Evaluation of mTOR as a chemoprevention target in skin cancer

mTOR 作为皮肤癌化学预防靶点的评估

• 批准号: 7475350
• 负责人: LISA M SHANTZ
• 金额: $ 7.68万
• 依托单位: PENNSYLVANIA STATE UNIV HERSHEY MED CTR
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-04-01 至 2010-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7475350
• 关键词: 1-Phosphatidylinositol 3-Kinase; 5' Untranslated Regions; Ablation; Affect; Alleles; Apoptosis; Basal Cell; Biogenesis; Biological; CCI-779; Calcineurin inhibitor; Catalytic Domain; Cell physiology; Cells; Chemoprevention; Chemopreventive Agent; Chimeric Proteins; Clinical Trials; Complex; Condition; Cyclin D1; Cytoskeletal Modeling; Development; Dimethyl Sulfoxide; Early Intervention; Epidermis; Epigenetic Process; Estrogen Receptors; Evaluation; Exons; Formalin; Future; Generations; Genes; Genetic; Hair follicle structure; Human; Immune; Incidence; Individual; Kidney Transplantation; Knock-out; Lead; Link; Macrolide Antibiotics; Malignant Neoplasms; Medicine; Messenger RNA; Mitogens; Mus; Mutate; Mutation; Numbers; Nutrient; Ornithine Decarboxylase; Outcome; Paraffin Embedding; Pathway interactions; Patients; Phosphorylation; Phosphotransferases; Pilot Projects; Plant Roots; Process; Protein Biosynthesis; Protein Kinase; Proteins; Protocols documentation; RNA; Ribosomes; Risk; Role; Sampling; Sirolimus; Site; Skin; Skin Cancer; Skin Carcinogenesis; Skin Carcinoma; Skin Neoplasms; Staging; Stem cells; Structure; System; Tamoxifen; Testing; Thinking; Time; Transgenic Organisms; Translations; Transplant Recipients; Tumor Promoters; Tumor Promotion; Untranslated Regions; Week; Work; analog; c-myc Genes; carcinogenesis; cell growth; college; design; epidermis cell; human FRAP1 protein; inhibitor/antagonist; keratin 14, K14; keratinocyte; mouse model; promoter; protective effect; protein expression; recombinase; research study; response; size; tumor; tumor initiation; tumorigenesis

DESCRIPTION (provided by applicant): Carcinogenesis is driven by tumor initiation, promotion and progression. There are a variety of epigenetic changes that occur with tumor promotion that are potential targets for early intervention by chemopreventive agents in high-risk individuals. The mammalian target of rapamycin (mTOR), which is downstream of both Rasand PI 3-kinase-controlled pathways, is activated by a large number of cancer-promoting mutations. The mTOR complexes mTORC1 and mTORC2 control diverse cellular processes, including ribosome biogenesis, cytoskeletal organization and protein synthesis, making mTOR a central regulator of cell growth. The naturally occurring macrolide antibiotic rapamycin and its analogues are highly specific inhibitors of mTORC1, and several rapamycin analogues are undergoing clinical trials against a variety of malignancies. While mTOR inhibition offers much promise as a chemotherapeutic strategy, the role of mTOR in chemoprevention has remained largely unexplored. Recent clinical trials establishing that renal transplant patients administered rapamycin as an immune suppressant suffer from significantly fewer non-melanoma skin cancers compared to patients taking calcineurin inhibitors have suggested mTOR as a valid target for chemoprevention of skin carcinogenesis. We present preliminary evidence that the reduction of mTOR in the skin of mTOR mice decreases 4EBP1 phosphorylation and blunts the induction of ornithine decarboxylase (ODC) in response to the tumor promoter TPA. Like many proliferation-associated genes, the ODC mRNA contains a long 5'- untranslated region that is regulated in a cap-dependent manner. Thus, changes in translation brought about by mTOR inhibition can dramatically affect total protein. The proposed experiments will knock out mTOR in the skin by conditional deletion using a CreLoxP approach. Mice containing a floxed mTOR allele, provided by our collaborator Dr. C.J. Lynch, will be crossed with commercially available K14CreERT mice, which express a tamoxifen-inducible Cre recombinase fused to the estrogen receptor, to generate K14CreERT/mTORlox/lox mice. Application of tamoxifen to the skin triggers expression of Cre driven by the keratin 14 promoter, which will ablate mTOR in the outer root sheath of the hair follicle and the basal cell layer of the interfollicular epidermis. Deletion of mTOR will inhibit both mTORC1- and mTORC2-dependent pathways, unlike rapamycin, which is thought to inhibit mTORC1 but activate mTORC2 in most systems. We will use the two-stage mouse model of DMBA/TPA-induced skin tumorigenesis in two Specific Aims designed to establish that mTOR is critical for the early response of proliferation-associated genes in tumor promotion, and mTOR ablation reduces skin tumor incidence and multiplicity. Aim 1 will generate and characterize K14CreERT/mTORlox/lox mice, while Aim 2 will test the effect of conditional mTOR ablation on protein expression and skin tumor development in DMBA/TPA-treated mice. These studies will validate mTOR as a target for chemoprevention, and lead to future work identifying the genetic and epigenetic changes necessary for tumor promotion that are controlled by mTOR.

