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: 8510388
• 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/8510388
• 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; 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例处于晚期，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。上皮交界处病变在恶性肿瘤中很常见。粘附连接蛋白E-钙粘蛋白的表达减少或错误定位长期以来与CRC有关。紧密连接蛋白-3、-4和-7（所有跨膜紧密连接蛋白）在卵巢癌、CRC和胃癌中过表达，并且在卵巢癌细胞系中敲低紧密连接蛋白-3和-4导致侵袭性降低。因此，紧密连接和粘附连接功能障碍都与CRC有关。BVES是一种紧密连接相关蛋白，当其被抑制时可诱导人角膜上皮细胞的间充质转化。由于间质转化发生在晚期结直肠癌，促进转移，我们推测BVES可能在结直肠癌中改变。我们发现BVES在CRC中的表达降低。BVES在腺瘤性息肉中表达不足，表明表达缺失是早期事件，可能暗示BVES除了可能促进转移外，还调节其他促肿瘤发生程序。我们确定低表达的机制是通过启动子高甲基化引起的转录沉默，发生在大部分临床样本中，几乎所有的CRC细胞系都被调查过。我们通过恢复BVES在LIM 2405细胞中的表达来测试BVES损失的功能相关性。这些细胞是弱粘附的、高度增殖的、迁移的和侵袭性的。BVES显着逆转了所有这些表型，导致细胞系“表型化”，转化为上皮形态，细胞角蛋白表达增加，波形蛋白相互损失。这些表型变化与E-钙粘蛋白表达增加相关，E-连环蛋白分布转移到细胞膜，WNT报告活性降低，暗示BVES调节已知的癌症信号传导途径。此外，在研究间充质表型时，我们观察到GEF-H1，BVES，Zo-1膜积累增加，RHOA活性降低;表明BVES可以调节Rho信号传导。此外，BVES在LIM 2405细胞中的表达减弱了裸鼠移植瘤的生长，并阻断了SW 620转移。总的来说，这些数据表明BVES在上皮恶性肿瘤中起肿瘤抑制剂的作用，因此BVES可能代表癌症的新的诊断标记物和/或治疗靶点。关于BVES在上皮性恶性肿瘤中的功能仍有许多未知之处。在这个建议中，我们构建了三个具体的目标，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在促进肿瘤发生中的合作，并测试BVES在炎性癌变中的作用。我们还提出了机械结构功能的研究，以映射BVES功能域有助于BVES依赖表型。我们已经进行了酵母双杂交筛选，并确定了一组BVES相互作用蛋白。我们建议表征BVES和BCAR 3之间的相互作用，BCAR 3是一种具有GEF活性的乳腺原癌基因，已知其调节RHO依赖性过程。最后，我们将确定使用表观遗传修饰剂恢复BVES表达是否逆转CRC细胞的恶性表型。这可以为使用BVES甲基化作为指导治疗的诊断标志物提供支持。通过这些研究，我们将从根本上了解BVES的丢失如何导致恶性进展，可能提供支持将BVES开发为结直肠癌治疗靶点或诊断标志物的证据，并解决NCI“…是否可以鉴定出可作为治疗功效的预测因子或替代物的生物标志物或特征？".