MOLECULAR ENGINEERING OF IL-2 FOR THE TREATMENT AND PREVENTION OF TYPE I DIABETES

用于治疗和预防 I 型糖尿病的 IL-2 分子工程

• 批准号: 8255168
• 负责人: Aaron Michael Ring
• 金额: $ 4.18万
• 依托单位: STANFORD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-09-30 至 2014-09-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8255168
• 关键词: Acute; Affect; Affinity; Alleles; Autoimmune Diseases; Autoimmune Process; Binding; Calorimetry; Cell physiology; Cells; Chronic; Clinical; Collaborations; Complex; Couples; Coupling; Crystallography; Cytokine Receptors; Cytokine Signaling; Cytometry; Development; Diabetes Mellitus; Disease; Distal; Dominant-Negative Mutation; Effector Cell; Engineering; Etiology; Goals; IL2RA gene; Immune; Immunity; In Vitro; Inbred NOD Mice; Insulin; Insulin-Dependent Diabetes Mellitus; Interleukin 2 Receptor; Interleukin-2; Kinetics; Laboratories; Ligands; Lymphocyte; MAP Kinase Gene; MAPK3 gene; Malignant Neoplasms; Measures; Mediating; Metabolic Control; Methodology; Methods; Molecular; Molecular Structure; Morbidity - disease rate; Natural Killer Cells; Output; PI3K/AKT; Pancreas; Pathway interactions; Patients; Prevention; Property; Proteins; Receptor Signaling; Regulatory T-Lymphocyte; Relative (related person); STAT5A gene; Screening procedure; Signal Transduction; Signaling Molecule; Site-Directed Mutagenesis; Solutions; Structure of beta Cell of islet; Surface; Surface Plasmon Resonance; T-Cell Proliferation; Testing; Therapeutic; Therapeutic immunosuppression; Titrations; Variant; X-Ray Crystallography; Yeasts; allograft rejection; analog; autoreactive T cell; base; cell type; combinatorial; cytokine; cytotoxicity; design; glycemic control; mortality; mouse model; multidisciplinary; novel; prevent; receptor; receptor binding; translational study

DESCRIPTION (provided by applicant): Type I diabetes (T1D) is a common and devastating disease characterized by autoimmune destruction of the insulin-producing beta cells of the pancreas. The ensuing condition leaves the affected patients unable to produce sufficient insulin for proper metabolic and glycemic control, resulting in acute and chronic complications that cause extensive morbidity and mortality. There is considerable hope, however, that T1D may be preventable, or even reversed, with the timely administration of immunosuppressive therapy. No such therapy has yet been approved, and considerable need exists for the development of novel T1D therapeutics. Interleukin-2 (IL-2) is an immunoregulatory cytokine with critical importance to the etiology of T1D: while IL-2 is essential to maintain the regulatory T cells (Tregs) that prevent the disease, it also stimulates autoreactive lymphocytes that attack the pancreas and cause T1D. This proposal describes the engineering of novel IL-2 based therapeutics tailored for the prevention and treatment of T1D by splitting the antithetical functions of IL-2 that occur in T1D. IL-2 analogues will be designed through rationale, site-directed mutagenesis (Aim 1) and through a combinatorial approach that couples yeast-surface display to phosphoflow cytometry to screen thousands of IL-2 variants on the basis of signaling output (Aim 2). Functional characterization of these synthetic IL-2 molecules will include in vitro asays of IL-2 dependent cellular functions, such as T cell proliferation and NK cell cytotoxicity. Biophysical characterization of these IL-2 analogues will employ surface plasmon resonance to measure affinities and binding kinetics to the IL-2 receptor subunits and x-ray crystallography to determine the molecular structures of the IL-2/IL-2 receptor complexes. Ultimately, and most importantly, candidate IL-2 therapeutics developed through the methods above will be assessed for efficacy in the prevention and treatment of diabetes in the non-obese diabetic (NOD) mouse model of T1D. Thus, through these multidisciplinary and translational studies, we will advance new candidate T1D therapies for clinical development. ! PUBLIC HEALTH RELEVANCE: Type I diabetes (T1D) is a common autoimmune disease with major impacts on morbidity and mortality. It is widely believed, however, that the disease could be prevented or reversed if an appropriate therapeutic were administered before irreversible damage to the pancreas occurred. The goal of this project is to engineer and characterize new immunosuppressive therapies targeting the protein Interleukin-2 (IL-2) that would be suitable for the treatment and prevention of T1D.

描述（由申请人提供）：I 型糖尿病 (T1D) 是一种常见的破坏性疾病，其特征是胰腺产生胰岛素的 β 细胞受到自身免疫性破坏。随后的情况使受影响的患者无法产生足够的胰岛素来进行适当的代谢和血糖控制，从而导致急性和慢性并发症，导致广泛的发病率和死亡率。然而，通过及时进行免疫抑制治疗，T1D 是可以预防甚至逆转的，这一点有很大的希望。此类疗法尚未获得批准，并且非常需要开发新型 T1D 疗法。白细胞介素-2 (IL-2) 是一种免疫调节细胞因子，对于 T1D 的病因学至关重要：虽然 IL-2 对于维持预防疾病的调节性 T 细胞 (Treg) 至关重要，但它也会刺激攻击胰腺的自身反应性淋巴细胞并导致 T1D。该提案描述了通过分解 T1D 中出现的 IL-2 的对立功能来设计用于预防和治疗 T1D 的新型基于 IL-2 的疗法。 IL-2 类似物将通过基本原理、定点诱变（目标 1）以及通过将酵母表面展示与磷流式细胞术相结合的组合方法来设计，以根据信号输出筛选数千种 IL-2 变体（目标 2） 。这些合成 IL-2 分子的功能表征将包括 IL-2 依赖性细胞功能的体外测定，例如 T 细胞增殖和 NK 细胞的细胞毒性。这些 IL-2 类似物的生物物理表征将采用表面等离子体共振来测量与 IL-2 受体亚基的亲和力和结合动力学，并采用 X 射线晶体学来确定 IL-2/IL-2 受体复合物的分子结构。最终，最重要的是，通过上述方法开发的候选 IL-2 疗法将在 T1D 的非肥胖糖尿病 (NOD) 小鼠模型中评估其预防和治疗糖尿病的功效。因此，通过这些多学科和转化研究，我们将推进新的 T1D 候选疗法的临床开发。 ！ 公共卫生相关性：I 型糖尿病 (T1D) 是一种常见的自身免疫性疾病，对发病率和死亡率有重大影响。然而，人们普遍认为，如果在胰腺发生不可逆转的损伤之前给予适当的治疗，则可以预防或逆转这种疾病。该项目的目标是设计和表征针对蛋白质白细胞介素 2 (IL-2) 的新型免疫抑制疗法，该疗法适用于治疗和预防 T1D。