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: 8698302
• 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/8698302
• 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; Deoxycytidine; 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; SW620; Sampling; 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; screening; 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)在美国的发病率接近15万例，占新诊断癌症的10%以上。2006年，在退伍军人管理局医疗保健系统内，有4500例新病例，其中650例已进入晚期，治疗选择非常有限。了解潜在恶性转化的基本生物学对于确定新的治疗靶点和可实施的策略至关重要。上皮性交界性病变在恶性肿瘤中很常见。E-钙粘蛋白是一种粘附性连接蛋白，其表达减少或定位错误一直与结直肠癌有关。Claudin-3、-4和-7都是跨膜紧密连接蛋白，在卵巢癌、结直肠癌和胃癌中都有过表达，并且在卵巢癌细胞系中下调Claudin-3和-4可以降低侵袭力。因此，紧密连接功能障碍和粘连连接功能障碍都与结直肠癌有关。BVES是一种紧密连接相关蛋白，抑制后可诱导人角膜上皮细胞间充质转化。由于间质转化发生在晚期结直肠癌，促进转移，我们推测结直肠癌中BVES可能发生改变。我们发现BVES在结直肠癌中的表达降低。BVES在腺瘤性息肉中低表达，表明BVES的表达缺失是一个早期事件，可能暗示BVES除了可能导致转移外，还参与调节其他促肿瘤程序。我们确定了低表达的机制是通过转录沉默通过启动子超甲基化，发生在很大一部分临床样本，几乎所有的结直肠癌细胞株调查。我们通过恢复BVES在LIM2405细胞中的表达来检验BVES缺失的功能相关性。这些细胞是弱黏附、高度增殖、迁移和侵袭性的。BVES显着逆转了所有这些表型，导致了转化为上皮细胞形态的“表型”，增加了细胞角蛋白的表达，相应地失去了波形蛋白。这些表型变化与E-钙粘附素表达增加、连环素分布转移到细胞膜以及相关的WNT报告活性降低有关，暗示BVES调节已知的癌症信号通路。此外，在对间充质细胞表型的研究中，我们观察到在RHOA活性降低的情况下，EF-H1、BVES、Zo-1膜积聚增加，表明BVES可能调节Rho信号转导。此外，BVES在LIM2405细胞中的表达抑制了裸鼠移植瘤的生长，并阻断了SW620的转移。综上所述，这些数据表明，BVES在上皮性恶性肿瘤中发挥肿瘤抑制作用，因此BVES可能是一种新的癌症诊断标记物和/或治疗靶点。BVES在上皮性恶性肿瘤中的作用尚不清楚。在这项提案中，我们构建了三个特定的目标，旨在了解BVES在肿瘤生物学中的作用。首先，我们将使用小鼠遗传学方法来测试BVES与APC在促进肿瘤形成方面的合作，并测试BVES在炎症性癌症发生中的作用。我们还建议进行机械性结构功能研究，以定位与BVES依赖表型相关的BVES功能结构域。我们已经进行了酵母双杂交筛选，并鉴定了一组BVES相互作用蛋白。我们建议描述BVES和BCAR3之间的相互作用，BCAR3是一种具有全球环境基金活性的乳腺原癌基因，已知可以调节Rho依赖的过程。最后，我们将确定使用表观遗传修饰剂恢复BVES的表达是否逆转了CRC细胞的恶性表型。这可能为使用BVES甲基化作为诊断标志物来指导治疗提供支持。通过这些研究，我们将获得对BVES缺失如何促进恶性进展的基本见解，潜在地提供证据支持将BVES发展为结直肠癌的治疗目标或诊断标记物，并解决NCI提出的挑衅性问题#20“……能否识别出可作为治疗效果预测或替代的生物标记物或签名？”