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.

项目摘要 肝病患者的很大一部分，范围从5％到30％，其原因未知 既定的病因。在肝脏中各种临床表型中观察到未知的病因 疾病，例如急性肝衰竭（ALF），肝炎，肝硬化和肝癌。这些是共同提及的 作为隐源性肝病（CLD）。长期以来一直假设存在其他人类病毒 原因CLD。在我们最近的血清病毒蛋白研究中，我们确定了387-NT DNA片段（GenBank MW468091）， 来自9名CLD患者中的1名，命名为SEQ260。在一系列基因走，酶消化的实验中， 和滚动圆的放大和分析，我们已经证明了SEQ260是线性的单链 脱氧核糖核酸。我们在409名受试者中筛选了SEQ260，包括健康的献血者（n = 200），乙型肝炎病毒感染 （n = 100），急性肝衰竭（ALF）患有不确定病因（n = 50）的患者和肝移植（LT） （n = 45）患者且没有已知病因（n = 14）。在5例CLD患者（1名ALF和 4 LT）和1 LT患有非酒精性脂肪性肝炎（NASH）相关的肝硬化患者。一位患者有SEQ260 在肝脏中可量化，显示肝脏中的滴度比血清高7.74倍（2.4x106拷贝/g vs。 3.1x105副本/毫升）。机器学习分析达到了SEQ260的高分（可能性）是真核生物 病毒序列。总体而言，这些数据导致了我们的假设，即SEQ260代表未被认可的 人类病毒伴有肝脏的向肝脏主义。为了确定SEQ260是否代表一种新型的肝炎病毒，我们带来了 当前提案中的研究计划包括三个主要实验。首先，我们将筛选SEQ260 在CLD患者以及对照组中。我们已获得从两个NIH-的患者标本的访问权限 赞助的临床试验，ALF研究小组（ALFSG）和成人到成年的供体肝移植 队列研究（A2ALL）。未知病因在ALFSG中分别占5.5％和29.5％， A2ALL。 SEQ260拷贝数将在SEQ260阳性患者的血清和肝脏中进行定量 可用的肝组织。其次，我们将确定含有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 将为假定病毒开发血清学检查。肽将分别评估其 两名病毒阳性患者的特异性和敏感性可用。选定 然后将肽合并到ELISA测试中，以测量抗体（IgG和IgM）反应 病毒阳性和病毒阴性患者。综上所述，拟议的研究将描述一个新颖的人类 病毒并从临床方面了解其与肝病的病因。它将扩大我们对 没有已知原因的人类病毒瘤以及肝病的病因。