Withaferin A and Withanolide D Analogues with Dual Heat-Shock-Inducing and Cytotoxic Activities: Semisynthesis and Biological Evaluation

Abstract

Withanolides constitute a valuable class of bioactive natural products because some members of the class are known to exhibit cytotoxic activity and also induce a cytoprotective heat-shock response. In order to understand the relationship between their structures and these dual bioactivities of the withanolide scaffold, we obtained 25 analogues of withaferin A (WA) and withanolide D (WD) including 17 new compounds by semisynthesis involving chemical and microbial transformations. Hitherto unknown 16β-hydroxy analogues of WA and WD were prepared by their reaction with triphenylphosphine/iodine, providing unexpected 5β-hydroxy-6α-iodo analogues (iodohydrins) followed by microbial biotransformation with Cunninghamella echinulata and base-catalyzed cyclization of the resulting 16β-hydroxy iodohydrins. Evaluation of these 25 withanolide analogues for their cytotoxicity and heat-shock-inducing activity (HSA) confirmed the known structure-activity relationships for WA-type withanolides and revealed that WD analogues were less active in both assays compared to their corresponding WA analogues. The 5β,6β-epoxide moiety of withanolides contributed to their cytotoxicity but not HSA. Introduction of a 16β-OAc group to 4,27-di- O-acetyl-WA enhanced cytotoxicity and decreased HSA, whereas introduction of the same group to 4- O-acetyl-WD decreased both activities.

Structural basis for species-selective targeting of Hsp90 in a pathogenic fungus

“…Non-tumorigenic mouse 3T3 fibroblasts were plated in 384-well format at 2500 per well. After overnight incubation, two-fold dilutions of test compounds were added to wells for 72 h followed by measurement of relative viable cell number using standard resazurin dye-reduction assay and microplate fluorometer (Tecan Infinite M1000Pro, 544 nm excitation and 590 nm emission) as previously published 74 .…”

“…Concentration-dependent activation of the heat-shock response in human cells by Hsp90 inhibitors was measured in 96-well format as previously published using 293T cells stably transduced with a reporter construct encoding GFP-luciferase fusion protein under transcriptional control of heat-shock elements from the HSP70B gene 74 . Activation of the response in C. albicans was also measured in 96-well format using a reporter strain (CaLC922) engineered with HSP70 promoter elements driving expression of the LacZ gene from Streptococcus thermophilus integrated at the HSP70 locus with the URA3 marker for selection.…”

Structure–Activity Relationships of Withanolides as Antiproliferative Agents for Multiple Myeloma: Comparison of Activity in 2D Models and a 3D Coculture Model

“…30 The WFA analogues 5−7, 10−16, and 32−35 and WND analogues 37−42, 49, and 51 were obtained as described previously. 51,52 The new analogues of WFA ( 8 52 Treatment of 15 with Ph 3 P:I 2 in CH 2 Cl 2 at 25 °C for 1 h 54 followed by usual workup and purification by RP-HPLC resulted in the isolation of 24. The structure of 24 was determined by the analyses of its 1 H, 13 C, and 2D NMR spectroscopic data.…”

Withanolide D Enhances Radiosensitivity of Human Cancer Cells by Inhibiting DNA Damage Non-homologous End Joining Repair Pathway

“…After WFA, WD highlighted the potential of withanolides to act as promising sources of radiosensitizers. Several studies investigated the relationship between the structure and the diverse bioactivities reported for withanolides to better understand their mechanism of action and modify the withanolide scaffold accordingly to enhance its effect (43)(44)(45)(46). Although WD-treated cells displayed a lower expression level of DNA damage repair proteins after irradiation than non-treated cells (maximal at 24 h), this inhibition is not complete.…”