Identification and Characterization of a Endogenous EGFR Regulatory Locus in Xiphophorus Genome

项目摘要： 表皮生长因子受体（EGFR）是一种重要的癌基因， 与多种癌症有关。抗EGFR小分子和 已经开发出单克隆抗体来阻断其激酶活性， 治疗然而，先天性和获得性抗性经常在 临床应用这些观察结果显著限制了抗EGFR药物 使用，也挑战了EGFR驱动癌症的现有知识。因此，我们认为， 因此，有必要从多个角度研究EGFR与肿瘤的关系， 不同的研究视角，不同的研究策略。 斑剑尾鱼编码一种突变的，自主的，失调的， 致癌EGFR，命名为xmrk。然而，致癌作用仅在 Xmrk阳性与Xmrk阴性剑尾鱼种间杂种回交 种，或者当xmrk在非剑尾鱼模型系统中异位表达时 (e.g.,青鳉，鼠）。这表明X. maculatus基因组还编码一种 调节基因，可能与xmrk共同进化，并能够抑制其 致癌活性因此，表征调节器，称为R（Diff） 抑制xmrk可能导致控制EGFR新的治疗策略。 我们最近的研究已经将R（Diff）肿瘤调节位点确定为101.7 kbp， 位于5号染色体上。这种小的候选大小和低的候选基因 数字使功能和机制的研究是不切实际的。因此，我们认为， 该建议旨在通过检查 以下目标： 目标1：我们将描述所有表达但未注释的遗传元件， 除了已知的编码基因，在新确定的候选R（Diff） 轨迹，以完全注释R（Diff）轨迹。 目的2：我们将通过以下方法确定携带R（Diff）活性的基因/元件： 在非荷瘤剑尾鱼中进行基因敲除， 杂种和携带肿瘤的转基因青鳉。 目标3：我们将分析xmrk和R（Diff）候选者，以及 几个剑尾鱼杂种，通过候选者的关联来定义R（Diff） 具有不同表型的序列。

Project Summary: The epidermal growth factor receptor (EGFR) is a leading oncogene firmly associated with many types of cancer. Both anti-EGFR small molecules and monoclonal antibodies have been developed to block its kinase activity for cancer treatment. However, innate and acquired resistance are frequently observed in clinical application. Such observations significantly limit anti-EGFR medicines usage, and also challenge current knowledge of EGFR-driver cancer. Therefore, it is necessary to study the relationship between EGFR and cancer from a different angle, with different research strategy. Xiphophorus maculatus encodes a mutant, autonomous, dysregulated and oncogenic EGFR, named xmrk. However, carcinogenesis is only observed in backcross interspecies hybrid between xmrk positive and xmrk-null Xiphophorus species, or when xmrk is ectopically expressed in non-Xiphophorus model system (e.g., medaka, murine). These suggest that X. maculatus genome also encode a regulator gene that may co-evolved with xmrk and is able to suppressing its oncogenic activity. Therefore, characterizing how the regulator, termed R(Diff) inhibit xmrk, may lead to novel therapeutic strategy in controlling EGFR. Our recent study has defined the R(Diff) tumor regulatory locus to a 101.7 kbp locus on chromosome 5. This small candidate size and low candidate gene number enables functional and mechanistic studies are impractical. Therefore, this proposal is designed to final determine the R(Diff) locus by examining the following aims: Aim 1: We will characterize all expressed but unannotated genetic elements, in addition to the known coding genes, within the newly identified candidate R(Diff) locus, to fully annotate the R(Diff) locus. Aim 2: We will determine the gene/element(s) carrying R(Diff) activity by performing gene knock-out in non-tumor-bearing Xiphophorus fish, tumor-bearing hybrids, and tumor-bearing transgenic medaka. Aim 3: We will profile xmrk and R(Diff) candidates, as well as phenotypes of several Xiphophorus hybrids, to define R(Diff) by association of candidate sequence with varied phenotypes.