Regulation of Autoimmune Type 1 Diabetes by Serpins B1 and A1 (Alpha 1-Antitrypsin)

全球糖尿病患者人数预计将从2015年的4.15亿人增加到2040年的6.42亿人。治疗这种疾病的卫生保健费用占全球卫生支出的12%，并且仍然是一个巨大的经济负担(IDF World Atlas 2015)。1型糖尿病(T1 DM)占疾病负担的10%-15%。这是一种自身免疫性疾病，被认为是由儿童早期的遗传或环境因素引发的，它会导致抗体介导的产生胰岛素的胰腺β细胞的破坏，导致危及生命的高血糖。相比之下，2型糖尿病(T2 DM)在生命后期发展，是靶组织中的胰岛素抵抗和炎症加上β细胞补偿不足的结果。有效治疗这两种疾病的一个主要限制是防止β细胞功能和/或质量的下降。最近，库尔卡尔尼实验室发现丝氨酸蛋白B1(一种细胞内丝氨酸蛋白酶抑制剂)是一种肝源性蛋白因子，它通过抑制弹性蛋白酶促进β细胞的代偿反应，并通过激活生长/生存因子信号通路在多种物种中诱导β细胞增殖。结构和功能相关的人血浆蛋白α1-抗胰蛋白酶(AAT，serpin A1)正在进行延迟T1 DM发病的晚期临床试验，此前大多数独立研究表明，它可以通过保留功能β细胞、提高同种异体胰岛移植存活和调节细胞免疫来恢复T1 DM动物模型的正常血糖。早期临床研究结果表明，AAT可降低HbA1c水平和抗胰岛细胞抗体效价。Serpin B1和AAT一样，是一种有效的抗炎弹性蛋白酶抑制剂，但与AAT不同的是，它还能诱导β细胞增殖，促进胰腺导管衬里上皮细胞中的胰岛素+细胞，抑制NETsis(另一个与糖尿病有关的促炎过程)，并调节Th17型细胞因子产生细胞的体外扩张。因此，我们建议SB1也可能在体内发挥作用，无论是单独或与AAT联合使用，以在T1 DM动物模型中提供更好的治疗效果。我们将测试SB1-/+AAT是否可以通过诱导功能性β细胞的增殖、促进导管衬里细胞产生胰岛素以及保护β细胞免受免疫反应的影响来更有效地预防或治疗NOD小鼠模型中的T1 DM。如果成功，我们将通过临床前和临床试验进一步开发这项技术，目的是改善T1 DM患者的长期结果。第一阶段的拟议研究将集中在(目标1)生产高纯度的活性重组蛋白和(目标2)在T1 DM动物模型中提供原理证明。这些研究的结果将指导未来在动物和人类身上进行疗效研究的设计。

The number of people with diabetes worldwide is predicted to increase from 415 million in 2015 to 642 million in 2040. The health care costs of treating the disease account for 12% of the global health expenditure and continue to be a huge economic burden (IDF World Atlas 2015). Type 1 diabetes mellitus (T1DM) constitutes 10-15% of the disease burden. It is an autoimmune disease, thought to be triggered by genetic or environmental factors in early childhood, which results in antibody- mediated destruction of insulin producing pancreatic β-cells causing life-threatening hyperglycemia. In contrast, type 2 diabetes mellitus (T2DM) develops later in life and results from insulin resistance in target tissues and inflammation coupled with an inadequate compensation by the β-cells. A major limitation in effectively treating both forms of the disease is preventing a decline in β- cell function and/or mass. Recently, the Kulkarni lab identified Serpin B1 (an intracellular SERine Proteinase Inhibitor) as a liver-derived protein factor that promotes a compensatory β-cell response by inhibiting elastase and inducing β-cell proliferation in multiple species by activating growth/survival factor signaling pathways. The structurally and functionally related human plasma protein alpha 1- antitrypsin (AAT, serpin A1) is in late stage clinical trials to delay the onset of T1DM after a majority of independent studies showed it could restore euglycemia in animal models of T1DM by preserving functional β-cells, enhancing islet allograft survival and modulating cellular immunity. Early stage clinical study results indicate that AAT reduces HbA1c levels and anti-islet cell antibody titers. Serpin B1 (sB1), like AAT, is an effective anti-inflammatory elastase inhibitor but unlike AAT, also induces β-cell proliferation, promotes insulin + cells in pancreatic ductal lining epithelia, inhibits NETosis (another pro-inflammatory process involved in diabetes) and regulates the expansion of Th17 cytokine producing cells in vitro. We therefore propose that sB1 may also work in vivo, either alone or in combination with AAT, to provide superior therapeutic effects in animal models of T1DM. We will test if sB1 -/+ AAT can more effectively prevent the onset of, or treat established, T1DM in the NOD mouse model by inducing proliferation of functional β-cells, enhancing production of insulin by ductal lining cells and protecting β-cells from the immune response. If successful, we will further develop this technology through preclinical and clinical testing with the intent to improve long-term outcome for patients with T1DM. The proposed research in phase I will focus on (Aim 1) producing highly purified active recombinant proteins and (Aim 2) providing proof-of-principle in animal models of T1DM. The outcome of these studies will guide the design of future efficacy studies in animals and humans.