Adam J. Gehring

Summary: Chronic hepatitis B is characterized by a decades-long evolving engagement between host immunity and the hepatitis B virus (HBV). Understanding the molecular characteristics of HBV-specific CD8 T cells linked to control of viral replication and antigenemia is essential to design effective immunotherapeutic modalities. Here we show that HBV-specific CD8 T cells, even during infection stages with extremely high viral loads, lack the features of terminally exhausted CD8 T cells observed in chronic HCV and HIV infection or cancer. Instead, we observe emerging gene expression programs over disease stages that correlate with increasing HBV control, which include a bona fide cytotoxic and T-cell localization program associated with low levels of viral replication, and a second NK-like T-cell program that combines expression of classical NK markers ( KIR s, KLR s, FCGR3A , TYROBP , IKZF2 ) with cytotoxic genes ( GZMB , GNLY , PRF1 ), which emerges with complete control of HBV viremia and antigenemia. We also found enrichment of both CD8 T-cell programs in HIV-specific CD8 T cells from HIV elite controllers, supporting a conserved role in controlling persistent viral infections with viral reservoirs.

Dysregulation of the adaptive immune system is a key feature of aging and is associated with age-related chronic diseases and mortality. Here, we find that T cell aging, especially in the CD4 subset, is controlled by B cells. B cells contributed to the age-related reduction of naive CD4 T cells, their differentiation toward immunosenescent T cell subsets, and age-associated T cell receptor clonal restriction. Concurrently, mice lacking B cells displayed improvements in health span and life span. We uncovered a role for B cell–intrinsic insulin receptor signaling in influencing age-related B cell phenotypes that in turn induces CD4 T cell dysfunction, a process that is in part driven by major histocompatibility complex class II. These results identify B cells as critical mediators driving age-associated adaptive immune dysfunction and health-span outcomes and suggest previously unrecognized modalities to manage aging and related health decline.

Hepatitis B virus (HBV) chronically infects 250 million people worldwide, making it a primary risk factor for progressive liver disease. The virus itself is not responsible for liver damage. HBV can replicate at very high levels and produces large amounts of viral antigen, but this does not lead to hepatocyte death or liver inflammation. Instead, pathogenesis of chronic hepatitis B (CHB) is driven by the interaction between the host immune system and the virus. In chronically infected individuals, the HBV-specific immune response is dysfunctional and not able to clear the infection. This inability to clear the virus leads to aberrant immune activation in the liver, causing hepatocellular damage that, over time, leads to fibrosis, cirrhosis, and liver cancer. This review covers two aspects of sequalae associated with CHB: ( a ) mechanisms of tissue damage leading to fibrosis and ( b ) dysfunctional features of HBV-specific immunity.

Background: Hepatitis C virus (HCV) diagnosis usually requires detection of antibody followed by HCV RNA. The requirement for 2 tests leads to major drop-offs in the cascade of care. Existing near-care HCV RNA tests have slow turnaround time and are expensive with limited availability. We aim to develop a cost-effective, rapid, and sensitive test for detection of HCV RNA to enhance screening, particularly in marginalized and remote populations. Methods: After RNA extraction from plasma, HCV RNA is reverse-transcribed and amplified using loop-mediated isothermal amplification with HCV-specific primers. The amplified HCV DNA is then detected via CRISPR-Cas12b with a fluorescence readout. Results: HCV RNA from patient samples with genotypes 1a, 1b, 2, 3a, and 4 was detected with high sensitivity and specificity. The lower limit of detection (LLOD) with HCV JFH1 plasmid (genotype 2) is 250 plasmid copies/mL (approximately 100 IU/mL). For clinical samples, we determined the LLOD for genotypes 1 and 3, the most common in North America. Using 500 μL of plasma, genotype 1 RNA ≥100 IU/mL was detected within 40–45 minutes, while genotype 3 had an LLOD of 5000 IU/mL. The clinical sensitivity was 100% in 72 HCV patient samples, including acute HCV and HCV/hepatitis B virus (HBV) coinfection. The specificity was 100%, with no false-positives in 33 HCV-negative samples, including those with HBV or human immunodeficiency virus/HBV coinfection. Conclusions: Our assay shows high specificity and sensitivity to detect HCV RNA directly from plasma within 45 minutes and hence could be used for efficient screening and diagnosis of HCV infection globally.

