Mousavi Tahoora, Valadan Reza, Rafiei Alireza, Abbasi Ali, Haghshenas Mohammad Reza
Molecular and Cell Biology Research Center (MCBRC), Faculty of Medicine, Mazandaran University of Medical Sciences, Sari, Iran.
Molecular and Cell Biology Research Center (MCBRC), Hemoglobinopathy Institute, Mazandaran University of Medical Sciences, Sari, Iran.
Biotechnol Lett. 2021 Sep;43(9):1933-1944. doi: 10.1007/s10529-021-03166-2. Epub 2021 Jul 27.
Human papillomavirus infection (HPV) is the most common viral infection which is causes of cervical, penal, vulvar, anal and, oropharyngeal cancer. E7 protein of HPV is a suitable target for induction of T cell responses and controlling HPV-related cancer. The aim of the current study was to designed and evaluated a novel fusion protein containing the different E7 proteins of the HPV 16, 18, 6 and 11, linked to the cell-penetrating peptide HIV-1 Tat 49-57, in order to improve cytotoxic immune responses in in-vitro and in-vivo.
In this study whole sequence of HPV16,18,6,11 E7-Tat (47-57) and HPV16,18,6,11 E7 cloned into the vector and expressed in E. coli (BL21). The purified protein was confirmed by SDS page and western blotting and then injected into the C57BL/6 mice. The efficiency of the fusion protein vaccine was assessed by antibody response assay, cytokine assay (IL-4 and IFN-γ), CD + 8 cytotoxicity assay and tumor challenge experiment. Result showed that fusion proteins containing Adjuvant (IFA,CFA) could express higher titer of antibody. Also, we showed that vaccination with E7-Tat and, E7-Tat-ADJ induced high frequencies of E7-specific CD8 + T cells and CD107a expression as well as IFN-γ level and enhanced long-term survival in the therapeutic animal models.
Our finding suggested that this novel fusion protein vaccine was able to induce therapeutic efficacy and immunogenicity by improving CD8 + T cell in TC-1 tumor bearing mice; so this vaccine may be appreciated for research against HPV and tumor immunotherapies.
人乳头瘤病毒感染(HPV)是最常见的病毒感染,是导致宫颈癌、阴茎癌、外阴癌、肛门癌和口咽癌的原因。HPV的E7蛋白是诱导T细胞反应和控制HPV相关癌症的合适靶点。本研究的目的是设计并评估一种新型融合蛋白,该蛋白包含HPV 16、18、6和11的不同E7蛋白,并与细胞穿透肽HIV-1 Tat 49-57相连,以提高体外和体内的细胞毒性免疫反应。
在本研究中,HPV16、18、6、11 E7-Tat(47-57)和HPV16、18、6、11 E7的全序列被克隆到载体中并在大肠杆菌(BL21)中表达。通过SDS-PAGE和蛋白质印迹法对纯化的蛋白进行了鉴定,然后将其注射到C57BL/6小鼠体内。通过抗体反应测定、细胞因子测定(IL-4和IFN-γ)、CD + 8细胞毒性测定和肿瘤攻击实验评估融合蛋白疫苗的效率。结果表明,含有佐剂(IFA、CFA)的融合蛋白能够表达更高滴度的抗体。此外,我们还表明,用E7-Tat和E7-Tat-ADJ进行疫苗接种可诱导高频率的E7特异性CD8 + T细胞和CD107a表达以及IFN-γ水平,并提高治疗性动物模型中的长期存活率。
我们的研究结果表明,这种新型融合蛋白疫苗能够通过改善荷TC-1肿瘤小鼠的CD8 + T细胞来诱导治疗效果和免疫原性;因此,这种疫苗可能在针对HPV和肿瘤免疫治疗的研究中受到重视。