CD44/VARIANT CYTOSKELETON IN BREAST CANCER PROGRESSION

乳腺癌进展中的 CD44/变异细胞骨架

• 批准号: 6376129
• 负责人: Lilly YW Bourguignon
• 金额: $ 0.74万
• 依托单位: UNIVERSITY OF MIAMI SCHOOL OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 1995
• 资助国家: 美国
• 起止时间: 1995-04-01 至 2001-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6376129
• 关键词: CD44 molecule; ankyrins; antisense nucleic acid; athymic mouse; biological signal transduction; breast neoplasms; gene expression; genetic markers; guanine nucleotide exchange factors; guanosinetriphosphatases; human tissue; immunocytochemistry; intermolecular interaction; laboratory rabbit; metastasis; neoplasm /cancer genetics; neoplastic process; oncoproteins; protein isoforms; protein structure function; site directed mutagenesis; tissue /cell culture

In this continuation research proposal, we plan to test the hypothesis that transmembrane interaction between CD44 variants (CD44V) and the cytoskeletal protein, ankyrin, plays an important role in promoting onocongenic [e.g. Tiam1-catalyzed RhoA activation and Rho-Kinase (ROK)-mediated cytoskeleton function] leading to tumor cell adhesion growth. invasion and migration during breast cancer progression. The results of this research will definitely provide a better understanding of the cellular and molecular mechanisms involved in human breast cancer metastasis and progression. Specifically, we plan to use a variety of biochemical, cell biological and molecular biological techniques to elucidate the binding site diversity on the cytoskeletal protein, ankyrin, required for CD44 variant (CD44V) association. An emphasis will be placed on the structural and functional relationships between the CD44V isoforms and the ankyrin repeat domain (ARD) in metastatic breast tumor cells. We will also explore a new CD44V and ankyrin-linked oncogenic signaling pathway with a special focus on the guanine nucleotide exchange factor, Tiam1 (T lymphoma invasion and metastasis)-catalyzed RhoA signaling and Rho-Kinase (ROK) activation in metastatic breast tumor cells. In addition, we will employ a novel signaling perturbation strategy to impair Tiam1-RhoA signaling and/or ROK-specific cytoskeleton function by first constructing various Tiam1 and/or ROK deletion and site-directed mutants lacking specific functional domains (e.g. dominant negative mutants). These dominant-negative Tiam1 and/or ROK mutant polypeptides will then be expressed in metastatic breast epithelial cells to test their ability to downregulate CD44V isoform-mediated metastatic tumor cell behaviors (e.g. tumor cell adhesion growth, migration and invasion). In the second part of the proposal, we will use immunohistochemistry to analyze the coexpression of CD44 variants (e.g. CD44V3 and CD44V10-containing isoforms) with ankyrin and certain oncogenic signaling molecules (e.g. Tiam1 or ROK) in human breast carcinomas in order to establish useful structure/function-related markers for breast cancer detection and prognosis. Finally, we plan to establish a therapeutic antisense strategy to specifically inhibit the mRNA encoding for certain CD44 isoforms (e.g. CD44V3/V10) in order to effectively block the expression of CD44V isoforms and their downstream oncogenic signaling events which lead to metastatic breast tumor progression. We believe the results of these proposed experiments will provide a much better understanding of the CD44V isoform interaction with ankyrin which regulates Tiam1-RhoA signaling and ROK activation required for cytoskeleton function and various metastatic tumor cell properties (e.g. tumor cell adhesion, growth, invasion and migration). The information obtained from this proposal may establish the coexpression of CD44V (e.g. CD44V3/CD44V 10-containing isoforms) with ankyrin and certain oncogenic signaling molecules (e.g. Tiam1 or ROK) as an important tumor marker for early detection and evaluation of oncogenic potential. Most importantly, our proposed new strategy using a novel Tiam1/ROK-related signaling perturbation, and specific antisense constructs of CD44V3/V10 isoforms together with certain oncogenic molecules, may identify new drug targets for the inhibition of breast tumor metastasis and cancer progression.

在这项持续研究建议中，我们计划检验以下假设：CD44变体（CD44V）和细胞骨架蛋白Ankyrin之间的跨膜相互作用在促进ancongonic中起着重要作用[例如。 TIAM1催化的RhoA激活和Rho--激酶（ROK）介导的细胞骨架功能，导致肿瘤细胞粘附生长。乳腺癌进展过程中的入侵和迁移。 这项研究的结果肯定会更好地了解与人类乳腺癌转移和进展有关的细胞和分子机制。 具体而言，我们计划使用各种生化，细胞生物学和分子生物学技术来阐明CD44变体（CD44V）关联所需的细胞骨架蛋白上的结合位点多样性。 将重点放在CD44V同工型与转移性乳腺肿瘤细胞中的Ankyrin重复结构域（ARD）之间的结构和功能关系上。 我们还将探索一种新的CD44V和氨基氨基蛋白连接的致癌信号通路，特别关注鸟嘌呤核苷酸交换因子TiAM1（T淋巴瘤侵袭和转移）催化的RhoA信号传导和Rho-kinase（ROK）和Rho-Kinase（ROK）激活。 此外，我们将采用一种新型的信号扰动策略来损害TIAM1-RHOA信号传导和/或ROK特异性细胞骨架功能，首先构建各种TIAM1和/或ROK缺失和缺乏特定功能域的位置指导突变体（例如，显性负突变体）。 然后将在转移性乳房上皮细胞中表达这些显性阴性TIAM1和/或ROK突变多肽，以测试其下调CD44V同工型介导的转移性肿瘤细胞行为的能力（例如肿瘤细胞粘附生长，迁移，迁移和入侵）。在该提案的第二部分中，我们将使用免疫组织化学分析与ANKYRIN的CD44变体（例如CD44V3和CD44V10含量的同工型）的共表达，并在人类乳腺癌中以癌症的癌症和癌症来建立有用的标志物（例如，TIAM1或ROK）（例如TIAM1或ROK）在有用的癌症中，以供您进行有用的标志。 最后，我们计划建立一种治疗性反义策略，以特异性抑制针对某些CD44同工型（例如CD44V3/V10）编码的mRNA，以有效地阻止CD44V同工型及其降低的致癌信号事件的表达，从而导致乳房乳腺肿瘤的转移性乳房肿瘤进展。我们认为，这些提出的实验的结果将为CD44V同工型与Ankyrin的相互作用提供更好的理解，该相互作用调节了TIAM1-RHOA信号传导和细胞骨架功能所需的ROK激活，以及各种转移性肿瘤细胞的特性（例如，肿瘤细胞粘附，生长，入侵，入侵和移植）。 从该提案中获得的信息可以建立用脚踝蛋白的CD44V（例如CD44V/CD44V 10含同工型）的共表达，并且某些致癌信号分子（例如TIAM1或ROK）是重要的肿瘤标记物，以早日检测和评估致癌潜力。最重要的是，我们提出的使用新型TIAM1/ROK相关的信号扰动以及CD44V3/V10同工型的特定反义构建体以及某些致癌分子的特定反义构建体可能会确定抑制乳腺肿瘤转移和癌症进展的新药物。