# Restore the Tumor-Suppressive Activities of p53 Mutants

> **NIH NIH R01** · VIRGINIA COMMONWEALTH UNIVERSITY · 2024 · $341,699

## Abstract

Project Summary/Abstract
This project is focused on reactivation of p53 mutants by peptidase D (PEPD). p53 tumor suppressor is the
most frequently mutated protein in cancer. Most p53 mutations are missense mutations, causing a single
amino acid change in each mutant, and are clustered within its DNA binding domain. p53 mutations nullify
its tumor suppressor functions and/or endow oncogenic functions. PEPD, also known as prolidase, is a
dipeptidase important for collagen metabolism. However, we recently found that PEPD binds to both wild-
type p53 (p53WT) and various mutants via their proline-rich domain (PRD) and that disrupting the binding by
PEPD knockdown (KD) not only activates p53WT but also reactivates its mutants. This is a novel function of
PEPD, which does not require its enzymatic activity. Our long-term goal is to advance the understanding of
regulation and function of p53WT and its mutants. The objective of the present proposal is to delineate the
reactivation of oncogenic hotspot p53 mutants by PEPD KD and to assess the tumor-suppressing activities
of the reactivated p53 mutants. The central hypothesis of the proposal is that PEPD binds to nearly half of
each p53 mutant in cells and that, while PEPD is not required for their oncogenic activities, disrupting PEPD
binding to p53 mutants induces post-translational modifications (PTMs) of the mutants that cause their
refolding and reactivation. We will test the hypothesis in three specific aims: 1) to determine binding of p53
mutants to PEPD and their reactivation by PEPD KD; 2) to determine the molecular mechanism by which
PEPD KD reactivates p53 mutants, focusing on the roles of PTMs, K373 acetylation in particular, in driving
refolding and reactivation of the mutants; 3) to determine the tumor-inhibitory activities of the reactivated
p53 mutants. We will pursue these aims by focusing on some of the most common oncogenic p53 mutants
in cancer, including conformation mutants (R175H, G245C, and R249S) and contact mutants (R248Q,
R273H, and R280K). Cell lines and mouse tumor models, including patient-derived xenografts, will be used.
PEPD KD will be achieved using siRNA and a doxycycline-regulated system. The proposed research is
significant, because it may bring about a paradigm shift in understanding of the biology and regulation of
p53 mutants, which in turn may offer innovative cancer treatment strategies. The expected outcome of this
project includes: 1) showing that PEPD binds to nearly half of each p53 mutant in the nucleus and cytosol;
2) showing that disrupting the PEPD-p53 mutant complex by PEPD KD frees the mutant for PTMs which
drive refolding and reactivation of the mutant, whether it is a conformation mutant or a contact mutant, and
K373 acetylation by p300/CBP is key to this process; and 3) showing that the tumor-suppressive activities
of the reactivated p53 mutants are similar to that of activated p53WT. As such, our research will bring to light
a critical intrinsic reacti...

## Key facts

- **NIH application ID:** 10851881
- **Project number:** 5R01CA282703-02
- **Recipient organization:** VIRGINIA COMMONWEALTH UNIVERSITY
- **Principal Investigator:** YUESHENG ZHANG
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2024
- **Award amount:** $341,699
- **Award type:** 5
- **Project period:** 2023-07-01 → 2028-06-30

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10851881

## Citation

> US National Institutes of Health, RePORTER application 10851881, Restore the Tumor-Suppressive Activities of p53 Mutants (5R01CA282703-02). Retrieved via AI Analytics 2026-05-25 from https://api.ai-analytics.org/grant/nih/10851881. Licensed CC0.

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