The role of the DNA damage response in the development and therapeutic sensitivity of malignant testicular germ cell tumors.

DNA 损伤反应在恶性睾丸生殖细胞肿瘤的发展和治疗敏感性中的作用。

• 批准号: 10065429
• 负责人: Amanda Loehr
• 金额: $ 4.25万
• 依托单位: CORNELL UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-01-01 至 2025-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10065429
• 关键词: Aftercare; Alleles; Anchorage-Independent Growth; Apoptosis; Breast Carcinoma; Candidate Disease Gene; Cell Cycle Arrest; Cell Differentiation process; Cells; Chemoresistance; Colon Carcinoma; Comet Assay; DNA; DNA Damage; DNA Repair; DNA Replication Timing; DNA Sequence Alteration; DNA biosynthesis; Development; Embryo; Embryonal Carcinoma; Embryonal Carcinoma Cell; Embryonic Development; Enterobacteria phage P1 Cre recombinase; Event; Exercise; Fiber; Fibroblasts; Generations; Genes; Genetic Recombination; Genetic Risk; Genetically Engineered Mouse; Genome; Germ Cells; Human; In Situ; In Vitro; Lesion; Malignant - descriptor; Malignant Neoplasms; Measures; Modeling; Mus; Mutagens; Mutate; Mutation; Nature; Neoplasms; Oncogene Activation; Oncogenes; Oncogenic; Pathology; Pathway interactions; Phenotype; Pluripotent Stem Cells; Property; Risk; Role; Signal Transduction; Somatic Cell; Structure of primordial sex cell; System; Teratocarcinoma; Teratoma; Testicular Germ Cell Tumor; Testicular Neoplasms; Testicular malignant germ cell tumor; Therapeutic; Tissue-Specific Gene Expression; Tissues; Tumor Suppressor Genes; Tumorigenicity; Undifferentiated; cell transformation; cell type; chemotherapy; experience; experimental study; genotoxicity; improved; in vitro Model; in vivo; induced pluripotent stem cell; lung Carcinoma; mouse model; neoplastic cell; pressure; prevent; repaired; replication stress; response; stem cell biomarkers; stem cell therapy; therapy development; transcriptome sequencing; treatment response; tumor; tumor initiation; tumor progression; tumorigenesis; young man

Project Summary Testicular germ cell tumors (TGCTs) are exceptionally sensitive to conventional genotoxic chemotherapy. This is likely due to the distinct DNA damage response (DDR) features of TGCTs and the germ cells from which they arise. Unlike somatic cells which often respond to DNA damage by arresting the cell cycle and conducting DNA repair, germ cells as well as long-lived pluripotent stem cells typically avoid the use of error-prone repair mechanisms and favor apoptosis, reducing the risk of genetic alterations in subsequent generations. Similarly, the TGCT precursor lesion, germ cell neoplasia in situ, does not show activation of a DDR, whereas precursor lesions of most somatic cancers express markers of an activated DDR in response to oncogene activation which serves as a barrier to tumor progression. To study TGCTs, our lab has developed the first genetically engineered mouse model of malignant TGCTs by conditional activation of Kras, an oncogene, and inactivation of Pten, a tumor suppressor gene, in germ cells. The malignant teratocarcinomas generated in these mice are composed of pluripotent embryonal carcinoma (EC) and differentiated teratoma tissue. Interestingly, EC cells, both in vivo and cultured in vitro, express stem cell markers, have tumor propagating activity, and are readily killed following chemotherapy treatment. Using cells derived from this model, the experiments proposed here will elucidate the DDR properties of TGCTs and the cells from which they arise, which will inform the mechanisms underlying their exceptional chemosensitivity. Specifically, this proposal aims to: understand how the cells that give rise to TGCTs respond to oncogenic events, apparently avoiding DDR activation (Aim 1), and determine the mechanism underlying the chemosensitivity of the embryonal carcinoma components of TGCTs and the chemoresistance of their differentiated counterparts (Aim 2). Aim 1 will be investigated by generating primordial germ cell-like cells (PGCLCs) and embryonic germ cell-like cells (EGCLCs) from induced pluripotent stem cells (iPSCs) derived from mouse embryonic fibroblasts (MEFs) with conditional Pten and Kras alleles and assessing malignant transformation, the degree of DNA replication stress, the extent of DNA damage, and the nature of the DDR in untransformed and transformed cells. For Aim 2, I will analyze differential gene expression in EC and differentiated cells before and after treatment with genotoxic chemotherapy and investigate the role of differentially regulated pathways in the chemosensitivity phenotype. It is critically important to study this curable cancer because understanding the basis of TGCT chemosensitivity will apply broadly to the development of treatments for the many other cancers that do not respond favorably to conventional chemotherapy. Additionally, understanding the malignant transformation of pluripotent cell types, including embryonic germ cells and iPSCs, will be important for the improvement of stem-cell based therapies, which carry the risk of tumorgenicity.

项目摘要 睾丸生殖细胞肿瘤（TGCTS）对常规的遗传毒性化疗非常敏感。这 可能是由于TGCT和它​​们的生殖细胞的独特DNA损伤响应（DDR）特征 出现。与通常通过阻止细胞周期和导电DNA响应DNA损伤的体细胞不同 修复，生殖细胞以及长寿命多能干细胞通常避免使用容易出错的修复 机制和有利于细胞凋亡，降低了随后世代的遗传改变的风险。相似地， TGCT前体病变，原位生殖细胞肿瘤，未显示DDR的激活，而前体 大多数体细胞癌的病变表达激活DDR的标记，以响应癌基因激活 充当肿瘤进展的障碍。为了研究TGCT，我们的实验室开发了第一个基因工程 通过条件激活Kras，癌基因和PTEN灭活的小鼠TGCT的小鼠模型 肿瘤抑制基因，在生殖细胞中。这些小鼠中产生的恶性变质瘤组成 多能性胚胎癌（EC）和分化的畸胎瘤组织的。有趣的是，EC细胞，都在体内 并在体外培养，表达干细胞标记物具有肿瘤传播活性，并且随后易于杀死 化学疗法治疗。使用从该模型得出的细胞，此处提出的实验将阐明 TGCT的DDR特性及其出现的细胞，这将告知其基础机制 异常的化学敏感性。具体而言，该建议的目的是：了解产生的细胞如何 TGCT对致癌事件做出反应，显然避免了DDR激活（AIM 1），并确定机制 TGCTS的胚胎癌成分的化学敏度和化学抗性的基础 他们的差异化对应物（AIM 2）。 AIM 1将通过产生原始生殖细胞样细胞来研究 （PGCLCS）和来自诱导多能干细胞（IPSC）的胚胎生殖细胞样细胞（EGCLC） 来自带有条件PTEN和KRAS等位基因的小鼠胚胎成纤维细胞（MEF）并评估恶性肿瘤 转化，DNA复制应力的程度，DNA损伤程度以及DDR中的性质 未转化和转化的细胞。对于AIM 2，我将分析EC和 通过遗传毒性化学疗法治疗前后分化细胞，并研究 化学敏感表型中的差异调节途径。研究这种可再生非常重要 癌症是因为了解TGCT化学敏感性的基础将广泛适用于 对许多对常规化学疗法反应不佳的癌症的治疗方法。此外， 了解多能细胞类型的恶性转化，包括胚胎生殖细胞和IPSC， 对于改善基于干细胞的疗法的疗法至关重要，这些疗法具有肿瘤性的风险。