5- or/and 20-O-alkyl-2,3-dehydrosilybins: Synthesis and biological profiles on prostate cancer cell models

Abstract

To investigate the effects of alkylation at 5-OH and 20-OH of 2,3-dehydrosilybin on prostate cancer cell proliferation, the synthetic approaches to 5- or/and 20-O-alkyl-2,3-dehydrosilybins, through a multi-step sequence from commercially available silybin, have been successfully developed. The first three reactions in the syntheses were completed through a one-pot procedure by managing anaerobic and aerobic conditions. With these synthetic methods in hand, twenty-one 2,3-dehydrosilybins, including seven 20-O-alkyl, seven 5,20-O-dialkyl, and seven 5-O-alkyl-2,3-dehydrosilybins, have been achieved for the evaluation of their biological profiles. Our WST-1 cell proliferation assay data indicate that nineteen out of the twenty-one 2,3-dehydrosilybins possess significantly improved antiproliferative potency as compared with silybin toward both androgen-sensitive (LNCaP) and androgen-insensitive prostate cancer cell lines (PC-3 and DU145). 5-O-Alkyl-2,3-dehydrosilybins were identified as the optimal subgroup that can consistently inhibit cell proliferation in three prostate cancer cell models with all IC₅₀ values lower than 8µM. Our flow cytometry-based assays also demonstrate that 5-O-heptyl-2,3-dehydrosilybin effectively arrests the cell cycle in the G₀/G₁ phase and activates PC-3 cell apoptosis.

Keywords: 2,3-Dehydrosilybin derivatives; Anti-proliferative activity; Cell apoptosis; Cell cycle regulation; Prostate cancer; Synthesis.

Figure 1

Structures of silybin and 2,3-dehydrosilybins

Figure 3

Cell cycle analysis of PC-3 cells. PC-3 cancer cells were untreated or treated with 5-O-heptyl-2,3-dehydrosilybin (35). Cells were harvested after 16 and 24 hours, fixed, stained, and analyzed for DNA content. The distribution and percentage of cells in G₀/G₁, and G₂/M phase of the cell cycle are indicated.

Figure 4

Evolution of viable, apoptotic, and necrotic PC-3 cells populations in response to increasing dosages of 5-O-heptyl-2,3-dehydrosilybin (35).

Scheme 1

Synthesis of 20-O-alkyl-2,3-dehydrosilybins (14–20) and 5,20-O-dialkyl-2,3-dehydrosilybins (21–27). Reactants and conditions: (i) BnBr (1.1 eq), K₂CO₃ (4 eq), acetone (0.1 M), argon, reflux for 4 h; (ii) BnBr (1.1 eq), K₂CO₃ (2 eq), DMF (0.25 M), Air, rt, 6 h; (iii) RI or RBr (2eq), K₂CO₃ (2 eq), DMF (0.25 M), rt, 24 h; (iv) ammonium formate (10 eq), MeOH/Ethyl Acetate (50/50, v/v), Pd/C, reflux, overnight, argon: (v) RI or RBr (4–10 eq), K₂CO₃ (10–20 eq), DMF (0.25 M), rt, 24–48 h.

Scheme 2

Synthesis of 5-O-alkyl-2,3-dehydrosilybins (29–35). Reactants and conditions: i) (a) BnBr (1.1 eq), K₂CO₃ (4 eq), acetone (0.1 M), argon, reflux overnight; (b) Remove acetone, DMF (0.25 M), BnBr (2.2 eq), K₂CO₃ (3 eq), rt, 4 h, air; ii) (a) RI or RBr (8 eq), K₂CO₃ (8 eq), DMF (0.2 M), rt, 4 h; (b) ammonium formate (10 eq), EtOAc and MeOH, Pd/C (20%), reflux, overnight, argon.