Background and Aim: PD‐1‐expressing T cells within the HBV‐infected liver constitute a target of novel immunotherapeutics. Our aim was to investigate the impact of viral suppression on PD‐1 expression on intrahepatic versus circulating lymphocyte populations from chronic hepatitis B (CHB) patients. Methods: Twenty‐two CHB patients, nine of them on nucleos(t)ide analogues (NUCs), had paired blood, liver fine needle aspirations (FNAs) and biopsies. A subset had a follow‐up FNA after treatment initiation ( n = 4) or discontinuation ( n = 4). Intrahepatic (iHBV‐DNA and cccDNA) and serum (HBV‐DNA, HBsAg, HBcrAg and cirB‐RNA) viral markers were quantified. Flow cytometry was used for immunophenotyping PBMCs and intrahepatic lymphocytes. An independent liver FNA scRNAseq dataset was used to consolidate our results. Results: PD‐1 expression on tissue‐resident memory CD8 T cells (T _RM ) correlated with both iHBV‐DNA and cccDNA, as well as surrogate markers of cccDNA transcriptional activity (cirB‐RNA and HBcrAg) in CHB patients with mild hepatitis. These associations were not reflected in circulating T cells. PD‐1 expression intensity on CD8 T _RM was lower in NUC‐treated than in naive patients, changes that were again not detectable in the circulation. Longitudinal analysis showed that viral load rebound induced by NUC discontinuation had the potential to drive re‐expression of high levels of PD‐1 on CD8 T _RM . Conversely, therapy initiation and subsequent viral suppression reversed these changes. scRNAseq results further extended the profiling of these PD‐1 + CD8 T _RM , showing a phenotype consistent with bystander activation in response to subclinical liver damage. Conclusions: Intrahepatic viral markers correlate with PD‐1 expression on global liver‐resident T cells of CHB patients with mild hepatitis, with a reduction after prolonged NUC therapy and re‐expression following treatment withdrawal.

Chronic hepatitis B virus (HBV) infection affects millions worldwide despite the availability of effective vaccines. The stability of HBV’s genomic minichromosome (cccDNA) within hepatocytes, the persistence of integrated viral sequences capable of producing viral antigens, and the ability of the virus to evade immune control all contribute to the difficulty in achieving a functional cure. Existing antiviral treatments have minimal impact on HBV transcription, allowing persistent viral antigen production and immune dysfunction. Emerging RNA interference (RNAi) therapies targeting HBV RNAs reduce viral replication, antigen expression, and, in turn, cccDNA activity, providing a potential path to a functional cure.

The natural history and response to therapy in chronic hepatitis B (CHB) infection have been defined by a combination of serological and virological biomarkers along with liver biochemistry and/or histology. A number of novel biomarkers, including HBV RNA, hepatitis B core-related antigen, hepatitis B core antigen, and quantitative HBsAg, have been developed and evaluated in different clinical settings. Novel immunological biomarkers have also been studied but have been less well characterized. In addition to providing insights into HBV biology, these novel biomarkers may significantly aid in the design, development, and assessment of novel antiviral strategies aiming for the cure of chronic hepatitis B. Biomarkers can be used to confirm the mechanism of action or target engagement of a novel agent but also may be used for patient selection for trials and clinical use. Ideally, biomarkers can be used to more accurately define stages of chronic hepatitis B, particularly degrees of virological control. In this review, the serological, virological, and immunological biomarkers are described with a focus on how they can be used to guide the development of HBV cure strategies. New terminology is proposed for clinical endpoints, including sustained control to replace the concept of partial cure and resolved chronic infection to replace functional cure, reserving the term cure for clearance or silencing of all covalently closed circular DNA and integrated HBV DNA.

Chronic liver injury triggers the activation and recruitment of immune cells, causing antigen-independent tissue damage and liver disease progression. Tissue inflammation can reshape macrophage composition through monocyte replacement. Replacement of tissue macrophages with monocytes differentiating in an inflammatory environment can potentially imprint a phenotype that switches the liver from an immune-tolerant organ to one predisposed to tissue damage. We longitudinally sampled the liver of patients with chronic hepatitis B who had active liver inflammation and were starting antiviral therapy. Antiviral therapy suppressed viral replication and liver inflammation, which coincided with decreased myeloid activation markers. Single-cell RNA-Seq mapped peripheral inflammatory markers to a monocyte-derived macrophage population, distinct from Kupffer cells, with an inflammatory transcriptional profile. The inflammatory macrophages (iMacs) differentiated from blood monocytes and were unique from macrophage found in healthy or cirrhotic liver. iMacs retained their core transcriptional signature after inflammation resolved, indicating inflammation-mediated remodeling of the macrophage population in the human liver that may affect progressive liver disease and immunotherapy.