Characterization of Human 3p Recessive Oncogenes

人类 3p 隐性癌基因的表征

• 批准号: 6608912
• 负责人: JOHN D. MINNA
• 金额: $ 35.1万
• 依托单位: UT SOUTHWESTERN MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 1996
• 资助国家: 美国
• 起止时间: 1996-09-13 至 2005-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6608912
• 关键词: DNA directed DNA polymerase; DNA footprinting; apoptosis; athymic mouse; autosomal recessive trait; cell growth regulation; enzyme activity; gene deletion mutation; gene expression; human genetic material tag; human tissue; lung neoplasms; molecular cloning; molecular pathology; neoplasm /cancer genetics; northern blottings; nucleic acid sequence; oncogenes; polymerase chain reaction; protein sequence; pulsed field gel electrophoresis; single strand conformation polymorphism; southern blotting; statistics /biometry; telomerase; telomere; tissue /cell culture

This project focuses on identification and functional characterization of human 3p recessive oncogenes (tumor suppressor genes, TSGs), with the initial emphasis being on discovery of TSGs in a 630 kb, completely sequenced, 3p21.3 region found to be homozygously deleted in lung and breast cancers. Our overall hypothesis is that biallelic inactivation or haploinsufficiency of one or more of these genes is of fundamental importance in the pathogenesis of many human cancers and that this inactivation occurs early in the multistep process of carcinogenesis. We have just completed the isolation and initial characterization of 25 genes in this area (23 of which are new) and found two, RASSF1A (Ras interacting protein with a DAG binding domain), and SEMA3B (secreted class III semaphorin) to have biallelic expression loss and potent tumor suppressing activity. We also found several others to have some characteristics of TSGs (loss of expression, mutation, suppression of the malignant phenotype) including CACNA2D2, 101F6, NPR21/G21, BLU, FUSI, HYAL1, and FUS2. The current proposal will: Confirm the tumor suppressing function of RASSF1A and SEMA3B and further implicate or dismiss the other candidate 3p21.3 TSGs in multiple human cancer cell lines with different genetic abnormalities by in vitro and in vivo xenograft assays (Aim 1); Confirm that tumor acquired loss of expression of RASSF1A and SEMA3B and some of the other 3p21.3 TSG candidates occurs in human cancers but not in normal tissues, and is explained by tumor acquired methylation of CpG islands in their promoter regions (Aim 2); Study the potential mechanisms for tumor suppression of RASSF1A and SEMA3B using human cancer cell lines and thus determine the pathways involved in RASSF1A and SEMA3B suppression of the tumor phenotype; in the process also determine if acquired and germline amino acid sequence alterations in these genes inactivate their function potentially predisposing to cancer development (Aim 3); Determine whether a Sema3b knockout mouse model challenged with carcinogens confirms SEMA3B as a TSG and also test whether SEMA3B haploinsufficiency is sufficient for aiding carcinogen induced lung cancer in the mouse. (Aim 4). The characterization of these gene(s) should ultimately have translational benefit for the development of new cancer diagnostics and therapeutics, including use in cancer early detection, prevention, and treatment.

该项目的重点是人类3 p隐性癌基因（肿瘤抑制基因，TSGs）的鉴定和功能表征，最初的重点是在630 kb，完全测序，3p21.3区域发现TSGs在肺癌和乳腺癌中被同源删除。 我们的总体假设是，这些基因中的一个或多个的双等位基因失活或单倍不足在许多人类癌症的发病机制中具有根本的重要性，并且这种失活发生在癌发生的多步骤过程的早期。 我们刚刚完成了该领域25个基因的分离和初步鉴定（其中23个是新的），并发现两个，RASSF 1A（Ras相互作用蛋白与DAG结合结构域）和SEMA 3B（分泌型III类信号蛋白）具有双等位基因表达丢失和有效的肿瘤抑制活性。 我们还发现其他几种具有TSGs的一些特征（表达缺失，突变，恶性表型抑制），包括CACNA 2D 2，101 F6，NPR 21/G21，BLU，FUSI，HYAL 1和FUS 2。 目前的提案将：通过体外和体内异种移植试验，确认RASSF 1A和SEMA 3B的肿瘤抑制功能，并进一步暗示或排除具有不同遗传异常的多种人类癌细胞系中的其他候选3p21.3 TSG（目的1）;确认肿瘤获得性RASSF 1A和SEMA 3B以及其他一些3p21.3 TSG候选物的表达缺失发生在人类癌症中，但不发生在正常组织中，并通过其启动子区域中CpG岛的肿瘤获得性甲基化来解释（目的2）;使用人癌细胞系研究RASSF 1A和SEMA 3B的肿瘤抑制的潜在机制，从而确定参与RASSF 1A和SEMA 3B肿瘤表型抑制的途径;在这个过程中，还确定这些基因中获得性和种系氨基酸序列的改变是否会破坏它们潜在的致癌功能，确定用致癌物攻击的Sema 3b敲除小鼠模型是否证实SEMA 3B为TSG，并且还测试SEMA 3B单倍不足是否足以帮助小鼠中致癌物诱导的肺癌。(Aim 4）。 这些基因的表征最终应该对开发新的癌症诊断和治疗具有转化益处，包括用于癌症早期检测、预防和治疗。