Identifying a nodal point for G alpha q signaling in eye disease

确定眼部疾病中 G α q 信号传导的节点

• 批准号: 9006784
• 负责人: DEAN Yaw LI
• 金额: $ 34.08万
• 依托单位: UNIVERSITY OF UTAH
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-01-01 至 2020-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9006784
• 关键词: BRAF gene; Binding; Biological Assay; Blood Vessels; Cause of Death; Cell Nucleus; Cell membrane; Choroid; Cutaneous Melanoma; Data; Disease; Eye; Eye Neoplasms; Eye diseases; FZD4 gene; G alpha q Protein; G(q) Alpha; GNAQ gene; Gene Silencing; Genetic Transcription; Glaucoma; Growth; Health; Human; In Vitro; Individual; Liver; Lung; MAP Kinase Gene; Malignant Neoplasms; Mediating; Melanoma Cell; Methods; Molecular Target; Monomeric GTP-Binding Proteins; Mutation; Neoplasm Metastasis; Nodal; Nuclear Translocation; Oncogenic; Organ; Pathway interactions; Patients; Pharmacotherapy; Phospholipase C; Play; Protein Kinase C; Proteins; RNA Interference; Research Personnel; Retina; Role; Running; Signal Pathway; Signal Transduction; Signaling Protein; Structure of lamina episcleralis; Sturge-Weber Syndrome; Testing; Therapeutic; Transcription Factor AP-1; Uveal Melanoma; WNT Signaling Pathway; WNT5A gene; Xenograft Model; Xenograft procedure; base; conjunctiva; drug development; effective therapy; in vivo; in vivo Model; knock-down; malformation; mouse model; small hairpin RNA; small molecule; small molecule inhibitor; therapeutic target; trafficking; tumor; tumorigenesis

DESCRIPTION (provided by applicant): Activating mutations in two Gα proteins of the q class (Gαq), GNAQ and GNA11, are the drivers of oncogenesis in approximately 80% of uveal melanomas. Similar activating mutations of GNAQ are also found in patients with Sturge-Weber syndrome, which can manifest itself as glaucoma and vascular malformations of the conjunctiva, choroid, retina, and episclera. Uveal melanoma is the most common primary ocular tumor, and in approximately 50% of the cases, the tumor will metastasize to other organs, primarily the liver. Once metastasized, the disease is invariably fatal. Activating GNAQ and GNA11 mutations drive uveal melanoma oncogenesis via the control of several recently identified signaling pathways, including phospholipase C-ß/protein kinase C (PLC-ß/PKC) and TRIO-RhoA/Rac1 pathways, which activate MAPK/ERK and YAP to induce AP1- and YAP-TEAD-mediated transcription. However, the mechanism(s) by which Gαq proteins activate multiple downstream pathways has not been completely elucidated. Preliminary data suggest that the small GTPase ARF6 may act as an immediate downstream effector of activated GNAQ/GNA11 to control all of the currently known oncogenic Gαq signaling pathways. Other preliminary data also suggest that Gαq may signal through ARF6 to activate ß-catenin signaling by promoting the relocalization of ß-catenin from the cell membrane to the nucleus where it can mediate gene transcription. Preliminary data also support the in vivo role of ARF6 in uveal melanoma. When ARF6 is silenced in uveal melanoma by shRNA or is inhibited with a small molecule inhibitor, tumor establishment and growth is significantly inhibited in an orthotopic xenograft mouse model. Based on these preliminary data, the following aims will be pursued. In Aim 1, we will investigate whether activated Gαq proteins induce ß-catenin signaling via activation of ARF6 in uveal melanoma. We will employ gene silencing via RNA interference and small molecule inhibition of selected targets to assess intracellular localization, transcriptional activity, and function of ß-catenin in uveal melanoma. In Aim 2, we will elucidate the role of ARF6 in orchestrating known downstream signaling pathways of oncogenic Gαq. The same strategies used in Aim 1 will be employed to determine whether ARF6 acts as an immediate downstream effector of activating GNAQ/GNA11 mutations to control (PLC-ß/PKC) and TRIORhoA/ Rac1 pathways and AP-1 and YAP-TEAD-mediated transcription. In Aim 3, we will assess the in vivo function of ARF6 in tumor establishment and growth by using orthotopic xenograft models of human uveal melanoma. The successful completion of these aims will allow us to determine whether ARF6 plays a critical role in Gq signaling, thus providing a promising therapeutic target for the development of drugs that could be used to treat uveal melanoma and possibly other Gαq-related disorders such as Sturge-Weber syndrome.

描述(由申请人提供)：在大约80%的葡萄膜黑色素瘤中，两个Q类Gα蛋白(GαQ)的激活突变是肿瘤发生的驱动因素。在斯特奇-韦伯综合征患者中也发现了类似的GNAQ激活突变，该综合征可表现为青光眼和结膜、脉络膜、视网膜和巩膜上的血管畸形。葡萄膜黑色素瘤是最常见的眼部原发肿瘤，在大约50%的病例中，肿瘤会转移到其他器官，主要是肝脏。一旦转移，这种疾病总是致命的。激活GNAQ和GNA11突变通过控制几个新近发现的信号通路来促进葡萄膜黑色素瘤的发生，这些信号通路包括磷脂酶C/蛋白激酶C(PLC-ç/PKC)和Trio-RhoA/rac1通路，它们激活MAPK/ERK和YAP，从而诱导AP1和YAP-tead介导的转录。然而，机制(S)由 G-α-Q蛋白激活多条下游通路的机制尚未完全阐明。初步数据表明，小分子GTP酶ARF6可能作为GNAQ/GNA11激活的直接下游效应器，控制目前已知的所有致癌G-α-Q信号通路。其他初步数据也表明，GαQ可能通过ARF6发出信号，通过促进？连环蛋白从细胞膜到细胞核的重新定位来激活？连环蛋白信号通路，在那里它可以介导基因转录。初步数据也支持ARF6在葡萄膜黑色素瘤中的体内作用。当ARF6在葡萄膜黑色素瘤中被shRNA沉默或被小分子抑制剂抑制时，在原位异种移植小鼠模型中，肿瘤的建立和生长受到显著抑制。在这些初步数据的基础上，将实现以下目标。在目标1中，我们将研究激活的G-αQ蛋白是否通过激活葡萄膜黑色素瘤中的ARF6来诱导？连环蛋白信号传导。我们将通过RNA干扰和小分子抑制所选靶点的基因沉默来评估葡萄膜黑色素瘤的细胞内定位、转录活性和ç-catenin的功能。在目标2中，我们将阐明ARF6在协调已知的致癌GαQ下游信号通路中的作用。目标1中使用的相同策略将被用来确定ARF6是否作为一个直接下游效应器，激活GNAQ/GNA11突变来控制(PLC-？/PKC)、TRIORhoA/RAC1通路以及AP-1和YAP-tead介导的转录。在目标3中，我们将通过人葡萄膜黑色素瘤的原位异种移植模型来评估ARF6在肿瘤建立和生长中的体内功能。这些目标的成功完成将使我们能够确定ARF6是否在GQ信号转导中发挥关键作用，从而为开发可用于治疗葡萄膜黑色素瘤和可能用于治疗其他GαQ相关疾病的药物提供一个有前途的治疗靶点，如Sturge-Weber综合征。