomedical preclinical research and could potentially give rise to improved alternatives to compliment or substitute current chemotherapy regimens.