Modulation of junctional signaling by BVES in colorectal carcinoma

BVES 对结直肠癌中连接信号的调节

• 批准号: 8332389
• 负责人: Christopher S. Williams
• 金额: --
• 依托单位: VETERANS HEALTH ADMINISTRATION
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-07-01 至 2016-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8332389
• 关键词: Address; Adenomatous Polyps; Adherens Junction; Adhesions; Adhesives; Affect; Anchorage-Independent Growth; Attenuated; BCAR3 gene; Binding; Biological Assay; Biological Markers; Biology; Blood Vessels; Breast; CDK4 gene; Cancer Control; Cancer Etiology; Cancer cell line; Cell Adhesion Molecules; Cell Culture Techniques; Cell Cycle; Cell Line; Cell Proliferation; Cell membrane; Cell-Cell Adhesion; Cells; Clinical; Colon; Colon Carcinoma; Colorectal Cancer; Complex; CpG Islands; Cyclin D1; Cytokeratin; Data; Decitabine; Development; Diagnosis; Diagnostic; Disease; Dominant-Negative Mutation; Dysmyelopoietic Syndromes; E-Cadherin; E-Cadherin Staining Method; Epigenetic Process; Epithelial; Epithelial Cells; Event; FDA approved; Functional disorder; Gene Activation; Genetic; Growth; Health Personnel; Healthcare Systems; Human; Hybrids; Hypermethylation; Incidence; Inflammatory; Large Intestine Carcinoma; Link; Malignant - descriptor; Malignant Epithelial Cell; Malignant Neoplasms; Malignant neoplasm of ovary; Maps; Mechanics; Mediating; Membrane; Mesenchymal; Methylation; Modeling; Molecular; Morphology; Mus; Myosin ATPase; Neoplasm Metastasis; Nuclear; Nude Mice; Ovarian; Pathology; Pathway interactions; Phenotype; Play; Process; Proteins; Proto-Oncogenes; RHOA gene; Regulator Genes; Regulatory Pathway; Reporter; Repression; Resistance; Role; Sampling; Screening procedure; Signal Pathway; Signal Transduction; Small Interfering RNA; Staging; Structure; Surveys; Testing; Tight Junctions; Transcriptional Activation; Treatment Efficacy; Tumor Biology; Tumor Suppressor Proteins; United States; United States Department of Veterans Affairs; Veterans; Vimentin; WNT Signaling Pathway; Wound Healing; Xenograft procedure; Yeasts; attenuation; base; blood vessel restoration; burden of illness; cancer cell; cancer diagnosis; carcinogenesis; cell motility; claudin 3; colorectal cancer prevention; corneal epithelium; design; in vivo; insight; knock-down; malignant phenotype; malignant stomach neoplasm; migration; monolayer; mortality; new therapeutic target; novel; novel diagnostics; overexpression; programs; promoter; rho; splenic capsule; therapeutic target; tumor; tumor growth; tumorigenesis; tumorigenic

DESCRIPTION (provided by applicant): Colorectal cancer (CRC) has a US incidence rate of almost 150,000 cases encompassing over 10% of new cancer diagnoses. In 2006 Within the Veterans Administration Healthcare System there were 4500 new cases and of those 650 were at advanced stage with very limited treatment options. Understanding the basic biology of underlying malignant transformation is critical in identifying new therapeutic targets and implementable strategies. Epithelial junctional pathology is common in malignancy. Decreased expression or mislocalization of E-cadherin, an adherens junction protein, has long been implicated in CRC. Claudin -3, -4, and -7, all transmembrane tight junction proteins, are overexpressed in ovarian, CRC, and gastric cancers and knock-down of claudin-3 and -4 in ovarian cancer cell lines leads to reduced invasiveness. Thus, both tight and adherens junctional dysfunction is implicated in CRC. BVES is a tight junction associated protein which when suppressed induces mesenchymal transformation in human corneal epithelial cells. Because mesenchymal transformation occurs in late stage CRC, facilitating metastasis, we postulated that BVES may be altered in CRC. We found decreased expression of BVES in CRC. BVES was underexpressed in adenomatous polyps indicating that loss of expression is an early event, likely implicating BVES in regulating additional pro- tumorigenic programs in addition to potentially contributing to metastasis. We determined the mechanism of underexpression was via transcriptional silencing via promoter hypermethylation, occurring in a large fraction of clinical samples, and virtually all CRC cell lines surveyed. We tested the functional relevance of BVES loss by restoring its expression in LIM2405 cells. These cells are weakly adherent, highly proliferative, migratory, and invasive. BVES strikingly reversed all of these phenotypes causing "epithealization" of the line with conversion to epithelial morphology and increased expression of cytokeratin and reciprocal loss of vimentin. These phenotypic changes were associated with increased E-cadherin expression with a shift in ¿-catenin distribution to the cell membrane with associated decreased WNT reporter activity, implicating BVES in regulating a known cancer-signaling pathway. Additionally, in investigating the mesenchymal phenotype, we observed increased GEF-H1, BVES, Zo-1 membrane accumulation with decreased RHOA activity; indicating BVES could regulate Rho signaling. Furthermore, BVES expression in LIM2405 cells attenuated tumor growth as nude mouse xenografts and blocked SW620 metastasis. Collectively, this data suggests that BVES functions as a tumor suppressor in epithelial malignancy, thus BVES may represent a new diagnostic marker and/or therapeutic target in cancer. Much remains to be known about BVES function in epithelial malignancy. In this proposal we structure three specific aims designed to understand the role of BVES in tumor biology. First we will use murine genetic approaches to test for BVES cooperation with APC in promoting tumorigenesis and to test for a role for BVES in inflammatory carcinogenesis. We also propose mechanistic structure function studies to map BVES functional domains contributing to BVES dependent phenotypes. We have performed a yeast 2-hybrid screen and identified a panel of BVES interacting proteins. We propose characterizing the interaction between BVES and BCAR3 a breast protooncogene with GEF activity known to regulate RHO dependent processes. Lastly, we will determine if restoration of BVES expression using epigenetic modifying agents reverses the malignant phenotype of CRC cells. This could provide support for using BVES methylation as a diagnostic marker to guide therapy. Through these studies we will gain fundamental insight into how loss of BVES contributes to malignant progression, potentially providing evidence that supports developing BVES as a therapeutic target or diagnostic marker in colorectal carcinoma and addressing Provocative Question #20 from the NCI "...can biomarkers or signatures be identified that can serve as predictors or surrogates of therapeutic efficacy?".

