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: 8413125
• 负责人: Aaron Michael Ring
• 金额: $ 4.22万
• 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/8413125
• 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. !

描述（由申请人提供）：I型糖尿病（T1D）是一种常见且毁灭性的疾病，其特征是胰腺产生胰岛素β细胞的自身免疫性破坏。随之而来的疾病使受影响的患者无法产生足够的胰岛素以进行适当的代谢和血糖控制，从而导致急性和慢性并发症，从而导致广泛的发病率和死亡率。但是，有很大的希望，可以及时进行免疫抑制疗法，可以预防甚至可以逆转T1D。尚未批准这种疗法，并且需要大量发展新型T1D治疗疗法。白介素-2（IL-2）是一种免疫调节性细胞因子，对T1D的病因至关重要：而IL-2对于维持预防疾病的调节性T细胞（TREG）至关重要，它也刺激自动诊断性淋巴细胞，攻击胰腺并引起T1D。该提案描述了针对预防和治疗T1D的新型基于IL-2的治疗剂的工程，通过分解T1D中IL-2的对立功能。 IL-2类似物将通过基本原理，定向的诱变（AIM 1）以及通过组合方法将酵母表面显示到磷酸流细胞仪，以基于信号输出的基础筛选数千个IL-2变体（AIM 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体外表征，例如T细胞增殖和NK细胞细胞毒性。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受体亚基和X射线晶体学的亲和力和结合动力学，以确定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治疗剂将在预防和治疗T1D的非肥胖糖尿病（NOD）小鼠模型中的预防和治疗中评估。因此，通过这些多学科和翻译研究，我们将推进新的候选T1D疗法用于临床开发。呢