Influence of integrant-derived HBV RNAs encoding the envelope proteins on HBV life cycle

编码包膜蛋白的整合体衍生的 HBV RNA 对 HBV 生命周期的影响

• 批准号: 10561639
• 负责人: Severin O Gudima
• 金额: $ 38.75万
• 依托单位: UNIVERSITY OF KANSAS MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-02-04 至 2026-01-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10561639
• 关键词: Affect; Aftercare; Antiviral Agents; Automobile Driving; Binding; Biological Assay; Cells; Chronic; Chronic Hepatitis B; Code; Complement; DNA Integration; Data; FDA approved; Genome; Genotype; HBV Genotype; Harvest; Hepatitis B Infection; Hepatitis B Surface Antigens; Hepatitis B Virus; Hepatitis Delta Virus; Hepatocyte; Human; Individual; Intervention; Knowledge; Length; Life Cycle Stages; Light; Liver; Liver diseases; Maintenance; Messenger RNA; Modeling; Mutation; Open Reading Frames; Pathogenesis; Patients; Persons; Pharmaceutical Preparations; Poly(A)+ RNA; Polyadenylation; Primary carcinoma of the liver cells; Production; Property; RNA; RNA Stability; Research; Reverse Transcriptase Polymerase Chain Reaction; Role; Sampling; Serum; Signal Transduction; Source; Surface Antigens; Suspensions; Testing; Tissues; Untranslated Regions; Viral; Virion; Virus; Work; analog; design; env Gene Products; interferon therapy; liver biopsy; novel; patient subsets; prevent; single-cell RNA sequencing; transcriptome sequencing; vector

Abstract There are more than 250 million of chronic HBV carriers worldwide. Chronic HBV infection is the main cause of hepatocellular carcinoma (HCC). There is no cure for HBV. Interferon therapy gives temporary benefits only to a subset of patients. Regardless that FDA-approved nucleos(t)ide analogs can greatly inhibit HBV replication, chronic HBV infection is not well under control. None of the drugs works in all patients, and is able to achieve the loss of the surface antigen (HBsAg, or HBV envelope proteins) in most cases. HBV frequently rebounds after treatment is stopped. Mechanism of HBV infection is not fully understood, which affects treatment options. This proposal will analyze the relationship between HBV integrant-derived RNAs (id-RNAs) coding for HBsAg and HBV life cycle. HBV DNA integration into host genome is not considered important for HBV life cycle, and id-RNAs are not well studied. Analyzing matching liver/HCC tissues from chronic HBV carrier humans using RT-PCR, we found that regardless of HBV replication: id-RNAs accumulated in all tissues and represented 45- 100% of all HBV RNAs found in 60% of tissues (40% livers/80% HCCs); RNAs polyadenylated via cryptic HBV poly(A) signal (could be integrant-derived or produced by HBV replication) were found in 60% livers/40% HCCs, and were the most abundant HBV RNAs in 20% livers/20% HCCs; and replication-derived RNAs (rd- RNAs) polyadenylated using conventional HBV poly(A) signal were the least abundant species in 70% of tissues (40% livers/100% HCCs). Furthermore, analysis of selected tissues by RNA sequencing also showed id-RNAs' abundance in most of the analyzed samples. The data suggest that id-RNAs may produce significant fraction of HBsAg in cells bearing HBV integrants. If the same cells also support HBV replication, then a great fraction of formed HBV virions likely will bear id-RNA-derived HBsAg (id-HBsAg) as a major component of their envelopes. Thus, infectivity of these HBV virions, and their ability to support virus spread will mostly depend on the properties of id-HBsAg, and not on those of HBV replication-derived HBsAg (rd-HBsAg). Thus, id-RNAs could regulate the maintenance of chronic HBV infection, which may call for revision of the current model of HBV infection that suggests that HBV life cycle is independent of the integration. It also became apparent that anti-HBV drugs fail to achieve loss of serum HBsAg, because major amounts of HBsAg can be produced from id-RNAs regardless of HBV replication. In light of our data, Aim 1 using a large set of liver/HCC tissues will find if id-RNAs' abundance is common during chronic HBV infection. Aims 2 and 3 will (i) find if large fraction of id- RNAs do not bear alterations in HBsAg-coding sequences, and thus can serve as sizeable source of functional HBsAg regardless of HBV replication, (ii) analyze how 3'-end untranslated regions of id-RNAs (including the host inserts) affect HBsAg synthesis, and (iii) find if id-HBsAg can efficiently support assembly and infectivity of HBV virions. The proposal will advance our understanding on how id-RNAs and id-HBsAg can impact HBV life cycle and HBV pathogenesis, and may justify id-RNAs as important targets for novel antiviral interventions.

