A Novel Approach to Restore Epithelial Barrier Homeostasis to Treat Inflammatory Bowel Disease

恢复上皮屏障稳态以治疗炎症性肠病的新方法

• 批准号: 9909608
• 负责人: W Vallen Graham
• 金额: $ 22.43万
• 依托单位: THELIUM THERAPEUTICS INC
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-09-15 至 2021-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9909608
• 关键词: Actomyosin; Acute; Acute Disease; Adverse effects; Affect; Affinity; Anti-Tumor Necrosis Factor Therapy; Binding; Biology; Biopsy; Cardiac Muscle Contraction; Celiac Disease; Cell Adhesion Molecules; Cell Line; Cellular Assay; Characteristics; Chemicals; Chronic; Chronic Disease; Clinical Drug Development; Clinical Trials; Crystallization; Data; Development; Diarrhea; Disease; Disease model; Docking; Endocytosis; Endothelium; Enteral; Enterocolitis; Enzymes; Epithelial; Epithelial Cells; Epithelium; Event; Experimental Animal Model; Experimental Models; Food Hypersensitivity; Foundations; Functional disorder; Gastrointestinal Diseases; Health; Homeostasis; Human; Hydrophobicity; Hypotension; Immune; Immunoglobulins; Immunosuppressive Agents; In Vitro; Inflammation; Inflammatory; Inflammatory Bowel Diseases; Interleukin-10; Intestinal Diseases; Intestines; Knockout Mice; Lead; Libraries; Light; Measures; Mediating; Medicine; Methods; Molecular; Mucous Membrane; Mus; Muscle; Muscle Contraction; Myosin Alkali Light Chains; Myosin Light Chain Kinase; Myosin Type II; Nature; Nutrient; Outcome; Permeability; Pharmaceutical Preparations; Phase; Phosphorylation; Phosphotransferases; Physics; Physiological; Positioning Attribute; Property; Publishing; RNA Splicing; Regulation; Reporting; Rho-associated kinase; Role; Signal Transduction; Site; Skeletal Muscle; Small Business Innovation Research Grant; Smooth Muscle; Specificity; Stimulus; Structure; Systemic disease; T-Lymphocyte; TNF gene; Tertiary Protein Structure; Therapeutic; Therapeutic Agents; Tight Junctions; Tissues; Toxic effect; Variant; Waste Products; Water; Work; Wound Healing; active method; base; clinical application; clinical candidate; cytokine; design; graft vs host disease; immune activation; immunomodulatory therapies; improved; in vivo; infection risk; insight; intestinal epithelium; lead candidate; lead optimization; non-muscle myosin; novel strategies; novel therapeutics; nutrient absorption; occludin; pathogen; phase 1 study; pre-clinical; preservation; prevent; recruit; repaired; restoration; screening; simulation; small molecule; standard of care; systemic toxicity; therapeutic target; tool

