THROMBOSPONDIN IN BREAST CANCER INVASION & ANGIOGENESIS

乳腺癌侵袭中的血小板反应蛋白

• 批准号: 2113754
• 负责人: GEORGE Paul TUSZYNSKI
• 金额: $ 15.77万
• 依托单位: ALLEGHENY UNIVERSITY OF HEALTH SCIENCES
• 依托单位国家: 美国
• 财政年份: 1995
• 资助国家: 美国
• 起止时间: 1995-05-23 至 1997-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2113754
• 关键词: angiogenesis; breast neoplasms; cell type; drug design /synthesis /production; extracellular matrix; fibroblasts; human tissue; laboratory rat; messenger RNA; metastasis; neoplastic process; northern blottings; nucleic acid sequence; plasminogen activator; smooth muscle; stromal cells; synthetic peptide; thromboplastin; transforming growth factors; vascular endothelium

Thrombospondin (TSP) is among the matrix molecules that have been implicated in angiogenesis. We recently reported that the extracellular matrix of breast cancer contains large amounts of TSP and that matrix- bound TSP promotes angiogenesis from vascular explants. TSP also induces and stabilizes capillary-like tubes formed by endothelial cells. In addition, we have shown that TSP promotes tumor cell invasion and up- regulates breast cancer cell production of plasminogen activator inhibitor-1 (PAI-1) and urokinase-type plasminogen activator (uPA) through a mechanism involving transforming growth factor beta-1 (TGF-beta). In this grant we will test the hypothesis that stromal cell-derived TSP promotes both breast cancer invasion and angiogenesis by modulating the activity of TGF-beta which in turn up-regulates production of PAI-1 and uPA. We will determine which stromal cells produce TSP in response to breast cancer cells by measuring TSP produced in tumor/stromal cell co- culture assays and by immunolocalizing TSP in tumor biopsies at the ultrastructural level. TSP will be measured by immunochemistry and Northern blot analysis. Functional involvement of stromal-derived TSP in tumor cell invasion will be assessed with specific antibodies against TSP and its CSVTCG-specific receptor using two different tumor/stromal cell co-culture assays. We will then determine the role of PAI-1 and uPA in TSP-induced tumor cell invasion and angiogenesis by evaluating the effect of anti-PAI-1 and uPA antibodies in our co-culture invasion and angiogenesis assays. We will also evaluate the effect of TSP on endothelial production of PAI-1 and uPA. To investigate how the TSP effects relate to TGF-beta, we will determine whether TSP up-regulates activation and/or production of TGF-beta by tumor and endothelial cells and whether TGF-beta in turn up-regulates TSP. These studies will be accomplished both at the protein and mRNA levels. We will also test the effect of anti-TGF-beta antibody on invasion and angiogenesis in TSP- treated cultures. Finally, we will design synthetic TSP antagonist peptides based on the TSP-adhesive amino acid sequences CSVTCG and CSTSCG and test their inhibitory activity in our angiogenesis and invasion assays. These studies will contribute to our long-term goal of developing new treatment for breast cancer based on the mechanisms whereby TSP modulates tumor cell invasion and angiogenesis.

血小板反应蛋白（TSP）是已经被发现的基质分子之一。 与血管生成有关。 我们最近报道细胞外 乳腺癌的基质中含有大量的TSP， 结合的TSP促进血管外植体的血管生成。 TSP还导致 并稳定由内皮细胞形成的毛细血管样管。 在 此外，我们已经表明TSP促进肿瘤细胞侵袭和上调- 调节乳腺癌细胞纤溶酶原激活物的产生 抑制剂-1（派-1）和尿激酶型纤溶酶原激活剂（uPA）， 一种涉及转化生长因子β-1（TGF-β）的机制。 在 本研究将验证基质细胞来源的TSP 通过调节乳腺癌的侵袭和血管生成来促进乳腺癌的侵袭和血管生成 TGF-β的活性，其反过来上调派-1的产生， uPA。 我们将确定哪些基质细胞产生TSP，以响应 乳腺癌细胞通过测量肿瘤/间质细胞共同产生的TSP， 培养测定和免疫定位肿瘤活检中的TSP， 超微结构水平 TSP将通过免疫化学法测量， 北方印迹分析。 间质来源的TSP在乳腺癌中的功能参与 肿瘤细胞的侵袭将用抗TSP的特异性抗体进行评估 及其CSVTCG特异性受体，使用两种不同的肿瘤/基质细胞 共培养测定。 然后我们将确定派-1和uPA在 通过评估TSP诱导肿瘤细胞侵袭和血管生成的作用 抗PAI-1和uPA抗体在我们的共培养侵袭中的表达， 血管生成测定。 我们亦会评估TSP对 内皮细胞产生派-1和uPA。 为了研究TSP是如何 影响与TGF-β有关，我们将确定TSP是否上调 肿瘤和内皮细胞活化和/或产生TGF-β 以及TGF-β是否反过来上调TSP。 这些研究报告将 在蛋白质和mRNA水平上完成。 我们还将测试 抗TGF-β抗体对TSP侵袭和血管生成影响- 处理的文化。 最后，我们将设计合成TSP拮抗剂， 基于TSP粘附氨基酸序列CSVTCG和CSTSCG的肽 并测试它们在血管生成和侵袭中的抑制活性 分析。 这些研究将有助于实现我们的长期目标， 基于TSP机制的乳腺癌新疗法 调节肿瘤细胞侵袭和血管生成。