Targeted cancer therapy relies on thorough validation of cancer targets. Our long-range goal is to discover a novel class of anticancer drugs that selectively target one type of E3 ubiquitin ligase, shown to be activated in human cancers. To this end, we have focused on SAG (Sensitive to Apoptosis Gene), also known as RBX2/ROC2, a RING component of SCF (Skp1-Cullin-F-box proteins) E3 ubiquitin ligases, required for its activity. Our strong published results and unpublished preliminary data, using mouse transgenic and knockout models and cell-based mechanistic studies, demonstrated that 1) SAG positively regulates angiogenesis: angiogenesis is enhanced by SAG transgenic expression in skin tumors, and inhibited by Sag knockout during embryonic development, in teratomas derived from ES cells, and in B16 melanoma tumorigenesis;2) SAG E3 promotes the degradation of a) neurofibromin 1 (NF1) to activate the RAS/MAPK, b) IkB to activate NFkB, c) DEPTOR to activate mTOR, leading to enhanced proliferation and survival;3) SAG inactivation by siRNA knockdown or by small molecule SAG E3 inhibitor MLN4924 selectively suppresses the growth of human squamous cell carcinoma (SCC) cells;4) Sag KO inhibits keratinocyte differentiation, and finally 5) SAG is overexpressed in all three developmental stages of human skin SCC (namely, actinic keratosis, SCC in situ, and invasive SCC). However, whether Sag plays an essential role in skin carcinogenesis induced by viral (HPV16 to inactivate p53 and RB), chemical (DMBA-TPA to activate Ras-AP1), or physical (UV) carcinogens, thus serving as a valid target for anti-skin cancer therapy, has not been systematically examined. The objectives of this application are to use tissue specific Sag knockout mouse models under physiological settings to investigate mechanistically the role of Sag in skin carcinogenesis triggered by various carcinogens. The central hypothesis is that SAG promotes angiogenesis, proliferation and tumorigenesis via targeted degradation of tumor suppressive substrates such as NF1, I?B and DEPTOR, leading to activation of the RAS, NF?B and mTOR pathways, respectively. Inactivation of SAG by genetic deletion or pharmaceutical inhibitor MLN4924 would cause the accumulation of these substrates to inactivate the RAS, NF?B, mTOR pathways leading to suppression of skin carcinogenesis.
Three specific aims are proposed to test our hypothesis by determining the effect of Sag deletion on 1) skin angiogenesis and carcinogenesis, induced by viral oncogenes;2) skin carcinogenesis, induced by DMBA/TPA or UV;and 3) mechanisms of Sag action. IMPACT: Our work uses skin-specific KO mouse models that recapitulate the development of human skin SCC to elucidate mechanistically that SAG E3 ligase is essential for skin carcinogenesis, thus serving as an attractive drug target for skin cancer. Our work is highly innovative and of significant impact with translational value by providing proof-of-concept evidence for future development of MLN4924 as a novel class of anti-cancer agent for the prevention and treatment of skin cancer.

Public Health Relevance

Targeted cancer therapy relies on thorough validation of cancer targets. We found that SAG E3 ubiquitin ligase is overexpressed in skin cancer and SAG inactivation selectively suppresses the growth of skin squamous cell carcinoma cells. In this study, we will mechanistically elucidate the role of Sag in skin carcinogenesis triggered by viral, chemical and physical carcinogens, using tissue-specific mouse KO models. We will also test the activity of MLN4924, a small molecule inhibitor of SAG E3 ligase against skin cancer. Thus, this study is of highly translational value by providing proof-of-concept evidence for future development of SAG inhibitor MLN4924 as a novel class of anti-cancer agent for the prevention and treatment of skin cancer.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
5R01CA171277-02
Application #
8602514
Study Section
Cancer Etiology Study Section (CE)
Program Officer
Johnson, Ronald L
Project Start
2013-01-03
Project End
2017-12-31
Budget Start
2014-01-01
Budget End
2014-12-31
Support Year
2
Fiscal Year
2014
Total Cost
$290,397
Indirect Cost
$103,647
Name
University of Michigan Ann Arbor
Department
Radiation-Diagnostic/Oncology
Type
Schools of Medicine
DUNS #
073133571
City
Ann Arbor
State
MI
Country
United States
Zip Code
48109
Mathewson, Nathan D; Fujiwara, Hideaki; Wu, Shin-Rong et al. (2016) SAG/Rbx2-Dependent Neddylation Regulates T-Cell Responses. Am J Pathol 186:2679-91
Tan, Mingjia; Xu, Jie; Siddiqui, Javed et al. (2016) Depletion of SAG/RBX2 E3 ubiquitin ligase suppresses prostate tumorigenesis via inactivation of the PI3K/AKT/mTOR axis. Mol Cancer 15:81
Zhou, Xiaochen; Tan, Mingjia; Nyati, Mukesh K et al. (2016) Blockage of neddylation modification stimulates tumor sphere formation in vitro and stem cell differentiation and wound healing in vivo. Proc Natl Acad Sci U S A 113:E2935-44
Zhou, Weihua; Xu, Jie; Li, Haomin et al. (2016) Neddylation E2 UBE2F promotes the survival of lung cancer cells by activating CRL5 to degrade NOXA via the K11 linkage. Clin Cancer Res :
Zhang, Qiang; Zhang, Yaqing; Parsels, Joshua D et al. (2016) Fbxw7 Deletion Accelerates Kras(G12D)-Driven Pancreatic Tumorigenesis via Yap Accumulation. Neoplasia 18:666-673
Zhang, Qiang; Karnak, David; Tan, Mingjia et al. (2016) FBXW7 Facilitates Nonhomologous End-Joining via K63-Linked Polyubiquitylation of XRCC4. Mol Cell 61:419-33
Zhou, Xiaochen; Wang, Gongxian; Sun, Yi (2015) A reliable parameter to standardize the scoring of stem cell spheres. PLoS One 10:e0127348
Wei, Dongping; Zhang, Qiang; Schreiber, Jason S et al. (2015) Targeting mcl-1 for radiosensitization of pancreatic cancers. Transl Oncol 8:47-54
Xie, Chuan-Ming; Wei, Dongping; Zhao, Lili et al. (2015) Erbin is a novel substrate of the Sag-βTrCP E3 ligase that regulates KrasG12D-induced skin tumorigenesis. J Cell Biol 209:721-37
Wang, Lizhong; Liu, Runhua; Ye, Peiying et al. (2015) Intracellular CD24 disrupts the ARF-NPM interaction and enables mutational and viral oncogene-mediated p53 inactivation. Nat Commun 6:5909

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