Dataset Information

A dendritic cell receptor-targeted chimeric immunotherapeutic protein (C-HBV) for the treatment of chronic hepatitis B.

ABSTRACT: In chronic Hepatitis B Virus (HBV) infections HBV-specific T cells are functionally impaired. Immunotherapy may restore HBV-specific T cell responses essential for sustained disease remission off-treatment and induction of a functional cure. Chimigen® Molecules are fusion proteins of antigen(s) with the Fc fragment of a xenotypic antibody designed to target specific receptors on dendritic cells (DCs). Here we describe the production and pre-clinical evaluation of Chimigen® HBV (C-HBV), containing HBV PreS1 and PreS2 peptide fragments, HBV core and murine Fc, produced in insect cells. C-HBV binding to immature DCs and internalization by endocytosis was FcγRII (CD32) and mannose receptor (CD206) dependent and led to increased MHC I and MHC II surface expression. Upon exposure of human T cells isolated from HBV un-infected healthy and chronically HBV-infected donors to C-HBV-pulsed mature DCs ex vivo, C-HBV induced vigorous T cell proliferation and enhanced expression of IFN-γ, TNF-α, perforin and granzyme B in both CD4⁺ and CD8⁺ T cell subsets. Re-stimulation of C-HBV-activated T cells from chronically infected donors with HBV PreS1/PreS2 and core overlapping peptides induced IFN-γ production in both CD4⁺ and CD8⁺ populations. C-HBV-activation of peripheral blood mononuclear cells (PBMCs) from chronically HBV-infected patients stimulated granzyme B production by CD4⁺CD25^- T responder (Tresp) cells, accompanied by an increase in Annexin V staining on CD4⁺CD25⁺ T regulatory (Treg) cell phenotype, consistent with apoptosis. The observed HBV-specific cellular responses induced by C-HBV ex vivo suggest that C-HBV is a promising immunotherapeutic candidate for the treatment of chronic HBV infections.

SUBMITTER: Ma A

PROVIDER: S-EPMC7227630 | biostudies-literature | 2020 Apr

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

A dendritic cell receptor-targeted chimeric immunotherapeutic protein (C-HBV) for the treatment of chronic hepatitis B.

Ma Allan A Motyka Bruce B Gutfreund Klaus K Shi Yuenian Eric YE George Rajan R

Human vaccines & immunotherapeutics 20191212 4

In chronic Hepatitis B Virus (HBV) infections HBV-specific T cells are functionally impaired. Immunotherapy may restore HBV-specific T cell responses essential for sustained disease remission off-treatment and induction of a functional cure. Chimigen® Molecules are fusion proteins of antigen(s) with the Fc fragment of a xenotypic antibody designed to target specific receptors on dendritic cells (DCs). Here we describe the production and pre-clinical evaluation of Chimigen® HBV (C-HBV), containin ...[more]

