Characterization of HIN1 Putative Tumor Suppressor

为了开发乳腺癌（BC）的预防性干预措施，确定乳腺肿瘤发生起始步骤的分子改变是很重要的。SAGE （Serial Analysis of Gene Expression）用于比较正常和DCIS（导管原位癌，浸润性BC的前体）乳腺上皮细胞的基因表达谱。该技术鉴定了HIN-1 (High In normal -1)，这是一种新的分泌蛋白，在正常乳腺上皮细胞中高度表达，但在90%的原发性侵袭前和侵袭性BC和BC细胞系中缺失。HIN- 1在这些早期肿瘤中的高频率失活表明它是一种新的肿瘤抑制因子。HIN-1在正常的肺、胰腺、前列腺和唾液腺中也有高表达，这些器官都含有分支上皮。提示HIN-1可能参与了细胞的形态发生或分化。本研究的总体目标是表征HIN-1的功能，并确定其在早期BC的起始步骤中的作用。特异性目的1是确定HIN-1对细胞增殖的影响。HIN-1将通过腺病毒和质粒载体或暴露于纯化的重组物中导入BC细胞系。使用集落生长试验和流式细胞术分析细胞周期进展来分析HIN-1及其作用。特异性目的2将在三维培养系统中探讨HIN-1表达对乳腺上皮细胞分支形态发生的影响。特异性目的3将利用转染四环素诱导的HIN-1载体的BC细胞系，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作为一种潜在的肿瘤抑制基因的作用，并且由于它在早期BC中经常失活，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.