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Cyclin C-CDK8/19 kinases in development and in cancer

发育和癌症中的细胞周期蛋白 C-CDK8/19 激酶

基本信息

批准号：
10579308
负责人：
Peter Sicinski
金额：
$ 51.55万
依托单位：
DANA-FARBER CANCER INST
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-03-01 至 2027-02-28
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10579308
关键词：
Ablation Address Affect Alanine Allografting Alpha Interleukin 2 Receptor Amino Acids Animals Antitumor Response Binding Biochemical CD8-Positive T-Lymphocytes CD8B1 gene CLN1 gene CLN2 gene CLN3 gene Cancer Model Cancer Patient Catalytic Domain Cell Culture Techniques Cell Lineage Cells Clinical Clinical Trials Combined Modality Therapy Complementary DNA Complex Cultured Cells Cyclin-Dependent Kinases Cyclins Cytotoxic T-Lymphocytes Development Endothelial Cells Enterobacteria phage P1 Cre recombinase Gatekeeping Genetic Transcription Growth Half-Life Human IL2RA gene IL2RB gene Immune Immune response Immune system Immunocompromised Host In Vitro Interleukin 2 Receptor Interleukin-2 Interleukin-4 Knockout Mice Laboratories Link LoxP-flanked allele Lymphoid Cell Malignant Neoplasms Maps Mediating Mediator Memory Modality Molecular Mus Mutation NKTR gene Oncogenes Outcome Patients Peripheral Phosphorylation Phosphotransferases Play Polyubiquitination Protein Complex Subunit Proteins RNA Polymerase II Regulatory T-Lymphocyte Research Role Saccharomyces cerevisiae Signal Transduction Site T cell differentiation T-Cell Development T-Lymphocyte T-Lymphocyte Subsets Testing Thymus Gland Transcript Tumor Immunity Tumor Promotion Up-Regulation Wild Type Mouse Work Xenograft procedure Yeasts ZNF145 gene adaptive immune response adaptive immunity anti-CTLA4 antibodies anti-PD-1 anti-cancer anti-tumor immune response antitumor effect cancer cell cancer therapy cyclin C cyclin G1 effector T cell experimental study genetic approach immune checkpoint blockade in vivo inhibitor mouse genetics novel overexpression paralogous gene peripheral lymphoid organ small molecule small molecule inhibitor systemic toxicity therapeutic target thymocyte transcription factor tumor tumorigenesis ubiquitin-protein ligase

项目摘要

Project Summary/Abstract Cyclin C was cloned, along with other G1 cyclins, as a cDNA which can rescue the proliferative block in yeast Saccharomyces cerevisiae lacking CLN1, CLN2 and CLN3 genes. Subsequent work revealed that cyclin C, acting together with its kinase partners, the cyclin-dependent kinase 8 (CDK8) and CDK19, plays a role in regulating gene transcription. Cyclin C serves as a regulatory partner of CDK8 and CDK19, and activates the kinase activity of these proteins. The two paralog kinases show substantial overall amino acid identity, particularly within their catalytic domains. Cyclin C, CDK8 and CDK19 represent parts of the mediator complex, a large multisubunit protein complex that regulates gene transcription by linking RNA polymerase II to sequence-specific transcription factors. In addition, cyclin C-CDK8/19 was shown to phosphorylate various transcription factors and to regulate their stability and activity. Growing evidence indicates that cyclin C-CDK8 and C-CDK19 kinases may represent potential anti-cancer targets. CDK8 has been identified as an oncogene in several human cancers. Cyclin C, CDK8 and CDK19 are overexpressed in a wide range of tumor types. Importantly, higher expression of these three proteins was found to be associated with poor clinical outcome. Consistent with tumor-promoting roles for cyclin C-CDK8/19, treatment of mice bearing xenografts of several human tumor types with small molecule CDK8/19 inhibitors resulted in a potent anti-tumor effect without apparent systemic toxicity. To understand the molecular function of cyclin C in vivo, we generated conditional cyclin C knockout mice (cyclin CF/F). To test the impact of cyclin C-CDK8/19 inhibition on T cell development and adaptive anti-tumor immunity, we crossed cyclin CF/F mice with CD4-Cre animals. The latter strain expresses Cre recombinase at an early stage of T cell development, and drives deletion of the ‘floxed’ sequences in all T cell subsets. In our preliminary analyses we established that cyclin C functions as a gate- keeper of T cell differentiation, likely by directly phosphorylating a lineage-specific transcription factor. By doing so, cyclin C affects signaling networks that regulate the immune response. Hence, this novel function of cyclin C may have a profound effect on the anti-tumor immune response and on tumorigenesis. In the proposed work we will extend these studies. In Aim 1, we will study the exact molecular function of cyclin C- CDK8 and C-CDK19 in the T cell lineage using a combination of biochemical in vitro studies, cell culture experiments, analyses of ex vivo cultured T cells as well as mouse genetic approaches. In Aim 2, we will study how ablation of cyclin C impacts tumorigenesis in vivo, using mouse cancer models and combination treatments with other anti-cancer modalities. In Aim 3, we will extend these cancer studies to cyclin C catalytic partners, using conditional Cdk19 knockout mice (Cdk19F/F), that my laboratory generated for this purpose. The expected overall impact of this proposal is that it will reveal a novel and important function of cyclin C- CDK8 and C-CDK19 in tumorigenesis, and will validate these kinases as attractive therapeutic targets.

