Cooper David, Wright Kevin J, Calderon Priscilla C, Guo Min, Nasar Farooq, Johnson J Erik, Coleman John W, Lee Margaret, Kotash Cheryl, Yurgelonis Irene, Natuk Robert J, Hendry R Michael, Udem Stephen A, Clarke David K
Wyeth, 401 N. Middletown Rd., Pearl River, NY 10965, USA.
J Virol. 2008 Jan;82(1):207-19. doi: 10.1128/JVI.01515-07. Epub 2007 Oct 17.
Recombinant vesicular stomatitis virus (rVSV) has shown great potential as a new viral vector for vaccination. However, the prototypic rVSV vector described previously was found to be insufficiently attenuated for clinical evaluation when assessed for neurovirulence in nonhuman primates. Here, we describe the attenuation, neurovirulence, and immunogenicity of rVSV vectors expressing human immunodeficiency virus type 1 Gag. These rVSV vectors were attenuated by combinations of the following manipulations: N gene translocations (N4), G gene truncations (CT1 or CT9), noncytopathic M gene mutations (Mncp), and positioning of the gag gene into the first position of the viral genome (gag1). The resulting N4CT1-gag1, N4CT9-gag1, and MncpCT1-gag1 vectors demonstrated dramatically reduced neurovirulence in mice following direct intracranial inoculation. Surprisingly, in spite of a very high level of attenuation, the N4CT1-gag1 and N4CT9-gag1 vectors generated robust Gag-specific immune responses following intramuscular immunization that were equivalent to or greater than immune responses generated by the more virulent prototypic vectors. MncpCT1-gag1 also induced Gag-specific immune responses following intramuscular immunization that were equivalent to immune responses generated by the prototypic rVSV vector. Placement of the gag gene in the first position of the VSV genome was associated with increased in vitro expression of Gag protein, in vivo expression of Gag mRNA, and enhanced immunogenicity of the vector. These findings demonstrate that through directed manipulation of the rVSV genome, vectors that have reduced neurovirulence and enhanced immunogenicity can be made.
重组水泡性口炎病毒(rVSV)作为一种新型疫苗病毒载体已显示出巨大潜力。然而,先前描述的原型rVSV载体在非人灵长类动物中进行神经毒力评估时,发现其减毒程度不足以用于临床评估。在此,我们描述了表达1型人类免疫缺陷病毒Gag的rVSV载体的减毒、神经毒力和免疫原性。这些rVSV载体通过以下操作组合进行减毒:N基因易位(N4)、G基因截短(CT1或CT9)、非细胞病变性M基因突变(Mncp)以及将gag基因定位到病毒基因组的第一个位置(gag1)。直接颅内接种后,所得的N4CT1 - gag1、N4CT9 - gag1和MncpCT1 - gag1载体在小鼠中显示出神经毒力显著降低。令人惊讶的是,尽管减毒程度非常高,但N4CT1 - gag1和N4CT9 - gag1载体在肌肉内免疫后产生了强大的Gag特异性免疫反应,等同于或大于毒性更强的原型载体产生的免疫反应。MncpCT1 - gag1在肌肉内免疫后也诱导了等同于原型rVSV载体产生的Gag特异性免疫反应。将gag基因置于VSV基因组的第一个位置与Gag蛋白的体外表达增加、Gag mRNA的体内表达以及载体免疫原性增强相关。这些发现表明,通过对rVSV基因组的定向操作,可以构建出神经毒力降低且免疫原性增强的载体。