Investigating mechanisms of epidermal cell death in Toxic Epidermal Necrolysis

研究中毒性表皮坏死松解症中表皮细胞死亡的机制

• 批准号: 10244900
• 负责人: Sherrie Jill Divito
• 金额: $ 22.38万
• 依托单位: BRIGHAM AND WOMEN'S HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-08-21 至 2024-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10244900
• 关键词: 3-Dimensional; Address; Affect; Apoptosis; Biological Assay; Biological Models; Blindness; Bulla; CD8-Positive T-Lymphocytes; Cell Death; Cells; Cessation of life; Clinical; Computer software; Coupled; Cutaneous; Cytoplasmic Granules; Data; Databases; Diagnosis; Disease; Drug Hypersensitivity; Epidermis; Esophageal Stenosis; Eye; FDA approved; Flow Cytometry; Formalin; Gene Expression Profiling; Genetic Transcription; Genital; Genitalia; Goals; Healthcare Systems; Helper-Inducer T-Lymphocyte; Hospitals; Human Engineering; Image Analysis; Immune; Immunoassay; Immunofluorescence Immunologic; Inflammatory; Intervention; Knowledge; Laboratories; Life; Ligands; Liquid substance; Lung diseases; Mediating; Mediator of activation protein; Microscopy; Modeling; Oral cavity; Organ; Paraffin Embedding; Pathogenesis; Pathway interactions; Patients; Pharmaceutical Preparations; Pharyngeal structure; Proteins; Proteomics; Publishing; Reaction; Reporting; Reproducibility; Research; Resources; Role; Sampling; Skin; Source; Specimen; Stains; Stevens-Johnson Syndrome; Survivors; TNF gene; TNFSF10 gene; Techniques; Technology; Testing; Thick; Tissue Engineering; Toxic Epidermal Necrolysis; clinical care; clinical imaging; cost; cost estimate; cytotoxic CD8 T cells; disorder prevention; imaging system; immunological synapse; improved; inhibitor/antagonist; innovation; innovative technologies; keratinocyte; mortality; nano-string; new technology; novel; pathology imaging; prospective; receptor; single-cell RNA sequencing; stem; systemic inflammatory response; tertiary care; transcriptome sequencing

Project Summary Stevens Johnson Syndrome/Toxic Epidermal Necrolysis (SJS/TEN) is a severe drug hypersensitivity reaction manifesting as sloughing of skin, eyes, mouth, throat and genitals, along with systemic inflammation and internal organ involvement. Disease can occur in otherwise healthy people and is largely unpredictable. Mortality is 20- 30% and there is no known treatment. SJS/TEN is a substantial burden on the US healthcare system, costing an estimated 129 million dollars per year. It is of paramount importance to elucidate the pathobiology of SJS/TEN to improve disease prevention, diagnosis and treatment. Despite this clear need, SJS/TEN is significantly under-researched. The dearth of high-quality research in this field stems from two major barriers: a lack of patient samples available for analysis and the absence of an easily attainable and reproducible model system to perform mechanistic studies. One major gap in our knowledge of the pathogenesis of SJS/TEN is the putative mechanism(s) by which immune cells mediate keratinocyte death. It is largely assumed that cytolytic granules generated by cytotoxic CD8+ T cells induce keratinocyte apoptosis. However, contradictory findings call this assumption into question. The central hypothesis of this proposal is that the inflammatory cell death pathways necroptosis and pyroptosis, rather than apoptosis, mediate keratinocyte death and epidermal destruction in SJS/TEN. The proposed study will directly test this hypothesis while overcoming current barriers in the field. Aim 1 employs two novel technologies to study a large bank of formalin-fixed paraffin embedded clinical specimens that until now were of limited research utility. Clinical specimens are drawn from a meticulously developed and thoroughly vetted patient database that spans three major tertiary care hospitals. These technologies are (i) OpalTM multispectral immunofluorescence staining with MantraTM Quantitative Pathology Imaging System and inFORM® Image Analysis Software and (ii) NanoString NCounter® platform for gene expression profiling. Aim 2 interrogates mechanism(s) of keratinocyte death by partnering with burn teams at two major centers to prospectively collect blister fluid and affected epidermis, a unique sample resource. Analysis utilizes RNAseq, high parameter flow cytometry, and multiplex immunoassay in concert with traditional laboratory techniques. Aim 3 generates a novel ex vivo model of SJS/TEN using patient blister fluid with commercially available 3D full thickness tissue engineered human skin that will be used to investigate the proposed hypothesis while also exploring potential novel treatments. The data generated through this innovative research has significant potential to advance understanding of fundamental disease pathobiology and clinical care.

