Cx43 phosphorylation modulates Kras mediated pancreas cancer progression

描述(申请人提供)：胰腺癌或胰腺导管腺癌(PDA)每年的发病率和死亡率约为38,000人，是美国癌症相关死亡的第四大原因。PDA患者的5年总生存率不到3%。缝隙连接蛋白43(Cx43)是一种肿瘤抑制基因，在胰腺导管细胞中表达，胰腺导管细胞是肿瘤的假定起源部位。我们有初步证据表明，Cx43在胰腺癌中的表达、定位和磷酸化异常调节。这项建议的重点是这些事件如何在体内相互作用，影响胰腺的癌变。缝隙连接(GJ)是一种特殊的膜域，它包含允许小分子(&lt；1000 Da)在相邻细胞之间交换的通道，包括离子、代谢物和第二信使(例如，Ca~(2+)和IP3)。与其他连接蛋白一样，连接蛋白也发挥着独立于通道功能的重要信号和生长控制作用。非常广泛的体内和细胞系相关证据表明，缝隙连接细胞间通讯(GJC)和连接蛋白的表达调节细胞增殖并发挥关键的肿瘤预防作用。到目前为止，Cx43是最广泛表达的连接蛋白(34个组织和46种细胞类型)，是在细胞质C末端区域发现的多个丝氨酸残基的磷酸化。Cx43的磷酸化可以调节蛋白质的转运、连接复合体的稳定性、缝隙连接通讯(GJC)以及与其他蛋白质的相互作用。我们发现突变的Cx43在酪蛋白激酶1(CK1)磷酸化位点S325、S328和S330(称为CK3*)发生S到A氨基酸替换的纯合子“敲入”(KI)导致依赖Cx43的GJC显著降低，并持续增加MAPK活性和减少组织对不同急性刺激的凋亡。我们的合作者最近通过将致癌基因KrasG12D定向表达到胰腺，开发了第一个侵袭前和侵袭性胰腺导管腺癌模型。切除的胰腺显示了患者所见的侵袭前病变的全部范围，随着时间的推移，病变在组织学上进展，最终形成完全侵袭性和转移性疾病。然而，就像患有胰腺癌的人类一样，即使在这些同基因模型中，疾病进展、症状表现和进展速度也不同。我们假设Cx43的磷酸化对细胞生长的控制至关重要，并且在胰腺癌的发生过程中受到调节，影响其进展。因此，阻止这些调节事件将导致胰腺癌发生过程的改变。 公共卫生相关性：我们假设Cx43的磷酸化对控制细胞生长至关重要，并且在胰腺癌发生过程中受到调节，影响其进展。由于影响Cx43活性和缝隙连接通讯的药物正在开发中，了解这些因素如何影响进展最终可能导致这种致命疾病的更好治疗。

DESCRIPTION (provided by applicant): With an annual incidence and mortality of ~38,000 people, pancreas cancer or pancreatic ductal adenocarcinoma (PDA) is the fourth leading cause of cancer-related death in the United States. The 5-year overall survival for patients with PDA is less than 3%. The gap junction protein connexin43 (Cx43) is a tumor suppressor gene that is expressed in pancreatic ductal cells, the putative site of cancer origin. We have preliminary evidence that Cx43 expression, localization and phosphorylation is dysregulated during pancreas cancer. This proposal focuses on how these events interplay in vivo to affect carcinogenesis of the pancreas. Gap junctions (GJ) are specialized membrane domains that contain channels that allow exchange of small molecules (<1000 Da) including ions, metabolites, and second messengers (e.g., Ca2+ and IP3) between neighboring cells. Connexins, like other junctional proteins, also play critical signaling and growth control roles that are independent of channel function. Very extensive correlative evidence in vivo and cell lines indicates that gap junctional intercellular communication (GJC) and connexin expression regulate proliferation and play key tumor prevention roles. Cx43, by far the most widely expressed connexin (> 34 tissues and 46 cell types), is phosphorylated at multiple serine residues found in the cytoplasmic, C-terminal region. Cx43 phosphorylation can modulate the levels of protein trafficking, stability of the junctional complex, gap junctional communication (GJC) and the interaction with other proteins. We have found that homozygous "knock-in" (KI) of mutant Cx43 bearing an S to A amino acid substitutions at casein kinase 1 (CK1) phosphorylation sites S325, S328 and S330 (termed CK3*) leads to dramatically reduced Cx43-dependent GJC and sustained increased MAPK activity and decreased apoptosis in tissues in response to different acute stimuli. Our collaborator has recently developed the first models of preinvasive and invasive pancreatic ductal adenocarcinoma through the targeted physiologic expression of oncogenic KrasG12D to the pancreas. Resected pancreata demonstrate the full spectrum of preinvasive lesions seen in patients, and the lesions progress histologically over time culminating in fully invasive and metastatic disease. However, like in humans with pancreas cancer, disease progression, symptom presentation and speed of progression varies even in these syngeneic models. We hypothesize that Cx43 phosphorylation is critical for the control of cell growth and is modulated during pancreas carcinogenesis affecting its progression. Consequently, prevention of these regulatory events will result in an altered course of carcinogenesis in pancreas cancer. PUBLIC HEALTH RELEVANCE: We hypothesize that Cx43 phosphorylation is critical for the control of cell growth and is modulated during pancreas carcinogenesis affecting its progression. Since drugs that affect Cx43 activity and gap junctional communication are under development, understanding how these factors affect progression could ultimately lead to better treatment of this lethal disease.