Evaluation of the effects of stem extract of Euphorbia poissonii on the gene expressions of BRCA1, BRCA2 and p53 in breast cancer cell line MCF-7

Abstract

Background: Breast cancer is the most frequently diagnosed cancer type and the leading cause of cancer death among females. Many Nigerian plants have been hypothesized to have anticancer potentials. However, a limited number have been subjected to acceptable scientific evaluation for their potential anticancer effects and mechanism of action. Euphorbia poissonii is a plant native to Northern Nigeria and it is known to possess cytotoxic properties.

Objective: The possibility of the crude ethanol extract, n-hexane, ethyl-acetate, chloroform, n-butanol and aqueous fractions of Euphorbia poissonii to modulate the tumour suppressor genes, BRCA1, BRCA2 and TP53 was investigated in the human breast cancer MCF-7 cell line.

Methods: The air-dried pulverized Euphorbia poissonii stem was soaked in 2 litres of 80% aqueous-ethanol at room temperature. Filtration was done after four days, using Muslin bag. Sequential fractionation of the resulting ethanol-free crude extract was carried out in a separating funnel using 150 ml each of n-butanol, ethyl acetate, nhexane and chloroform, in increasing order of polarity. The fractions: crude ethanol (ET), n-butanol (BU), ethyl acetate (EA), n-hexane (HE), chloroform (CH) and aqueous (AQ) were collected, separately into pre-weighed crucibles and dried in the hot air oven. Qualitative and quantitative phytochemical screenings were performed on the 6 panel of solvent fractions. MCF-7 cells were grown in a 25-cm² flask with Dulbecco's Modified Eagle Media (DMEM) supplemented with 10% fetal bovine serum (FBS), Streptomycin and Penicillin. Breast cancer MCF-7 cell line was treated separately with each of the 6 fractions of the plant extract and the gene expressions of BRCA1, BRCA2 and TP53 were measured using semi-quantitative Reverse Transcription Polymerase Chain Reaction (RT-PCR) analysis following extraction of the RNA from the treated cells.

Results: Qualitative and quantitative phytochemical analyses revealed the presence of tannin (80.50%), saponin (27.85%), phenol (4.84%), flavonoid (4.20%) and alkaloid (2.00%). The results from the gene expression
study showed down-regulation of BRCA1. BRCA2 was up-regulated by ET, BU, EA, and AQ; TP53, by HE, CH, and EA.

Conclusion:  Euphorbia poissonii demonstrates modulatory potentials on BRCA2 and TP53 gene expression in MCF-7 cell lines. The up-regulation of BRCA2 and TP53 suggests that the fractions contain bioactive compounds that restored the loss of functions of BRCA2 and P53 in the human breast cancer cell line, MCF-7.

Keywords: Euphorbia poissonii, phytochemicals, BRCA1, BRCA2, P53, gene expression, breast cancer