PHLPP1/2 phosphatases as new target for beta-cell-directed therapy in diabetes

1型糖尿病（T1 D）和2型糖尿病（T2 D）均由胰腺β细胞功能和/或质量下降引起。迫切需要开发治疗方法来恢复糖尿病患者的β细胞群。 细胞凋亡和胰岛素分泌β细胞功能受损是β细胞衰竭的标志，也是糖尿病的根本原因。β细胞凋亡的调节和/或再生能力的增强代表了治疗糖尿病的有吸引力的治疗方法。丝氨酸-苏氨酸磷酸酶（STP）如PHLPP 1/2（PH结构域富含亮氨酸重复蛋白磷酸酶-1/2）调节细胞死亡，并作为药物开发的潜在靶标。我们对PHLPPs在β细胞死亡调节中的功能的理解具有重要的生物学和临床意义。我们表明，PHLPP 1和PHLPP 2在体外致糖尿病条件下，在人T1 D和T2 D以及糖尿病小鼠模型中的β细胞中高度上调。PHLPP 1/2过表达损害β细胞存活和功能。PHLPP 1/2诱导的β-细胞凋亡是通过AKT促存活信号转导的失活和促细胞凋亡MST 1激酶的活化介导的，这两种激酶是公认的PHLPP底物。在最初的DFG资助期间，我们发现：（i）PHLPP、AKT和MST 1形成控制β细胞凋亡的自抑制三角形。(ii)在糖尿病条件下，mTORC 1超活化调节PHLPP上调、MST 1活化和β细胞死亡。(iii)PHLPP的遗传和药理学抑制恢复了体外和体内糖尿病小鼠模型中的β细胞存活和胰岛素分泌。在本发明中，我们继续通过评估PHLPP在β细胞中如何上调以及mTORC 1是否在翻译水平控制PHLPP来充分发现糖尿病条件下PHLPP上调的机制。我们还将进行胰岛移植研究;将来自PHLPP 1-KO小鼠和野生型对照的胰岛移植到糖尿病小鼠中，以揭示和区分体内PHLPP 1缺失的胰岛特异性作用和全身代谢益处。最终，我们将研究实验室中可用的特定化学PHLPP抑制剂的效用、药代动力学和基于MALDI的药物定位，随后在临床前研究中测试它们在体外恢复β细胞存活和功能以及在糖尿病小鼠模型中使胰岛β细胞、β细胞存活和体内功能正常化的功效。本申请的目的是进一步阐明PHLPP在胰岛中的调节和作用的细胞和分子机制，以便将我们最初非常有希望的结果向前推进到转化研究中;建立以前未表征的PHLPP 1/2信号通路作为β细胞治疗性药理学干预的新靶点，以恢复糖尿病中的功能性胰腺β细胞群。

Both type 1 diabetes (T1D) and type 2 diabetes (T2D) result from a decline in pancreatic beta-cell function and/or mass. There is a critical need to develop therapeutic approaches to restore beta-cell mass in patients with diabetes. Apoptosis as well as impaired function of insulin producing beta-cell are hallmarks of beta-cell failure and the fundamental cause of diabetes. Modulation of beta-cell apoptosis and/or enhancement of regenerative capacity represent an attractive therapeutic approach to the treatment of diabetes. Serine-threonine phosphatases (STPs) such as PHLPP1/2 (PH domain leucine-rich repeat protein phosphatase-1/2) regulate cell death and serve as potential targets for drug development. Our understanding of the function of PHLPPs in beta-cell death regulation has important biological and clinical significance. We show that PHLPP1 and PHLPP2 are highly up-regulated in beta-cells under diabetogenic conditions in vitro, in human T1D and T2D and in mouse models of diabetes. PHLPP1/2 overexpression impairs beta-cell survival and function. PHLPP1/2 induced beta-apoptosis is mediated through the inactivation of AKT pro-survival signaling and the activation of pro-apoptotic MST1 kinase, two well-established PHLPP substrates. During the initial DFG-funded period, we have found: (i) PHLPP, AKT and MST1 form an auto-inhibitory triangle that controls beta-cell apoptosis. (ii) mTORC1 hyper-activation regulates PHLPPs up-regulation, MST1 activation and beta-cell death under diabetic conditions. (iii) Genetic and pharmacological inhibition of PHLPP restores beta-cell survival and insulin secretion in vitro and in in vivo mouse models of diabetes.In the present proposal, we continue to fully discover the mechanism of PHLPP upregulation under diabetic conditions by assessing how PHLPP is upregulated in the -cell and whether mTORC1 controls PHLPPs at the translation level. We will also perform islet transplantation studies; islets from PHLPP1-KO mice and wild-type controls are transplanted into diabetic mice in order to uncover and distinguish an islet specific action and a systemic metabolic benefit of PHLPP1 deletion in vivo. Ultimately, we will investigate the utility, pharmacokinetics and MALDI-based drug localization of specific chemical PHLPP inhibitors available in the lab, and subsequently test their efficacy to restore beta-cell survival and function in vitro and to normalize glycemia, beta-cell survival and function in vivo in a mouse model of diabetes within a preclinical study. The purpose of this application is to further elucidate the cellular and molecular mechanisms of PHLPP regulation and action in pancreatic islets in order to move our initial highly promising results forward into translational research; to establish the previously uncharacterized PHLPP1/2 signaling pathway as a novel target for beta-cell curative pharmacological intervention to restore a functional pancreatic beta-cell mass in diabetes.