# Cellular activity upregulation of the thermolabile p53 cancer mutant Y220C by small molecule indazole derivatives

## TL;DR

This study identifies small molecule indazole derivatives that reactivate a cancer-causing p53 mutant, potentially offering a new approach to cancer therapy.

## Contribution

The study introduces novel indazole derivatives that selectively reactivate the p53-Y220C mutant in cancer cells.

## Key findings

- JC16 and JC36 show selective cytotoxicity and pro-apoptotic activity in p53-Y220C mutant cancer cells.
- The compounds induce a conformational shift in p53-Y220C, activating p53 target genes like BBC3 and MDM2.
- The reactivation occurs without increasing total p53 protein levels, suggesting a conformation-based mechanism.

## Abstract

The TP53 gene is one of the most frequently mutated genes in human cancers. Mutations often result in loss of tumor-suppressive functions and acquisition of oncogenic properties by p53, contributing to tumor progression and resistance to therapy. Among structural p53 mutations, Y220C is particularly notable for creating a surface-exposed hydrophobic pocket that destabilizes the protein while preserving partial function, making it a promising target for pharmacological reactivation. In this study, we performed a structure-guided phenotypic screen of an in-house heterocyclic compound library to identify novel small-molecule modulators of p53-Y220C. This led to the identification of a series of (1H-pyrrol-1-yl)indazole derivatives (JC16, JC36, JC65), structurally inspired by known Y220C binders. JC16 and JC36 exhibited selective cytotoxicity and pro-apoptotic activity in p53-Y220C mutant cancer cell lines, with minimal effects in wild-type or p53-null cells. These compounds induced a mutant-to-wild-type conformational shift in cellular p53-Y220C, accompanied by transcriptional activation of canonical p53 target genes, including BBC3 (PUMA) and MDM2. Western blot analysis revealed that in HUH7 cells, these effects occurred without a corresponding increase in total p53 protein levels, suggesting a mechanism based on conformational reactivation. Our findings position JC16 and JC36 as early-stage chemical leads with potential to restore mutant p53 function in a context-dependent manner. While their exact mechanism of action remains to be fully elucidated, these results provide a foundation for further development of indazole-based scaffolds as reactivators of the p53-Y220C mutant in cancer therapy.