The vast majority of p53 mutations are missense mutations in the DNA-binding domain (mutp53) that generate conformationally aberrant proteins with broadly abrogated functions. Importantly, missense mutp53 proteins not only lose their tumor suppressor function, but often acquire oncogenic gain-of-function (GOF) to drive tumorigenesis. GOF contributes to malignant progression with higher proliferation, invasion, metastatic ability and chemoresistance. A central feature of GOF is that nearly all mutp53 proteins exhibit massive constitutive stabilization, and that stabilization is the prerequisite for exerting GOF. Currently, ~ 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 p53-null tumors, historically the premier preclinical model used. We find that mutp53 cancers have developed a strong dependency on high levels of mutp53 for survival. Thus, its acute withdrawal triggers strong spontaneous cytotoxicity in xenografts. We identified that the heat shock protein 90 chaperone (HSP90) is a major determinant of mutp53 stabilization. This identifies acute depletion of mutp53 as a very promising clinical strategy, and a potentially rather rapidly attainable goal in p53-based therapy. HSP90 is highly upregulated specifically in cancers but not in normal tissues. Importantly, the HSP90 machinery is a crucial facilitator of cancer cell survival by supporting proper folding of conformationally aberrant oncogenes including mutp53. Mechanistically, HSP90 protects mutp53 via stable complex ('caging') from its E3 ligases. Thus, goals of this project are to determine whether established tumors require continued expression of stabilized mutp53 for their maintenance in vivo, and to explore whether destabilizing mutp53 - by attacking the chaperone support on which they depend - is a promising new mutp53-specific anticancer strategy. This is an immensely important unsolved problem that is understudied and urgently needs more attention, with a high translational potential to achieve true clinical impact.
Aim 1 determines if continued expression of high levels of mutant p53 is essential for maintenance of established tumors in vivo. We will perform genetic proof-of-principle experiments using acute ablation in mice. To this end, we generated a novel humanized inactivatable mutp53 KI model and will test it in a spontaneous and induced cancer context in vivo.
Aim 2 asks whether small molecule inhibitors of the HSP90-HDAC6 chaperone have a preferential therapeutic response in mutp53 cancer cells in vivo. This will be extensively tested in xenografts and two different hotspot mutp53 KI mouse models. We will also test if mutp53 is a predictive biomarker for response to HSP90i-based therapy in three retrospective/ prospective clinical trials of lung and breast cancer patients. Also, causality studies on mechanism of drug action will be performed.
Aim 3 is based on a novel unsuspected link between aberrant mutp53 stabilization and the tumor-associated HSP27 chaperone system that we uncovered. We will test if the HSP27 chaperone is a mechanistically independent second determinant of mutp53 stabilization.

Public Health Relevance

Compelling new evidence from mutant p53 knockin mice, modeling human tumors more faithfully than p53-/- mice, strongly argue that we need a mutant p53-specific therapeutic strategy for the millions of patients whose tumors harbor a p53 missense mutation (mutp53) in the DNA binding domain. Many of these mutp53 proteins acquire gain-of-function (GOF) which actively contributes to malignant progression. A central feature of GOF is that nearly all mutp53 proteins exhibit massive constitutive stabilization, which is a prerequisite for exerting GOF. The main goal of this project is to determine whether established tumors require continued expression of stabilized mutant p53 for their maintenance in vivo, and to explore whether destabilizing mutant p53 - by attacking the chaperone support on which they depend - is a promising novel mutant p53-specific anticancer strategy for such patients.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
5R01CA176647-02
Application #
8640903
Study Section
Tumor Cell Biology Study Section (TCB)
Program Officer
Watson, Joanna M
Project Start
2013-04-01
Project End
2018-03-31
Budget Start
2014-04-01
Budget End
2015-03-31
Support Year
2
Fiscal Year
2014
Total Cost
$345,545
Indirect Cost
$126,672
Name
State University New York Stony Brook
Department
Pathology
Type
Schools of Medicine
DUNS #
804878247
City
Stony Brook
State
NY
Country
United States
Zip Code
11794
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
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
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
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, 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
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

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