Scatter factor induced carcinoma cell migration

描述(申请人提供)：基质细胞分泌的生长因子、细胞外基质及其在癌细胞上的受体之间的相互作用导致信号转导，是了解癌细胞迁移和侵袭机制的关键。这样的理解需要识别由这些相互作用诱导的基因和信号通路的汇聚。分散因子，也称为肝细胞生长因子(SF)及其酪氨酸激酶原癌基因Met，在大多数转移性癌症中表达上调，与预后不良有关。SF导致癌细胞迁移，这一过程需要从头开始基因转录。然而，这种新转录表达的早期基因尚不清楚。我们的长期目标是确定抑制转移的癌细胞特异性靶点。我们在人子宫内膜癌细胞系RL-95[和HECIA，以及胰腺癌细胞系FG]中分离到一种新的c DNA，我们称之为Mig-7，它是由SF在发生迁移之前诱导的。同源性搜索表明，Mig-7与两个人类EST序列有99%的同源性。然而，没有发现全长的cDNA、基因或蛋白质同源性。经逆转录和聚合酶链式反应检测，该基因在正常组织中缺乏表达。相比之下，它在各种转移性人类肿瘤中都能检测到，如卵巢、结肠、子宫内膜和鳞状细胞。此外，SF对Mig-7的诱导作用也受整合素的调节。这项建议的重点是了解Mig-7在癌细胞迁移中的表达作用。我们的假设是Mig-7的表达在癌细胞的迁移中发挥了作用。为了验证这一假说并实现这一应用的目的，我们将追求以下两个特定的目标：目的1是确定细胞迁移的程度是否依赖于癌细胞中Mig-7的表达水平，以验证我们的假设，即Mig-7表达导致癌细胞迁移。[我们的假设基于我们的初步结果，与无关的寡核苷酸相比，我们可以在体外使用针对Mig-7的反义寡核苷酸抑制癌细胞迁移83.50%+/-2.77%(p&lt；0.05)。]我们将在癌细胞系中过表达Mig-7，并与使用迁移[和侵袭]分析的对照相比，测量迁移和侵袭能力。目的2是为了确定Met和Alphavbeta5整合素信号转导通路之间相互作用的信号机制，因为我们的研究表明Met和Alphavbeta5整合素信号通路都需要诱导Mig-7的表达。已有研究表明，MET激活可激活粘着斑激酶(FAK)。此外，酪氨酸激酶受体诱导FG癌细胞迁移需要αvbeta5整合素结合。然而，这种串扰信号导致的基因表达尚未确定。]MiG-7是在该信号系统中表达的一个新基因。我们预计，这些研究将导致对癌细胞如何迁移的认识的重大进步，并可能导致新的抗转移治疗的靶点。

DESCRIPTION (provided by applicant): Interactions between growth factors secreted by the stroma cells, extracellular matrix, and their receptors on carcinoma cells resulting in signal transduction are key to understanding the mechanisms underlying carcinoma cell migration and invasion. Such understanding requires identification of the genes induced by these interactions and the convergence of signaling pathways. Scatter factor, also known as Hepatocyte growth factor, (SF) and its tyrosine kinase protooncogene, Met, are upregulated in most metastatic cancers and is associated with a poor prognosis. SF causes migration of cancer cells, a process that requires de novo gene transcription. However, the early genes expressed by this new transcription are not known. Our long range goal is to determine carcinoma cell specific targets to inhibit metastasis. We have isolated a novel cDNA we call Mig-7 that is induced by SF in the human endometrial carcinoma cell lines, RL-95 [and HECIA, as well as the pancreatic carcinoma cell line FG] before migration occurs. Homology searches show that Mig-7 cDNA sequence possesses 99% homology with two human ESTs. However, no full length cDNA, gene or protein homologies were found. Expression of this gene is lacking in normal tissue as determined by reverse transcription and polymerase chain reaction. In contrast, it is detected in various metastatic human tumors such as ovary, colon, endometrial and squamous cell. Furthermore, the Mig-7 induction by SF is also regulated by ?v?5 integrin ligation. The focus of this proposal is to understand the role of Mig-7 in expression in carcinoma cell migration. Our hypothesis is that Mig-7 expression plays a role in migration of cancer cells. To test this hypothesis and accomplish the objective of this application we will pursue the following two specific aims: Aim 1 is to determine if the degree of cell migration is dependent upon the level of Mig-7 expression in carcinoma cells to test our hypothesis that Mig-7 expression leads to migration of carcinoma cells. [We base our hypothesis on our preliminary results showing that we can inhibit carcinoma cell migration by 83.50% +/- 2.77% (p<0.05) in vitro using antisense oligonucleotides specific to Mig-7 as compared to irrelevant oligonucleotide.] We will overexpress Mig-7 in carcinoma cell lines and measure migration and invasion capabilities as compared to controls using migration [and invasion] assays. Aim 2 is to determine the signaling mechanism underlying the cross talk between the Met and alphavbeta5 integrln signal transduction pathways because our studies indicate that both Met and alphavbeta5 integrin signaling pathways are required to induce Mig-7 expression. [Met activation has been shown to activate focal adhesion kinase (FAK). Further, tyrosine kinase receptor induction of FG carcinoma cell migration requires alphavbeta5 integrin binding. However, gene expression resulting from this cross-talk signaling has not been determined.] Mig-7 is a novel gene expressed in this signaling system. We expect that these studies will lead to a significant advance in the knowledge of how carcinoma cells migrate and could lead to a target for new anti-metastatic therapy.