抽象的 全球范围内有超过2.5亿个慢性HBV运营商。慢性HBV感染是 肝细胞癌（HCC）。无法治愈HBV。干扰素治疗仅给 一部分患者。无论是FDA批准的核（T）IDE类似物都可以极大地抑制HBV的复制， 慢性HBV感染不能很好地控制。这些药物在所有患者中都没有作用，并且能够达到 在大多数情况下，表面抗原（HBSAG或HBV包膜蛋白）的丧失。 HBV经常反弹 治疗后停止。 HBV感染的机制尚不完全了解，这会影响治疗选择。 该提案将分析HBV整体衍生的RNA（ID-RNA）编码HBSAG之间的关系 和HBV生命周期。 HBV DNA整合到宿主基因组中并不被认为对HBV生命周期不重要，并且 ID-RNA的研究不好。分析使用慢性HBV载体人类的肝脏/HCC组织使用 RT-PCR，我们发现，无论HBV复制如何：累积在所有组织中，代表45-- 在60％的组织中发现的所有HBV RNA中有100％（40％肝脏/80％HCC）； RNA通过隐性HBV进行聚腺苷酸化 在60％的肝脏中发现聚（a）信号（可以是HBV复制的整合性衍生或通过HBV复制产生）/40％ HCC，是20％肝脏/20％HCC中最丰富的HBV RNA；和复制衍生的RNA（rd- RNA）使用常规HBV聚（A）信号进行聚腺苷酸化是70％的最少丰富物种 组织（40％肝脏/100％HCC）。此外，通过RNA测序对选定组织的分析也显示 在大多数分析样本中，ID-RNA的丰度。数据表明ID-RNA可能会产生重要的 带有HBV整合体的细胞中HBSAG的一部分。如果相同的单元格也支持HBV复制，那么 形成的HBV病毒体的一部分可能会带有ID-RNA衍生的HBSAG（ID-HBSAG）作为其主要组成部分 信封。因此，这些HBV病毒体的感染性及其支持病毒扩散的能力将主要取决于 ID-HBSAG的属性，而不是HBV复制衍生的HBSAG（RD-HBSAG）的属性。因此，ID-RNA 可以调节慢性HBV感染的维持，这可能需要修改当前模型 HBV感染表明HBV生命周期与整合无关。也很明显 抗HBV药物无法实现血清HBSAG的损失，因为可以从中产生大量的HBSAG ID-RNA不管HBV复制如何。鉴于我们的数据，AIM 1使用大量肝脏/HCC组织会发现 如果ID-RNA的丰度在慢性HBV感染过程中很常见。目标2和3将（i）发现是否大部分ID- RNA在HBSAG编码序列中没有改变，因此可以用作功能的大量来源 HBSAG不管HBV复制如何 宿主插入）影响HBSAG合成，（iii）发现ID-HBSAG是否可以有效地支持组装和感染力 HBV病毒体。该提案将提高我们对ID-RNA和ID-HBSAG如何影响HBV生活的理解 循环和HBV发病机理，并可能证明ID-RNA是新型抗病毒干预措施的重要目标。