c-Src/EGF Receptors Interactions and Therapeutic Resistance in Breast Cancer

描述(申请人提供)：这项建议的目标是确定乳腺癌患者使用一种新的信号通路，该通路源于表皮生长因子受体(EGFR)，它介导了药物治疗的耐药性，并确定了该通路的下游靶点。我们最近的数据表明，配体激活的EGFR可能通过移位到线粒体并与线粒体酶COX II(细胞色素C氧化酶II)结合而导致耐药性。环氧合酶II是结合细胞色素c的电子传递链的关键组成部分，推测与EGFR的结合可增强环氧合酶II的活性和细胞色素c在线粒体中的滞留，从而减少药物诱导的细胞凋亡。COX II结合了EGFR的磷酸酪氨酸845(PY845)，这是一个新的位点，在EGF处理后被c-Src磷酸化。连接蛋白p130Cas(Cas)可能通过激活c-Src和非依赖于配体的方式促进Y845的磷酸化而在此过程中发挥作用。Y845突变对受体的催化活性没有影响，但阻断了EGF诱导的DNA合成和药物治疗后EGF介导的乳腺癌细胞的存活。目前尚不清楚COX II与pY845的结合如何影响COX II活性或线粒体的其他特性。这一机制适用的全部治疗剂也是未知的。将采取三种方法来解决这些问题。首先，我们将检查选定的可诱导或瞬时表达wt或突变的Y845F EGFR的乳腺癌细胞株对目前临床使用的一组药物的敏感性。预计表达wt EGFR的细胞将表现出增强的抵抗力，而表达突变受体的细胞将更加敏感。从pY845发出的通路和雌激素受体(ER)之间的串扰也将在这些相同的细胞系的背景下进行研究。其次，我们将测量药物治疗后表达wt或突变EGFR的细胞中线粒体成分和功能的变化。最后，我们将探索Cas通过其激活c-Src的能力，独立于或协同EGF促进Y845的磷酸化的可能性。在每一种情况下，培养细胞和异种移植瘤的结果都将与人类乳腺癌样本进行比较。相关性：尽管最近在预防和治疗乳腺癌方面取得了重大进展，但每年仍有大约40,000名妇女死于转移性或复发性疾病。许多人的治疗失败是因为他们的肿瘤对细胞毒性或细胞抑制药物的作用具有抵抗力。EGFR家族在耐药中发挥着重要作用，我们推测，通过EGFR上的pY845发出的生存信号是该过程的关键组成部分。我们的研究旨在更好地了解哪些癌症表现出pY845依赖的治疗耐药，以便我们能够针对这一途径进行干预。

DESCRIPTION (provided by applicant): The goals of this proposal are to determine in breast cancer patients the utilization of a novel signaling pathway emanating from the epidermal growth factor receptor (EGFR) that mediates resistance to drug therapy and to identify the downstream targets of this pathway. Our recent data suggest that ligand-activated EGFR may contribute to drug resistance by translocating to the mitochondria and binding to the mitochondrial enzyme, Cox II (cytochrome c oxidase II). Cox II is a key component of the electron transport chain that binds cytochrome c. It is postulated that binding of EGFR enhances Cox II activity and retention of cytochrome c in the mitochondria, thus reducing drug-induced apoptosis. Cox II binds phospho-Tyr 845 (pY845) of the EGFR, a novel site that is phosphorylated by c-Src following EGF treatment. The adaptor protein, p130Cas (Cas), may play a role in this process by activating c-Src and promoting phosphorylation of Y845 independently of ligand. Mutation of Y845 has no effect on the catalytic activity of the receptor but ablates EGF-induced DNA synthesis and EGF-mediated survival of breast cancer cells following drug treatment. It is not currently known how Cox II binding to pY845 affects Cox II activity or other properties of the mitochondria. The full array of therapeutic agents for which this mechanism applies is also not known. Three approaches will be taken to address these questions. First, we will examine selected breast cancer cell lines that inducibly or transiently express wt or mutant Y845F EGFR for their sensitivity to a panel of drugs that are currently in clinical use. It is expected that cells expressing the wt EGFR will exhibit enhanced resistance, while those expressing the mutant receptor will be more sensitive. Crosstalk between pathways emanating from pY845 and the estrogen receptor (ER) will also be examined in the context of these same cell lines. Second, we will measure alterations in mitochondrial components and functions following drug treatment in cells expressing either the wt or mutant EGFR. Finally, we will explore the possibility that, through its ability to activate c-Src, Cas can promote phosphorylation of Y845 independently of, or in concert with, EGF. In each case, findings from cultured cells and xenograft tumors will be compared to those of human breast cancer samples. Relevance: In spite of significant, recent advances in prevention and treatment of breast cancer, approximately 40,000 women succumb to metastatic or recurrent disease each year. Many fail therapy because their tumors are resistant to the action of cytotoxic or cytostatic drugs. The EGFR family plays an important role in resistance, and we hypothesize that survival signaling through pY845 on the EGFR is a key component of the process. Our studies aim to achieve a better understanding of which cancers demonstrate pY845-dependent resistance to therapy, so that we can target this pathway for intervention.