Discovery of autoimmune antigens from non-coding sequences

从非编码序列发现自身免疫抗原

• 批准号: 10286063
• 负责人: Miriam Isaaca Rosenberg
• 金额: $ 18.08万
• 依托单位: HEBREW UNIVERSITY OF JERUSALEM
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-07-13 至 2023-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10286063
• 关键词: Acute; Address; Adult; Affinity; Antibodies; Antigens; Autoantibodies; Autoantigens; Autoimmune; Autoimmune Diseases; Autoimmunity; Bacteriophage T7; Bacteriophages; Base Sequence; Binding; Biological Assay; Biological Markers; Brain; Cells; Cerebrospinal Fluid; Characteristics; Childhood; Clinical; Complex; DNA Transposable Elements; Diagnosis; Disease; Disease Marker; Embryo; Encephalitis; Epitopes; Genes; Genome; Goals; HLA Antigens; Human; Immune Targeting; Immune system; Immunoprecipitation; Immunosuppression; Knowledge; Lead; Length; Libraries; Light; Link; Mass Spectrum Analysis; Methods; Molecular; Monitor; Multiple Sclerosis; N-Methyl-D-Aspartate Receptors; Neuroimmune; Neuromyelitis Optica; Neurons; Oligonucleotides; Open Reading Frames; Opsoclonus-Myoclonus Syndromes; Orphan; Pathway interactions; Patients; Peptide Library; Peptides; Phage Display; Phage ImmunoPrecipitation Sequencing; Positioning Attribute; Process; Proteins; Proteome; Random Peptide Libraries; Reporting; Risk; Sampling; Serum; Source; Specificity; Testing; Translating; Translations; Tumor Antigens; Untranslated RNA; Untranslated Regions; Variant; Virion; Work; adaptive immunity; antigen diagnostic; base; cohort; combinatorial; design; expression vector; high risk; immunogenic; improved; mimetics; neglect; next generation sequencing; novel diagnostics; patient screening; pediatric patients; population based; screening; specific biomarkers; success; tool; tumor

7. Project Summary. This project addresses an acute clinical need for the many neuroimmune diseases with no known causative antigen, and those with variation that remains unexplained by single defining antigens. Antibodies in NMDA-receptor encephalitis (α-NMDAR) have revolutionized diagnosis and treatment of that paraneoplastic disease (PND), while other PNDs, like Opsoclonus Myoclonus Syndrome (OMS), await these groundbreaking discoveries. Several methods, such as Phage immunoprecipitation and sequencing (PhIP-seq) have been used successfully to pan for biomarkers and defining autoantigens. However, for some diseases, these methods have failed. One possibility is that defining antigens have been missed by conventional screens because of unconventional origins. We hypothesize that some important autoantigens may reside within “non-coding” sequences. These include the vast landscape of small open reading frames (sORFs), or “micropeptides”, that originate in regions of the genome, like UTRs and long “non-coding” RNAs, that may have been erroneously overlooked in past screens. Importantly for this work, their proven translation is temporally and spatially regulated, they can be tumor specific antigens and are presented to the immune system. A second source of untested proteins in these assays is transposable elements. The proteins they naturally encode, and those that arise from their transposition into active genes, are also potential sources of immunogenic material. While few are active, their expression in embryos, adult brain and in tumor suggest their potential relevance for paraneoplastic autoimmunity. Cryptic peptides generated by a host of mechanisms remain a further challenge to screens that rely on defined target reference peptidomes. Our study aims to improve upon previous PhIPseq searches, by designing and building new libraries that cover this unconventional set of peptides. In Aim 1, we will design and build two new T7 PhIP-seq libraries covering this defined “non-coding” peptidome: one with targets represented as arrays of oligos encoding overlapping peptides, and a second in which full-length targets are expressed, to mitigate risk of disrupting epitopes by sequence fragmentation. In Aim 2, we will screen samples from pediatric neuroimmune disease patients (and controls) against these gene-specific libraries, to identify distinctive signatures of reactivity. To capture information about cryptic antigenic peptides, Aim 3 will test the same samples against an extant random peptide library, which reports on convergent feature specificity of patient samples from each disorder. This high-risk approach, if successful, will lead quickly to revolutionary new diagnostic paradigms for the neuroimmune diseases tested, as it did for NMDAR encephalitis, with paradigm-shifting potential for other autoimmune diseases, as well.

7。项目摘要。 该项目解决了许多神经免疫性疾病的急性临床需求 因定义抗原而导致的抗原和具有变化的抗原。 NMDA受体脑炎（α-NMDAR）的抗体已彻底改变了诊断和治疗 那种副塑性疾病（PND），而其他PND，例如Opsoclonus肌阵挛综合征（OMS）， 等待这些开创性的发现。几种方法，例如噬菌体免疫沉淀和 测序（PHIP-SEQ）已成功用于生物标志物和定义自身抗原。 但是，对于某些疾病，这些方法失败了。一种可能性是定义抗原 由于非常规的起源，传统屏幕遗漏了。我们假设这一点 一些重要的自身抗原可能位于“非编码”序列中。这些包括疫苗 小型开放式阅读框（SORF）或“微肽”的景观，起源于区域 基因组，例如UTR和长期的“非编码” RNA，可能被错误地忽略了 屏幕。重要的是，对于这项工作，他们经过验证的翻译经过暂时和空间的监管，他们 可以是肿瘤特异性抗原，并呈现给免疫系统。第二个来源 这些测定中未经测试的蛋白质是转座元件。它们自然编码的蛋白质，以及 那些由将其转置分为活性基因的人也是免疫原性的潜在来源 材料。虽然很少有活性，但它们在胚胎，成人大脑和肿瘤中的表达表明它们 与副塑性自身免疫的潜在相关性。寄主产生的神秘胡椒 机制仍然是依赖定义目标参考肽组的筛查的进一步挑战。 我们的研究旨在通过设计和构建新图书馆来改善以前的Phipseq搜索 这涵盖了这组非常规的肽。在AIM 1中，我们将设计和构建两个新的T7 PHIP-SEQ 涵盖此定义的“非编码”胡椒的库：一个具有目标的目标 编码重叠肽的寡核体，第二个表达全长目标， 通过序列碎片化来减轻破坏表位的风险。在AIM 2中，我们将从 针对这些基因特异性文库的小儿神经免疫性疾病患者（和对照），以鉴定 反应性的独特签名。为了捕获有关加密抗原肽的信息，AIM 3将 针对额外的随机胡椒库测试相同的样品，该库报告了收敛功能 来自每种疾病的患者样本的特异性。如果成功的话，这种高风险的方法将领导 迅速转化为已测试神经免疫性疾病的革命性新诊断范例 NMDAR脑炎，也具有其他自身免疫性疾病的范式转移潜力。