Novel human gene-IG20-potential role in cancer therapy

描述(申请人提供)：IG20基因是动物生存所必需的，在人类肿瘤和转化细胞系中高度表达。它可以编码IG20、MADD、IG20-SV2和Denn-SV剪接变体。Denn-SV在所测试的所有组织和细胞中都有成分表达，在肿瘤组织和细胞中高表达。Denn-SV过表达可增强肿瘤坏死因子-α、TRAIL、依托泊苷、长春花碱和伽玛射线诱导的细胞增殖和抗凋亡作用。相反，IG20的过度表达减缓了细胞的增殖，并增强了对上述药物诱导的凋亡的敏感性。另外两个变体对这些属性影响很小或没有影响。IG20通过促进caspase 8和FADD到死亡诱导信号复合体(DISC)的募集来促进细胞凋亡。相反，Denn-SV似乎激活了以核因子-kappaB激活为特征的细胞存活。一旦引入IG20，它们就表现出NFKB活性降低，对TRAIL诱导的细胞凋亡敏感，体外复制缓慢，无法在裸鼠体内形成肿瘤。此外，siRNA介导的Denn-SV而不是IG20的敲除很可能诱导了癌细胞的自发凋亡。这些观察结果表明Denn-SV和IG20的功能分别类似于“癌基因”和“肿瘤抑制因子”，并且参与了细胞增殖和死亡的调节。基于这些观察结果，我们假设“IG20和Denn-SV剪接变异体的不同表达可以分别使卵巢癌细胞对诱导的凋亡更敏感或更具抵抗力，IG20变异体或靶向siRNA的促凋亡特性可用于使原本具有耐药性的卵巢癌细胞变得对TRAIL敏感。”在Aim-1中，我们将检测IG20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IG20在TRAIL/TNFa诱导的细胞凋亡中的作用机制，并绘制功能相关区域图；在Aim-3中，我们将研究Denn-SV如何增强细胞增殖和抗凋亡能力，并寻找功能相关的结构域和接头蛋白；在Aim-4中，我们将利用野生型和敲除小鼠来源的小鼠胚胎成纤维细胞，研究选择性表达IG20和/或Denn-SV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IG20/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.