Role of SAG/RBX2 E3 Ubiquitin Ligase in Skin Carcinogenesis

SAG/RBX2 E3 泛素连接酶在皮肤癌发生中的作用

• 批准号: 8602514
• 负责人: YI SUN
• 金额: $ 31.3万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-01-03 至 2017-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8602514
• 关键词: Actinic keratosis; Adenoviruses; Alleles; Antineoplastic Agents; Apoptosis; Apoptosis Inhibitor Gene; Benz(a)Anthracenes; Biotin; Carcinogens; Carcinoma in Situ; Cell model; Chemicals; Chemoprevention; Complement Factor B; Cullin Proteins; Cyclin-Dependent Kinases; Cycloheximide; DNA Nucleotidylexotransferase; Data; Development; Drug Targeting; Embryo; Embryonic Development; Enzymes; Fibroblasts; Fluorescence-Activated Cell Sorting; Future; Gene Silencing; Genes; Genetic; Goals; Growth; Head and Neck Squamous Cell Carcinoma; Histocytochemistry; Human; Human Development; Human Papillomavirus; Human papillomavirus 16; Immunoblotting; Immunoprecipitation; In Situ Nick-End Labeling; In Vitro; Keratin; Knock-out; Knockout Mice; Label; LoxP-flanked allele; MAP Kinase Gene; Malignant Epithelial Cell; Malignant Neoplasms; Modeling; Mus; Neurofibromatosis Type 1 Protein; Nuclear; Pathway interactions; Pharmacologic Substance; Physical Carcinogens; Physiological; Play; Prevention; Proteins; Proteomics; Publishing; RBX1 gene; Reactive Oxygen Species; Role; Skin; Skin Cancer; Skin Carcinogenesis; Skin Neoplasms; Skp1-Cullin-F-Box Proteins; Small Interfering RNA; Squamous cell carcinoma; Staging; Teratoma; Testing; Tetradecanoylphorbol Acetate; Tissues; Transcription Factor AP-1; Transgenic Mice; Transgenic Model; Transgenic Organisms; Tumor Angiogenesis; UV induced; Validation; Viral; Viral Oncogene; Work; angiogenesis; antiangiogenesis therapy; base; benzanthracene; cancer therapy; cancer viral etiology; carcinogenesis; chemical carcinogen; dimethylbenzanthracene; embryonic stem cell; human FRAP1 protein; human RBX1 protein; in vivo; inhibitor/antagonist; innovation; interest; keratin 5; keratinocyte differentiation; melanoma; mouse model; novel; overexpression; public health relevance; recombinase; skin squamous cell carcinoma; small molecule; tumor; tumorigenesis; ubiquitin ligase; ubiquitin-protein ligase; ultraviolet irradiation

DESCRIPTION (provided by applicant): 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.

描述（由申请人提供）：有针对性的癌症治疗依赖于对癌症靶标的彻底验证。我们的远程目标是发现一种新型的抗癌药物，该药物有选择地靶向一种类型的E3泛素连接酶，该类型在人类癌症中被激活。为此，我们专注于SAG（对凋亡基因敏感），也称为RBX2/ROC2，这是其活性所需的SCF（SKP1-Cullin-F-box蛋白）E3泛素连接酶的环。我们使用小鼠转基因和基因敲除模型以及基于细胞的机械研究的强烈发表的结果和未发表的初步数据表明，1）SAG阳性地调节血管生成：血管生成通过皮肤肿瘤中的SAG转基因表达增强了血管生成，并在胚胎中脱离eSAG敲除，在胚胎中，在Teratomas中被SAG敲除抑制，从Es eSTORIG中抑制了eS.66 Melan neran byan Melan nuran tuman tuman tuman tuman numan numan numan tuman nuran tuman tuman inturoman inturomas。 2）SAG E3促进了A）神经纤维蛋白1（NF1）激活Ras/Mapk的降解，b）IKB激活NFKB，C）激活MTOR的deptor，从而导致增强和存活增强； 3）通过siRNA敲低或小分子SAG E3抑制剂MLN4924的下垂灭活，选择性地抑制了人类鳞状细胞癌（SCC）细胞的生长； 4）SAG KO抑制角质形成细胞的分化，最后5）在人类皮肤SCC的所有三个发育阶段中均过表达SAG（即，光化性角化病，SCC，原位和侵入性SCC）。但是，SAG是否在病毒（HPV16失活p53和RB）诱导的皮肤癌发生中起着至关重要的作用，化学（DMBA-TPA激活RAS-AP1）还是物理（UV）致癌物，因此是对抗皮肤癌症的有效靶标，因此是抗皮肤癌症的有效靶标。该应用的目标是在生理环境下使用组织特定的SAG敲除小鼠模型来机械地研究SAG在各种致癌物触发的皮肤致癌中的作用。中心假设是SAG通过靶向抑制性抑制性底物（如NF1，I？b和Deptor）的靶向降解促进血管生成，增殖和肿瘤发生，从而分别激活RAS，NF？B和MTOR途径。通过遗传缺失或药物抑制剂MLN4924对SAG的失活会导致这些底物的积累使RAS失活NF？B，MTOR途径，导致抑制皮肤致癌。提出了三个特定的目的来检验我们的假设，通过确定SAG缺失对1）由病毒癌基因引起的皮肤血管生成和致癌作用的影响； 2）皮肤致癌作用，由DMBA/TPA或紫外线诱导； 3）SAG作用的机制。影响：我们的工作使用皮肤特异性的KO小鼠模型，这些模型概括了人类皮肤SCC的发展，以机械地阐明SAG E3连接酶对于皮肤癌变至关重要，因此是皮肤癌的有吸引力的药物靶标。我们的工作具有很高的创新性，具有重大影响，具有转化价值，通过为MLN4924的未来开发提供概念验证证据，作为预防和治疗皮肤癌的一种新型抗癌剂。