Vaccine Development Application and Research Center, Ege University, 35100, İzmir, Türkiye.
Department of Bioengineering, Faculty of Engineering, Ege University, İzmir, Türkiye.
Sci Rep. 2024 Jun 15;14(1):13865. doi: 10.1038/s41598-024-64690-5.
Severe acute respiratory syndrome coronavirus 2 had devastating consequences for human health. Despite the introduction of several vaccines, COVID-19 continues to pose a serious health risk due to emerging variants of concern. DNA vaccines gained importance during the pandemic due to their advantages such as induction of both arms of immune response, rapid development, stability, and safety profiles. Here, we report the immunogenicity and protective efficacy of a DNA vaccine encoding spike protein with D614G mutation (named pcoSpikeD614G) and define a large-scale production process. According to the in vitro studies, pcoSpikeD614G expressed abundant spike protein in HEK293T cells. After the administration of pcoSpikeD614G to BALB/c mice through intramuscular (IM) route and intradermal route using an electroporation device (ID + EP), it induced high level of anti-S1 IgG and neutralizing antibodies (P < 0.0001), strong Th1-biased immune response as shown by IgG2a polarization (P < 0.01), increase in IFN-γ levels (P < 0.01), and increment in the ratio of IFN-γ secreting CD4 (3.78-10.19%) and CD8 (5.24-12.51%) T cells. Challenging K18-hACE2 transgenic mice showed that pcoSpikeD614G administered through IM and ID + EP routes conferred 90-100% protection and there was no sign of pneumonia. Subsequently, pcoSpikeD614G was evaluated as a promising DNA vaccine candidate and scale-up studies were performed. Accordingly, a large-scale production process was described, including a 36 h fermentation process of E. coli DH5α cells containing pcoSpikeD614G resulting in a wet cell weight of 242 g/L and a three-step chromatography for purification of the pcoSpikeD614G DNA vaccine.
严重急性呼吸系统综合症冠状病毒 2 给人类健康带来了毁灭性的影响。尽管已经推出了几种疫苗，但由于关注的变种的出现，COVID-19 仍然构成严重的健康风险。在大流行期间，由于 DNA 疫苗具有诱导免疫反应的两个方面、快速开发、稳定性和安全特性等优势而变得重要。在这里，我们报告了一种编码带有 D614G 突变的刺突蛋白的 DNA 疫苗（命名为 pcoSpikeD614G）的免疫原性和保护效力，并定义了一个大规模生产过程。根据体外研究，pcoSpikeD614G 在 HEK293T 细胞中表达丰富的刺突蛋白。通过肌肉内（IM）途径和使用电穿孔装置（ID + EP）经皮途径向 BALB/c 小鼠给药 pcoSpikeD614G 后，它诱导了高水平的抗 S1 IgG 和中和抗体（P < 0.0001），强烈的 Th1 偏向免疫反应，如 IgG2a 极化（P < 0.01），IFN-γ 水平增加（P < 0.01），以及 IFN-γ 分泌 CD4（3.78-10.19％）和 CD8（5.24-12.51％）T 细胞的比例增加。挑战 K18-hACE2 转基因小鼠表明，通过 IM 和 ID + EP 途径给药的 pcoSpikeD614G 赋予 90-100％的保护，并且没有肺炎的迹象。随后，pcoSpikeD614G 被评估为有前途的 DNA 疫苗候选物，并进行了扩大规模的研究。因此，描述了一个大规模生产过程，包括含有 pcoSpikeD614G 的 E. coli DH5α 细胞的 36 小时发酵过程，导致湿细胞重量为 242g/L 和三步层析法用于纯化 pcoSpikeD614G DNA 疫苗。
