Bioactivity of fractions and constituents of Piper capense fruits towards a broad panel of cancer cells

Abstract

Ethnopharmacological relevance: Piper capense is a medicinal spice whose fruits are traditionally used as aqueous decoction to heal several ailments such as trypanosomiasis, helminthic infections, and cancer. Aim of the study. (1) To perform phytochemical investigation of the methanol extract of Piper capense; (2) to evaluate the cytotoxicity of botanicals (PCF, fractions PCFa-e), isolated phytochemicals on a broad panel of animal and human cancer cell lines; (3) to evaluate the induction of apoptosis of the most active samples. Material and methods: Resazurin reduction assay (RRA) was used to determine the cytotoxicity of the studied samples. Cell cycle distribution (PI staining), apoptosis (annexin V/PI staining), mitochondrial membrane potential (MMP; JC-1) and reactive oxygen species (ROS; H2DCFH-DA) were measured by flow cytometry. Column chromatography (CC) was used for the purification of PCF, whilst nuclear magnetic resonance (NMR) spectroscopic and mass spectrometric (MS) analyses were applied for structural elucidation. Results: The phytochemical investigation of PCF led to the isolation of 11 compounds: licarin B (1), licarin A (2), 7-(1,3-benzodioxol-5-yl)-7,8-dihydro-8-methyl-5-(2-propenyl)-furo [3,2-e]-1,3-benzodioxole (3), nitidine isocyanate (4), 5-hydroxy-7,4'-dimethoxyflavone (5), cardamomin (6), sitosteml (7) and stigmasteml (8), beta-sitosterol 3-O-beta-D-glucopyranoside (9), oleanolic acid (10) and lupeol (11). Fraction PCFb, compound 2 and doxorubicin (as positive control drug) revealed cytotoxic effects towards the 18 tested cancer cell lines. The IC50 values ranged from 6.1 mu g/mL (against CCRF-CEM cells) to 44.2 mu g/mL (against BRAF-V600E homozygous mutant melanoma cells) for PSCb; from 4.3 mu M (against CCRF-CEM cells) to 21.8 mu M (against HCT116 p53(-/-)) for compound 2 and from 0.02 mu M (against CCRF-CEM cells) to 123.0 mu M (against CEM/ADR5000 cells) for doxorubicin. PCFb and compound 2 induced apoptosis in CCRF-CEM cells mediated by activation of caspase 3/7, 8 and 9, MMP alteration and increased ROS production. Conclusion: Piper capense is a source of potent cytotoxic botanicals and phytochemicals that could help to fight various types of cancer including multidrug resistance phenotypes. PCFb and compound 2 should further be explored to develop new drugs to fight malignancies.