Wild type p53 function in p53Y220C mutant harboring cells by treatment with Ashwagandha derived anticancer withanolides: bioinformatics and experimental evidence.

Tumor suppressor p53 protein is frequently mutated in a large majority of cancers. These mutations induce local or global changes in protein structure thereby affecting its binding to DNA. The structural differences between the wild type and mutant p53 thus provide an opportunity to selectively target mutated p53 harboring cancer cells. Restoration of wild type p53 activity in mutants using small molecules that can revert the structural changes have been considered for cancer therapeutics.

We used bioinformatics and molecular docking tools to investigate the structural changes between the wild type and mutant p53 proteins (p53^V143A, p53^R249S, p53^R273H and p53^Y220C) and explored the therapeutic potential of Withaferin A and Withanone for restoration of wild type p53 function in cancer cells. Cancer cells harboring the specific mutant p53 proteins were used for molecular assays to determine the mutant or wild type p53 functions.

We found that p53^V143A mutation does not show any significant structural changes and was also refractory to the binding of withanolides. p53^R249S mutation critically disturbed the H-bond network and destabilized the DNA binding site. However, withanolides did not show any selective binding to either this mutant or other similar variants. p53^Y220C mutation created a cavity near the site of mutation with local loss of hydrophobicity and water network, leading to functionally inactive conformation. Mutated structure could accommodate withanolides suggesting their conformational selectivity to target p53^Y220C mutant. Using human cell lines containing specific p53 mutant proteins, we demonstrated that Withaferin A, Withanone and the extract rich in these withanolides caused restoration of wild type p53 function in mutant p53^Y220C cells. This was associated with induction of p21^WAF-1-mediated growth arrest/apoptosis.

The study suggested that withanolides may serve as highly potent anticancer compounds for treatment of cancers harboring a p53^Y220C mutation.