描述（由申请人提供）： 致癌作用是由肿瘤起始、促进和进展驱动的。有多种表观遗传学的变化，发生与肿瘤的促进，是潜在的目标，早期干预的化学预防剂在高风险的个人。哺乳动物雷帕霉素靶蛋白（mTOR）位于Ras和PI 3-激酶控制通路的下游，可被大量促癌突变激活。mTOR复合物mTORC 1和mTORC 2控制不同的细胞过程，包括核糖体生物发生、细胞骨架组织和蛋白质合成，使mTOR成为细胞生长的中心调节因子。天然存在的大环内酯类抗生素雷帕霉素及其类似物是mTORC 1的高度特异性抑制剂，几种雷帕霉素类似物正在进行针对各种恶性肿瘤的临床试验。虽然mTOR抑制作为化疗策略提供了很多希望，但mTOR在化学预防中的作用在很大程度上仍未被探索。最近的临床试验表明，与服用钙调磷酸酶抑制剂的患者相比，给予雷帕霉素作为免疫抑制剂的肾移植患者患非黑色素瘤皮肤癌的人数显著减少，这表明mTOR是皮肤癌发生化学预防的有效靶点。我们目前的初步证据表明，mTOR小鼠皮肤中mTOR的减少降低了4 EBP 1磷酸化，并减弱了响应于肿瘤促进剂TPA的鸟氨酸脱羧酶（ODC）的诱导。与许多增殖相关基因一样，ODC mRNA含有一个长的5 '非翻译区，其以帽依赖性方式调节。因此，由mTOR抑制引起的翻译变化可以显著影响总蛋白。所提出的实验将通过使用CreLoxP方法的条件删除来敲除皮肤中的mTOR。由我们的合作者Dr. C. J. Lynch提供的含有floxed mTOR等位基因的小鼠将与市售的K14 CreERT小鼠杂交，所述K14 CreERT小鼠表达与雌激素受体融合的他莫昔芬诱导型Cre重组酶，以产生K14 CreERT/mTORlox/lox小鼠。将他莫昔芬应用于皮肤触发由角蛋白14启动子驱动的Cre的表达，这将消融毛囊的外根鞘和毛囊间表皮的基底细胞层中的mTOR。mTOR的缺失将抑制mTORC 1和mTORC 2依赖性途径，不像雷帕霉素，雷帕霉素被认为在大多数系统中抑制mTORC 1但激活mTORC 2。我们将在两个特定目标中使用DMBA/TPA诱导的皮肤肿瘤发生的两阶段小鼠模型，旨在确定mTOR对于肿瘤促进中增殖相关基因的早期反应至关重要，而mTOR消融可降低皮肤肿瘤的发生率和多样性。目标1将产生和表征K14 CreERT/mTORlox/lox小鼠，而目标2将测试条件性mTOR消融对DMBA/TPA处理的小鼠中蛋白质表达和皮肤肿瘤发展的影响。这些研究将验证mTOR作为化学预防的靶点，并导致未来的工作，确定由mTOR控制的肿瘤促进所必需的遗传和表观遗传变化。