Targeted cancer therapy relies on a 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, which is activated in human cancer. Selective targeting one ubiquitin ligase pathway will reduce normal cell toxicity associated with overall inhibition of protein degradation, as seen in Velcade (also known as Bortezomib or PS-341), the first (and only) class of general proteasome inhibitor, approved by FDA for the treatment of relapsed/refractory multiple myeloma and mantle cell lymphoma. To this end, we have focused on SCF (Skp1-Cullin-F-box proteins) E3 ubiquitin ligases, also known as CRLs (Cullin-RING ubiquitin ligases). SCF E3 ligases, the largest E3 ligase family consisting of Skp1, cullins, F-box proteins, and a RING protein, ROC or RBX, promotes the ubiquitination of a subset of key regulatory proteins for targeted degradation, thus governing important biological processes, including cell cycle progression, signal transduction and DNA replication. While cullin-ROC constitutes the core ligase component, cullin needs to be neddylated via Nedd8-Activating Enzyme (NAE) for its enzymatic activity. Our strong preliminary data showed that ROC1 is over-expressed in a number of human cancers, and inactivation of SCF E3 ubiquitin ligase activity via ROC1 siRNA silencing or by an NAE inhibitor, MLN4924, triggers DNA double strand breaks (DSB) and the DNA damage response (DDR), and induces tumor cell killing via autophagy, senescence and apoptosis. Inhibition of SCF E3 ligase also enhances radiation-induced DDR, leading to radiosensitization. The objective of this proposed study is to elucidate the underlying mechanisms by which inactivation of ROC1-SCF E3 ligase triggers these biochemical and biological changes, leading to various types of cell death, and to validate ROC1-SCF E3 ligase as an anticancer and radiosensitizing target. Our central hypothesis is that inactivation of ROC1-SCF E3 ubiquitin ligase causes accumulation of several key substrates, which triggers DSB and DDR and induces cell death via autophagy, senescence and apoptosis in a sequential or parallel order. These triggered cell killing mechanisms could also enhance radiation effects, leading to radiosensitization of cancer cells.
Three specific aims are proposed 1) to determine how inactivation of ROC1-SCF E3 ligase triggers DNA damage and DDR, and induces cell death via different mechanisms;2) to determine how inactivation of ROC1-SCF E3 ligase blocks mTOR to induce autophagy;and 3) to validate ROC1-SCF E3 ligase as a novel radiosensitizing target. IMPACT: This work is highly innovative and of significant impact with translational value by validating SCF E3 ligase as an anticancer target and by paving the ground for future development of MLN4924 or its analogues as a novel class of radiosensitizers.

Public Health Relevance

Proteasome inhibitor, Velcade is used in the clinic for the treatment of relapsed/refractory multiple myeloma. The drug is in general toxic to normal cells due to its overall inhibition of protein degradation. One way to reduce toxic side-effect is to inhibit a particular E3 ligase that is activated in cancer cells. This study will validate SCF E3 ubiquitin ligase as an anticancer and radiosensitizing target and MLN4924, a newly discovered SCF E3 ligase inhibitor, as a tumor-selective and radiosensitizing agent for cancer therapy.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
5R01CA156744-04
Application #
8625717
Study Section
Basic Mechanisms of Cancer Therapeutics Study Section (BMCT)
Program Officer
Bernhard, Eric J
Project Start
2011-05-11
Project End
2016-03-31
Budget Start
2014-04-01
Budget End
2015-03-31
Support Year
4
Fiscal Year
2014
Total Cost
$312,983
Indirect Cost
$111,708
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
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Li, Hua; Zhou, Weihua; Li, Lihui et al. (2017) Inhibition of Neddylation Modification Sensitizes Pancreatic Cancer Cells to Gemcitabine. Neoplasia 19:509-518
Zhou, Haibin; Lu, Jianfeng; Liu, Liu et al. (2017) A potent small-molecule inhibitor of the DCN1-UBC12 interaction that selectively blocks cullin 3 neddylation. Nat Commun 8:1150
Xu, Jie; Zhou, Weihua; Yang, Fei et al. (2017) The ?-TrCP-FBXW2-SKP2 axis regulates lung cancer cell growth with FBXW2 acting as a tumour suppressor. Nat Commun 8:14002
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Zhang, Qiang; Karnak, David; Tan, Mingjia et al. (2016) FBXW7 Facilitates Nonhomologous End-Joining via K63-Linked Polyubiquitylation of XRCC4. Mol Cell 61:419-433
Lan, Huiyin; Tang, Zaiming; Jin, Hongchuan et al. (2016) Neddylation inhibitor MLN4924 suppresses growth and migration of human gastric cancer cells. Sci Rep 6:24218
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
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
Xie, Chuan-Ming; Wei, Dongping; Zhao, Lili et al. (2015) Erbin is a novel substrate of the Sag-?TrCP E3 ligase that regulates KrasG12D-induced skin tumorigenesis. J Cell Biol 209:721-37

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