Department of Pathology, The Johns Hopkins University, Baltimore, Maryland, USA.
Ph.D. Program for Cancer Biology and Drug Discovery, College of Medical Science and Technology, Taipei Medical University and Academia Sinica, Taipei, Taiwan.
J Virol. 2018 Jul 17;92(15). doi: 10.1128/JVI.00572-18. Print 2018 Aug 1.
To address how L2-specific antibodies prevent human papillomavirus (HPV) infection of the genital tract, we generated neutralizing monoclonal antibodies (MAbs) WW1, a rat IgG2a that binds L2 residues 17 to 36 (like mouse MAb RG1), and JWW3, a mouse IgG2b derivative of Mab24 specific for L2 residues 58 to 64. By Western blotting, WW1 recognized L2 of 29/34 HPV genotypes tested, compared to only 13/34 for RG1 and 25/34 for JWW3. WW1 IgG and F(ab') bound HPV16 pseudovirions similarly; however, whole IgG provided better protection against HPV vaginal challenge. Passive transfer of WW1 IgG was similarly protective in wild-type and neonatal Fc receptor (FcRn)-deficient mice, suggesting that protection by WW1 IgG is not mediated by FcRn-dependent transcytosis. Rather, local epithelial disruption, required for genital infection and induced by either brushing or nonoxynol-9 treatment, released serum IgG in the genital tract, suggesting Fc-independent exudation. Depletion of neutrophils and macrophages reduced protection of mice upon passive transfer of whole WW1 or JWW3 IgGs. Similarly, IgG-mediated protection by L2 MAbs WW1, JWW3, and RG1 was reduced in Fc receptor knockout compared to wild-type mice. However, levels of neutralization by WW1 IgG were similar in TRIM21 knockout and wild-type cells, indicating that Fc does not contribute to antibody-dependent intracellular neutralization (ADIN). In conclusion, the Fc domain of L2-specific IgGs is not active for ADIN, but it opsonizes bound extracellular pseudovirions for phagocytes in protecting mice from intravaginal HPV challenge. Systemically administered neutralizing IgG can access the site of infection in an abrasion via exudation without the need for FcRn-mediated transcytosis. At least 15 alpha HPV types are causative agents for 5% of all cancers worldwide, and beta types have been implicated in nonmelanoma skin cancer, whereas others produce benign papillomas, such as genital warts, associated with considerable morbidity and health systems costs. Vaccines targeting the minor capsid protein L2 have the potential to provide broad-spectrum immunity against medically relevant HPVs of divergent genera via the induction of broadly cross-neutralizing serum IgG. Here we examine the mechanisms by which L2-specific serum IgG reaches the viral inoculum in the genital tract to effect protection. Abrasion of the vaginal epithelium allows the virus to access and infect basal keratinocytes, and our findings suggest that this also permits the local exudation of neutralizing IgG and vaccine-induced sterilizing immunity. We also demonstrate the importance of Fc-mediated phagocytosis of L2 antibody-virion complexes for humoral immunity, a protective mechanism that is not detected by current neutralization assays.
为了解 L2 特异性抗体如何预防人类乳头瘤病毒（HPV）感染生殖道，我们生成了中和单克隆抗体（MAbs）WW1，这是一种与人乳头瘤病毒 29 型结合的大鼠 IgG2a，结合 L2 残基 17 至 36（与小鼠 MAb RG1 类似），以及针对 L2 残基 58 至 64 的小鼠 IgG2b 衍生物 JWW3。通过 Western blot，WW1 识别 29/34 种测试的 HPV 基因型的 L2，而 RG1 仅识别 13/34，JWW3 仅识别 25/34。WW1 IgG 和 F(ab') 类似地结合 HPV16 假病毒；然而，完整的 IgG 提供了更好的 HPV 阴道挑战保护。WW1 IgG 的被动转移在野生型和新生儿 Fc 受体（FcRn）缺陷型小鼠中同样具有保护作用，表明 WW1 IgG 的保护作用不是由 FcRn 依赖性转胞吞介导的。相反，局部上皮破坏是生殖道感染所必需的，无论是通过刷洗还是非诺酮-9 处理诱导的，都会将血清 IgG 释放到生殖道中，这表明 Fc 不依赖外渗。耗尽中性粒细胞和巨噬细胞会降低 WW1 或 JWW3 完整 IgG 被动转移时小鼠的保护作用。同样，与野生型小鼠相比，L2 MAb WW1、JWW3 和 RG1 的 IgG 介导的保护作用在 Fc 受体敲除小鼠中降低。然而，WW1 IgG 的中和水平在 TRIM21 敲除和野生型细胞中相似，表明 Fc 不参与抗体依赖性细胞内中和（ADIN）。总之，L2 特异性 IgG 的 Fc 结构域对于 ADIN 没有活性，但它可以调理结合的细胞外假病毒，以便吞噬细胞吞噬，从而保护小鼠免受阴道内 HPV 挑战。系统给予的中和 IgG 可以通过渗出作用进入通过擦伤到达感染部位，而无需 FcRn 介导的转胞吞作用。至少有 15 种 alpha HPV 类型是全球 5%所有癌症的致病因素，而 beta 类型与非黑色素瘤皮肤癌有关，而其他类型则产生良性乳头瘤，如生殖器疣，这些疾病会导致相当大的发病率和卫生系统成本。针对次要衣壳蛋白 L2 的疫苗有可能通过诱导广泛交叉中和血清 IgG，为具有不同属的医学相关 HPV 提供广谱免疫。在这里，我们研究了 L2 特异性血清 IgG 到达生殖道中病毒接种部位以发挥保护作用的机制。阴道上皮的擦伤允许病毒进入并感染基底角质形成细胞，我们的发现表明，这也允许中和 IgG 的局部渗出和疫苗诱导的绝育免疫。我们还证明了 Fc 介导的 L2 抗体-病毒复合物的吞噬作用对于体液免疫很重要，这是当前中和测定未检测到的保护性机制。
