Beijing Institute of Biological Products Co., Ltd, Beijing, 100176, China.
Virol J. 2024 Jul 8;21(1):154. doi: 10.1186/s12985-024-02416-9.
Rabies is a fatal zoonotic disease whose pathogenesis has not been fully elucidated, and vaccination is the only effective method for protecting against rabies virus infection. Most inactivated vaccines are produced using Vero cells, which are African green monkey kidney cells, to achieve large-scale production. However, there is a potential carcinogenic risk due to nonhuman DNA contamination. Thus, replacing Vero cells with human diploid cells may be a safer strategy. In this study, we developed a novel 2BS cell-adapted rabies virus strain and analysed its sequence, virulence and immunogenicity to determine its application potential as a human diploid cell inactivated vaccine.
The 2BS cell-adapted rabies virus strain 2aG4-B40 was established by passage for 40 generations and selection of plaques in 2BS cells. RNA sequence analysis revealed that mutations in 2BS cell-adapted strains were not located at key sites that regulate the production of neutralizing antibodies or virulence in the aG strain (GQ412744.1). The gradual increase in virulence (remaining above 7.0 logLD50/ml from the 40th to 55th generation) and antigen further indicated that these mutations may increase the affinity of the adapted strains for human diploid cells. Identification tests revealed that the 2BS cell-adapted virus strain was neutralized by anti-rabies serum, with a neutralization index of 19,952. PrEP and PEP vaccination and the NIH test further indicated that the vaccine prepared with the 2aG4-B40 strain had high neutralizing antibody levels (2.24 to 46.67 IU/ml), immunogenicity (protection index 270) and potency (average 11.6 IU/ml).
In this study, a 2BS cell-adapted strain of the 2aG4 rabies virus was obtained by passage for 40 generations. The results of sequencing analysis and titre determination of the adapted strain showed that the mutations in the adaptive process are not located at key sequence regions of the virus, and these mutations may enhance the affinity of the adapted strain for human diploid cells. Moreover, vaccines made from the adapted strain 2aG4-B40 had high potency and immunogenicity and could be an ideal candidate rabies virus strain for inactivated vaccine preparation.
狂犬病是一种致命的人畜共患病，其发病机制尚未完全阐明，疫苗接种是预防狂犬病病毒感染的唯一有效方法。大多数灭活疫苗是使用非洲绿猴肾细胞（Vero 细胞）生产的，以实现大规模生产。然而，由于非人类 DNA 污染，存在潜在的致癌风险。因此，用人二倍体细胞替代 Vero 细胞可能是一种更安全的策略。本研究中，我们开发了一种新型 2BS 细胞适应的狂犬病病毒株，并分析了其序列、毒力和免疫原性，以确定其作为人二倍体细胞灭活疫苗的应用潜力。
通过在 2BS 细胞中进行 40 代传代和蚀斑选择，建立了 2BS 细胞适应的狂犬病病毒株 2aG4-B40。RNA 序列分析表明，2BS 细胞适应株的突变并不位于调节中和抗体产生或 aG 株毒力的关键位点（GQ412744.1）。毒力（从第 40 代到第 55 代逐渐增加，保持在 7.0logLD50/ml 以上）和抗原的逐渐增加进一步表明，这些突变可能增加了适应株与人二倍体细胞的亲和力。鉴定试验表明，2BS 细胞适应的病毒株被狂犬病血清中和，中和指数为 19952。PrEP 和 PEP 疫苗接种和 NIH 试验进一步表明，用 2aG4-B40 株制备的疫苗具有高中和抗体水平（2.24 至 46.67IU/ml）、免疫原性（保护指数 270）和效力（平均 11.6IU/ml）。
本研究通过 40 代传代获得了 2aG4 狂犬病病毒的 2BS 细胞适应株。适应性株测序分析和滴度测定结果表明，适应过程中的突变并不位于病毒的关键序列区域，这些突变可能增强了适应株与人二倍体细胞的亲和力。此外，由适应株 2aG4-B40 制备的疫苗具有高效力和免疫原性，可能是狂犬病病毒灭活疫苗制备的理想候选株。
