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Serine Protease Modulation of ErbB Receptors

ErbB 受体的丝氨酸蛋白酶调节

基本信息

批准号：
8099942
负责人：
KARL X CHAI
金额：
$ 31.14万
依托单位：
UNIVERSITY OF CENTRAL FLORIDA
依托单位国家：
美国
项目类别：
财政年份：
2011
资助国家：
美国
起止时间：
2011-04-01 至 2015-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8099942
关键词：
Address Aggressive behavior American Anchorage-Independent Growth Breast Cancer Cell Cancer Patient Cancer cell line Cell Line Cells Cleaved cell Clinical Combined Modality Therapy Complementary DNA Data Development Diagnosis Distant Metastasis Drug Delivery Systems Drug resistance Effectiveness Engineering Environment Enzymes Evaluation Extracellular Domain Functional disorder Goals Her2/erbb2/neu Staining Method Human In Vitro Intervention Investigation Laboratories Malignant Neoplasms Maps Mediating Membrane Modeling Monoclonal Antibodies Outcome Patients Peptide Hydrolases Pharmaceutical Preparations Phenotype Physiology Plasmin Plasminogen Inactivators Play Prevention Protease Inhibitor Receptor Protein-Tyrosine Kinases Research Resistance Roche brand of trastuzumab Role Serine Protease Signal Transduction Site Subfamily lentivirinae Testing Transfection Trastuzumab Tyrosine Kinase Inhibitor Up-Regulation Urokinase Urokinase Plasminogen Activator Receptor Woman chemotherapy clinically relevant effective therapy established cell line extracellular hepsin human PRSS8 protein inhibitor/antagonist lapatinib malignant breast neoplasm matriptase meetings migration novel prevent receptor research study resistance mechanism small molecule success treatment strategy tumor vector

项目摘要

DESCRIPTION (provided by applicant): The Her2+/uPA+ breast cancers are clinically the most aggressive, more so than the Her2+/uPA-, Her2-/uPA+, and Her2-/uPA- subtypes. Proteolytic modulation of Her2/neu/ErbB2 receptor tyrosine kinase signaling via extracellular domain/ectodomain (ECD) shedding by the urokinase-type plasminogen activator (uPA) and a novel network of extracellular membrane serine proteases is hypothesized to play a mechanistic role in the aggressive behaviors of the Her2+/uPA+ breast cancers and in resistance to anti-Her2 therapies using monoclonal antibody or small-molecule tyrosine kinase inhibitor drugs. Targeting Her2 and uPA simultaneously in Her2+/uPA+ breast cancers could therefore be a more effective strategy of treating these cancers; and potentially all Her2+ breast cancers. This hypothesis will be tested in this project. With the following specific aims, we will be creating novel cell-line models of Her2+/uPA+ breast cancer and testing the effectiveness of a novel treatment strategy by targeting Her2 and uPA simultaneously. We will also be investigating novel functional roles of several extracellular membrane serine proteases associated with breast cancer, and many other cancers. Specific Aim 1. To establish Her2+/uPA+ human breast cancer cell lines for evaluation of enhanced aggressiveness. We will establish Her2+/uPA+ human breast cancer cell lines by over-expressing uPA/uPAR in the Her2-amplified SK-BR-3 and BT-474 cell lines or over-expressing Her2 in the uPA+ but Her2-negative (unamplified) MDA-MB-231 cell line; and perform in vitro evaluations of tumor aggressiveness in proliferation, migration, invasion, and anchorage-independent growth. Specific Aim 2. To treat Her2+/uPA+ human breast cancer cell lines with Herceptin, lapatinib, and WX-UK1 for evaluation of effectiveness of targeting Her2 and uPA together. With our engineered Her2+/uPA+ model cell lines, the following clinically relevant questions will be addressed: 1). whether Her2+/uPA+ breast cancers present de novo drug resistance to Herceptin or even lapatinib and whether uPA inhibition with a selective uPA inhibitor WX-UK1 overcomes the resistance; 2) whether Her2+/uPA+ breast cancers acquire resistance to anti- Her2 therapies more aggressively and whether uPA inhibition with WX-UK1 delays the acquisition of resistance. Specific Aim 3. To determine the specific actions of matriptase, prostasin, and uPA on Her2 ECD. We will use the FT-293 cell line in co-transfection experiments to determine if uPA is a Her2 ECD sheddase and if plasmin plays a role in Her2 ECD shedding; to purify protease-cleaved Her2 for mapping the cleavage sites; and to determine if the protease-cleaved Her2 is responsive to Herceptin-induced turnover. We will use the SK-BR-3 cell line to determine if the Her2 ECD shedding associated with uPA induction in breast cancer cells also involves or requires matriptase and prostasin by using specific inhibitors. PUBLIC HEALTH RELEVANCE: Resistance to chemotherapy drugs targeting Her2/neu/ErbB2 limits the success of these drugs on select groups of breast cancer patients and could render patients with acquired resistance untreatable. Proteolytic shedding of the Her2 extracellular domain/ectodomain (ECD) by proteases is a mechanism of this resistance. Our preliminary studies have identified new candidate proteases involved in Her2 ECD shedding and the proposed research will reveal if specific protease inhibitors should be considered for combination therapy to reduce or prevent the resistance to the anti-Her2 drugs.

