Jafarpour Nazli, Memarnejadian Arash, Aghasadeghi Mohammad Reza, Kohram Fatemeh, Aghababa Haniyeh, Khoramabadi Nima, Mahdavi Mehdi
Department of Biology, Science and Research Branch, Islamic Azad University, Tehran, Iran.
Mol Biol Rep. 2014 Aug;41(8):5207-14. doi: 10.1007/s11033-014-3388-y. Epub 2014 May 20.
Despite a huge number of studies towards vaccine development against human immunodeficiency virus-1, no effective vaccine has been approved yet. Thus, new vaccines should be provided with new formulations. Herein, a new DNA vaccine candidate encoding conserved and immunogenic epitopes from HIV-1 antigens of tat, pol, gag and env is designed and constructed. After bioinformatics analyses to find the best epitopes and their tandem, nucleotide sequence corresponding to the designed multiepitope was synthesized and cloned into pcDNA3.1+ vector. Expression of pcDNA3.1-tat/pol/gag/env plasmid was evaluated in HEK293T cells by RT-PCR and western-blotting. Seven groups of BALB/c mice were intramuscularly immunized three times either with 50, 100, 200 µg of plasmid in 2-week intervals or with similar doses of insert-free plasmid. Two weeks after the last injection, proliferation of T cells and secretion of IL4 and IFN-γ cytokines were evaluated using Brdu and ELISA methods, respectively. Results showed the proper expression of the plasmid in protein and mRNA levels. Moreover, the designed multiepitope plasmid was capable of induction of both proliferation responses as well as IFN-γ and IL-4 cytokine production in a considerable level compared to the control groups. Overall, our primary data warranted further detailed studies on the potency of this vaccine.
尽管针对人类免疫缺陷病毒1型开展了大量疫苗研发研究,但尚未有有效的疫苗获批。因此,需要为新型疫苗提供新的配方。在此,设计并构建了一种新的DNA候选疫苗,其编码来自HIV-1抗原tat、pol、gag和env的保守且具有免疫原性的表位。在通过生物信息学分析找到最佳表位及其串联体后,合成了与设计的多表位相对应的核苷酸序列,并将其克隆到pcDNA3.1+载体中。通过RT-PCR和蛋白质印迹法在HEK293T细胞中评估pcDNA3.1-tat/pol/gag/env质粒的表达。将七组BALB/c小鼠每隔2周分别用50、100、200μg质粒或相同剂量的无插入片段质粒进行三次肌肉注射免疫。在最后一次注射后两周,分别使用Brdu和ELISA方法评估T细胞增殖以及IL4和IFN-γ细胞因子的分泌。结果表明该质粒在蛋白质和mRNA水平上均有适当表达。此外,与对照组相比,设计的多表位质粒能够在相当程度上诱导增殖反应以及IFN-γ和IL-4细胞因子的产生。总体而言,我们的初步数据表明有必要对该疫苗的效力进行进一步详细研究。