KCNK9 Imprinting in Breast Cancer Progression and Metastasis

描述（申请人提供）：三阴性乳腺癌[ER/PR-/-， HER2/ newwt， EGFR+]多见于年轻女性，预后较差。虽然并非所有的三阴性乳腺癌都是致命的，但三阴性乳腺癌在绝经前的非裔美国妇女中有86%的5年死亡率。由于许多三阴性乳腺癌在诊断时具有化疗耐药性，因此非常需要早期发现。在这里，我们的目标是研究一种新的信号通路，有望早期发现三阴性乳腺癌。TASK3是一种pH敏感的钾通道蛋白，调节线粒体膜电位。过表达的TASK3增加m，从而促进细胞凋亡抵抗和缺氧耐受。在初步数据中，我们发现TASK3在1)化疗耐药转移性三阴性乳腺癌和2)高危非裔美国女性的癌前乳腺癌中过表达。KCNK9 （TASK3基因）受印迹调控：印迹的缺失预示着三阴性乳腺癌的化疗耐药和随后的转移：编码TASK3蛋白的基因KCNK9受甲基化印迹调控。印迹是一个正常的调控过程，其中基因的一个拷贝失活导致单等位基因表达。正常印记的缺失导致二倍体的功能状态和靶基因的过度表达。KCNK9基因的一个拷贝正常印迹，使KCNK9单倍体的正常功能状态（一个拷贝表达，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），该区域在正常乳腺上皮细胞中有印迹，但在原发性化疗耐药三阴性乳腺癌中没有。DMR印迹的缺失预示着TASK3过表达、化疗耐药和随后的转移。假设：KCNK9 DMR甲基化印迹的缺失和缺氧协同促进三阴性乳腺癌中TASK3的过表达和转移。目的1将测试缺氧是否在缺乏KCNK9 DMR印记的三阴性乳腺癌中转录激活TASK3。我们将在常氧和缺氧条件下对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体内多参数分析。目的2将测试正常KCNK9 DMR印迹的缺失是否促进三阴性乳腺癌的发生、进展和转移。意义：成功完成本提案的目标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非裔美国妇女中死亡率很高。我们建立的高风险队列将使我们能够快速测试KCNK9 DMR印记的丢失是否会促进化疗耐药的转移性三阴性乳腺癌。

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