项目概要 史蒂文斯约翰逊综合征/中毒性表皮坏死松解症 (SJS/TEN) 是一种严重的药物过敏反应 表现为皮肤、眼睛、口腔、喉咙和生殖器脱落，并伴有全身炎症和内部炎症 器官受累。疾病可能发生在其他健康的人身上，并且在很大程度上是不可预测的。死亡率为20- 30%，并且没有已知的治疗方法。 SJS/TEN 是美国医疗保健系统的沉重负担，成本高昂 估计每年1.29亿美元。阐明其病理学至关重要 SJS/TEN 改善疾病预防、诊断和治疗。尽管有这个明确的需求，SJS/TEN 研究严重不足。该领域缺乏高质量研究源于两个主要障碍： 缺乏可用于分析的患者样本，并且缺乏易于获得和可重复的模型 系统进行机械研究。我们对 SJS/TEN 发病机制了解的一个主要空白是 免疫细胞介导角质形成细胞死亡的推定机制。人们普遍认为，溶细胞作用 细胞毒性 CD8+ T 细胞产生的颗粒诱导角质形成细胞凋亡。然而，矛盾的发现呼吁 这个假设受到质疑。该提案的中心假设是炎症细胞死亡 坏死性凋亡和细胞焦亡（而不是细胞凋亡）介导角质形成细胞死亡和表皮细胞凋亡 SJS/TEN 中的破坏。拟议的研究将直接检验这一假设，同时克服当前的障碍 在外地。目标 1 采用两种新技术来研究大量福尔马林固定石蜡包埋的 迄今为止，研究用途有限的临床标本。临床标本是从精心采集的 开发并彻底审查了涵盖三个主要三级护理医院的患者数据库。这些 技术包括 (i) OpalTM 多光谱免疫荧光染色与 MantraTM 定量病理学 成像系统和 inFORM® 图像分析软件和 (ii) NanoString NCounter® 基因平台 表达分析。目标 2 通过与烧伤团队合作探究角质形成细胞死亡的机制 两个主要中心前瞻性地收集水疱液和受影响的表皮，这是一种独特的样本资源。分析 与传统实验室相结合，利用 RNAseq、高参数流式细胞术和多重免疫分析 技术。目标 3 使用患者水疱液生成 SJS/TEN 的新型离体模型 可用的 3D 全厚度组织工程人体皮肤，将用于研究所提出的假设 同时也在探索潜在的新疗法。通过这项创新研究产生的数据 促进对基础疾病病理学和临床护理的理解具有巨大潜力。

项目成果

Physical and mental health impact of Stevens-Johnson syndrome/toxic epidermal necrolysis and post-hospital discharge care: Identifying practice gaps.

• DOI: 10.1016/j.jdin.2023.01.012
• 发表时间: 2023-06
• 期刊: JAAD international
• 作者: Coromilas, Alexandra J.;Divito, Sherrie J.;Phillips, Elizabeth J.;Micheletti, Robert G.
• 通讯作者: Micheletti, Robert G.

Advances in the Pathomechanisms of Delayed Drug Hypersensitivity.

• DOI: 10.1016/j.iac.2022.01.002
• 发表时间: 2022-03
• 期刊: Immunology and allergy clinics of North America
• 影响因子: 2.6
• 作者: Chuang-Wei Wang;S. Divito;W. Chung;S. Hung

Assessment of Need for Improved Identification of a Culprit Drug in Stevens-Johnson Syndrome/Toxic Epidermal Necrolysis.

史蒂文斯-约翰逊综合征/中毒性表皮坏死松解症罪魁祸首药物鉴定需要改进的评估。

• DOI: 10.1001/jamadermatol.2023.1693
• 发表时间: 2023
• 期刊: JAMA dermatology
• 影响因子: 10.9
• 作者: Li,DayanJ;Velasquez,GustavoA;Romar,GeorgeA;Schunkert,ElisaM;Foreman,RuthK;Divito,SherrieJ
• 通讯作者: Divito,SherrieJ