Novel human gene-IG20-potential role in cancer therapy

描述（由申请人提供）：IG 20基因对动物的生存至关重要，并在人肿瘤和转化细胞系中高度表达。它可以编码IG 20、MADD、IG 20-SV 2和DENN-SV剪接变体。DENN-SV在所有测试的组织和细胞中组成型表达，并且在肿瘤组织和细胞中高度表达。DENN-SV的过表达增强了增殖和对TNF-α、TRAIL、依托泊苷、长春碱和γ-辐射诱导的凋亡的抗性。相反，IG 20的过表达减缓了细胞增殖并增强了对由上述试剂诱导的细胞凋亡的易感性。其他两种变体对这些特性几乎没有影响。IG 20增强的细胞凋亡是通过增强半胱天冬酶8和FADD向死亡诱导信号复合物（DISC）的募集介导的。相反，DENN-SV似乎激活以NF-κ B激活为特征的细胞存活。一旦IG 20被引入，它们显示出降低的NF κ B活性，变得对TRAIL诱导的细胞凋亡敏感，在体外缓慢复制并且不能在裸鼠中形成肿瘤。此外，siRNA介导的DENN-SV而不是IG 20的敲低最有可能诱导癌细胞的自发凋亡。这些观察结果表明，DENN-SV和IG 20的功能分别类似于“癌基因”和“肿瘤抑制因子”，并且参与细胞增殖和死亡的调节。 基于这些观察结果，我们假设“IG 20和DENN-SV剪接变体的差异表达可以使卵巢癌细胞分别对诱导的凋亡更敏感或更耐受，并且IG 20变体或靶向siRNA的促凋亡特性可以用于使原本耐受的卵巢癌细胞变得对TRAIL的杀伤敏感。“在Aim-1中，我们将测试IG 20和DENN-SV对卵巢癌细胞增殖和凋亡的影响;在Aim-2中，我们将确定IG 20在TRAIL/TNFa诱导的凋亡中的作用机制，并绘制功能相关结构域;在Aim-3中，我们将研究DENN-SV如何增强细胞增殖和抗凋亡，并鉴定功能相关结构域和接头蛋白;在Aim-4中，我们将使用来自野生型和敲除小鼠的小鼠胚胎成纤维细胞研究IG 20和/或DENN-SV的选择性表达对细胞增殖和凋亡的影响。这些研究将有助于更清楚地了解IG 20/DENN-SV在卵巢癌中的作用机制。

DESCRIPTION (provided by applicant): The IG20 gene is essential for survival of the animal and is highly expressed in human tumors and transformed cell lines. It can encode IG20, MADD, IG20-SV2 and DENN-SV splice variants. DENN-SV is constitutively expressed in all tissues and cells tested and highly expressed in tumor tissues and cells. Over-expression of DENN-SV enhances proliferation and resistance to apoptosis induced by TNF-alpha, TRAIL, etoposide, vinblastine and gamma-radiation. In contrast, over- expression of IG20 slows cell proliferation and enhances susceptibility to apoptosis induced by the above agents. Other two variants have little or no effect on these properties. IG20 enhanced apoptosis is mediated by enhanced recruitment of caspase 8 and FADD to the Death Inducing Signaling Complex (DISC). In contrast, DENN-SV appears to activate cell survival characterized by NF-kappaB activation. Once IG20 is introduced, they show reduced NFKB activity, become susceptible to TRAIL induced apoptosis, replicate slowly in vitro and fail to form tumors in nude mice. In addition, siRNA mediated knock-down of, most likely, DENN-SV, and not IG20, induces spontaneous apoptosis of cancer cells. These observations show that DENN-SV and IG20 function like an "oncogene" and a "tumor suppressor" respectively, and are involved in the regulation of cell proliferation and death. Based on these observations, we hypothesize that "differential expression of IG20 and DENN-SV splice variants could render ovarian carcinoma cells either more susceptible or resistant to induced apoptosis respectively, and that the pro-apoptotic property of IG20 variant or targeted siRNA can be used to render ovarian cancer cells that are otherwise resistant become susceptible to killing by TRAIL." In Aim-1, we will test the effects of IG20 and DENN-SV on proliferation and apoptosis of ovarian cancer cells; In Aim-2, we will determine the mechanism of action of IG20 in TRAIL/TNFa induced apoptosis and map functionally relevant domains; In Aim-3, we will investigate how DENN-SV enhances cell proliferation and resistance to apoptosis, and identify functionally relevant domains and adaptor proteins; and in Aim-4, we will study the effects of selective expression of IG20 and/or DENN-SV on cell proliferation and apoptosis using mouse embryonic fibroblasts derived from wild type and knockout mice. These studies will lead to a clearer understanding of the mechanism of action of IG20/DENN-SV in ovarian cancer.