Surgical Studies of GI Peptides - Mechanisms of Action

描述（由申请人提供）：胃肠道（GI）癌症仍然是一个重要的和具有挑战性的临床问题。手术切除是治疗消化道恶性肿瘤的主要方法；然而，只有肿瘤是局部的，没有扩散到淋巴结和远处器官，才能治愈。需要增加对控制胃肠道癌细胞生长的分子机制的理解，以开发与手术切除相结合的新治疗策略。胃肠肽激素可以刺激正常和肿瘤肠道组织的生长。多年来，我们的研究主要集中在确定肠道肽，胃泌素（G-17）及其同源受体调节细胞生长的分子机制。最近，我们发现了一种新的CCK-B/胃泌素受体的剪接变体，称为CCK-BRi4sv，它在结肠癌和胰腺癌中表达，但在正常组织中不表达。与先前表征的野生型CCK-BRwt相比，CCK-BRi4sv表现出明显不同的信号特性，包括不依赖g -17刺激细胞生长、调节细胞内ca2 +和亚细胞运输。此外，我们发现丝裂原活化蛋白激酶（MAPKs）在激动剂刺激前后cck - br介导的信号传导中都起着关键作用。MAPK激酶（MEK）调节CCK-BRwt对G-17刺激的敏感性，并介导G-17刺激对下游效应物的影响。最后，我们发现CCK-BRi4sv和CCK-BRwt介导g -17诱导COX-2基因表达。基于我们的研究结果，我们假设CCK-BR变体通过激动剂依赖性和非依赖性机制调节GI细胞的生长，并且MAPKs通过调节受体对激动剂刺激的敏感性以及作为激动剂诱导的信号转导的下游效应物，在受体介导的细胞生长调节中发挥核心作用。为了检验这些假设，我们计划了三个具体目标的实验。目的1：明确CCK-BRwt和CCK-BRi4sv内化和细胞内受体转运的机制。目的2：明确MAPKs在CCK-BRwt-和cck - bri4v -介导的细胞内信号转导中的作用。目的3：确定CCK-BRi4sv和CCK-BRwt表达对基因表达的影响。

DESCRIPTION (provided by applicant): Gastrointestinal (GI) cancers continue to be a significant and challenging clinical problem. Surgical resection is the mainstay for cure of GI malignancies; however, cure can only be achieved if the tumors are localized and have not spread to lymph nodes and distant organs. Increased understanding of the molecular mechanisms controlling GI cancer cell growth is required for the development of novel thera-peutic strategies to be used in combination with surgical resection. GI peptide hormones can stimulate the growth of normal and neoplastic gut tissues. For years, our studies have focused on determining the molecular mechanisms by which the gut peptide, gastrin (G-17), and its cognate receptors, regulate cell growth. Recently, we have discovered a novel splice variant of the CCK-B/gastrin receptor called CCK-BRi4sv that is expressed in colonic and pancreatic cancers, but not the normal tissues. CCK-BRi4sv exhibit distinctly different signaling properties when compared to the previously characterized wild-type CCK-BR (CCK-BRwt) including G-17-independent stimulation of cell growth, regulation of intracellular Ca 2+and subcellular trafficking. Also, we found that mitogen-activated protein kinases (MAPKs) play a key role in CCK-BR-mediated signaling both before and after agonist stimulation. MAPK kinase (MEK) regulates CCK-BRwt sensitivity to G-17 stimulation and mediates the effects of G-17 stimulation on downstream effectors. Finally, we found that CCK-BRi4sv and CCK-BRwt mediate G-17-induction of COX-2 gene expression. Based on our findings, we hypothesize that CCK-BR variants regulate GI cell growth by both agonist-dependent and -independent mechanisms, and that MAPKs play a central role in receptor-mediated regulation of cell growth by modulating the sensitivity of the receptor to agonist stimulation and by acting as downstream effectors of agonist-induced signal transduction. To examine these hypotheses, we have planned experiments with three Specific Aims. Aim 1: To define the mechanisms of CCK-BRwt and CCK-BRi4sv internalization and intracellular receptor trafficking. Aim 2: To define the role of MAPKs in CCK-BRwt- and CCK-BRi4sv-mediated intracellular signal transduction. Aim 3: To determine the effects of CCK-BRi4sv and CCK-BRwt expression on gene expression.