This study aims to evaluate the in-vitro cytotoxic and anti-migratory effects of Ficus deltoidea L. plant on prostate cancer cells, identify the active compound/s and characterise their mechanism of actions. The crude methanolic extract was partitioned into n-hexane (FD1h, FD2h) chloroform (FD1c, FD2c) and aqueous extracts (FD1a, FD2a). Active fractions were further fractionated. Active compound/s were dereplicated based on UHPLC-HRMS/MS. In-vitro mechanistic studies on PC3 cells included: Annexin V-FITC staining, MMP depolarization measurements, activity of caspases 3/7, nuclear DNA fragmentation, cell cycle analysis, modulation of Bax, Bcl-2, Smac/Diablo and Alox-5 mRNA gene expression by RT-PCR. 2D migration and 3D invasion were tested by exclusion assays and modified Boyden chamber [1]. The expression of VEGF-A, CXCR4 and CXCL12 in PC3 cells were analysed by RT-PCR. FD1c and FD2c extracts induced cell death (P<0.05) via apoptosis as evidenced by nuclear DNA fragmentation accompanied by an increase in MMP depolarization (P < 0.05), and activation of caspases 3/7 (P<0.05). All active plant extracts up-regulated Bax and Smac/DIABLO, and down-regulated Bcl-2 (P<0.05). Active plant extracts inhibited both migration and invasion of PC3 cells (P<0.05). These effects were accompanied by down-regulation of both VEGF-A and CXCL-12 gene expressions (P<0.001). UHPLC-HRMS/MS dereplication identified isovitexin in FD1c; and oleanolic acid, moretenol, betulin, lupenone and lupeol in FD2c. In conclusion, FD1c and FD2c were able to induce apoptosis by activating the intrinsic pathway, inhibit both migration and invasion by modulating the CXCL12-CXCR4 axis, and inhibit angiogenesis by modulating VEGF-A expression in PC3 cell line.
Acknowledgement
I would like to thank the Foundation Plants for Health for awarding me with a research subsidy and giving me the opportunity to collaborate with Prof Jean-Luc Wolfender in completing this project.
[1] Ridley, Anne J., et al. "Cell migration: integrating signals from front to back." Science 302.5651 (2003): 1704-1709.