编码SARS-CoV-2受体结合域(RBD)的DNA疫苗的电穿孔递送

D. N. Kisakov, L. A. Orlova, Сергей V. Sharabrin, A. Rudometov, M. Borgoyakova, E. Starostina, L. Karpenko, A. Ilyichev
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摘要

背景:基于核酸的预防工具为开发疫苗提供了一个有前景的平台,包括针对COVID-19的疫苗。之前,我们开发了编码SARS-CoV-2受体结合域(RBD)的pVAXrbd DNA疫苗,当给动物肌肉注射时,诱导了相对较弱的免疫反应。研究的下一阶段是增加免疫反应,特别是使用电穿孔作为增加DNA疫苗免疫原性的方法之一。目的:用电穿孔法评价pVAXrbd免疫小鼠后的免疫反应。材料与方法:采用正反极性方波直流电穿孔法对BALB/c小鼠进行pVAXrbd免疫,脉冲为12 V,间隔为950 ml,电流限为45 mA。结果:BALB/c小鼠用100 g DNA免疫2次,间隔3周。ELISA检测结果显示,pVAXrbd电穿孔免疫组的rbd特异性抗体滴度为1:109350,是仅肌内注射DNA疫苗组(滴度为1:6750)的16倍。IFN ELISpot分析显示,电穿孔pVAXrbd免疫动物组在RBD蛋白肽刺激下产生IFN的细胞数量最多(2434个斑点/脾细胞,百万)。相比之下,对照组的细胞数量减少了6.5倍:380个斑点/脾细胞,百万。结论:与裸DNA疫苗肌肉注射相比,通过电穿孔给药pVAXrbd DNA疫苗可显著增强实验动物的体液和细胞特异性免疫反应。
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Delivery of a DNA vaccine encoding SARS-CoV-2 receptor-binding domain (RBD) by electroporation
BACKGROUND: Nucleic acid-based prevention tools provide a promising platform for developing vaccines, including those against COVID-19. Previously, we developed the pVAXrbd DNA vaccine encoding the receptor-binding domain (RBD) of SARS-CoV-2, which, when administered intramuscularly to animals, induced a relatively weak immune response. The next stage of the study is to increase the immune response, in particular, using electroporation as one of the methods for increasing the immunogenicity of DNA vaccines. AIM: The aim of this article is to evaluate the immune response using electroporation in mice after immunization with pVAXrbd. MATERIALS AND METHODS: BALB/c mice were immunized with pVAXrbd using direct and reverse polarity square wave direct current electroporation with three pulses of 12 V for 30 ms and an interval of 950 ml with a current limit of 45 mA. RESULTS: BALB/c mice were immunized twice with an interval of three weeks with a dose of 100 g of DNA. Using ELISA, the titers of RBD-specific antibodies in the group of animals immunized with pVAXrbd using electroporation were 1:109350, which is 16 times higher than in the group of animals that received the DNA vaccine only intramuscularly (titers 1:6750). IFN ELISpot analysis showed that the largest number of cells (2434 spots/splenocytes, million) producing IFN in response to stimulation with peptides from the RBD protein was registered in the group of animals immunized with pVAXrbd using electroporation. For comparison, in the control group, the number of cells is 6.5 times lower: 380 spots / splenocytes, mln. CONCLUSIONS: Administration of the pVAXrbd DNA vaccine to laboratory animals by electroporation significantly enhances both the humoral and cellular specific immune response compared to intramuscular administration of the naked DNA vaccine.
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