Scatter factor induced carcinoma cell migration

描述（申请人提供）：间质细胞、细胞外基质分泌的生长因子与癌细胞上的受体之间的相互作用导致信号转导是理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趋同。分散因子，也被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上调，并与不良预后相关。SF引起癌细胞的迁移，这一过程需要从头开始的基因转录。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癌细胞的特异性靶点来抑制转移。我们在迁移发生前，在人子宫内膜癌细胞系RL-95[和HECIA]以及胰腺癌细胞系FG中分离出了一种被SF诱导的新型cDNA，我们称之为Mig-7。同源性分析表明，Mig-7 cDNA序列与2条人类ESTs序列具有99%的同源性。但未发现全长cDNA、基因或蛋白同源性。通过逆转录和聚合酶链反应确定，正常组织中缺乏该基因的表达。相反，在卵巢、结肠、子宫内膜和鳞状细胞等各种转移性人类肿瘤中均可检测到。此外，SF对米格-7的诱导也受到？v？整合素连接。本研究的重点是了解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起作用。为了验证这一假设并实现这一应用的目标，我们将追求以下两个具体目标：目的1是确定细胞迁移的程度是否依赖于癌细胞中米格-7的表达水平，以验证我们的假设，即米格-7的表达导致癌细胞迁移。[我们的假设基于我们的初步结果，与不相关的寡核苷酸相比，我们使用米格-7特异性的反义寡核苷酸可以体外抑制癌细胞迁移83.50% +/- 2.77% （p<0.05）。我们将在癌细胞系中过表达米格-7，并使用迁移[和侵袭]试验来测量与对照相比的迁移和侵袭能力。目的2是确定Met和alphavbeta5整合素信号转导通路之间串扰的信号传导机制，因为我们的研究表明Met和alphavbeta5整合素信号转导通路都是诱导米格-7表达所必需的。[Met激活已被证明可以激活局灶黏附激酶（FAK）。]此外，酪氨酸激酶受体诱导FG癌细胞迁移需要alphavbeta5整合素结合。然而，由这种串音信号引起的基因表达尚未确定。米格-7是在这个信号系统中表达的一种新基因。我们期望这些研究将在了解癌细胞如何迁移方面取得重大进展，并可能导致新的抗转移治疗的目标。

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.