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: 10450060
• 负责人: Miriam Isaaca Rosenberg
• 金额: $ 11.6万
• 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 2024-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10450060
• 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.项目总结。 该项目解决了许多未知的神经免疫疾病的迫切临床需求。 致病抗原，以及那些仍无法用单一定义抗原解释的变异。 N-甲基-D-天冬氨酸受体脑炎抗体(α-NMDAR)使脑炎的诊断和治疗发生了革命性的变化 副肿瘤性疾病(PND)，而其他PND，如眼阵挛肌阵挛综合征(OMS)， 等待着这些开创性的发现。几种方法，如噬菌体免疫沉淀和 测序(PhIP-seq)已经成功地用于PAN生物标记物和定义自身抗原。 然而，对于某些疾病，这些方法都失败了。一种可能性是定义抗原 因为非传统的起源而被传统的银幕遗漏。我们假设 一些重要的自身抗原可能存在于“非编码”序列中。其中包括广大的 小开放阅读框(SORF)，或“微肽”的景观，起源于 基因组，如UTRs和长的“非编码”RNA，这在过去可能被错误地忽视 屏幕。对这项工作来说，重要的是，他们被证明的翻译是在时间和空间上受到调节的，他们 可以是肿瘤特异性抗原，并被呈递到免疫系统。第二个来源 这些检测中未检测的蛋白质是转座元件。它们自然编码的蛋白质，以及 那些通过转座进入活性基因而产生的基因，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、成人脑和肿瘤中的表达表明 副肿瘤自身免疫的潜在相关性。由大量的 对于依赖于已定义的靶标参考肽的筛查来说，机制仍然是一个进一步的挑战。 我们的研究旨在通过设计和建立新的图书馆来改进以前的PhIPseq搜索 涵盖了这一组非传统的多肽。在目标1中，我们将设计和构建两个新T7 PhIP-seq 涵盖这个已定义的“非编码”多肽的库：一个目标表示为 编码重叠多肽的寡核苷酸，以及表达全长靶标的第二个寡核苷酸，以 降低通过序列碎片破坏表位的风险。在目标2中，我们将筛选来自 儿科神经免疫性疾病患者(和对照)对照这些基因特异性文库，以确定 反应性的独特特征。为了获取有关隐蔽抗原肽的信息，Aim 3将 用现有的随机多肽文库测试相同的样本，该文库报告收敛特征 每种疾病患者样本的特异性。这种高风险的方法，如果成功，将导致 迅速为测试的神经免疫性疾病提供革命性的新诊断范例，就像它对 NMDAR脑炎，也有可能导致其他自身免疫性疾病。