BRIT 1: Roles in Immortalization, Checkpoints and Cancer

描述(由申请人提供)：失去细胞周期检查点控制和获得无限增殖的能力是癌症发展所需的两个基本变化。BRIT1(BRCT-Repeat Inhibitor of hTERT Expression，BRCT-Repeat Inhibitor of hTERT Expression)是通过我们的增强型逆转录病毒突变筛查发现的一个新的hTERT功能抑制基因，与细胞永生化有关。我们的初步数据表明，BRIT1具有额外的功能--调节S内部的检查点和G2/M检查点--部分原因可能是它调节BRCA1和Chk1的表达，以及它在DNA损伤信号传递中的直接作用。此外，BRIT1位于染色体8p23.1上，根据杂合性缺失和CGH分析，该染色体与包括乳腺癌和卵巢癌在内的几种恶性肿瘤的发生有关。通过DNA阵列分析，BRIT1在这两种类型的癌症中的表达也显著降低。因此，我们推测BRIT1可能通过抑制细胞永生化和维持基因组稳定性的双重功能发挥肿瘤抑制基因的功能。 这一假设将在以下步骤中得到检验。(1)我们将通过有条件地表达抗BRIT1的小发夹RNA的正常人细胞来确定BR T1在防止细胞永生化中的作用，并探讨BRIT1的S功能对端粒酶逆转录酶表达的影响机制。(2)我们将利用BRIT1缺失的细胞研究BRIT1在应对遗传毒性应激、维持基因组完整性和细胞转化中的作用；我们将通过比较基因组杂交(CGH)和光谱核型分析(SKY)检测染色体变化来评估基因组的不稳定性；我们将研究BRIT1如何调节BRCA1和Chk1的启动子活性以及BRIT1如何在DNA损伤信号的传递中发挥直接作用；我们还将建立BRIT1条件基因敲除小鼠模型，以评估BRIT1在体内维持基因组完整性和抑制肿瘤形成中的S功能。(3)我们将对乳腺癌和卵巢癌标本中BRIT1的异常进行鉴定和表征，以确定BRIT1是否可以作为癌症的预后标志物或治疗靶点。因此，分析BRIT1在癌症患者中的功能及其异常将有助于更好地理解癌症发生和发展中的关键病理变化，并有助于开发新的、有效的癌症治疗方法。

DESCRIPTION (provided by applicant): Loss of cell cycle checkpoint control and acquisition of the ability to proliferate indefinitely constitute two of the fundamental changes required for the development of cancer. BRIT1 (BRCT-Repeat Inhibitor of hTERT expression), a novel gene identified through our enhanced retroviral mutation screen as a repressor of hTERT function, is implicated in cellular immortalization. Our preliminary data indicate that BRIT1 has additional functions-regulating both the intra-S and G2/M checkpoints-that, in part, may result from its regulation of BRCA1 and Chk1 expression and its direct role in the transmission of DNA damage signals. Moreover, BRIT1 is located on chromosome 8p23.1, which has been implicated in the development of several malignancies, including breast cancer and ovarian cancer, based on LOH and CGH analyses. The expression of BRIT1 also was significantly reduced in these two cancer types when analyzed by DNA arrays. Thus, we hypothesize that BRIT1 may function as a tumor suppressor gene through dual functions: repressing cellular immortalization and maintaining genomic stability. This hypothesis will be tested in the following steps. (1) We will determine the role of BR T1 in preventing cellular immortalization by using normal human cells that conditionally express small hairpin RNA against BRIT1, and we will investigate the mechanisms mediating BRIT1's function on hTERT expression. (2) We will use BRIT 1-depleted cells to study the role of BRIT1 in the response to genotoxic stress, maintenance of genomic integrity, and cellular transformation; we will assess genomic instability by detecting chromosome alterations using comparative genomic hybridization (CGH) and spectral karyotyping (SKY); we will study how BRIT1 regulates the promoter activities of BRCA1 and Chk1 and how BRIT1 may play a direct role in transmission of DNA damage signal; and we will develop a BRIT1 conditional knockout mouse model to assess the BRIT1's function in vivo in maintaining genomic integrity and suppressing tumor formation. (3) We will identify and characterize the aberrations of BRIT1 in breast and ovarian cancer samples to determine if BRIT1 serves as a prognostic marker or as a therapeutic target in cancer. As a corollary, an analysis of the function of BRIT1 and its aberrations in cancer patients will contribute to an improved understanding of the key pathological alterations in cancer initiation and progression and to the development of novel, effective therapeutic approaches for cancer.