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 相当一部分肝病患者，从5%到30%不等，除了原因之外，还有其他未知的原因。 已经确定的病因。在肝脏的多种临床表型中观察到未知的病因。 疾病，如急性肝功能衰竭(ALF)、肝炎、肝硬变和肝癌。这些被统称为 归因于隐源性肝病(CLD)。长期以来，人们一直假设存在更多的人类病毒 原因是慢性阻塞性肺疾病。在我们最近的血清病毒研究中，我们发现了一个387-NT的DNA片段(GenBank MW468091)， 命名为Seq260，取自9名CLD患者中的1名。在一系列的基因行走、酶消化、 并对滚动圆进行了放大和分析，证明了Seq260是一种线性单链 DNA我们对409名受试者进行了序列260筛查，其中包括健康献血者(n=200)，丙型肝炎病毒感染 (n=100)，病因不明的急性肝功能衰竭(ALF)患者(n=50)，以及肝移植(LT) 有(n=45)和不明原因(n=14)患者。在5例CLD患者中检测到Seq260(1例ALF和 4例LT)和1例LT合并非酒精性脂肪性肝炎(NASH)相关性肝硬变。一名患者的序列为260 在肝脏中可定量，显示肝脏中的滴度是血清中滴度的7.74倍(2.4×106拷贝/克比 3.1×105拷贝/毫升)。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。总而言之，这些数据导致了我们的假设，即序列260代表一个未被识别的 嗜肝的人类病毒。为了确定Seq260是否代表一种新的肝炎病毒，我们带来了一种 研究计划在目前的提案中由三个主要实验组成。首先，我们将对序列260进行筛选 在CLD患者和对照组中。我们已经获准接触到两个NIH的病人样本- 赞助临床试验、ALF研究组(ALFSG)和成人对成人活体供肝移植 队列研究(A2ALL)。不明原因分别占ALFSG和ALFSG的5.5%和29.5% A2ALL。Seq260阳性患者的血清和肝脏中的Seq260拷贝数将进行定量 肝组织可用。其次，我们将确定含有Seq260的推定病毒的全基因组。 最后，我们将评估病毒阳性患者和对照组的抗体反应。一种基于多肽的 将对推定的病毒进行血清学测试。多肽将被单独评估其 在有大量血清可用的两名病毒阳性患者中具有特异性和敏感性。已选择 然后将多肽结合到ELISA测试中，以测量抗体(Ig G和Ig M)的反应。 病毒阳性和病毒阴性患者。综上所述，这项拟议的研究将描述一种新的人类 并从临床角度了解其与肝病的病因学联系。它将扩大我们对 人类病毒体以及不明原因的肝病的病因学。