KCNK9 Imprinting in Breast Cancer Progression and Metastasis

描述(申请人提供)：三阴性乳腺癌[ER/PR-/-，HER2/neu wt，EGFR+]常见于年轻女性，预后不良。虽然并不是所有的三阴性乳腺癌都是致命的，但在绝经前的非裔美国女性中，三阴性乳腺癌有86%的5年死亡率。由于许多三阴性乳腺癌在确诊时对化疗耐药，因此非常有必要及早发现。在这里，我们旨在研究一种新的信号通路，有望用于三阴性乳腺癌的早期检测。TASK3是一种调节线粒体膜电位的pH敏感钾通道蛋白。TASK3的过表达增加了m，从而促进了细胞的抗凋亡和耐缺氧。在初步数据中，我们发现TASK3在1)化疗耐药的转移性三阴性乳腺癌和2)高危非裔美国女性的癌前病变中过度表达。KCNK9(TASK3基因)受印记调控：印记缺失预测化疗耐药和三阴性乳腺癌的后续转移：编码TASK3蛋白的基因KCNK9受甲基化印记调控。印记是一个正常的调控过程，基因的一个副本被灭活，导致单等位基因表达。正常印迹的丧失会导致功能二倍体状态和靶基因的过度表达。正常情况下，KCNK9基因的一个拷贝被印迹，使KCNK9的正常功能状态成为单倍体(一个基因拷贝表达，一个拷贝不表达)。在初步数据中，我们在KCNK9启动子中发现了一个差异甲基化区域(DMR)，该区域印记在正常乳腺上皮细胞中，但在原发化疗耐药的三阴性乳腺癌中不存在。DMR印迹缺失可预测TASK3过表达、化疗耐药和随后的转移。假设：在三阴性乳腺癌中，KCNK9 DMR的甲基化缺失和缺氧协同促进TASK3的过度表达和转移。目标1将测试在缺乏KCNK9 DMR印记的三阴性乳腺癌中，低氧是否转录激活TASK3。我们将在体内对常氧和低氧条件下KCNK9 HRE活性、TASK3表达、肿瘤生长和转移以及血红蛋白饱和度进行多参数分析。目的2检测正常KCNK9 DMR印迹缺失是否促进三阴性乳腺癌的发生、发展和转移。意义：这项建议的目标的成功完成将提高我们识别生物侵袭性转移性三阴性乳腺癌的能力。三阴性乳腺癌在绝经前的非裔美国女性中死亡率很高。我们建立的高危队列将使我们能够快速测试KCNK9 DMR印记缺失是否会促进化疗耐药的转移性三阴性乳腺癌的发生。 公共卫生相关性：非裔美国人和高加索女性雌激素受体阴性(ER-)乳腺癌的分子起源尚不清楚。重要的是，我们不知道ER(-)乳腺癌侵袭行为背后的分子途径是否可以在癌前病变中检测到。这些信息对于制定成功的早期发现和有针对性的预防战略至关重要。在这里，我们采用跨学科的方法，在我们的Duke乳腺癌组织库和世界级高危绝经前高加索和非裔美国女性的预期队列中调查ER(-)乳腺癌的表观遗传学起源。PHS 398/2590(09/04版，2006年4月4日重新发布)页面续格式页面

DESCRIPTION (provided by applicant): Triple-negative breast cancers [ER/PR-/-, HER2/neu wt, EGFR+] frequently occur in young women and carry a poor prognosis. While not all triple-negative breast cancers are lethal, triple-negative breast cancers have 86% 5-year mortality in pre-menopausal African American women. Since many triple-negative breast cancers are chemotherapy-resistant at diagnosis, there is a great need for early detection. Here we aim to investigate a novel signaling pathway that holds promise for early detection of triple-negative breast cancer.TASK3 is a pH sensitive potassium channel protein that regulates mitochondrial membrane potential m). Overexpression of TASK3 increases m, and thereby promotes apoptosis resistance and tolerance of hypoxia. In Preliminary Data, we show that TASK3 is overexpressed in 1) chemotherapy-resistant metastatic triple-negative breast cancers and 2) premalignant breast disease in high-risk African American women. KCNK9 (TASK3 gene) is regulated by imprinting: loss of imprinting predicts chemotherapy resistance and subsequent metastasis of triple-negative breast cancer: The gene coding for the TASK3 protein, KCNK9, is regulated by methylation imprinting. Imprinting is a normal regulatory process where one copy of the gene is inactivated resulting in mono-allelic gene expression. Loss of normal imprinting results in a functional diploid state and overexpression of the target gene. One copy of the KCNK9 gene is normally imprinted, making the normal functional state of KCNK9 haploid (one gene copy expressed, one copy not expressed). In Preliminary Data, we identified a differentially methylated region (DMR) in the KCNK9 promoter that was imprinted in normal mammary epithelial cells but not in primary chemotherapy-resistant triple-negative breast cancers. Loss of DMR imprinting predicted TASK3 overexpression, chemotherapy-resistance, and subsequent metastasis. Hypothesis: Loss of methylation imprinting of KCNK9 DMR and hypoxia synergistically promote TASK3 overexpression and metastasis in triple-negative breast cancer. Aim 1 will test whether hypoxia transcriptionally activates TASK3 in triple-negative breast cancers that lack imprinting of the KCNK9 DMR. We will perform in vivo multi-parametric analysis of KCNK9 HRE activity, TASK3 expression, tumor growth and metastasis, and hemoglobin saturation in normoxic and hypoxic conditions. Aim 2 will test whether loss of normal KCNK9 DMR imprinting promote initiation, progression, and metastasis of triple-negative breast cancer. Significance: Successful completion of the aims of this proposal will improve our ability to identify biologically aggressive metastatic triple-negative breast cancer. Triple-negative breast cancers have a high mortality in premenopausal African American women. Our established high-risk cohort will allow us to rapidly test whether loss of KCNK9 DMR imprinting promotes chemotherapy-resistant metastatic triple- negative breast cancers. PUBLIC HEALTH RELEVANCE: The molecular origins of estrogen receptor-negative (ER-) breast cancers in African American and Caucasian women are poorly understood. Importantly, we do not understand whether the molecular pathways that underlie the aggressive behavior of ER(-) breast cancers can be detected in premalignant lesions. This information is crucial for developing successful early detection and targeted-prevention strategies. Here we take trans-disciplinary approach to investigate the epigenetic origins of ER(-) breast cancer in our Duke breast cancer tissue bank and world-class prospective cohort of high-risk pre- menopausal Caucasian and African American women.. PHS 398/2590 (Rev. 09/04, Reissued 4/2006) Page Continuation Format Page