Hepatitis B virus envelope proteins can serve as therapeutic targets embedded in the host cell plasma membrane

Abstract

Hepatitis B virus (HBV) infection is a major health threat causing 880,000 deaths each year. Available therapies control viral replication but do not cure HBV, leaving patients at risk to develop hepatocellular carcinoma. Here, we show that HBV envelope proteins (HBs)-besides their integration into endosomal membranesbecome embedded in the plasma membrane where they can be targeted by redirected T-cells. HBs was detected on the surface of HBV-infected cells, in livers of mice replicating HBV and in HBV-induced hepatocellular carcinoma.Staining with HBs-specific recombinant antibody MoMab recognising a conformational epitope indicated that membrane-associated HBs remains correctly folded in HBV-replicating cells in cell culture and in livers of HBV-transgenic mice in vivo. MoMab coated onto superparamagnetic iron oxide nanoparticles allowed to detect membrane-associated HBs after HBV infection by electron microscopy in distinct stretches of the hepatocyte plasma membrane. Last but not least, we demonstrate that HBs located on the cell surface allow therapeutic targeting of HBV-positive cells by T-cells either engrafted with a chimeric antigen receptor or redirected by bispecific, T-cell engager antibodies.

Hepatitis B Virus Neutralization with DNA Origami Nanoshells

“…An additional distinctive feature is the achievement of neutralization enhancement of the efficacy of the recombinant IgG antibody MoMAb, which we used as a coating inside the shells to specifically trap HBV. TheIC 50 decreased significantly from around 0.5 μmol/L per free anti-HBs IgG to ∼5 nmol/L when the IgG is mounted inside the shells. This corresponds to an IC 50 of approximately 55 pmol/L of HBV-capturing shells.…”

“…The anti-S scFV-Linker-hIgG1Fcmut antibodies (MoMAb) used to capture subviral particles and HBV virions were previously described and characterized . Large-scale production was contracted to InVivo Biotech Services GmbH.…”

Hepatitis B virus neutralization with DNA origami nanoshells

“…26 We included 9 single-stranded DNA ( ssDNA ) handles that protrude from each of the triangular subunits, creating 90 attachment sites in the shell’s internal cavity. We selected a synthetic immunoglobulin G ( IgG ) antibody against HBsAg , referred to as MoMAb , as a binder, 25 and labeled it with thiolated DNA single-strands that were complementary to the handles displayed on the interior surface of the shells. To tag the antibodies with DNA, we used a sulfosuccinimidyll 4-(N-maleimidomethyl) cyclohexan-1-carboxylat ( sulfo-SMCC ) crosslinker to target surface-exposed lysine residues on the IgG (Supplementary Figure 2, 3).…”

“…Markers such as HBeAg and cccDNA can be used to analyze the level of HBV infection. We used a synthetic antibody ( MoMAb ) that consists of 2 copies of a single chain antibody fragment fused to a mutated fragment crystallizable ( Fc) domain of an immunoglobulin G1 (IgG1) with reduced Fc -receptor binding affinity 25 to coat DNA nanoshells and test their neutralization capacity relative to that of the free antibodies.…”

Activation of distinct antiviral T-cell immunity: A comparison of bi- and trispecific T-cell engager antibodies with a chimeric antigen receptor targeting HBV envelope proteins

“…For specific targeting of HBVenv, we chose the binder C8, since it recognizes all HBV geno- and subtypes ( 19 ) and therefore is an optimal candidate with broad applicability in the majority of CHB patients. HBVenv proteins are exposed on HBV-infected cells ( 39 ), but are also expressed in about 30% of HBV-related tumor tissue ( 40 ), which might additionally allow specific targeting of certain HCCs.…”