SAG (Sensitive to Apoptosis Gene), also known as RBX2/ROC2, is a stress-responsive RING component of SCF (Skp1, Cullins, F-box proteins) E3 ubiquitin ligase, required for its activity. Our strong preliminary data as well as published results showed that 1) Sag is a gene essential for growth during mouse embryonic development, and Sag-null MEFs or ES cells are growth retarded when cultured in vitro or grown in nude mice, respectively;2) SAG transgenic expression accelerates the growth of DMBA/TPA-induced skin papilloma by inhibiting apoptosis;3) SAG is overexpressed in multiple human cancers, particularly lung cancer, and 4) SAG siRNA silencing suppresses cancer cell growth via inducing apoptosis. However, whether Sag is required for immortalization, transformation, or carcinogenesis, thus serving as a valid cancer target for chemoprevention and therapy, has not been examined. The objectives of this application are to use tissue specific Sag knockout mouse models at physiological settings to study the role of Sag in immortalization and transformation of primary cultures and in carcinogenesis of lung and skin tissues. The central hypothesis is that Sag, upon overexpression, promotes cell growth and inhibits apoptosis via promoting the degradation of tumor suppressive and apoptosis-inducing substrates such as IkB, Nf1, p27, and Noxa, leading to activation of the NFkB and Ras pathways. On the other hand, Sag, upon inactivation by genetic deletion or treatment with small molecule inhibitor MLN4924, causes an accumulation of these substrates to inactivate the NFkB and Ras pathways, resulting in suppression of proliferation and carcinogenesis.
Three specific aims are proposed to elucidate the role of Sag in 1) immortalization and transformation of mouse embryonic fibroblasts by E1A/Hras;2) lung carcinogenesis induced by KrasG12D, and 3) skin carcinogenesis induced by DMBA/TPA. IMPACT: our work is highly innovative and of significant impact with translational value by validating SAG E3 ubiquitin ligase as an attractive target for cancer chemoprevention and therapy, thus providing some proof-of-concept evidence for future development of MLN4924, a newly discovered small molecule inhibitor of SCF E3 ligase, as a novel class of chemoprevention and anticancer agents.

Public Health Relevance

Targeted cancer therapy relies on thorough validation of cancer targets. We found that Sag E3 ubiquitin ligase is overexpressed and activated in human cancer, particularly lung cancer. In this study, we will mechanistically elucidate the role of Sag in neoplastic transformation and in carcinogenesis of lung (induced by Kras) and skin (induced by chemical carcinogens). We will test MLN4924, a small molecule inhibitor of Sag E3 ligase against lung cancer at the early and later stages. Thus, this study is of highly translational value by validating Sag E3 ubiquitin ligase and its inhibitor MLN4924 as an attractive chemoprevention and anticancer target and drug, respectively.

National Institute of Health (NIH)
Research Project (R01)
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Cancer Etiology Study Section (CE)
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Hildesheim, Jeffrey
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University of Michigan Ann Arbor
Schools of Medicine
Ann Arbor
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Zhou, Weihua; Xu, Jie; Zhao, Yongchao et al. (2014) SAG/RBX2 is a novel substrate of NEDD4-1 E3 ubiquitin ligase and mediates NEDD4-1 induced chemosensitization. Oncotarget 5:6746-55
Wan, Lixin; Tan, Mingjia; Yang, Jie et al. (2014) APC(Cdc20) suppresses apoptosis through targeting Bim for ubiquitination and destruction. Dev Cell 29:377-91
Xiong, Xiufang; Zhao, Yongchao; Tang, Fei et al. (2014) Ribosomal protein S27-like is a physiological regulator of p53 that suppresses genomic instability and tumorigenesis. Elife 3:e02236
Li, Hua; Tan, Mingjia; Jia, Lijun et al. (2014) Inactivation of SAG/RBX2 E3 ubiquitin ligase suppresses KrasG12D-driven lung tumorigenesis. J Clin Invest 124:835-46
Sun, Yi; Li, Hua (2013) Functional characterization of SAG/RBX2/ROC2/RNF7, an antioxidant protein and an E3 ubiquitin ligase. Protein Cell 4:103-16
Xie, Chuan-Ming; Wei, Wenyi; Sun, Yi (2013) Role of SKP1-CUL1-F-box-protein (SCF) E3 ubiquitin ligases in skin cancer. J Genet Genomics 40:97-106
Zhou, Weihua; Wei, Wenyi; Sun, Yi (2013) Genetically engineered mouse models for functional studies of SKP1-CUL1-F-box-protein (SCF) E3 ubiquitin ligases. Cell Res 23:599-619
Zhao, Yongchao; Sun, Yi (2013) Cullin-RING Ligases as attractive anti-cancer targets. Curr Pharm Des 19:3215-25
Yang, Dong; Zhao, Yongchao; Li, Amy Y et al. (2012) Smac-mimetic compound SM-164 induces radiosensitization in breast cancer cells through activation of caspases and induction of apoptosis. Breast Cancer Res Treat 133:189-99
Yang, Dong; Tan, Mingjia; Wang, Gongxian et al. (2012) The p21-dependent radiosensitization of human breast cancer cells by MLN4924, an investigational inhibitor of NEDD8 activating enzyme. PLoS One 7:e34079

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