PMID: 31687879

Similar Datasets

Project description:Background & aimsHBV infects over 257 million people worldwide and is associated with the development of hepatocellular carcinoma (HCC). Integration of HBV DNA into the host genome is likely a key driver of HCC oncogenesis. Here, we utilise targeted long-read sequencing to determine the structure of HBV DNA integrations as well as full isoform information of HBV mRNA with more accurate quantification than traditional next generation sequencing platforms.MethodsDNA and RNA were isolated from fresh frozen liver biopsies collected within the GS-US-174-0149 clinical trial. A pan-genotypic panel of biotinylated oligos was developed to enrich for HBV sequences from sheared genomic DNA (∼7 kb) and full-length cDNA libraries from poly-adenylated RNA. Samples were sequenced on the PacBio long-read platform and analysed using a custom bioinformatic pipeline.ResultsHBV-targeted long-read DNA sequencing generated high coverage data spanning entire integrations. Strikingly, in 13 of 42 samples (31%) we were able to detect HBV sequences flanked by 2 different chromosomes, indicating a chromosomal translocation associated with HBV integration. Chromosomal translocations were unique to each biopsy sample, suggesting that each originated randomly, and in some cases had evidence of clonal expansion. Using targeted long-read RNA sequencing, we determined that upwards of 95% of all HBV transcripts in patients who are HBeAg-positive originate from cccDNA. In contrast, patients who are HBeAg-negative expressed mostly HBsAg from integrations.ConclusionsTargeted lso-Seq allowed for accurate quantitation of the HBV transcriptome and assignment of transcripts to either cccDNA or integration origins. The existence of multiple unique HBV-associated inter-chromosomal translocations in non-HCC CHB patient liver biopsies suggests a novel mechanism with mutagenic potential that may contribute to progression to HCC.Lay summaryFresh frozen liver biopsies from patients infected with HBV were subjected to targeted long-read RNA and DNA sequencing. Long-read RNA sequencing captures entire HBV transcripts in a single read, allowing for resolution of overlapping transcripts from the HBV genome. This resolution allowed us to quantify the burden of transcription from integrations vs. cccDNA origin in individual patients. Patients who were HBeAg-positive had a significantly larger fraction of the HBV transcriptome originating from cccDNA compared with those who were HBeAg-negative. Long-read DNA sequencing captured entire integrated HBV sequences including multiple kilobases of flanking host sequence within single reads. This resolution allowed us to describe integration events flanked by 2 different host chromosomes, indicating that integrated HBV DNA are associated with inter-chromosomal translocations. This may lead to significant transcriptional dysregulation and drive progression to HCC.

Project description:Background & aimsPatterns of liver HBV antigen expression have been described but not quantified at single-cell resolution. We applied quantitative techniques to liver biopsies from individuals with chronic hepatitis B and evaluated sampling heterogeneity, effects of disease stage, and nucleos(t)ide (NUC) treatment, and correlations between liver and peripheral viral biomarkers.MethodsHepatocytes positive for HBV core and HBsAg were quantified using a novel four-plex immunofluorescence assay and image analysis. Biopsies were analysed from HBeAg-positive (n = 39) and HBeAg-negative (n = 75) participants before and after NUC treatment. To evaluate sampling effects, duplicate biopsies collected at the same time point were compared. Serum or plasma samples were evaluated for levels of HBV DNA, HBsAg, hepatitis B core-related antigen (HBcrAg), and HBV RNA.ResultsDiffusely distributed individual HBV core+ cells and foci of HBsAg+ cells were the most common staining patterns. Hepatocytes positive for both HBV core and HBsAg were rare. Paired biopsies revealed large local variation in HBV staining within participants, which was confirmed in a large liver resection. NUC treatment was associated with a >100-fold lower median frequency of HBV core+ cells in HBeAg-positive and HBeAg-negative participants, whereas reductions in HBsAg+ cells were not statistically significant. The frequency of HBV core+ hepatocytes was lower in HBeAg-negative participants than in HBeAg-positive participants at all time points evaluated. Total HBV+ hepatocyte burden correlated with HBcrAg, HBV DNA, and HBV RNA only in baseline HBeAg-positive samples.ConclusionsReductions in HBV core+ hepatocytes were associated with HBeAg-negative status and NUC treatment. Variation in HBV positivity within individual livers was extensive. Correlations between the liver and the periphery were found only between biomarkers likely indicative of cccDNA (HBV core+ and HBcrAg, HBV DNA, and RNA).Impact and implicationsHBV infects liver hepatocyte cells, and its genome can exist in two forms that express different sets of viral proteins: a circular genome called cccDNA that can express all viral proteins, including the HBV core and HBsAg proteins, or a linear fragment that inserts into the host genome typically to express HBsAg, but not HBV core. We used new techniques to determine the percentage of hepatocytes expressing the HBV core and HBsAg proteins in a large set of liver biopsies. We find that abundance and patterns of expression differ across patient groups and even within a single liver and that NUC treatment greatly reduces the number of core-expressing hepatocytes.

Dataset Information

A dendritic cell receptor-targeted chimeric immunotherapeutic protein (C-HBV) for the treatment of chronic hepatitis B.

Publications

A dendritic cell receptor-targeted chimeric immunotherapeutic protein (C-HBV) for the treatment of chronic hepatitis B.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets