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

描述（由申请人提供）：本提案的目标是确定乳腺癌患者对介导药物治疗耐药性的表皮生长因子受体（EGFR）发出的新型信号通路的利用，并确定该通路的下游靶点。我们最近的数据表明，配体激活的EGFR可能通过转运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。Cox II是结合细胞色素c的电子传递链的关键组成部分。据推测，EGFR的结合增强了Cox II的活性和线粒体中细胞色素c的保留，从而减少了药物诱导的细胞凋亡。Cox II结合EGFR的phospho-Tyr 845 (pY845)，这是一个在EGF处理后被c-Src磷酸化的新位点。适配蛋白p130Cas （Cas）可能通过激活c-Src并独立于配体促进Y845的磷酸化而在这一过程中发挥作用。Y845的突变对受体的催化活性没有影响，但会破坏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Cox II与pY845结合如何影响Cox II活性或线粒体的其他特性。这一机制适用的治疗剂的全部种类也尚不清楚。将采取三种方法来解决这些问题。首先，我们将检查选定的乳腺癌细胞系，诱导或瞬时表达wt或突变型Y845F EGFR，以检测它们对目前临床使用的一组药物的敏感性。我们预计，表达wt型EGFR的细胞会表现出更强的耐药性，而表达突变受体的细胞会更敏感。从pY845和雌激素受体（ER）发出的通路之间的串扰也将在这些相同细胞系的背景下进行研究。其次，我们将在表达wt或突变EGFR的细胞中测量药物治疗后线粒体成分和功能的变化。最后，我们将探索通过其激活c-Src的能力，Cas可以独立或协同EGF促进Y845的磷酸化的可能性。在每种情况下，培养细胞和异种移植肿瘤的结果将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。许多人治疗失败，因为他们的肿瘤对细胞毒性或细胞抑制剂的作用有抵抗力。EGFR家族在耐药性中发挥重要作用，我们假设通过pY845在EGFR上传递的生存信号是这一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依赖的治疗抗性，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.