INTEGRIN ACTIVITIES DURING TUMOR PROGRESSION BY GALECTIN-3

Galectin-3 在肿瘤进展过程中的整合素活性

• 批准号: 6485272
• 负责人: JOSIAH OCHIENG
• 金额: $ 17.91万
• 依托单位: MEHARRY MEDICAL COLLEGE
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-08-01 至 2002-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6485272
• 关键词: breast neoplasms; confocal scanning microscopy; extracellular matrix; flow cytometry; immunoprecipitation; integrins; laminin; lectin; neoplastic cell; neoplastic process; prostate neoplasms; tissue /cell culture; transfection

The proposal seeks to establish the significance of galectin-3 in integrin functions during malignant progression of breast and prostate carcinomas. The progression of tumors if normally accompanied by the alterations in the interactions of the transformed cells with extracellular matrix proteins, such as increased invasiveness and motility. These processes are regulated by integrins, which are in turn modulated by other cellular proteins. The ability of integrins to interact with galectin-3 prompted us to question the biological significance of this association. Specifically, we are interested in testing the hypothesis that the expression of galectin-3 is necessary for the modulation of the activities of integrins responsible for cell to extracellular matrix interactions. The study is designed to; a) determine the importance of galectin-3 in cell to extracellular matrix interactions, b) demonstrate that galaectin3 is a critical molecular partner of integrins which regulate their functions, c) to identify integrin heterodimers which associate with galectin-3 in the breast and prostate carcinomas and d) to analyze the interaction of galectin3- expressing and null expressing cell lines including those transfected with antisense oligos, with bone and lung extracellular matrices. Antisense DNA technologies will be employed to disrupt galectin-3 gene expression in the carcinoma cells. The interaction of cells with the diminished galectin- 3 expression (antisense transfectants) as well as sense and vector transfected controls with extracellular matrix proteins will then be monitored in vitro. It is expected that lack of galectin-3 expression will result in diminished interaction of these cells with the extracellular matrix proteins. The interactions between purified galectin-3 expression will result in diminished interaction of these cells with the extracellular matrix proteins. The interaction between purified galectin-3 and integrins, particularly alpha 1 beta1 will be analyzed by light scattering methodology to obtain binding parameters. Using fluorescence activated cell sorting techniques (FACS(, we will seek to determine whether galectin-3 alters the expression of specific cellular integrins. Using a simplified galectin-3/integrin binding assay, we will determine how galectin-3 modulates the binding interaction between alpha1 beta1 integrins and laminin-1. We will also determine how the modifications of the biological functions of galectin-3 affects its interaction with integrins and vice vera. Immunoprecipitation methodology and confocal laser microscopy will be used to identify integrins which are associated with galectin-3 on the surfaces of the breast and prostate epithelial tumor cells. Lastly, the adhesion potential of galectin-3 expressing and null expressing cells to bone extracellular matrix and to lung tissue secretions will be evaluated. Both breast and prostate carcinoma cells preferentially metastasize to the bones and to a lesser extent, the lungs. These studies will provide new avenues which may be exploited for the control of the dissemination and growth of metastatic breast and prostate tumor cells.

该提案旨在确定半乳糖凝集素-3在乳腺癌和前列腺癌恶性进展期间整合素功能中的重要性。肿瘤的进展通常伴随着转化细胞与细胞外基质蛋白相互作用的改变，如侵袭性和运动性增加。这些过程由整合素调节，而整合素又由其他细胞蛋白调节。整合素与半乳糖凝集素-3相互作用的能力促使我们质疑这种关联的生物学意义。具体而言，我们感兴趣的假设，半乳糖凝集素-3的表达是必要的负责细胞与细胞外基质相互作用的整合素的活动的调制。该研究旨在; a）确定半乳糖凝集素-3在细胞与细胞外基质相互作用中的重要性，B）证明半乳糖凝集素3是调节其功能的整联蛋白的关键分子伴侣，c）鉴定在乳腺癌和前列腺癌中与半乳糖凝集素-3缔合的整联蛋白异二聚体，和d）分析半乳糖凝集素3表达和无效表达细胞系（包括用反义寡核苷酸转染的那些）的相互作用，与骨和肺细胞外基质混合反义DNA技术将被用来破坏癌细胞中galectin-3基因的表达。然后将在体外监测半乳糖凝集素-3表达减少的细胞（反义转染子）以及正义和载体转染的对照与细胞外基质蛋白的相互作用。预期半乳糖凝集素-3表达的缺乏将导致这些细胞与细胞外基质蛋白的相互作用减少。纯化的半乳糖凝集素-3表达之间的相互作用将导致这些细胞与细胞外基质蛋白的相互作用减弱。将通过光散射方法分析纯化的半乳糖凝集素-3和整联蛋白，特别是α 1 β 1之间的相互作用，以获得结合参数。使用荧光激活细胞分选技术（FACS），我们将试图确定半乳糖凝集素-3是否改变特定细胞整合素的表达。使用简化的半乳糖凝集素-3/整合素结合试验，我们将确定半乳糖凝集素-3如何调节α 1 β 1整合素和层粘连蛋白-1之间的结合相互作用。我们还将确定半乳糖凝集素-3的生物学功能的修饰如何影响其与整合素和副vera的相互作用。免疫沉淀方法和共聚焦激光显微镜将用于鉴定与乳腺和前列腺上皮肿瘤细胞表面上的半乳糖凝集素-3相关的整合素。最后，将评价半乳糖凝集素-3表达和无效表达细胞对骨细胞外基质和肺组织分泌物的粘附潜力。乳腺癌和前列腺癌细胞都优先转移到骨骼，其次转移到肺部。这些研究将为控制转移性乳腺和前列腺肿瘤细胞的扩散和生长提供新的途径。