A Homodimer of Withaferin A Formed by Base-Promoted Elimination of Acetic Acid from 27- O -Acetylwithaferin A Followed by a Diels-Alder Reaction.

Treatment of 27- O -acetylwithaferin A ( 2 ) with the non-nucleophilic base, 1,8-diazabicyclo[5,4,0]undec-7-ene (DBU), afforded 5β,6β-epoxy-4β-hydroxy-1-oxo-witha-2(3),23(24),25(27)-trienolide ( 3 ) and 4 , a homodimer of withaferin A resulting from a Diels-Alder [4 + 2] type cycloaddition of the intermediate α,β-dimethylene-δ-lactone ( 9 ). Structures of 3 and 4 were elucidated using HRMS and 1D and 2D NMR spectroscopic data. The structure of 4 was also confirmed by single crystal X-ray crystallographic analysis of its bis -4- O - p -nitrobenzoate ( 8 ). Formation of withaferin A homodimer ( 4 ) as the major product suggests regio- and stereoselectivity of the Diels-Alder [4 + 2] cycloaddition reaction of 9 . Acetylation of 2 - 4 afforded their acetyl derivatives 5 - 7 , respectively. Compounds 2 - 4 and 6 - 8 were evaluated for their cytotoxic activities against four prostate cancer (PC) cell lines (LNCaP, 22Rv1, DU-145, and PC-3) and normal human foreskin fibroblast (HFF) cells. Significantly, 4 exhibited improved activity compared to the other compounds for most of the tested cell lines.