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Molecular Pathogenesis and Immune Biology of Intestinal MSI Tumors in a DNA Mismatch Repair-Deficient Mouse Model

DNA 错配修复缺陷小鼠模型中肠道 MSI 肿瘤的分子发病机制和免疫生物学

基本信息

批准号：
290363600
负责人：
Professor Dr. Peer Bork
金额：
--
依托单位：
Europäisches Laboratorium für Molekularbiologie (EMBL)
依托单位国家：
德国
项目类别：
Research Grants
财政年份：
2016
资助国家：
德国
起止时间：
2015-12-31 至 2019-12-31
项目状态：
已结题

来源：
https://gepris.dfg.de/gepris/projekt/290363600?language=en
关键词：
Molecular Pathogenesis Immune Biology Intestinal

项目摘要

Microsatellite instability (MSI) occurs in almost all Lynch syndrome cancers and in about 15% of sporadic colorectal cancers due to functional inactivation of the cellular DNA mismatch repair (MMR) system. MSI colorectal tumorigenesis in Lynch syndrome patients is assumed to proceed through multiple steps which in comparison to the sporadic forms include an accelerated progression from adenomas to carcinomas. MMR deficiency manifests early during adenoma development and even occurs in normal-appearing mucosa. Instability at coding mononucleotide repeats (cMNRs) leads to frameshift mutations in some genes that can drive MSI tumorigenesis.The functional role and clinical utility of cMNR instability in murine MMR deficient tumors has not been examined yet. Previous MMR-deficient mice do not accurately recapitulate the human situation because homozygous mutant MMR mice predominantly develop lymphomas and only few intestinal tumors. However, recently established conditional MMR gene knockout mice for the first time provide the opportunity to address research questions related to the specific implications of defined cMNR mutations in vivo. In preliminary work cMNR mutations in mouse genes have been identified. We plan to extend our initial experiments to a conditional MMR k.o. mouse line (VCMsh2loxP), a preclinical model of Lynch syndrome. These mice lack intestine-specific Msh2 expression and develop intestinal tumors. Whole intestines from VCMsh2loxP mice at different ages will be examined to identify preneoplastic and neoplastic lesions as well as potential early histological alterations in tumor-adjacent and -distant normal mucosa. Histological and MMR immunostaining analyses of whole intestines from heterozygous VCMsh2loxP mice will resolve whether MMR-deficient crypt foci as recently observed in human Lynch syndrome patients also exist in these mice. For all histologically characterized normal mucosa and tumor tissues, mutation profiles of bioinformatically identified cMNR candidate genes will be determined and based on these mutation data tumor-relevant MSI target genes be sought. Potential correlations between mutations in MSI target genes and age-dependent development of MSI neoplasms and preneoplasia will be investigated. Comparison of human and mouse mutation profiles in MSI target genes and corresponding signaling pathways will define differences and overlaps in murine and human MSI carcinogenesis. Immunohistochemical analyses will determine the type and pattern of immune infiltration with emphasis on the identity of the cells involved. These studies will provide important mechanistic insights into the etiology and progression of MSI tumors. Uncovering tumor-specific murine MSI target genes enables prediction of frameshift peptides derived thereof and a molecular basis to test preventive frameshift peptide-based vaccination strategies for MSI tumors in an animal model.

由于细胞DNA错配修复（MMR）系统的功能失活，几乎所有Lynch综合征癌症和约15%的散发性结直肠癌都存在微卫星不稳定性（MSI）。林奇综合征患者的MSI结直肠肿瘤发生被认为是通过多个步骤进行的，与散发形式相比，包括从腺瘤到癌的加速进展。MMR缺陷在腺瘤发展的早期就表现出来，甚至出现在正常的粘膜中。编码单核苷酸重复序列（cMNR）的不稳定性导致某些基因的移码突变，从而导致MSI肿瘤的发生，cMNR不稳定性在小鼠MMR缺陷型肿瘤中的功能作用和临床应用尚未得到研究。以前的MMR缺陷小鼠不能准确地再现人类的情况，因为纯合子突变MMR小鼠主要发生淋巴瘤，只有少数肠道肿瘤。然而，最近建立的条件性MMR基因敲除小鼠的第一次提供了机会，以解决与定义的cMNR突变在体内的具体影响的研究问题。在初步工作中，已经鉴定了小鼠基因中的cMNR突变。我们计划将我们的初步实验扩展到条件MMR k.o.。小鼠系（VCMsh2loxP），Lynch综合征的临床前模型。这些小鼠缺乏精氨酸特异性Msh2表达，并发生肠道肿瘤。将检查来自不同年龄的VCMsh2loxP小鼠的全肠，以鉴定肿瘤前病变和肿瘤病变以及肿瘤邻近和远端正常粘膜中的潜在早期组织学改变。对来自杂合子VCMsh2loxP小鼠的全肠进行组织学和MMR免疫染色分析将解决最近在人类Lynch综合征患者中观察到的MMR缺陷型隐窝病灶是否也存在于这些小鼠中。对于所有组织学表征的正常粘膜和肿瘤组织，将确定生物信息学鉴定的cMNR候选基因的突变谱，并基于这些突变数据寻找肿瘤相关MSI靶基因。MSI靶基因突变与MSI肿瘤和癌前病变的年龄依赖性发展之间的潜在相关性将被研究。人类和小鼠MSI靶基因突变谱和相应信号通路的比较将确定小鼠和人类MSI致癌作用的差异和重叠。免疫组织化学分析将确定免疫浸润的类型和模式，重点是所涉及的细胞的身份。这些研究将为MSI肿瘤的病因和进展提供重要的机制见解。揭示肿瘤特异性鼠MSI靶基因使得能够预测其衍生的移码肽和在动物模型中测试针对MSI肿瘤的基于移码肽的预防性疫苗接种策略的分子基础。