Anticancer and Apoptotic Potentials of Gliricidia sepium Leaf Extract on Breast HCC1395, Prostate DU145 and Colorectal CT26 Cell Lines

Background: Plants have become a household name in the quest for effective and safe cancer chemotherapy in the Pharmaceutical industry. We studied the Brine shrimps lethality, antiproliferative and apoptotic potentials of Gliricidia sepium leaf extract against some selected cancer cell lines.

Methodology: Plant leaves extraction was done with 70% ethanol and petroleum ether. Twenty four hours old shrimp’s larvae exposure to different concentrations (1 mg/ mL, 100 µg/ mL, 10 µg/ mL and 1 µg/ mL) of the extracts were used to evaluate the cytotoxicity in Brine shrimps lethality assay (BSLA). Three cancer cell lines: Breast (HCC1395); Prostate (DU145); Colorectal (CT26) and one normal cell line (Vero E6) were used for the in-vitro cytotoxicity testing using MTT assay. IC50, CC50 and Selectivity Index (SI) were determined respectively, using standard methods. Quantitative reverse transcription-polymerase chain reaction (qRT-PCR) was carried out on HCC1395 and DU145 cell lines. Primers for the genes p53 and BAX were generated and amplified for apoptotic evaluation. The fold change relative to GADPH, the house­keeping gene, was calculated using double delta Ct analysis [2ˉΔΔCT].

Results: The results of BSLA showed that both extracts of G. sepium are strongly toxic (LC50 < 100 µg/ml). However, petroleum ether extract (PGS) exhibited the highest toxicity to the shrimps, with LC50 of 11.95 µg/ml. This is about 8 times more cytotoxic than cyclophosphamide (LC50 of 98.76 µg/ml). On cancer cells, PGS exhibited varying antiproliferative activities; it was high on prostate (IC50 = 12.76 µg/ml), antiproliferative (IC50 = 23.55 µg/ml) and moderately antiproliferative (IC50 = 77.58 µg/ml) on breast and colorectal cancer respectively. PGS CC50 value was greater than 100 µg/ml. Ethanol extract (EGS) showed very high toxicity to all the tested cancer cells (IC50 <20µg/ml) with low cytotoxicity (CC50 = 41.81 µg/ml) to the normal cells. It exhibited a significant difference and high selectivity index across all the cell lines used in this study. EGS also upregulated both p53 and BAX biomarkers in the qRT-PCR apoptotic study.

Conclusion: The leaf extracts of G. sepium is a potential anticancer agent. The 70% ethanol extract selectively induced antiproliferative activities on cancer cells and upregulated apoptotic genes. The Petroleum ether extract also showed very low cytotoxicity to the normal cells. The plant should be considered a novel candidate for further studies.