利用微环DNA载体在肌肉中持续表达HBV特异DART清除cccDNA

根治慢性乙肝有待清除HBV cccDNA。DART(双亲和重定向分子)是一种结构紧凑的新型双特异性抗体，可重定向T细胞至靶细胞并介导T细胞对靶细胞的高效杀伤。HBV表面抗原(HBs)和核心抗原(HBc)胞内表达并加工成多肽后，可经MHC分子递呈至细胞表面，形成HBs/MHC或HBc/MHC复合物；它们有别于游离抗原，是感染细胞表面特有的靶标。本项目拟设计相应的DART，精准靶向感染细胞表面的HBs/MHC或HBc/MHC复合物，介导T细胞杀灭感染细胞，从而间接清除cccDNA。由于DART半衰期短、需连续给药，且成本高昂，本项目提出了体内持续表达DART的低成本基因治疗方案：即利用独特的微环DNA肌肉转染技术实现HBV特异DART在肌肉中的持续表达，达到“一次注射、长久治疗”的效果。总之，利用微环DNA在肌肉中持续表达DART有望成为高效方便、可负担的慢性乙肝基因治疗体系。

To cure chronic HBV infection requires the permanent removal of viral cccDNA. Dual-affinity re-targeting (DART) molecule, a novel rigid and compact format of bispecific antibody that bridges target cells and effector T cells, can induce robust redirected T-cell cytotoxicity to target cells. The peptides derived from the surface antigen (HBs) and core antigen (HBc) are able to presented onto cell surface by the major histocompatibility complex (MHC), where form the MHC/HBV peptides complexes (HBs/MHC and HBc/MHC). The HBs/MHC and HBc/MHC complexes differ from the excessive intact viral antigens in circulation and subviral particles, and they can distinguish the infected cells from the uninfected normal cells, by which they may serve as the attractive specific targets for immunotherapies. In this proposal, we are planning to design the HBs/MHC or HBc/MHC targeted DARTs to clear cccDNA indirectly via a redirected T-cell cytotoxicity on infected cells. However, apart from the high cost, the short half-life of DARTs inconveniences their clinical application, as they were often intravenously administrated with a pump. Therefore, we propose a convenient and affordable gene therapy strategy: durable producing HBV-specific DARTs in skeletal muscle with our established minicircle (MC)/MusFX system. In summary, the minicircle-mediated prolonged expression in muscle of DARTs may serve as an effective, convenient and affordable gene therapy for chronic HBV infection treatment.