SUMMARY Intestinal barrier function is compromised in enteric and systemic diseases, including infectious enterocolitis, food allergy, celiac disease, graft versus host disease (GvHD), and inflammatory bowel disease (IBD). The co-founders of Thelium Therapeutics discovered the central role of myosin light chain kinase (MLCK) in barrier regulation and demonstrated that targeted intestinal epithelial MLCK inhibition limits experimental IBD and GvHD. Unfortunately, severe toxicities associated with barrier-independent MLCK functions in epithelia and other tissues, e.g., smooth muscle, preclude therapeutic targeting of MLCK enzymatic activity. We recently reported (Graham et al., Nature Medicine, 2019) that a specific MLCK splice variant, MLCK1, is central to barrier regulation and depends on interactions mediated by immunoglobulin-cell adhesion molecule domain 3 (IgCAM3). We solved the IgCAM3 crystal structure, identified a drug binding pocket unique to IgCAM3, and screened a library of ~140,000 drug-like molecules. One compound, Divertin, bound IgCAM3, prevented cytokine-induced MLCK1 recruitment, myosin II regulatory light chain phosphorylation, and barrier dysfunction. Critically, Divertin did not inhibit MLCK enzymatic function, epithelial wound healing, or smooth muscle contraction, and in vivo toxicity studies failed to identify adverse effects. Divertin prevented acute TNF-induced barrier loss in vivo (mice) and ex vivo (human intestinal biopsies) and restored immune-mediated barrier loss in vivo (IL-10 knockout mice). Finally, Divertin delayed onset and prevented progression of experimental immune-mediated (T cell transfer) IBD, as indicated by barrier preservation and restoration, reduced mucosal immune activation, and enhanced survival. Although useful as a tool compound, Divertin lacks characteristics required for a lead compound. This proposal will overcome that obstacle to clinical application of Divertin by discovering lead compounds with similar activities. Cutting edge, physics-based rational design and molecular dynamic simulation methods has already been used to probe an expansive compound library and identify those with predicted high-affinity binding to the MLCK1 IgCAM3 crystal structure. Those molecules will be subjected to a rank order screening funnel to identify compounds with suitable MLCK1 binding affinities, efficacy in preserving and restoring epithelial barrier function, and absence of enzymatic inhibitory activity. These lead compounds will be suitable for optimization and development to facilitate IND-enabling studies for a first-in-class barrier-restorative therapy to manage gastrointestinal and systemic diseases.

摘要 肠道和全身疾病，包括传染性疾病，肠道屏障功能受损 小肠结肠炎、食物过敏、腹腔疾病、移植物抗宿主病(GvHD)和炎症性疾病 肠道疾病(IBD)。钍治疗公司的联合创始人发现了 肌球蛋白轻链激酶(MLCK)在屏障调节和靶向肠道中的作用 上皮MLCK抑制可限制实验性IBD和GvHD。不幸的是，严重的毒副作用 与上皮和其他组织中的非屏障MLCK功能有关，如Smooth 肌肉，排除MLCK酶活性的治疗靶点。我们最近报道了 (Graham等人，自然医学，2019)认为特定的MLCK剪接变异体MLCK1是 屏障调节及其依赖于免疫球蛋白-细胞黏附的相互作用 分子结构域3(IgCAM3)。我们解析了IgCAM3的晶体结构，鉴定了药物结合 Pocket是IgCAM3独有的，并筛选了一个约140,000个类药物分子的文库。一 Divertin，结合IgCAM3的化合物，阻止细胞因子诱导的MLCK1募集，肌球蛋白II 调节轻链磷酸化和屏障功能障碍。关键的是，Divertin并没有抑制 MLCK酶功能，上皮伤口愈合，或平滑肌收缩，以及体内 毒性研究未能确定不良反应。Divertin预防急性肿瘤坏死因子诱导的屏障 体内(小鼠)和体外(人类肠道活检)丢失和恢复的免疫介导性 体内屏障丧失(IL-10基因敲除小鼠)。最后，Divertin延迟起效并预防 屏障提示实验性免疫介导(T细胞转移)IBD的研究进展 保存和修复，降低粘膜免疫活性，提高存活率。 尽管Divertin作为一种工具化合物很有用，但它缺乏铅所需的特征 化合物。该建议将通过以下方式克服Divertin临床应用的障碍 发现具有相似活性的先导化合物。前沿、基于物理的理性设计 分子动力学模拟方法已经被用来探索一种可扩展的 并鉴定预测与MLCK1 IgCAM3高亲和力结合的化合物文库 晶体结构。这些分子将接受排序筛选漏斗以识别 具有合适的MLCK1结合亲和力的化合物，在保存和修复上皮细胞方面的有效性 屏障功能，缺乏酶抑制活性。这些先导化合物将是 适合于优化和开发，以促进一流的IND支持研究 屏障修复疗法，用于管理胃肠道和全身疾病。