Department of Microbiology and Immunology, Peter Doherty Institute for Infection and Immunity, The University of Melbourne, Melbourne, Victoria 3000, Australia.
Viral Entry and Vaccines Laboratory, Disease Elimination, Burnet Institute, Melbourne, Victoria 3004, Australia.
Viruses. 2019 Jun 2;11(6):507. doi: 10.3390/v11060507.
A prophylactic vaccine eliciting both broad neutralizing antibodies (bNAbs) to the HIV-1 envelope glycoprotein (Env) and strong T cell responses would be optimal for preventing HIV-1 transmissions. Replication incompetent HIV-1 virus-like particles (VLPs) offer the opportunity to present authentic-structured, virion-associated Env to elicit bNAbs, and also stimulate T cell responses. Here, we optimize our DNA vaccine plasmids as VLP expression vectors for efficient Env incorporation and budding. The original vector that was used in human trials inefficiently produced VLPs, but maximized safety by inactivating RNA genome packaging, enzyme functions that are required for integration into the host genome, and deleting accessory proteins Vif, Vpr, and Nef. These original DNA vaccine vectors generated VLPs with incomplete protease-mediated cleavage of Gag and were irregularly sized. Mutations to restore function within the defective genes revealed that several of the reverse transcriptase (RT) deletions mediated this immature phenotype. Here, we made efficient budding, protease-processed, and mature-form VLPs that resembled infectious virions by introducing alternative mutations that completely removed the RT domain, but preserved most other safety mutations. These VLPs, either expressed from DNA vectors in vivo or purified after expression in vitro, are potentially useful immunogens that can be used to elicit antibody responses that target Env on fully infectious HIV-1 virions.
一种能诱导产生广谱中和抗体（bNAbs）和强烈 T 细胞反应的预防性疫苗，对于预防 HIV-1 传播将是最佳选择。复制缺陷型 HIV-1 病毒样颗粒（VLPs）提供了机会，可以呈现出具有天然结构的、病毒相关的包膜Env，从而诱导产生 bNAbs，并刺激 T 细胞反应。在这里，我们对 DNA 疫苗质粒进行了优化，使其成为高效表达包膜的 VLP 表达载体，并促进其出芽。最初用于人体试验的载体不能有效地产生 VLPs，但通过使 RNA 基因组包装、整合到宿主基因组所需的酶功能失活，以及删除辅助蛋白 Vif、Vpr 和 Nef，最大限度地提高了安全性。这些原始的 DNA 疫苗载体产生的 VLPs 中 Gag 的蛋白酶介导切割不完全，并且大小不规则。恢复有缺陷基因内功能的突变表明，几种逆转录酶（RT）缺失介导了这种不成熟的表型。在这里，我们通过引入完全去除 RT 结构域的替代突变，但保留了大多数其他安全性突变，制造了高效出芽、蛋白酶处理和成熟形式的 VLPs，这些 VLPs 类似于具有感染性的 HIV-1 病毒粒子。这些 VLPs 无论是通过体内表达 DNA 载体还是体外表达后纯化，都可以作为潜在的有用免疫原，用于诱导针对完全感染性 HIV-1 病毒粒子上的Env 的抗体反应。
