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

项目总结： 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 已经开发出单抗来阻断其在癌症中的激酶活性 治疗。然而，先天和后天的抵抗力经常被观察到 临床应用。这些观察结果大大限制了抗EGFR药物的使用 用法，也挑战了目前对EGFR驱动癌症的认识。因此， 从分子水平研究EGFR与肿瘤的关系十分必要。 不同的视角，不同的研究策略。 剑尾鱼编码一种突变体，自治的，失调的和 致癌的EGFR，命名为xmrk。然而，致癌只在 Xmrk阳性与Xmrk缺失剑鱼的回交种间杂交 物种，或者当xmrk在非剑鱼模型系统中异位表达时 (例如，青竹、小鼠)。这些结果表明，斑纹伊蚊基因组还编码一种 可能与xmrk共同进化的调节基因，并能够抑制其 致癌活性。因此，刻画调节器，称为R(Diff) 抑制Xmrk，可能为控制EGFR提供新的治疗策略。 我们最近的研究已经将R(DIFF)肿瘤调节点定义为101.7 kBP 位于5号染色体上的基因。这个小的候选大小和低候选的基因 数字使功能和机制研究变得不切实际。因此， 该建议旨在通过检查R(Diff)轨迹来最终确定 以下目标： 目标1：我们将描述所有表达但未注释的遗传元素，在 除了已知的编码基因，在新确定的候选R(Diff)内 轨迹，以完全注释R(Diff)轨迹。 目的2：我们将确定携带R(Diff)活性的基因/元件(S) 在非荷瘤剑鱼中进行基因敲除 杂交种，以及携带肿瘤的转基因青竹。 目标3：我们将分析xmrk和R(Diff)候选者，以及 几个剑尾鱼杂交种，用候选组合来确定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.