3,3'-Di-O-methylellagic Acid Isolated from Euphorbia humifusa Willd Suppresses Prostate Cancer Cell Viability via Regulating VDAC1 Protein Expression.

Background

Prostate cancer (PCa) is the leading male urinary malignancy globally. Our previous article demonstrated the anti-PCa activity of Euphorbia humifusa Willd water extract (EHW) and some of its compounds via downregulating AR expression, but the anti-PCa active compounds from Euphorbia humifusa Willd (EH) and their mechanisms of action are yet to be clarified. Thus, the current article studied the in vitro anti-PCa effects of 3,3'-di-O-methylellagic acid (3,3'-di-O-Me-EA) derived from EHW and the related mechanism involved.

Methods

3,3'-di-O-Me-EA was isolated from EHW applying bioassay-guided fractionation. The spectroscopic methods were used to determining the structure of 3,3'-di-O-Me-EA. The drug-likeness and ADMET properties (absorption, distribution, metabolism, excretion, and toxicity) of 3,3'-di-O-Me-EA were analyzed in silico. Molecular docking and real-time surface plasmon resonance (SPR) analysis were performed to measure the interaction of 3,3'-di-O-Me-EA and VDAC1 protein. The viability and apoptosis of 22RV-1 and DU145 PCa cells were determined using MTT and Annexin V-FITC staining assay, respectively. q-PCR and Western blot experiments were used to analyzing the gene and protein expressions of VDAC1.

Results

3,3'-di-O-Me-EA was isolated and purified from EHW with a purity of ≥90.06%, and its structure was identified by HRTOF mass, NMR, and an authentic standard. In silico ADMET analysis indicated its favorable drug-like and pharmacokinetic properties. Molecular docking and SPR results confirmed that 3,3'-di-O-Me-EA could bind with the VDAC1 protein. Moreover, 3,3'-di-O-Me-EA dose- and time-dependently inhibited 22RV-1 and DU145 PCa cell viability, and induced apoptosis in a dose-dependent manner (p < 0.05). RT-qPCR and Western blot results showed that 3,3'-di-O-Me-EA dose-dependently up-regulated VDAC1 gene and protein expression levels in 22RV-1 and DU145 cells (p < 0.05). Meanwhile, in VDAC1-depleted 22RV-1 and DU145 cells, 3,3'-di-O-Me-EA down-regulated VDAC1 gene and protein expression levels, increased cell viability, and inhibited apoptosis compared to 22RV-1 and DU145 cells (p < 0.05). Furthermore, 3,3'-di-O-Me-EA enhanced VDAC1 gene and protein expression levels, inhibited cell viability, and induced apoptosis in VDAC1-overexpressed 22RV-1 and DU145 cells compared with 22RV-1 and DU145 cells (p < 0.05). Overall, EH active compound 3,3'-di-O-Me-EA may inhibit viability and induce apoptosis of 22RV-1 and DU145 PCa cells via up-regulating VDAC1 gene and protein expression levels.

Conclusion

The results indicated that the 22RV1 and DU145 PCa cell viability inhibitory effects of 3,3'-di-O-Me-EA isolated from EH may be mediated by induction of apoptosis through up-regulation of VDAC1 gene and protein expression levels.