Regulation of Apical Polarity in Breast Cancer

描述（由申请人提供）：癌症与组织结构和功能的改变有关。值得注意的是，组织极性的改变可能是上皮性癌症进展的决定性因素。在乳腺中，基底-根尖组织的极性表现为膜蛋白在基底膜接触细胞的基极和上皮结构或腺泡的管腔细胞的根极之间的不对称分布。基底极性的改变伴随着侵袭性表型。虽然在乳腺癌中经常报道根尖极性的改变，但对这种改变的控制机制、后果以及诊断和预后价值知之甚少。紧密连接是在顶端极性的建立和维持中起关键作用的粘附复合物。我们的初步结果表明，紧密连接组织者ZO-1的分布在浸润性癌和某些类型的原位癌中发生了改变，证实了文献报道的顶端极性的丧失伴随着肿瘤的进展。此外，我们有证据表明DNA甲基化是表观遗传控制顶端极性的主要机制。长期的假设是，确定参与根尖极性控制的基因将为乳腺肿瘤的诊断和预后带来宝贵的信息，并建立新的策略来帮助预防乳腺癌的进展。提出了两个目标：(1)利用Affymetrix微阵列分析非肿瘤性乳腺上皮SI细胞的三维（3D）培养，确定具有统计学意义的根尖极性潜在调节基因，这些细胞在基质存在下形成表型正常的乳腺腺泡，在5-氮胞苷dna低甲基化、胶原培养或用间隙连接阻滞剂治疗后形成根尖极性损伤的腺泡。与对照相比，将使用3种不同的基于模型的微阵列数据统计分析来选择在所有顶端极性受损的腺泡细胞模型中表达改变的基因。我们将通过改变这些基因在不同乳腺细胞模型形成的极化和顶端极性受损结构中的表达来研究这些基因对顶端极性控制的影响。(2)探讨正常乳腺组织和肿瘤组织档案活检中已知根尖极性标志物的分布和CAP基因的表达谱，并将CAP基因的表达谱与根尖极性标志物分布的改变和肿瘤进展联系起来。非肿瘤性、癌前和侵袭性乳腺病变将进行一系列紧密连接和根尖下CRB复合物蛋白的免疫染色，并评估这些根尖标志物的分布模式与根尖极性丧失的组织学分级或组织标准之间的相关性。将CAP基因在非侵袭性和侵袭性乳腺病变中的表达与组织学分类和根尖标记物的分布图进行比较。该项目将更好地了解根尖极性丧失在乳腺癌进展中的作用，并为预后相关研究和预防肿瘤进展的靶点确定新的标志物。

DESCRIPTION (provided by applicant): Cancer is associated with alterations in tissue organization and function. Notably, the alteration of tissue polarity may be determinant for the progression of epithelial cancers. In the mammary gland, baso-apical tissue polarity is illustrated by the asymmetrical distribution of membrane proteins between the basal pole of cells in contact with the basement membrane, and the apical pole of cells that delineates the lumen of the epithelial structures or acini. Alterations in basal polarity accompany the invasive phenotype. Although alterations in apical polarity have been frequently reported in breast cancer, little is known regarding the control mechanisms, and the consequences as well as diagnosis and prognosis values of such alterations. Tight junctions are adhesion complexes that play a critical role in the establishment and maintenance of apical polarity. Our preliminary results show that the distribution of tight junction organizer ZO-1 is altered in invasive carcinomas and in certain types of carcinoma in situ, confirming literature reports that suggest that loss of apical polarity accompany tumor progression. In addition, we have evidence to show that DNA methylation is a major mechanism of epigenetic control of apical polarity. The long-term hypothesis is that the identification of genes involved in the control of apical polarity will bring invaluable information for the diagnosis and prognosis of breast neoplasias, and the establishment of new strategies to help prevent breast cancer progression. 2 aims are proposed: (1) to identify statistically significant genes that are potential regulators of apical polarity using Affymetrix microarray analysis of three-dimension (3D) culture of non-neoplastic mammary epithelial SI cells that form phenotypically normal breast acini in the presence of matrigel, and apical polarity-impaired acini upon DNA-hypomethylation by 5-azacytidine, culture in collagen I, or treatment with a gap junction blocker. Genes showing altered expression in all models of apical polarity-impaired acini compared to controls will be selected using 3 different model-based statistical analyses of the microarray data. The influence of the selected genes on the control of apical polarity (CAP genes) will be investigated by altering their expression in polarized and apical polarity-impaired structures formed by different breast cell models. (2) To explore the distribution of known apical polarity markers and the expression profile of CAP genes on archival biopsies of normal and neoplastic breast tissues and relate the expression profile of CAP genes to alterations in the distribution of apical polarity markers and tumor progression. Non-neoplastic, pre-malignant and pre-invasive breast lesions will be immunostained for a series of tight junction and subapical CRB complex proteins and the correlation between the distribution patterns of these apical markers and histological grade or tissue criteria for apical polarity loss will be assessed. CAP genes expression in non-invasive and invasive breast lesions will be compared to histological classification and distribution maps of apical markers. This project should bring a better knowledge of the involvement of apical polarity loss in breast cancer progression and identify novel markers for prognosis-related studies and targets to prevent tumor progression.