HSV-2 immune evasion as a virulence factor

描述（由申请人提供）：全球有5亿人感染单纯疱疹病毒2型（HSV-2），这具有重要的公共卫生意义，因为HSV-2生殖器溃疡疾病使感染和传播艾滋病毒的风险增加了3倍。我们的工作重点是研究HSV-2糖蛋白C （gC2）和E （gE2）的免疫逃避策略。HSV-2 gC2结合补体成分C3b抑制补体激活。HSV-2 gE2通过结合F(ab')2结构域与HSV抗原结合的IgG抗体分子的Fc结构域来阻断Fc介导的活性，从而抑制补体活化和抗体依赖性细胞毒性（ADCC）。在Aim 1中，我们将评估gC2和gE2免疫逃避通过抑制抗体和补体大大降低HSV-2糖蛋白D （gD2）疫苗在人类中的有效性的假设，并且gC2和gE2免疫逃避有助于自然获得性感染后复发性生殖器疾病。我们将通过评估接种GSK gD2亚单位疫苗的受试者的血清来检验我们的假设，以确定gC2和gE2免疫逃避是否抑制抗体和补体中和以及ADCC，并通过评估生殖器HSV-2多次复发或无复发的受试者的血清来评估gC2和gE2免疫逃避对复发性生殖器溃疡疾病的贡献。在Aim 2中，我们将评估在小鼠和豚鼠阴道感染模型中存在免疫逃避缺陷的HSV-2 gC2和gE2突变株的毒性比野生型或复发型病毒低log10个数量级的假设。我们将通过构建单一gC2和gE2突变株和双gC2/gE2突变株来验证我们的假设，这些突变株在免疫逃避方面存在缺陷，但在复制、蛋白表达和其他功能上完好无损。我们将在小鼠和豚鼠中确定免疫逃避的影响程度，并使用补体和NK缺陷动物确定机制。在目标3中，我们将评估gC2/gD2/gE2疫苗完全预防生殖器溃疡疾病并显着减少或完全预防潜伏期的假设。我们将确定gC2和gE2抗原诱导高滴度抗体阻断免疫逃避和gD2抗原诱导高滴度中和抗体的最佳剂量。然后，我们将评估三价疫苗是否比任何单一抗原或双抗原组合提供更好的保护，以防止生殖器疾病和潜伏感染。为了提高动物模型对人类研究的可翻译性，我们将修改动物模型中的终点确定以匹配人类试验，包括通过血清转化、PCR和培养确认疾病评分。我们将确定在没有生殖器疾病或潜伏感染的免疫动物中，阴道组织中的局部病毒复制是否足以诱导血清转化。在这些条件下检测血清转换将提供信息，因为它可能不能作为人体试验中延迟的有用标记。这些研究采取了一种新的疫苗开发方法，并可能导致新一代疫苗的产生。

DESCRIPTION (provided by applicant): Herpes simplex virus type 2 (HSV-2) infects a half-billion individuals worldwide, which has important public health implications since HSV-2 genital ulcer disease increases the risk of acquiring and transmitting HIV by 3-fold. Our work focuses on immune evasion strategies of HSV-2 glycoproteins C (gC2) and E (gE2). HSV-2 gC2 binds complement component C3b to inhibit complement activation. HSV-2 gE2 blocks Fc- mediated activities by binding the Fc domain of an IgG antibody molecule that is bound by its F(ab')2 domain to HSV antigen, which inhibits complement activation and antibody-dependent cellular cytotoxicity (ADCC). In Aim 1 we will evaluate the hypothesis that gC2 and gE2 immune evasion greatly reduce the effectiveness of an HSV-2 glycoprotein D (gD2) vaccine in humans by inhibiting antibody and complement, and that gC2 and gE2 immune evasion contribute to recurrent genital disease after naturally acquired infection. We will test our hypothesis by evaluating sera from subjects immunized with the GSK gD2 subunit vaccine to determine whether gC2 and gE2 immune evasion inhibit antibody and complement neutralization and ADCC, and by assessing sera obtained from subjects with multiple recurrences of genital HSV-2 or with no recurrences to evaluate the contribution of gC2 and gE2 immune evasion to recurrent genital ulcer disease. In Aim 2 we will assess the hypothesis that HSV-2 gC2 and gE2 mutant strains defective in immune evasion will be log10 orders of magnitude less virulent than wild-type or recue virus in mouse and guinea pig vaginal infection models. We will test our hypothesis by constructing single gC2 and gE2 mutant strains and a double gC2/gE2 mutant strain that are defective in immune evasion but intact for replication, protein expression and other functions ascribed to them. We will determine the magnitude of the impact of immune evasion in mice and guinea pigs and define mechanisms using complement and NK deficient animals. In Aim 3 we will evaluate the hypothesis that a gC2/gD2/gE2 vaccine totally protects against genital ulcer disease and markedly reduces or totally prevents latency. We will define the optimum dose of gC2 and gE2 antigen that induce high titers of antibodies that block immune evasion and of gD2 antigen that induces high titers of neutralizing antibodies. We will then evaluate whether the trivalent vaccine provides better protection against genital disease and latent infection than any single antigen or double antigen combination. To improve the translatability of animal models to human studies, we will modify endpoint determinations in animal models to match human trials, including confirming disease scores by seroconversion, PCR and culture. We will determine if local virus replication in vaginal tissues is sufficient to induce seroconversion in immunized animals that have no genital disease or latent infection. Detecting seroconversion under these conditions would be informative since it is may not serve as a useful marker for latency in human trials. These studies take a novel approach to vaccine development and may lead to a new generation of vaccines.