CDK6 in T cell development and cancer

CDK6 在 T 细胞发育和癌症中的作用

• 批准号: 8214597
• 负责人: Philip W. Hinds
• 金额: $ 32.41万
• 依托单位: TUFTS MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-01-15 至 2013-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8214597
• 关键词: Accounting; Adult; Alleles; Animal Testing; Animals; Apoptosis; Apoptotic; Biological Assay; Bone Marrow; Bone Marrow Aspiration; Bone Marrow Cells; CD44 gene; CD8B1 gene; CDKN2A gene; Cell Count; Cell Culture Techniques; Cell Cycle; Cell Cycle Regulation; Cell division; Cells; Cellularity; Control Animal; Cyclin D1; Cyclins; Data; Defect; Development; Enzymes; Event; Fetal Liver; Fibroblasts; Financial compensation; Future; Generations; Genetic; Genetic Epistasis; Genetic Techniques; Genetic Transcription; Growth; Harvest; Health; Hyperactive behavior; IL2RA gene; IL7R gene; Immune System Diseases; In Vitro; Intervention; Knock-out; Knockout Mice; Learning; Malignant Neoplasms; Mammary Neoplasms; Masks; Measures; Mediating; Mediator of activation protein; Modeling; Mouse Strains; Mus; Mutant Strains Mice; Mutation; Oncogenes; Other Genetics; Partner in relationship; Pharmacologic Substance; Phosphorylation; Phosphotransferases; Play; Population; Pre-Clinical Model; Predisposition; Process; Production; Proteins; Proto-Oncogene Proteins c-akt; Reporting; Resistance; Role; Seeds; Signal Transduction; Source; Staging; Staining method; Stains; Stem cells; System; T-Cell Development; T-Cell Leukemia; T-Cell Proliferation; T-Lymphocyte; T-Lymphocyte Subsets; Testing; Therapeutic; Thymoma; Thymus Gland; Thymus Neoplasms; Tissues; Transgenes; Transgenic Organisms; Transplantation; Tumor Stem Cells; Tumor Suppressor Proteins; Tumor-Derived; cell transformation; cell type; cohort; design; developmental plasticity; human disease; leukemia/lymphoma; loss of function; mimetics; mutant; neoplastic cell; notch protein; novel; prevent; progenitor; research study; response; stem; thymocyte; tool; tumor; tumorigenesis

DESCRIPTION (provided by applicant): Cdk4 and cdk6 have been widely thought to be essential but redundant for initiation of the cell cycle in response to growth regulatory signals. In contradiction to this concept, recent genetic evidence indicates that cdk4 and cdk6 are not essential for early development, nor for proliferation of primary cells in culture. However, this insensitivity to cdk4/6 loss may be ascribed to compensation, since "free" D-type cyclins in cdk4/6 knockout cells appear able to activate cdk2 to allow proliferation. Further, considerable evidence now exists to support a crucial role of cdk4/6 in transformed and tumor cells. For example, cdk4 knockout fibroblasts resist transformation and mice lacking the ability of cyclin D1 to activate cdk4/6 are resistant to ErbB2-induced mammary tumors. The role of cdk6 in tumorigenesis is poorly understood, however recent studies have revealed a novel function of cdk6 in the differentiation of some cell types. For example, mice lacking cdk6 show defects in T cell proliferation. Importantly, conventional knockout experiments may underestimate the impact of specific subunits on development and cancer, due to promiscuity of D cyclin subunits that activate different cdks when their "proper" partners are lost. To clarify the role of cdk6 in cell cycle control and tumorigenesis using refined genetic tools, we have generated mice with targeted mutations in Cdk6. These knockin alleles generate hyperactive or inactive kinase subunits that may better mimic hyperactivation of cdk6 in tumor cells or model pharmaceutical intervention, respectively. We have found that loss of cdk6 or specific inactivation of cdk6 kinase function greatly reduces thymocyte numbers and significantly reduces proliferation in DN2 cells while concomitantly increasing the DN3 fraction. Conversely, the INK4-insensitive, hyperactive CDK6R31C allele increases thymic cellularity, consistent with increased proliferation of most T cell subsets, most notably the DN2 and CD8+ fractions. However, this increase in proliferation is countered by increased apoptosis, which may limit thymocyte expansion and tumorigenesis. Notably, preliminary analysis of LCK-myrAKT;cdk6-/- mice shows a complete absence of tumors, in contrast to LCK-myrAKT;cdk6WT mice, which quickly succumb to massive T cell tumors. Together, these data demonstrate that cdk6 is an important regulator of T cell proliferation and development, and may be crucial in some forms of T cell leukemia and lymphoma. Cdk6 may thus prove to be a valuable therapeutic candidate for treatment of both T cell tumors and immune disorders. To further characterize the role of cdk6 mutants in T cell development in mice, with particular emphasis on early progenitor function and formation of T cell cancers, we propose to (1) Assess the role of Cdk6 in extra-thymic T cell progenitors and determine the source of alterations in CD25 and CD44 expression in Cdk6-mutant thymocytes; (2) Determine the impact of cdk6 loss or mutation on AKT-induced tumors in the thymus; and (3) Determine if loss of p53 collaborates with cdk6 hyperactivity in R31C thymocytes and study the effects of cdk6 loss of function on tumor formation in p53 mutant mice. PUBLIC HEALTH RELEVANCE: It has become increasingly clear that the enzymes that control the ability of cells to divide play key roles in stem and progenitor cells that act in cellular differentiation, the process whereby cells acquire their specific functions in adult tissue, and also in tumor formation. Here we describe our extensive preliminary studies of cdk6, a cell division control enzyme that we show is key to the formation of normal T cells in the thymus and we find that loss of this enzyme prevents T cell tumor formation. The studies we propose herein are aimed at using unique, genetically altered mice that we have produced to understand the role of cdk6 in T cell development and tumorigensis in an effort to learn how to best target this enzyme with future therapeutics.

