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

描述（由申请人提供）：本提案的目的是确定乳腺癌患者中表皮生长因子受体（EGFR）产生的新型信号传导途径的利用，该途径介导药物治疗耐药性，并确定该途径的下游靶点。我们最近的数据表明，配体激活的EGFR可能通过转移到线粒体并结合线粒体酶考克斯II（细胞色素c氧化酶II）而导致耐药性。考克斯II是结合细胞色素c的电子传递链的关键组分。据推测，EGFR的结合增强了线粒体中的考克斯II活性和细胞色素c的保留，从而减少了药物诱导的细胞凋亡。考克斯II结合EGFR的磷酸-Tyr 845（pY 845），这是EGF处理后被c-Src磷酸化的新位点。衔接蛋白p130 Cas（Cas）可能通过激活c-Src和促进Y845的磷酸化而独立于配体在此过程中发挥作用。Y845的突变对受体的催化活性没有影响，但消除了EGF诱导的DNA合成和EGF介导的乳腺癌细胞在药物治疗后的存活。目前尚不清楚考克斯II与pY 845的结合如何影响考克斯II活性或线粒体的其他性质。这种机制所适用的全部治疗剂也是未知的。将采取三种办法来解决这些问题。首先，我们将研究选定的乳腺癌细胞系，诱导或瞬时表达野生型或突变型Y845 F EGFR的敏感性，目前在临床使用的药物面板。预期表达wt EGFR的细胞将表现出增强的抗性，而表达突变受体的细胞将更敏感。还将在这些相同细胞系的背景下检查源自pY 845和雌激素受体（ER）的途径之间的串扰。其次，我们将测量在表达野生型或突变型EGFR的细胞中药物治疗后线粒体组分和功能的改变。最后，我们将探讨的可能性，通过其激活c-Src的能力，Cas可以促进磷酸化的Y845独立，或与EGF。在每种情况下，将培养细胞和异种移植肿瘤的结果与人类乳腺癌样本的结果进行比较。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，但每年约有40，000名妇女死于转移性或复发性疾病。许多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有抵抗力。EGFR家族在耐药性中起着重要作用，我们假设通过EGFR上的pY 845的生存信号传导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pY 845依赖性的耐药性，以便我们可以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.