The vast majority of p53 mutations are missense mutations in the DNA-binding domain (termed ?mutp53?) that generate conformationally aberrant proteins with broadly abrogated functions. Importantly, in the previous grant cycle we generated new mouse models that definitively proved that certain hotspot missense mutant p53 proteins not only lose their tumor suppressor function, but acquire broad oncogenic gain-of-function (GOF) activities (?mutp53GOF?). Our humanized p53R248Q knockin mice (termed ?Q? mice) provided the long-sought compelling phenotype of faster onset of all spontaneously arising tumor types and significantly shorter survival compared to p53null littermates. Importantly, our finding translates to human cancers. In Li-Fraumeni patients harboring p53 germline mutations, the Q allele dramatically accelerates tumor onset by 10.5 years and leads to increased mortality compared to p53null-like Li-Fraumeni patients. Moreover, accumulating evidence from TCGA data suggests that sporadic cancer patients harboring specific GOF alleles have higher death rates than patients with p53 mutations that are functionally null. GOF contributes to malignant progression with increased proliferation, invasion, metastasis, chemoresistance, stroma remodeling and reprogrammed metabolism. A central feature of GOF is that mutp53 proteins exhibit massive constitutive stabilization, and that stabilization is the prerequisite for exerting GOF. We identified the HSP90 chaperone machinery, which protects mutp53 from its E3 ubiquitin ligases, as a major determinant of stabilization in vivo. Globally about 11 million people are living with tumors expressing highly stabilized mutp53. Importantly, our findings indicate that the oncogenic wiring of mutp53 tumors fundamentally differs from p53null tumors, which historically was the premier preclinical model used. Notably, we established that autochthonous mutp53GOF cancers develop a strong dependency on continued expression of high levels of mutp53 for tumor growth, maintenance and metastasis. Consequently, acute genetic (via floxQ) or pharmacologic (via Hsp90 inhibitors) ablation of mutp53 triggers strong tumor cytotoxicity in two distinct GOF mice, translating to major gains in survival by up to 59%, even in the absence of wildtype p53. These paradigm-shifting results identify mutp53 as an actionable cancer-specific drug target.
Aim 1 So far we demonstrated GOF resulting in mutp53 tumor dependency - and its therapeutic exploitability - in the context of lymphoma and colorectal carcinoma. We will evaluate the therapeutic potential of targeting mutp53 in other major tumor types, specifically in epithelial-derived models of liver and pancreatic carcinomas.
Aim 2 will determine the in vivo core network of pathways and interaction partners mediating mutp53GOF. We will use ChIPseq/RNAseq and functional proteome analyses to directly observe the dynamic events that occur upon p53 mutation during transformation in primary mutp53GOF-driven lymphoma.
Aim 3 will use genome-wide transcriptional perturbation via CRISPR-mediated gene repression and activation to identify critical genetic sensitizers as novel therapeutic targets specific for mutp53GOF cancer cells.

Public Health Relevance

In contrast to other tumor suppressors, the vast majority of p53 alterations in human cancers are very discrete mutations that incorporate a single faulty amino acid in the central DNA-binding part of the molecule and generate abnormally stabilized mutant proteins (called ?mutp53?). Compelling evidence from mutp53 knockin mice as well as emerging evidence from human cancers indicate that many of these mutp53 alleles acquire new oncogenic gain-of-function (GOF) which actively contribute to tumor progression. Using genetic and pharmacological removal, we demonstrated that such tumors exhibit a strong dependency on continued expression of mutp53 for tumor growth, maintenance and metastasis. mutp53 depletion induces tumor cell death and leads to major survival gains in preclinical mouse models. Thus, mutp53 represents an exploitable cancer-specific drug target. The main goal of this project is to build on these findings and explore mutp53GOF in a broader cancer context, determine the core molecular mechanisms underlying GOF activity and identify novel druggable targets specific for mutp53 cancers.

National Institute of Health (NIH)
National Cancer Institute (NCI)
Research Project (R01)
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Tumor Cell Biology Study Section (TCB)
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Watson, Joanna M
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State University New York Stony Brook
Schools of Medicine
Stony Brook
United States
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Alexandrova, Evguenia M; Moll, Ute M (2017) Depleting stabilized GOF mutant p53 proteins by inhibiting molecular folding chaperones: a new promise in cancer therapy. Cell Death Differ 24:3-5
Alexandrova, Evguenia M; Xu, Sulan; Moll, Ute M (2017) Ganetespib synergizes with cyclophosphamide to improve survival of mice with autochthonous tumors in a mutant p53-dependent manner. Cell Death Dis 8:e2683
Alexandrova, Evguenia M; Mirza, Safia A; Xu, Sulan et al. (2017) p53 loss-of-heterozygosity is a necessary prerequisite for mutant p53 stabilization and gain-of-function in vivo. Cell Death Dis 8:e2661
Kramer, Daniela; Stark, Nadine; Schulz-Heddergott, Ramona et al. (2017) Strong antitumor synergy between DNA crosslinking and HSP90 inhibition causes massive premitotic DNA fragmentation in ovarian cancer cells. Cell Death Differ 24:300-316
Lebedev, Ivan; Nemajerova, Alice; Foda, Zachariah H et al. (2016) A Novel In Vitro CypD-Mediated p53 Aggregation Assay Suggests a Model for Mitochondrial Permeability Transition by Chaperone Systems. J Mol Biol 428:4154-4167
Alexandrova, E M; Yallowitz, A R; Li, D et al. (2015) Improving survival by exploiting tumour dependence on stabilized mutant p53 for treatment. Nature 523:352-6
Yallowitz, A R; Hanel, W; Moll, U M (2015) Expansion of the hematopoietic stem cell compartment is necessary but not sufficient for gain-of-function mutant p53 R248Q to accelerate lymphomagenesis. Cell Death Differ 22:1397
Alexandrova, E M; Yallowitz, A R; Li, D et al. (2015) Corrigendum: Improving survival by exploiting tumour dependence on stabilized mutant p53 for treatment. Nature 527:398
Schulz, R; Streller, F; Scheel, A H et al. (2014) HER2/ErbB2 activates HSF1 and thereby controls HSP90 clients including MIF in HER2-overexpressing breast cancer. Cell Death Dis 5:e980
Schulz, Ramona; Moll, Ute M (2014) Targeting the heat shock protein 90: a rational way to inhibit macrophage migration inhibitory factor function in cancer. Curr Opin Oncol 26:108-13

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