TUMOR CELL ADHESIVE GLYCOPROTEINS AND METASTASIS

随着原发部位控制和复发控制的最新进展 疾病，转移性疾病仍然是阻碍 改善癌症治疗。转移级联是一个复杂的多步骤 肿瘤间同型和异型相互作用的过程 细胞和宿主细胞，除了肿瘤细胞与基质的黏附 蛋白质。肿瘤细胞具有多种表型特征 这使它们能够完成转移过程并形成新的病变。 然而，转移级联的某些步骤可能是速率限制的，并且 可能是治疗干预的目标。大部分(如果不是全部)步骤 转移级联涉及细胞表面受体，如 整合素。许多整合素黏附受体已被发现在 各种人和啮齿动物肿瘤细胞系。其中，有Vitronectin 受体α-v-β-3和α-II-b-β-3。α-II-的表达 B-β-3最初被认为仅限于血小板和 巨核细胞系细胞。最近，我们已经演示了，由Southern 印迹、Northern印迹和免疫沉淀，α-蛋白的存在 II-b-β-3在几种人和小鼠肿瘤系中的表达。这项研究旨在 测定肿瘤细胞α-II-b和β-3的基因序列 逆转录合成第一链c DNA，并利用 聚合酶链式反应技术扩增α-II-b和β-3 并对扩增产物进行测序。接下来，α-II-b-β-3受体 与指示变形潜力的表型性状有关，如 在内皮、血小板和肺集落上黏附和扩散 将确定编队，试图建立一个可能的 肿瘤细胞中α-II-β-3的表达与肿瘤的关系 转移。为了进一步确定α-II-β-3在小鼠肿瘤中的作用 用抗-α-II-b和/或抗-α-B基因转染细胞系。 Beta-3结构。将对α-II-b-β-3阴性细胞进行检测 它们经历转移所必需的相互作用的能力(例如， 血小板聚集、黏附、扩散和肺部定植)。 此外，细胞粘附素整合素的相对贡献(即， α-II-b-β-3和α-v-β-3)对肿瘤细胞的黏附和 扩散于基质配体和内皮细胞，肿瘤细胞诱导的血小板 将对聚合进行研究。一种表达α-II-b的肿瘤细胞系 但不包括α-v(即B16无色素性黑色素瘤)和另一种细胞系 (Lewis肺癌)同时表达α-II-b和α-v的Will 用于比较每种整合素对转移的相对贡献 潜力。针对α-II-b和/或α-V的反义结构 在这些对比实验中将被引入这些细胞系 在黏附、血小板聚集和肺集落形成试验前。在……里面 此外，细胞因子转化生长因子-β、肿瘤坏死因子和白介素1， 以及二十烷类12-羟基二十碳四烯酸，它们已经被 证明对某些整合素的表达有调节作用，将进行研究 它们对转录和/或翻译控制的影响 II-b-β-3和α-v-β-3的表达及其功能。 细胞因子和/或二十烷类化合物对整合素功能的影响可能通过 磷酸化或增加它们与细胞的结合 细胞骨架。α-II-b、α-v和β-β的磷酸化模式 3，并将检查受体与细胞骨架的关联。在……里面 这个提案，我们将综合运用包括分子， 生化和细胞生物学技术研究亲缘关系 细胞粘附素整合素(即α-II-b-β-3和α-β-3)的作用 V-β-3)与高转移潜能相关的细胞功能。

With the recent advances in primary site control and control of recurrent disease, metastatic disease remains one of the principle impediments to improved cancer treatment. The metastatic cascade is a complex multi-step process which involves homotypic and heterotypic interactions among tumor cells and host cells, in addition to tumor cell adhesion to matrix proteins. Tumor cells possess a variety of phenotypic characteristics which enable them to complete the metastatic process and form a new lesion. However, some steps of the metastatic cascade may be rate limiting and possibly targeted for therapeutic intervention. Most, if not all, steps of the metastatic cascade involve cell surface receptors such as the integrins. A number of integrin adhesion receptors have been found on various human and rodent tumor cell lines. Among them, are the vitronectin receptor alpha-v-beta-3 and alpha-II-b-beta-3. The expression of alpha-II- b-beta-3 was originally thought to be confined to platelets and megakaryocyte lineage cells. Recently, we have demonstrated, by Southern blotting, Northern blotting and immunoprecipitation, the presence of alpha- II-b-beta-3 in several human and murine tumor lines. This study is aimed to determine cDNA sequences of tumor cell alpha-II-b and beta-3 by synthesizing the first strand cDNA by reverse transcription and using the polymerase chain reaction technique to amplify the alpha-II-b and beta-3 cDNAs and sequence the PCR products. Next, the alpha-II-b-beta-3 receptor involvement in phenotypic traits indicative of metastic potential such as adhesion to and spreading on endothelium, platelets and lung colony formation will be determined, in an attempt to establish a possible correlation between alpha-II-beta-3 expression in tumor cells and metastasis. To further define the role of alpha-II-beta-3, murine tumor cell lines have been transfected with anti-alpha-II-b and/or anti-alpha- beta-3 constructs. The alpha-II-b-beta-3 negative cells will be tested for their ability to undergo interactions necessary for metastasis (e.g., platelet aggregation, adhesion, spreading and lung colonization). Moreover, the relative contribution of the cytoadhesin integrins (i.e., alpha-II-b-beta-3 and alpha-v-beta-3) to tumor cell adhesion to and spreading on matrix ligands and endothelium, tumor cell induced platelet aggregation will be studied. A tumor cell line which expresses alpha-II-b but not alpha-v (i.e., B 16 amelanotic melanoma) and another cell line (Lewis lung carcinoma) which expresses both the alpha-II-b and alpha-v will be used to compare the relative contribution of each integrin to metastatic potential. The anti-sense constructs against alpha-II-b and/or alpha-v will be introduced into these cell lines in these comparison experiments prior to adhesion, platelet aggregation and lung colony forming assays. In addition, the cytokines TGF-beta, tumor necrosis factor and interleukin 1, as well as the eicosanoid 12-hydroxyeicosatetraenoic acid, which have been demonstrated to regulate the expression of some integrins, will be studied for their effects on transcriptional and/or translational control of alpha- II-b-beta-3 and alpha-v-beta-3 expression and on their functions. Cytokines and/or eicosanoid effect on integrin function may be regulated by phosphorylation or increase in their association with the cellular cytoskeleton. The phosphorylation pattern of alpha-II-b, alpha-v and beta- 3 and the receptor association with cell skeleton will be examined. In this proposal, we will use a comprehensive approach including molecular, biochemical, and cell biological techniques to study the relative contribution of cytoadehesin integrins (i.e., alpha-II-b-beta-3 and alpha- v-beta-3 ) to cellular functions correlated with high metastatic potential.