Anti-HBs-based Functionalized Cellular Membrane Nanovesicles For HBV Immunotherapy

Chronic hepatitis B caused by persistent hepatitis B virus infection is a major public health problem in the world.However,the current antiviral drugs are difficult to induce active immune response.There are problems such as long treatment time,high cost,incomplete clearance and rebound after drug withdrawal.How to activate antiviral immune response based on effectively reducing the HBsAg level is of great significance to achieve the immunotherapy of HBV.As a novel drug delivery carrier,membrane-derived nanovesicles have become the focus of nanomedicine research area in recent years.By genetic engineering,a variety of targeted protein ligands can be displayed on the surface of cell membrane vesicles,which endured cell membrane vesicles with targeting and biological activity.In this study,we constructed anti-HBs-based functionalized cellular membrane vesicles displayed anti-HBs and immune-stimulating molecule CD40L.A(Anti-HBs)-CD40L-MV could reduce viral load and activate immune cells.First,A-CD40L-MV was prepared by differential centrifugation after ultrasound and characterization.MV with anti-HBs and CD40L on the surface was verified.The particle size was about 130nm.Secondly,we verified that A-CD40L-MV could recognize and bind the virus surface antigen(HBsAg)secreted by HBV cell lines in vitro,reduce the level of HBsAg in the cell supernatant.Third,C57BL/6 mice were intramuscular injected with A low dose of A complex formed by A-CD40L-MV and HBsAg.The mice produced antigen specific antibodies.A-CD40L-MV and HBsAg induced the specific immune response.Then,the HBsAg was restimulated to C57BL/6 mice.Mice produced a high level of protective antibodies.And the proportion of B cells in mice spleen cells was up-regulated.A-CD40L-MV and HBsAg induced an effective immune memory response.In addition,C57BL/6 mice intramuscular injected with a complex formed by high dose of A-CD40L-MV and HBsAg were challenged with HBV recombinant virus.The level of HBsAg in mice reduced,HBsAb increased.A-CD40L-MV and HBsAg induced an immune protection response.Finally,intravenous injection of MV reduced the load of HBsAg in HBV transgenic mice,and induced HBsAb secretion.A-CD40L-MV had the therapeutic effect on HBV transgenic mice.In summary,this study prepared anti-HBs-based functionalized cellular membrane vesicles,A-CD40L-MV.On the one hand,MV reduced the load of HBsAg rely on high specific antibody.On the other hand,MV proved immune cells stimulate signal,promoted cell activation and proliferation,enhanced antiviral response,improved the efficacy of immunotherapy for chronic HBV infection.