A Novel Human Virus in Patients with Cryptogenic Liver Disease

隐源性肝病患者中的一种新型人类病毒

• 批准号: 10636331
• 负责人: XIAOFENG FAN
• 金额: $ 22.73万
• 依托单位: SAINT LOUIS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-08-01 至 2025-07-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10636331
• 关键词: Acute Hepatitis; Acute Liver Failure; Adult; Age; Antibodies; Antibody Response; Autoimmunity; Blood donor; Capsid Proteins; Cause of Death; Chronic Hepatitis; Cirrhosis; Classification; Clinical; Clinical Trials; Cloning; Cohort Studies; DNA; Data; Dideoxy Chain Termination DNA Sequencing; Digestion; Double Stranded DNA Virus; Enzyme-Linked Immunosorbent Assay; Etiology; Exclusion; GB virus C; Genbank; Genes; Genetic Diseases; Genome; Grant; Hepatitis; Hepatitis A; Hepatitis C; Hepatitis C virus; Hepatitis Viruses; Human; Immunoglobulin G; Immunoglobulin M; Individual; Knowledge; Lead; Life; Ligation; Link; Liver; Liver diseases; Living Donor Liver Transplantation; Machine Learning; Malignant neoplasm of liver; Measurement; Mediating; Membrane; Methods; Morphologic artifacts; Names; Nature; Nucleic Acids; Patients; Peptides; Prevention; Primer Extension; Probability; Reagent; Reporting; Research; Scanning; Sensitivity and Specificity; Series; Serology test; Serum; Single-Stranded DNA; Specimen; Structure; Surface; Testing; Tissues; Transfusion-Transmitted Virus; Transplant Recipients; Tropism; United States; United States National Institutes of Health; Venous blood sampling; Viral; Viral Genome; Virus; Vital Statistics; Walking; chronic liver disease; clinical phenotype; design; drug induced liver injury; ds-DNA; experimental study; human virome; hydropathy; in silico; liver transplantation; next generation sequencing; nonalcoholic steatohepatitis; novel; problem drinker; response; sequencing platform; virome; whole genome

Project Summary A substantial portion of patients with liver disease, ranging from 5% to 30%, have unknown causes beyond the established etiologies. Unknown etiology is observed across a wide array of clinical phenotypes in liver disease, such as acute liver failure (ALF), hepatitis, cirrhosis, and liver cancer. These are collectively referred to as cryptogenic liver disease (CLD). It has long been hypothesized there exist additional human viruses that cause CLD. In our recent serum virome study, we identified a 387-nt DNA fragment (GenBank MW468091), named Seq260, from 1 of 9 CLD patients. In a series of experiments of gene-walking, enzymatic digestion, and rolling circle amplification and analyses, we have demonstrated that Seq260 is a linear single-stranded DNA. We screened Seq260 in 409 subjects, including healthy blood donors (n=200), hepatitis C virus infection (n=100), Acute liver failure (ALF) patients with indeterminate etiology (n=50), and liver transplantation (LT) patients with (n=45) and without known etiology (n=14). Seq260 was detected in 5 CLD patients (1 ALF and 4 LT) and 1 LT patient with nonalcoholic steatohepatitis (NASH)-associated cirrhosis. One patient had Seq260 quantifiable in liver, showing a titer in the liver 7.74 times higher than that in serum (2.4x106 copies/g vs. 3.1x105 copies/mL). Machine learning analysis reached a high score (likelihood) of Seq260 being a eukaryotic viral sequence. Aggregately, these data lead to our hypothesis that Seq260 represents an unrecognized human virus with liver tropism. To determine if Seq260 represents a novel hepatitis virus, we bring about a research plan in the current proposal that consists of three major experiments. First, we will screen Seq260 in CLD patients as well as the controls. We have been granted access to patient specimens from two NIH- sponsored clinical trials, ALF study group (ALFSG), and the adult-to-adult living donor liver transplantation cohort study (A2ALL). Unknown etiology accounted for 5.5% and 29.5% respectively in the ALFSG and A2ALL. Seq260 copy numbers will be quantitated in both serum and liver in Seq260-positive patients with liver tissue available. Second, we will determine the full genome of the putative virus containing Seq260. Finally, we will evaluate antibody responses in virus-positive patients and the controls. A peptide-based serological test will be developed for the putative virus. Peptides will be individually assessed for their specificity and sensitivity in two virus-positive patients with large volumes of serum available. Selected peptides will then be combined to ELISA tests for the measurement of antibody (IgG and IgM) responses in virus-positive and virus-negative patients. Taken together, the proposed study will characterize a novel human virus and understand its etiological link to liver disease from a clinical aspect. It will expand our knowledge of the human virome as well as the etiology of liver disease without a known cause.

项目概要 很大一部分肝病患者（5% 至 30%）的病因不明 已确定的病因。在肝脏的多种临床表型中观察到未知的病因 疾病，如急性肝衰竭（ALF）、肝炎、肝硬化和肝癌。这些统称为 称为隐源性肝病（CLD）。长期以来人们一直假设存在其他人类病毒 导致慢性肺病。在我们最近的血清病毒组研究中，我们鉴定了一个 387-nt DNA 片段 (GenBank MW468091)， 命名为 Seq260，来自 9 名 CLD 患者中的 1 名。在一系列基因行走、酶消化实验中， 以及滚环扩增和分析，我们证明了Seq260是一个线性单链 脱氧核糖核酸。我们在 409 名受试者中筛选了 Seq260，其中包括健康献血者 (n=200)、丙型肝炎病毒感染者 (n=100)、病因不明的急性肝衰竭 (ALF) 患者 (n=50) 和肝移植 (LT) 患有（n = 45）和未知病因（n = 14）的患者。在 5 名 CLD 患者（1 名 ALF 和 1 名 ALF）中检测到 Seq260 4 名 LT）和 1 名患有非酒精性脂肪性肝炎 (NASH) 相关肝硬化的 LT 患者。一名患者进行了 Seq260 可在肝脏中定量，显示肝脏中的滴度比血清中高 7.74 倍（2.4x106 拷贝/克 vs. 3.1x105 拷贝/mL）。机器学习分析达到了Seq260是真核生物的高分（可能性） 病毒序列。总的来说，这些数据导致我们假设 Seq260 代表了一个未被识别的 人类病毒具有肝向性。为了确定 Seq260 是否代表一种新型肝炎病毒，我们进行了 当前提案中的研究计划由三个主要实验组成。首先，我们将筛选Seq260 CLD 患者以及对照组。我们已获准使用来自两个 NIH 的患者样本 赞助的临床试验、ALF 研究组 (ALFSG) 和成人活体肝移植 队列研究（A2ALL）。 ALFSG 和 ALFSG 中病因不明分别占 5.5% 和 29.5% A2ALL。对 Seq260 阳性患者的血清和肝脏中的 Seq260 拷贝数进行定量 可用肝组织。其次，我们将确定包含 Seq260 的假定病毒的完整基因组。 最后，我们将评估病毒阳性患者和对照组的抗体反应。基于肽的 将为假定的病毒开发血清学检测。肽将被单独评估 两名有大量血清的病毒阳性患者的特异性和敏感性。已选择 然后将肽与 ELISA 测试相结合，以测量抗体（IgG 和 IgM）反应 病毒阳性和病毒阴性患者。总而言之，拟议的研究将描述一种新的人类特征 病毒并从临床角度了解其与肝病的病因学联系。它将扩展我们的知识 人类病毒组以及不明原因的肝脏疾病的病因学。