描述（由申请人提供）：Her2+/uPA+ 乳腺癌在临床上是最具侵袭性的，比 Her2+/uPA-、Her2-/uPA+ 和 Her2-/uPA- 亚型更具侵袭性。尿激酶型纤溶酶原激活剂 (uPA) 和新型细胞外膜丝氨酸蛋白酶网络通过胞外域/胞外域 (ECD) 脱落对 Her2/neu/ErbB2 受体酪氨酸激酶信号传导进行蛋白水解调节，推测在 Her2+/uPA+ 乳腺癌的侵袭行为和使用单克隆抗体或小分子酪氨酸激酶抑制剂药物的抗 Her2 疗法产生耐药性。因此，在 Her2+/uPA+ 乳腺癌中同时靶向 Her2 和 uPA 可能是治疗这些癌症的更有效策略；以及可能所有 Her2+ 乳腺癌。这个假设将在这个项目中得到检验。为了实现以下具体目标，我们将创建 Her2+/uPA+ 乳腺癌的新型细胞系模型，并通过同时靶向 Her2 和 uPA 来测试新型治疗策略的有效性。我们还将研究与乳腺癌和许多其他癌症相关的几种细胞外膜丝氨酸蛋白酶的新功能作用。具体目标 1. 建立 Her2+/uPA+ 人乳腺癌细胞系，用于评估增强的侵袭性。我们将通过在Her2扩增的SK-BR-3和BT-474细胞系中过表达uPA/uPAR或在uPA+但Her2阴性（未扩增）的MDA-MB-231细胞系中过表达Her2来建立Her2+/uPA+人乳腺癌细胞系；并对肿瘤在增殖、迁移、侵袭和不依赖贴壁的生长方面的侵袭性进行体外评估。具体目标 2. 用赫赛汀、拉帕替尼和 WX-UK1 处理 Her2+/uPA+ 人乳腺癌细胞系，以评估同时靶向 Her2 和 uPA 的有效性。通过我们设计的 Her2+/uPA+ 模型细胞系，将解决以下临床相关问题：1)。 Her2+/uPA+ 乳腺癌是否对赫赛汀甚至拉帕替尼产生新的耐药性，以及用选择性 uPA 抑制剂 WX-UK1 抑制 uPA 是否可以克服耐药性； 2) Her2+/uPA+乳腺癌是否会更积极地获得抗Her2疗法的耐药性，以及WX-UK1抑制uPA是否会延迟耐药性的获得。具体目标 3. 确定 matriptase、prostasin 和 uPA 对 Her2 ECD 的具体作用。我们将在共转染实验中使用 FT-293 细胞系来确定 uPA 是否是 Her2 ECD 脱落酶以及纤溶酶是否在 Her2 ECD 脱落中发挥作用；纯化蛋白酶切割的 Her2 以绘制切割位点图谱；并确定蛋白酶切割的 Her2 是否对赫赛汀诱导的周转有反应。我们将使用 SK-BR-3 细胞系，通过使用特定的抑制剂来确定与乳腺癌细胞中 uPA 诱导相关的 Her2 ECD 脱落是否也涉及或需要 matriptase 和 prostasin。公共卫生相关性：针对 Her2/neu/ErbB2 的化疗药物的耐药性限制了这些药物在特定乳腺癌患者群体中的成功，并可能使获得性耐药的患者无法治疗。蛋白酶对 Her2 胞外域/胞外域 (ECD) 的蛋白水解脱落是这种耐药性的机制之一。我们的初步研究已经确定了参与 Her2 ECD 脱落的新候选蛋白酶，拟议的研究将揭示是否应考虑使用特定蛋白酶抑制剂进行联合治疗，以减少或预防抗 Her2 药物的耐药性。