THE MOLECULAR TARGET OF GAP JUNCTION ENHANCERS

间隙连接增强剂的分子靶点

• 批准号: 8364918
• 负责人: DUY H HUA
• 金额: $ 12.36万
• 依托单位: UNIVERSITY OF KANSAS LAWRENCE
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-06-01 至 2012-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8364918
• 关键词: Ablation; Animals; Attenuated; Binding; Breast Cancer Cell; Cancer Center; Cancer Patient; Cell Communication; Cell Proliferation; Cell Survival; Cells; Clinical; Code; Collaborations; Connexin 43; Connexins; Data; Defect; Disease; Disease Progression; Drug Delivery Systems; EGF gene; Enhancers; Epithelial; Estrogen Therapy; Estrogens; Etiology; Exhibits; Fibroblast Growth Factor; Funding; Gap Junctions; Genes; Goals; Grant; Growth; Growth Factor; Homeostasis; Hormones; Inhibitory Concentration 50; Intercellular Junctions; Ions; Laboratories; Lead; Link; Maintenance; Malignant Neoplasms; Mammary Neoplasms; Mammary gland; Mediating; Molecular Target; Molecular Weight; Mus; Names; National Center for Research Resources; Neoplasm Metastasis; Normal Cell; Pathology; Phosphorylation; Principal Investigator; Proteins; RNA; Reporting; Research; Research Infrastructure; Resources; Signal Transduction; Source; Surface; Surface Plasmon Resonance; T47D; Therapeutic; Time; Tissues; Tumorigenicity; United States National Institutes of Health; Woman; Xenograft procedure; animal tissue; antitumor agent; cancer cell; carcinogenesis; cell growth regulation; cost; dalton; drug sensitivity; extracellular; gap junction channel; high throughput screening; intercellular communication; malignant breast neoplasm; mortality; neoplastic; neoplastic cell; novel; quinoline; small molecule; therapy development; trafficking; tumor; tumor growth; ubiquitin-protein ligase

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Breast cancer is the most common cancer in women worldwide and mortality from breast cancer is consistent due to tumor metastasis. Breast cancer patients initially respond to estrogen ablation therapy, but estrogen-independent cells almost always aggressively emerge. The disease eventually progresses to estrogen-independent breast cancer. Tumor is no longer responsive to estrogen ablation therapy and unrestrained progression of the disease is inevitable. Progress in understanding the etiology of this disease and developing therapies has been slow due to multiple deregulations of various genes. The additive effects against mammary tumor cells might be achieved by combining antitumor agents directed against one or more altered mechanisms in cancer. Cancer cells exhibit many defects in cell communication that contribute to the loss of tissue homeostasis (excess cell proliferation, invasion, and metastasis). Cell communication is important in the cooperation between neighboring cells. It is mediated through extracellular signals such as hormones (i.e. estrogen) and growth factors (i.e. IGF, EGF, and FGF), or through cell to cell interaction between adjacent cells. One type of cell communication is gap junctional intercellular communication (GJIC), considered to be of fundamental importance. Gap junction (GJ) channels allow the transfer of small molecules, which involve in the regulation of cell growth, differentiation, and function. Gap junctions are the only communicating junctions found in animal tissues, in all species, which are responsible for the direct traffic of ions and molecules with molecular weights less than 1,200 Daltons. These traffic ways are formed by the interaction between two hemichannels on the surface of opposing cells. These hemichannels are formed by the association of six proteins, the connexins. Because of the importance of intercellular junctions in the maintenance of the cellular homeostasis, the modulation of intercellular junctions and expression of connexin is involved in carcinogenesis. Most normal cells have functional GJIC, while most, if not all, tumors cells have dysfunctional GJIC. It is believed that restoring GJIC is linked to drug sensitivity and reduction of tumorigenicity. Thus, increasing gap junction activity or enhancing GJIC in tumor cells provides the targets to enhance anti-neoplastic therapies. Several GJIC enhancers have been reported; however, an effective clinical drug targeting gap junction is not available at this time. Recently, we synthesized a new class of substituted quinolines (code name: PQ) and found that they possess potent inhibitory activities against T47D breast cancer cells (IC50 value of PQ7 is 16 nM and PQ1 is 119 nM) through the enhancement of GJIC. Our data showed that PQ1 significantly increases gap junction activity and inhibits cell viability and colony growth of T47D breast cancer cells. Moreover, PQ1 and PQ7 decrease 71% and 99%, respectively, of xenograft breast tumors bearing mice. Interestingly, PQ-treated animals have a normal histological pathology. Furthermore, PQ1 and PQ7 have no effect on normal primary epithelial mammary cells. We have found that PQ1 has a very strong binding with Nedd4 (from surface plasmon resonance studies), an E3 ubiquitin ligase, and decreases the interaction of Nedd4 and connexin 43 (Cx43) and the phosphorylation of Cx43, which lead to the enhancement of GJIC. Thus, the principle hypothesis of this proposal is that PQs can 1) increase gap junction activities, 2) inhibit the interaction of Nedd4 and Cx43 and phosphorylation of Cx43, and 3) attenuate tumor growth. The goal of this application is to identify the molecular target(s) of this novel class of molecules through Si RNA high-throughput screening and study the morphological changes of cells treated with PQs through a collaboration with Dr. Rathnam Chaguturu at the KU High-Throughput Screening Laboratory.

