Mechanisms of suppression of colon cancer by receptor tyrosine phosphatase PTPRT

描述(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)作为肿瘤抑制因子的分子机制。在高通量突变分析中，我们发现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也在肺癌、胃癌和卵巢癌、头颈部鳞状细胞癌和黑色素瘤中发生突变。在之前的资助期间，我们证明了Ptprt基因敲除(KO)小鼠容易患结肠癌，从而提供了关键的体内证据，支持了PTPRT作为肿瘤抑制因子的前提。使用尖端的磷酸蛋白质组学方法，我们确定STAT3和巴西林为PTPRT底物。PTPRT使关键的STAT3激活位点Y705去磷酸化。我们的初步研究表明，与野生型乳鼠相比，Ptprt KO小鼠的肠道干细胞更具增殖能力，并且pY705 STAT3在Ptprt KO小鼠的肠道干细胞区段中上调。鉴于最近一些令人兴奋的研究表明肠道干细胞是肠道肿瘤的起源，我们将在目标1中确定PTPRT如何调节肠道干细胞的稳态，以及PTPRT调节的STAT3信号是否在肠道干细胞稳态和PTPRT的体内肿瘤抑制功能中发挥关键作用。此外，PTPRT还使一个以前未被研究过的巴西林磷酸化酪氨酸位点Y88去磷酸化(PY88)。我们发现pY88 paxlin在大多数人类结肠癌标本中表达上调。有趣的是，我们的初步研究表明，非常高水平的pY88帕克西林与晚期结肠癌有关，这表明pY88帕克西林可能被用作预测预后的生物标志物。此外，我们还设计了突变型Paxlin Y88F结肠癌细胞系，并证明它们在体外能减少肿瘤的形成，并且不能在裸鼠体内形成异种移植瘤。总之，我们的研究表明，pY88帕克西林在结肠癌中介导了关键的致癌信号。在目标2中，我们将重点阐明pY88帕克西林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 paxlin通路是否可以被用作结肠癌的预后标志物。这些研究的成功将导致描述肠道干细胞稳态和结肠肿瘤发生的新机制，为指导这种致命疾病的临床治疗提供新的预后工具，并确定潜在的重要新的治疗靶点。由于PTPRT在多种人类癌症中发生突变，从这些研究中获得的知识可能会对癌症治疗产生广泛的影响。

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.