描述（由申请人提供）：Cdk4和cdk6已被广泛认为对于响应生长调节信号的细胞周期的启动是必需的但是多余的。与这一概念相反，最近的遗传证据表明 cdk4 和 cdk6 对于早期发育和培养中原代细胞的增殖都不是必需的。然而，这种对 cdk4/6 丢失的不敏感性可能归因于补偿，因为 cdk4/6 敲除细胞中的“游离”D 型细胞周期蛋白似乎能够激活 cdk2 以允许增殖。此外，现在存在大量证据支持 cdk4/6 在转化细胞和肿瘤细胞中的关键作用。例如，cdk4 敲除的成纤维细胞抵抗转化，缺乏细胞周期蛋白 D1 激活 cdk4/6 能力的小鼠对 ErbB2 诱导的乳腺肿瘤具有抵抗力。 cdk6 在肿瘤发生中的作用尚不清楚，但最近的研究揭示了 cdk6 在某些细胞类型分化中的新功能。例如，缺乏 cdk6 的小鼠表现出 T 细胞增殖缺陷。重要的是，传统的敲除实验可能会低估特定亚基对发育和癌症的影响，因为细胞周期蛋白 D 亚基的混杂性，当它们的“适当”伴侣丢失时，会激活不同的 cdk。为了利用精细的遗传工具阐明 cdk6 在细胞周期控制和肿瘤发生中的作用，我们培育了具有 Cdk6 靶向突变的小鼠。这些敲入等位基因产生过度活跃或失活的激酶亚基，可以分别更好地模拟肿瘤细胞中 cdk6 的过度激活或模型药物干预。我们发现，cdk6 的缺失或 cdk6 激酶功能的特异性失活会大大减少胸腺细胞数量，并显着减少 DN2 细胞的增殖，同时同时增加 DN3 分数。相反，INK4 不敏感、过度活跃的 CDK6R31C 等位基因会增加胸腺细胞结构，这与大多数 T 细胞亚群（尤其是 DN2 和 CD8+ 部分）增殖的增加一致。然而，这种增殖的增加被细胞凋亡的增加所抵消，这可能限制胸腺细胞的扩张和肿瘤发生。值得注意的是，LCK-myrAKT;cdk6-/- 小鼠的初步分析显示完全没有肿瘤，而 LCK-myrAKT;cdk6WT 小鼠则很快死于大量 T 细胞肿瘤。总之，这些数据表明 cdk6 是 T 细胞增殖和发育的重要调节因子，并且可能在某些形式的 T 细胞白血病和淋巴瘤中至关重要。因此，Cdk6 可能被证明是治疗 T 细胞肿瘤和免疫疾病的有价值的候选药物。为了进一步表征 cdk6 突变体在小鼠 T 细胞发育中的作用，特别强调早期祖细胞功能和 T 细胞癌症的形成，我们建议 (1) 评估 Cdk6 在胸腺外 T 细胞祖细胞中的作用并确定Cdk6 突变胸腺细胞中 CD25 和 CD44 表达变化的来源； (2)确定cdk6缺失或突变对AKT诱导的胸腺肿瘤的影响； (3)确定p53的缺失是否与R31C胸腺细胞中的cdk6过度活跃有关，并研究cdk6功能缺失对p53突变小鼠肿瘤形成的影响。 公共健康相关性：越来越清楚的是，控制细胞分裂能力的酶在干细胞和祖细胞中发挥着关键作用，这些干细胞和祖细胞在细胞分化中发挥着关键作用，细胞在成体组织和肿瘤中获得其特定功能的过程形成。在这里，我们描述了对 cdk6 的广泛初步研究，cdk6 是一种细胞分裂控制酶，我们证明它是胸腺中正常 T 细胞形成的关键，并且我们发现这种酶的缺失可以防止 T 细胞肿瘤形成。我们在此提出的研究旨在使用我们培育的独特的基因改造小鼠来了解 cdk6 在 T 细胞发育和肿瘤发生中的作用，以了解如何在未来的治疗中最好地靶向这种酶。