描述（由申请人提供）： 结直肠癌（CRC）的发病率近15万例病例，其中包括超过10％的新癌症诊断。 2006年，在退伍军人管理系统中，有4500例新病例，其中650例处于高级阶段，治疗方案非常有限。了解潜在的恶性转化的基本生物学对于确定新的治疗靶标和可实施策略至关重要。上皮连接病理在恶性肿瘤中很常见。 E-钙粘蛋白（一种粘附连接蛋白）的表达降低或错误定位已在CRC中长期暗示。 claudin -3，-4和-7，所有跨膜紧密连接蛋白在卵巢，CRC和胃癌中均过表达，而Claudin -3和-4在卵巢癌细胞系中的敲除导致侵入性降低。 CRC中暗示着紧密和粘附的连接功能障碍。 BVE是一种紧密的相关蛋白，当抑制诱导人角膜上皮细胞中的间充质转化时。由于间充质转化发生在晚期CRC，支持转移，因此我们发布了BVE可能会改变CRC中的BVE。我们发现BVE在CRC中的表达降低。 BVE在腺瘤息肉中不受欢迎，表明表达丧失是早期事件，除了潜在地导致转移外，还可能在调节性额外的非肿瘤性计划中隐含BVE。我们确定了通过启动子高甲基化的转录沉默，确定了不渗透的机制，发生在很大一部分的临床样本中，几乎所有的CRC细胞系都进行了调查。我们通过恢复其在LIM2405细胞中的表达来测试BVE丢失的功能相关性。这些细胞是弱的，高度增殖，迁移和侵入性的。 BVE显着逆转了所有这些表型，导致线路上的“上皮化”，并转化为上皮形态，并增加了细胞角蛋白的表达和波形蛋白的相互损失。这些表型的变化与E-钙粘蛋白表达的增加有关，随着 - 钙蛋白分布向细胞膜的转移，与相关的Wnt报告降低了，这暗示了BVE在确定已知的癌症信号途径时暗示了BVE。此外，在研究间充质表型时，我们观察到GEF-H1，BVE，ZO-1膜的积累增加，RHOA活性降低。表明BVE可以调节RHO信号传导。此外，BVE在LIM2405细胞中的表达使肿瘤的生长减弱，因为裸小鼠异种移植物并阻断了SW620转移。总的来说，这些数据表明BVE在上皮恶性肿瘤中起肿瘤的抑制作用，因此BVE可能代表癌症中新的诊断标记和/或治疗靶标。关于BVE在上皮恶性肿瘤中的功能还有很多尚待了解。在此提案中，我们构建了三个旨在了解BVE在肿瘤生物学中的作用的特定目标。首先，我们将使用鼠遗传方法来测试BVE与APC合作，以促进肿瘤发生，并测试BVE在炎症性癌中的作用。我们还提出了机械结构功能研究，以绘制BVE的功能域，这些功能域促成依赖BVE的表型。我们已经进行了酵母2杂交筛选，并鉴定了一组BVE的相互作用蛋白。我们提出表征BVE和BCAR3 A乳腺原子元与已知调节RHO依赖过程的GEF活性的相互作用。最后，我们将确定使用表观遗传修饰剂恢复BVE表达是否会逆转CRC细胞的恶性表型。这可以为使用BVE甲基化作为指导治疗的诊断标记提供支持。通过这些研究，我们将获得基本的洞察力，了解BVE的损失如何有助于恶性进展，并提供证据支持将BVE作为结直肠癌的治疗目标或诊断标记，并从NCI中解决挑衅性问题。