Transplantation of B cells engineered ex vivo to secrete broadly neutralizing antibodies (bNAbs) has shown efficacy in disease models. However, clinical translation of this approach would require specialized medical centers, technically demanding protocols and major histocompatibility complex compatibility of donor cells and recipients. Here we report in vivo B cell engineering using two adeno-associated viral vectors, with one coding for Staphylococcus aureus Cas9 (saCas9) and the other for 3BNC117, an anti-HIV bNAb. After intravenously injecting the vectors into mice, we observe successful editing of B cells leading to memory retention and bNAb secretion at neutralizing titers of up to 6.8 µg ml⁻¹. We observed minimal clustered regularly interspaced palindromic repeats (CRISPR)–Cas9 off-target cleavage as detected by unbiased CHANGE-sequencing analysis, whereas on-target cleavage in undesired tissues is reduced by expressing saCas9 from a B cell-specific promoter. In vivo B cell engineering to express therapeutic antibodies is a safe, potent and scalable method, which may be applicable not only to infectious diseases but also in the treatment of noncommunicable conditions, such as cancer and autoimmune disease.

体外工程改造的 B 细胞移植可分泌广泛中和抗体 (bNAb)，已在疾病模型中显示出疗效。然而，这种方法的临床转化需要专门的医疗中心、技术要求高的协议以及供体细胞和受体的主要组织相容性复杂相容性。在这里，我们报告使用两种腺相关病毒载体进行体内 B 细胞工程，一种编码金黄色葡萄球菌Cas9 (saCas9)，另一种编码 3BNC117，一种抗 HIV bNAb。将载体静脉注射到小鼠体内后，我们观察到 B 细胞的成功编辑导致记忆保留和 bNAb 分泌，中和滴度高达 6.8 µg ml ^-1. 我们观察到通过无偏 CHANGE 测序分析检测到的最小成簇规律间隔回文重复 (CRISPR)–Cas9 脱靶切割，而通过从 B 细胞特异性启动子表达 saCas9 可减少不需要组织中的靶向切割。体内 B 细胞工程表达治疗性抗体是一种安全、有效且可扩展的方法，不仅适用于传染病，还适用于治疗非传染性疾病，如癌症和自身免疫性疾病。