Anticancer potential of polyphenolic compounds of Barhi date palm kernels through induction of apoptosis in cancer cells

Abstract

Cancer is a major burden of disease worldwide, with an estimated 6.9 million cancer deaths in 2018. The number of new cases is expected to increase by about 70% over the next two decades. Polyphenolic compounds (extractable, EPPs and non-extractable, NEPPs), which are most abundantly present as natural bioactive compounds, have recently shown to be a powerful in-vitro anticancer activity through inducing apoptosis in cancer cells. Date palm kernels have gained interest as a valuable by-product “waste” of the date fruit industry and have been identified as a rich source of EPP and NEPP. Therefore, this study aimed to investigate the anticancer potentiality of polyphenols from Barhi date palm kernels (BDPK) extracts through inducing apoptosis in cancer cells. In the first stage of the study, chemical composition of BDPK was performed before the extraction process to identify its nutritional quality. Cytotoxic and antioxidant activities were screened in aqueous and organic extracts of BDPK by using in-vitro assays, and accordingly, the optimisation of a set of extraction conditions for high-yield recovery of EPP and NEPP, with high anticancer and antioxidant activities, was performed using response surface methodology (RSM). From the obtained results, it can be revealed that EPP exhibited more promising cytotoxic and antioxidant properties when compared to NEPP. Therefore, screening phenolic profile by LC-QTOF/MS to identify new cytotoxic compounds in the active EPP crude extract was done in the next stage. The isolation process was performed through successive steps of various chromatographic methods which had been assisted by the cytotoxic and antioxidant activities of the separated fractions and sub-fractions. Three new cytotoxic flavonoids nobiletin (C1, NOB), tectorigenin (C2, TEC) and persicogenin (C3; PERS) were successfully isolated and identified from BDPK for the first time. The final structures of the isolated flavonoids were established with the aid of spectroscopic analysis including one- and two-dimension at nuclear magnetic resonance (NMR), melting points (m.p.), preparative thin layer chromatography (TLC) and mass spectrometry (MS). The three isolated flavonoids exhibited cytotoxic effects against two human cancer cell lines; lung (A549) and colon (HT29), but not normal cells, with higher cell death in A549 than in HT29 after 72 hours treatment, where PERS was found to be the most potent compound, followed by NOB and TEC (IC50=5.75μg/mL, 8.56μg/mL, 10.48μg/mL, respectively). Subsequently, the mechanism of cell death induced by BDPK crude extracts (EPP and NEPP), and the isolated compounds was demonstrated in the last stage of the research. After performing various morphological, biochemical and molecular assessments, it was established that the BDPK extracts induced late stages of apoptosis as there was evidence of the DNA degradation, and large percent of the cells population being in the sub-G0 phase induced by the two extracts. Furthermore, EPP and NEPP exhibited dependent mitochondrial signalling pathway as seen with caspase-9 and induced receptor-mediated (extrinsic) apoptotic pathway as seen with caspase-8. In conclusion, the present study has revealed the anticancer potentials of BDPK extracts and the three new cytotoxic isolated compounds may be new promising preventive/therapeutic candidates in treating lung and colon cancers.