Epi)Genomic drivers of primary and metastatic pancreatic islet cell carcinoma

描述(申请人提供)：胰腺神经内分泌肿瘤相对罕见，但由于肿瘤谱系的复杂性和不确定的组织发生，对这些实体仍有相当大的兴趣。不幸的是，能够忠实地复制人类疾病并能够更彻底地调查疾病过程的病因的动物模型很少。我们已经建立了一种非常有用的新的胰岛细胞癌动物模型，这是由于细胞特异性地删除了胰腺肾素表达的细胞室内的Rb和P53定位。观察到的主要肿瘤是胰岛细胞癌，表达胰高血糖素，表现为高外显性，症状和深度转移到代表人类疾病的部位，导致5-6个月死亡。在人类疾病中，主要的转移部位是区域淋巴结和肝脏，这与我们的模型相似。这一模型展示了一些独特的特征，使我们能够更好地了解胰岛细胞的发育、肾素-血管紧张素系统在其中的作用、胰岛细胞的癌变以及相关的转移过程，目的是最终确定这些过程的遗传特征。我们假设肾素在早期的胰高血糖素谱系分化中瞬时表达，并且在发生随机合作遗传和表观遗传学异常时，p53和Rb的丢失使这些胰岛细胞易于直接致癌。此外，我们认为细胞需要额外或特定的基因组损伤才能获得转移表型。我们的假设得到了我们的初步发现的支持，即表达肾素的细胞与表达胰高血糖素的阿尔法胰岛细胞共存，并且随机发生的原发和转移性肿瘤继续表达胰高血糖素。最近的文献进一步支持了这一点，即人胰腺前体细胞在向各种类型的胰岛细胞分化的过程中，肾素-血管紧张素系统的各种成分都表现出高度调控的表达。当前提案将使用Next 对外显子组、所有转录本和甲基组进行世代测序，以确定与随机出现的原发肿瘤和相应的肝转移相关的正常胰岛Rb、P53、肾素表达细胞的直系后代中发生的合作突变、基因表达变化和DNA甲基化变化。此外，使用荧光记者追踪细胞谱系的能力有助于 对于疾病的发生和发展提供了一个独特的机会来剖析疾病病理的时间线。我们还期望五彩纸屑记者将有助于识别每个原发肿瘤胰岛中单个有色细胞的克隆性扩张，并有助于正确配对有色转移肿瘤和相同颜色的原发肿瘤。这将增加原发肿瘤和转移肿瘤之间的成对比较的能力，并最终将允许解剖转移扩散的多个分子路线。

DESCRIPTION (provided by applicant): Neuroendocrine tumors of the pancreas are relatively rare but there remains considerable interest in these entities because of the complexity of the tumor spectrum and the uncertain histogenesis. Unfortunately, animal models that can faithfully replicate human disease and enable a more thorough investigation of the etiology of the disease process are few. We have created what appears to be a remarkably useful new animal model for pancreatic islet cell carcinoma as a result of cell-specifically deleting floxed Rb and p53 loc within the renin- expressing cell compartment of pancreas. The predominant neoplasia observed is an islet cell carcinoma, expressing glucagon, which arises with high penetrance and exhibits symptoms and profound metastatic spread to sites representative of the human disease, resulting in death by 5-6 months. In the human disease the primary metastatic sites are regional lymph nodes and liver, which is mimicked by our model. This model exhibits unique features which will allow us to foster better understanding of pancreatic islet cell development, the role the renin-angiotensin system therein, islet cell carcinogenesis, and the associated metastatic process, with the aim of ultimately identifying genetic signatures for these processes. We hypothesize that renin is transiently expressed in early glucagon lineage differentiation and that the loss of p53 and Rb predisposes these islet cells to frank carcinogenesis upon the occurrence of stochastic co-operating genetic and epigenetic abnormalities. Furthermore, we propose that additional or specific genomic insults are required for the cells to acquire a metastatic phenotype. Our hypothesis is supported by our preliminary findings that the renin expressing cells co-localize with the alpha islet cells expressing glucagon and that both primary and metastatic tumors, stochastically arising, continue to express glucagon. This is further supported by recent literature demonstrating that human pancreatic progenitor cells exhibit highly regulated expression of various components of the renin-angiotensin system throughout the progenitor cell differentiation to various islet cell types. The current proposal will use Next Generation Sequencing of the exome, all transcripts, and of the methylome to identify co-operating mutations, gene expression changes and DNA methylation changes occurring within lineal descendants of the Rb-, p53-, renin-expressing cells of normal pancreatic islets that are associated with the stochastically arising primary tumors and corresponding liver metastases. Furthermore, the ability to use fluorescent reporters for lineage tracing of the cells contributing to disease initiation and progression provides a unique opportunity to dissect the timeline of disease pathology. We further expect that the Confetti reporter will assist in the identification o clonal expansion of individual colored cells in the islet for each primary tumor, and facilitate correctly pairing the colored metastatic tumor with the same colored primary tumor. This will increase the power of the pairwise comparisons between primary and metastatic tumors and will ultimately allow for dissection of multiple molecular routes of metastatic spread.