Characterization of HIN1 Putative Tumor Suppressor

为了开发乳腺癌(BC)的预防干预措施，确定表征乳腺肿瘤发生的起始步骤的分子变化是很重要的。采用SAGE(基因表达序列分析)软件，比较正常乳腺上皮细胞和乳腺浸润性癌前体导管癌(DCIS)的基因表达谱。这项技术鉴定了HIN-1(High in Normal-1)，一种新的分泌蛋白，在正常乳腺上皮细胞中高度表达，但在90%的原代侵袭前和侵袭BCS和BC细胞系中缺失。在这些早期肿瘤中，HIN-1失活的频率很高，这表明它是一种新的肿瘤抑制因子。Hin-1在正常肺、胰腺、前列腺和唾液腺也有高表达，所有器官都有分支上皮细胞。这表明HIN-1可能参与了形态发生或分化。。拟议研究的总体目标是确定HIN-1的功能，并确定其在公元前早期启动步骤中的作用。具体目的1是确定HIN-1对细胞增殖的影响。Hin-1将使用腺病毒和质粒载体或暴露于纯化的重组体而导入BC细胞系。用集落生长实验和细胞周期进程的流式细胞仪分析其作用。特定目的2将探讨HIN-1表达对三维培养体系中乳腺上皮细胞分支形态发生的影响。具体目标3将利用四环素诱导的HIN-1载体转染的BC细胞株，研究HIN-1抑制裸鼠肿瘤细胞生长的能力。因此，这些研究的结果将确定HIN-1作为一个潜在的肿瘤抑制基因的作用，并且由于它在BC早期经常被失活，因此可能为预防性干预提供一个很好的靶点。

To develop preventive interventions for breast cancer (BC), it is important to determine the molecular alterations that characterize the initiating steps of breast tumorigenesis. SAGE (Serial Analysis of Gene Expression) was used to compare the gene expression profiles of normal and DCIS (Ductal Carcinoma In-Situ, a precursor of invasive BC) mammary epithelial cells. This technique identified HIN-1 (High In Normal-1), a novel secreted protein that is highly expressed in normal breast epithelial cells, but lost in >90% of primary pre-invasive and invasive BCs and BC cell lines. The high frequency at which HIN- 1 is inactivated in these early tumors suggests that it acts as a novel tumor suppressor. HIN-1 is also highly expressed in normal lung, pancreas, prostate, and salivary gland, all organs containing branching epithelia. This suggests that HIN-1 may be involved in morphogenesis or differentiation. . The overall objective of the proposed research is to characterize the function of HIN-1 and to determine its role in the initiating steps of early BC. Specific aim 1 is to determine the effect of HIN-1 on cell proliferation. HIN-1 will be introduced into BC cell lines using adenoviral and plasmid vectors or by exposure to purified recombinant. HIN-1 and the effects analyzed using colony growth assays and flow cytometric analysis of cell cycle progression. Specific aim 2 will explore the effects of HIN-1 expression on branching morphogenesis of mammary epithelial cells in a three-dimensional culture system. Specific aim 3 will address the ability of HIN-1 to suppress tumor cell growth in nude mice using BC cell lines transfected with a tetracycline inducible HIN-1 vector. The results of these studies will thus determine the role of HIN-1 as a potential tumor suppressor gene and, since it is inactivated frequently in early BC, may provide an excellent target for preventive intervention.