Mechanisms of suppression of colon cancer by receptor tyrosine phosphatase PTPRT

描述（由申请人提供）：我们提案的长期目标是阐明蛋白酪氨酸磷酸酶T（PTPRT）作为肿瘤抑制剂发挥作用的分子机制。在高通量突变分析中，我们确定PTPRT是人类结肠癌中最常见的突变磷酸酶。最近的外显子组测序研究发现，PTPRT在肺癌、胃癌和卵巢癌、头颈部鳞状细胞癌和黑色素瘤中也发生突变。在之前的资助期间，我们证明了Ptprt敲除（KO）小鼠易于发生结肠癌，从而提供了关键的体内证据来支持PTPRT作为肿瘤抑制因子发挥作用的前提。使用尖端的磷酸化蛋白质组学方法，我们确定了STAT3和桩蛋白作为PTPRT底物。PTPRT使关键STAT3活化位点Y705去磷酸化。我们的初步研究表明，与野生型同窝仔相比，Ptprt KO小鼠中的肠干细胞更具增殖性，并且在Ptprt KO小鼠的肠干细胞区室中pY705 STAT3上调。鉴于其他人最近令人兴奋的研究表明肠干细胞是肠肿瘤的起源，我们将在目标1中确定PTPRT如何调节肠干细胞稳态以及PTPRT调节的STAT3信号传导是否在肠干细胞稳态和PTPRT的体内肿瘤抑制功能中起关键作用。此外，PTPRT使以前未研究的桩蛋白磷酸酪氨酸位点Y88（pY88）去磷酸化。我们发现pY88桩蛋白在大多数人结肠癌标本中上调。有趣的是，我们的初步研究表明，非常高水平的pY88桩蛋白与晚期结肠癌相关，这表明pY88桩蛋白可用作预后生物标志物。此外，我们工程化桩蛋白Y88F突变体敲入结肠癌细胞系，并证明它们在体外表现出减少的肿瘤形成，并且在裸鼠中不能形成异种移植肿瘤。总之，我们的研究表明pY88桩蛋白介导结肠癌中的关键致癌信号传导。在目标2中，我们将专注于阐明pY88桩蛋白信号促进肿瘤发生的机制。在目标3中，我们将确定PTPRT调节的pY88桩蛋白通路是否可以用作结肠癌的预后标志物。这些研究的成功将有助于阐明肠道干细胞稳态和结肠肿瘤发生的新机制，提供一种新的预后工具来指导这种致命疾病的临床治疗，并确定一个潜在的重要新治疗靶点。由于PTPRT在多种人类癌症中发生突变，因此从这些研究中获得的知识可能对癌症治疗具有广泛的意义。

DESCRIPTION (provided by applicant): The long-term goal of our proposal is to elucidate the molecular mechanisms by which protein tyrosine phosphatase T (PTPRT) functions as a tumor suppressor. In a high-throughput mutational analysis, we identified PTPRT as the most frequently mutated phosphatase in human colon cancers. Recent exome sequencing studies uncovered that PTPRT is also mutated in lung, gastric, and ovarian cancers, in head & neck squamous cell carcinomas, and in melanomas. In the previous funding period, we demonstrated that Ptprt knockout (KO) mice are prone to develop colon cancer, thereby providing critical in vivo evidence to support the premise that PTPRT functions as a tumor suppressor. Using a cutting-edge phospho-proteomic approach, we identified STAT3 and paxillin as PTPRT substrates. PTPRT dephosphorylates the key STAT3 activation site Y705. Our preliminary studies showed that, compared to wild-type littermates, intestinal stem cells in Ptprt KO mice are more proliferative and that pY705 STAT3 is up-regulated in the intestinal stem cell compartments of Ptprt KO mice. In light of recent exciting studies by others demonstrating that intestinal stem cells are the origin of intestinal tumors, we will determine in Aim 1 how PTPRT regulates intestinal stem cell homeostasis and whether PTPRT-regulated STAT3 signaling plays a critical role in intestinal stem cell homeostasis and the in vivo tumor suppressor function of PTPRT. In addition, PTPRT dephosphorylates a previously unstudied paxillin phospho-tyrosine site, Y88 (pY88). We found that pY88 paxillin is up-regulated in a majority of human colon cancer specimens. Interestingly, our preliminary studies showed that very high levels of pY88 paxillin associate with advanced-stage colon cancers, suggesting that pY88 paxillin may be exploited as a prognostic biomarker. Moreover, we engineered paxillin Y88F mutant knock-in colon cancer cell lines and demonstrated that they exhibit reduced tumor formation in vitro and fail to form xenograft tumors in nude mice. Together, our studies suggest that pY88 paxillin mediates pivotal oncogenic signaling in colon cancer. In Aim 2, we will focus on elucidating the mechanisms by which pY88 paxillin signaling promotes oncogenesis. In Aim 3, we will determine if the PTPRT-regulated pY88 paxillin pathway can be exploited as a prognostic marker for colon cancer. Success in these studies will result in delineation of novel mechanisms underlying intestinal stem cell homeostasis and colon tumorigenesis, provide a novel prognostic tool to guide clinical management of this deadly disease, and define a potentially important new therapeutic target. Because PTPRT is mutated in multiple human cancers, knowledge gained from these studies may have broad implications for cancer therapy.