该子项目是利用资源的众多研究子项目之一 由 NIH/NCRR 资助的中心拨款提供。子项目的主要支持 并且子项目的主要研究者可能是由其他来源提供的， 包括其他 NIH 来源。 子项目可能列出的总成本 代表子项目使用的中心基础设施的估计数量， NCRR 赠款不直接向子项目或子项目工作人员提供资金。 乳腺癌是全世界女性最常见的癌症，由于肿瘤转移，乳腺癌的死亡率是一致的。乳腺癌患者最初对雌激素消融疗法有反应，但雌激素非依赖性细胞几乎总是积极地出现。该疾病最终发展为不依赖雌激素的乳腺癌。肿瘤不再对雌激素消融治疗有反应，并且疾病的不受限制的进展是不可避免的。由于各种基因的多重失调，了解这种疾病的病因和开发治疗方法的进展缓慢。 通过联合针对一种或多种癌症改变机制的抗肿瘤药物，可以实现针对乳腺肿瘤细胞的附加效应。 癌细胞在细胞通讯中表现出许多缺陷，导致组织稳态丧失（细胞过度增殖、侵袭和转移）。 小区通信对于相邻小区之间的合作非常重要。 它通过细胞外信号（例如激素（即雌激素）和生长因子（即 IGF、EGF 和 FGF））或通过相邻细胞之间的细胞间相互作用介导。 细胞通讯的一种类型是间隙连接细胞间通讯（GJIC），被认为具有根本重要性。 间隙连接 (GJ) 通道允许小分子的转移，参与细胞生长、分化和功能的调节。 间隙连接是在所有物种的动物组织中发现的唯一通讯连接，负责分子量小于 1,200 道尔顿的离子和分子的直接交通。这些交通通道是由相对细胞表面上的两个半通道之间的相互作用形成的。 这些半通道由六种蛋白质（连接蛋白）结合形成。 由于细胞间连接在维持细胞稳态中的重要性，细胞间连接的调节和连接蛋白的表达参与癌发生。 大多数正常细胞具有功能性 GJIC，而大多数（如果不是全部）肿瘤细胞具有功能失调的 GJIC。 据信恢复 GJIC 与药物敏感性和降低致瘤性有关。 因此，增加间隙连接活性或增强肿瘤细胞中的GJIC提供了增强抗肿瘤治疗的靶标。 已报道了几种 GJIC 增强剂；然而，目前临床上尚无针对间隙连接的有效药物。 最近，我们合成了一类新的取代喹啉（代号：PQ），发现它们通过增强GJIC对T47D乳腺癌细胞具有有效的抑制活性（PQ7的IC50值为16 nM，PQ1为119 nM）。我们的数据表明，PQ1 显着增加间隙连接活性并抑制 T47D 乳腺癌细胞的细胞活力和集落生长。此外，携带异种移植乳腺肿瘤的小鼠中，PQ1 和 PQ7 分别减少了 71% 和 99%。有趣的是，PQ 治疗的动物具有正常的组织病理学。此外，PQ1和PQ7对正常的原代乳腺上皮细胞没有影响。 我们发现PQ1与E3泛素连接酶Nedd4（来自表面等离子共振研究）具有非常强的结合，并降低Nedd4和连接蛋白43（Cx43）的相互作用以及Cx43的磷酸化，从而导致GJIC的增强。因此，该提议的主要假设是 PQ 可以 1) 增加间隙连接活性，2) 抑制 Nedd4 和 Cx43 的相互作用以及 Cx43 的磷酸化，以及 3) 减弱肿瘤生长。 该应用的目标是通过 Si RNA 高通量筛选来鉴定此类新型分子的分子靶标，并通过与 KU High-Rathnam Chaguturu 博士合作研究用 PQ 处理的细胞的形态变化。吞吐量筛选实验室。