项目总结/摘要细胞周期蛋白C与其他G1期细胞周期蛋白沿着被克隆，作为一种可以拯救酵母增殖阻滞的cDNA 缺乏CLN 1、CLN 2和CLN 3基因的酿酒酵母。随后的研究表明，细胞周期蛋白C，与其激酶伴侣细胞周期蛋白依赖性激酶8（CDK 8）和CDK 19一起作用，在调节基因转录。细胞周期蛋白C作为CDK 8和CDK 19的调节伙伴，并激活细胞周期蛋白C。这些蛋白质的激酶活性。这两种帕金森病激酶显示出基本的总体氨基酸同一性，特别是在它们的催化域内。细胞周期蛋白C、CDK 8和CDK 19代表介导因子的一部分复合物，一种大的多亚基蛋白复合物，通过连接RNA聚合酶II与序列特异性转录因子。此外，细胞周期蛋白C-CDK 8/19被证明磷酸化各种细胞周期蛋白。转录因子，并调节其稳定性和活性。越来越多的证据表明细胞周期蛋白C-CDK 8 C-CDK 19激酶可能是潜在的抗癌靶点。CDK 8已被鉴定为癌基因在几种人类癌症中。细胞周期蛋白C、CDK 8和CDK 19在广泛的肿瘤类型中过表达。重要的是，发现这三种蛋白质的较高表达与较差的临床结果相关。与细胞周期蛋白C-CDK 8/19的肿瘤促进作用一致，对携带几种细胞周期蛋白C-CDK 8/19的异种移植物的小鼠进行治疗，使用小分子CDK 8/19抑制剂的人肿瘤类型导致有效的抗肿瘤作用，明显的全身毒性。为了了解细胞周期蛋白C在体内的分子功能，我们产生了条件性的细胞周期蛋白C敲除小鼠（细胞周期蛋白CF/F）。测试细胞周期蛋白C-CDK 8/19抑制对T细胞发育的影响和适应性抗肿瘤免疫，我们将cyclin CF/F小鼠与CD 4-Cre动物杂交。后一种菌株在T细胞发育的早期阶段表达Cre重组酶，并驱动"floxed“ 所有T细胞亚群中的序列。在我们的初步分析中，我们确定了细胞周期蛋白C的功能是一个门- T细胞分化的保持者，可能通过直接磷酸化谱系特异性转录因子。通过这样，细胞周期蛋白C就影响调节免疫反应的信号网络。因此，细胞周期蛋白C可能对抗肿瘤免疫应答和肿瘤发生具有深远的影响。在我们会继续进行这些研究。在目的1中，我们将研究细胞周期蛋白C的确切分子功能， CDK 8和C-CDK 19在T细胞谱系中使用生物化学体外研究、细胞培养实验，离体培养的T细胞的分析以及小鼠遗传方法。在目标2中，我们将研究使用小鼠癌症模型和组合，与其他抗癌方式的治疗。在目标3中，我们将这些癌症研究扩展到细胞周期蛋白C催化合作伙伴，使用条件性Cdk 19敲除小鼠（Cdk 19 F/F），我的实验室为此目的产生。这项提议的预期总体影响是，它将揭示细胞周期蛋白C的一个新的和重要的功能， CDK 8和C-CDK 19在肿瘤发生中的作用，并将验证这些激酶作为有吸引力的治疗靶点。