FA pathway activities in the normal and transformed epidermis

正常和转化表皮中的 FA 途径活性

• 批准号: 9767108
• 负责人: Susanne I Wells
• 金额: $ 35.28万
• 依托单位: CINCINNATI CHILDRENS HOSP MED CTR
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-08-20 至 2023-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9767108
• 关键词: 3-Dimensional; Actins; Adhesions; Age; Anemia; Basal Cell; Biological Markers; Biology; Biopsy; Bulla; Cell Adhesion; Cell Compartmentation; Cell Death; Cell Proliferation; Cell membrane; Cell model; Cell physiology; Cells; Chemicals; Childhood; Clinical Research; Coupled; Crosslinker; Cytoskeleton; DNA Damage; DNA Interstrand Crosslinking; DNA Repair; DNA-dependent protein kinase; Data; Defect; Desmosomes; Development; Diagnosis; Disease; Environment; Enzymes; Epidermis; Epithelial; Etiology; Evolution; Exhibits; Fanconi Anemia pathway; Fanconi' s Anemia; Ganglioside Biosynthesis Pathway; Gangliosides; Genes; Genetic; Genomic Instability; Goals; Growth; Guanosine Triphosphate Phosphohydrolases; Health; Hematopoietic stem cells; Hemidesmosomes; Homeostasis; Human; Human Papillomavirus; Human Pathology; Human papilloma virus infection; Impairment; In Vitro; Inherited; Integrin alpha6; Isotopes; Knowledge; Link; Lipids; Malignant Epithelial Cell; Mass Spectrum Analysis; Mechanics; Mediator of activation protein; Membrane; Membrane Microdomains; Mesenchymal; Methods; Molecular; Monomeric GTP-Binding Proteins; Mutation; Names; Neoplasm Metastasis; Nuclear; Oncogenic; Oral cavity; Pancytopenia; Pathogenesis; Pathology; Pathway interactions; Patients; Pediatric Hospitals; Pharmaceutical Preparations; Phenotype; Phosphotransferases; Play; Polynucleotide 5' -Hydroxyl-Kinase; Predisposition; Prevention; Production; Publishing; Radiation; Regulation; Research; Resistance to infection; Risk; Role; Signal Pathway; Signal Transduction; Skin; Skin Cancer; Squamous cell carcinoma; Stem cells; Structural defect; Structure; Testing; Therapeutic; Tissues; Work; Xenograft procedure; base; cancer cell; cancer prevention; cancer therapy; cell motility; cell transformation; cell type; cellular engineering; crosslink; desmoplakin; experience; experimental study; genome integrity; human model; improved; in vivo; induced pluripotent stem cell; keratinocyte; leukemia; loss of function mutation; malignant mouth neoplasm; mouse model; novel; patient population; prevent; reconstitution; sensor; skin cancer prevention; stem; therapeutic target; tool; translational study; tumor

ABSTRACT Human epidermis serves as a critical barrier against mechanical, chemical, radiation and infectious insults in the external environment. Keratinocytes are the main cell type in this stratified tissue, and are organized into basal stem and progenitor cells, and the more superficial differentiated progeny. Epidermal homeostasis and integrity are tightly regulated and critical for human health. Squamous cell carcinomas (SCCs) are perhaps a prime example of epidermal homeostasis gone awry. Both sporadic and inherited forms of SCC are associated with mutations in the 21 genes of the Fanconi anemia (FA) pathway, which participate in the repair of DNA crosslinks. Close associations between the FA pathway and SCC development point to important roles for this pathway in keratinocytes and epidermis – roles that have not yet been studied, but will be investigated here using FA patient populations and tools to study keratinocyte biology. These tools include human iPSC-derived keratinocytes and 3D organotypic epithelial rafts isogenic for FA genes, as well as skin biopsies from FA patients (vs. controls) that are uniquely available at Cincinnati Children's Hospital. Our translational studies build on recent data demonstrating cell adhesion, cytoskeletal and membrane defects in FA-deficient keratinocytes, skin biopsies, and 3D organotypic rafts. The defective keratinocytes were also hyper-motile, and invasive particularly when transformed – and the invasive phenotype depended on the activities of the DNA damage sensor kinase DNA-PK and the small GTPase Rac1, as well as ganglioside accumulation. The overarching hypothesis is that FA pathway loss leads to the assembly of a Rac1 signalosome in membrane domains enriched for gangliosides (eg lipid rafts), and consequent phenotypes that are reminiscent of epithelial-mesenchymal-transition (EMT-like). Aim 1 tests the hypothesis that the FA pathway regulates adhesion/integrity and cell motility in the normal epidermis through ganglioside accumulation and DNA- PK/Rac1 signaling. Aim 2 directly tests the impact of FA loss on transformation, defines the FA-pathway- dependent flux of ganglioside biosynthesis and its role in sustaining SCC growth and progression. The results will identify a) signaling and lipid-based mechanisms whereby nuclear DNA repair machineries control the integrity of human epidermis, and b) biomarkers and therapeutic targets for the prevention and treatment of SCC development in patients with acquired and inherited FA pathway deficiencies.

摘要 人表皮在免疫系统中充当抵抗机械、化学、辐射和感染性损伤的关键屏障。 外部环境。角质形成细胞是该分层组织中的主要细胞类型，并且被组织成 基底干细胞和祖细胞，以及更表层的分化后代。表皮稳态和 诚信受到严格监管，对人类健康至关重要。鳞状细胞癌（SCC）可能是一种 表皮稳态失调的典型例子散发性和遗传性SCC都与 范可尼贫血（FA）途径的21个基因发生突变，这些基因参与DNA修复， 交叉连接。FA途径和SCC发展之间的密切联系表明了这一重要作用。 角质形成细胞和表皮中的通路-尚未研究的作用，但将在这里进行研究 使用FA患者群体和工具来研究角质形成细胞生物学。这些工具包括人类iPSC衍生的 角质形成细胞和3D器官型上皮筏与FA基因同源，以及来自FA的皮肤活检 辛辛那提儿童医院唯一可用的患者（与对照组）。我们的翻译研究 建立在最近的数据表明，细胞粘附，细胞骨架和膜缺陷，在FA缺乏 角质形成细胞、皮肤活检和3D器官型筏。有缺陷的角质形成细胞也是超运动的， 侵袭性，特别是转化时-侵袭性表型取决于DNA的活性 损伤传感器激酶DNA-PK和小GT3 Rac 1，以及神经节苷脂积累。的 总体假设是FA途径的丧失导致Rac 1信号体在膜中的组装 富含神经节苷脂的结构域（如脂筏），以及随后的表型， 上皮-间充质转化（EMT样）。目的1检验FA途径调节 粘附/完整性和细胞运动性在正常表皮通过神经节苷脂积累和DNA- PK/Rac 1信号传导。目标2直接测试FA损失对转化的影响，定义FA途径- 神经节苷脂生物合成的依赖性流量及其在维持SCC生长和进展中的作用。结果 将确定a）信号和脂质为基础的机制，从而核DNA修复机制控制 人表皮的完整性，和B）用于预防和治疗 获得性和遗传性FA通路缺陷患者的SCC发展