Development of Neutralizing Monoclonal Antibodies for Oncogenic Human Papillomavirus Types 31, 33, 45, 52, and 58

Abstract

bHuman papillomavirus (HPV) is the etiological agent for all cervical cancers, a significant number of other anogenital cancers, and a growing number of head and neck cancers. Two licensed vaccines offer protection against the most prevalent oncogenic types, 16 and 18, responsible for approximately 70% of cervical cancer cases worldwide and one of these also offers protection against types 6 and 11, responsible for 90% of genital warts. The vaccines are comprised of recombinantly expressed major capsid proteins that self-assemble into virus-like particles (VLPs) and prevent infection by eliciting neutralizing antibodies. Adding the other frequently identified oncogenic types 31, 33, 45, 52, and 58 to a vaccine would increase the coverage against HPV-induced cancers to approximately 90%. We describe the generation and characterization of panels of monoclonal antibodies to these five additional oncogenic HPV types, and the selection of antibody pairs that were high affinity and type specific and recognized conformation-dependent neutralizing epitopes. Such characteristics make these antibodies useful tools for monitoring the production and potency of a prototype vaccine as well as monitoring vaccine-induced immune responses in the clinic. This major capsid protein self-assembles into virus-like particles (VLPs) that mimic the structure of the native virion (2, 3). The two currently licensed vaccines offer protection against HPV types 16 and 18, which account for 70% of cervical cancer cases (4); the quadrivalent vaccine also offers protection against HPV types 6 and 11, which account for 90% of genital warts (5). A second-generation HPV vaccine was developed by Merck to provide extended coverage for the next five most prevalent oncogenic HPV types: 31, 33, 45, 52, and 58. Inclusion of VLPs to the additional five oncogenic HPV types will potentially increase the coverage to approximately 90% (4).Monitoring HPV type-specific neutralizing epitopes is an essential aspect of the development of this vaccine. A meaningful and rapid evaluation of serological responses facilitates decisions regarding the optimum vaccine and assists the collection of data for regulatory approval. Potency assays conducted in vivo (typically mouse potency assays) and functional in vitro assays such as pseudovirus and plaque assays can be laborious and difficult to standardize. Assays that utilize antibody binding and competition can act as valuable surrogates and can be readily qualified and adapted for high-throughput and automated platforms (6, 7). A critical requirement is that the antibodies that are chosen for the assays reflect the biological properties of the original potency and neutralization assays (8).Neutralizing antibodies to HPV L1 VLPs are primarily type specific (9) and bind to surface-exposed conformational loops (10). Despite high degrees of L1 sequence homology among members of the same family, neutralizing antibodies to cross-reactive epitopes have rarely been identified (11). Panels of monoclonal antibodies (MAbs) have be...

Immunogenicity and safety of the 9-valent HPV vaccine in men

“…MSM appeared to have substantially more prior exposure to HPV than HM (as judged by the HPV seroposivity rates at day 1) which might influence their immune response to subsequent HPV vaccination. Specifically, there is a high amino acid identity between HPV L1 proteins from the same HPV species [31,34]. Thus, lower antibody responses to HPV vaccination in MSM (a population with wide prior exposure to HPV) are reminiscent of previous observations that a prior antigenic encounter can alter a subsequent response to a related antigen [35][36][37].…”

“…At enrollment, serum antibody titers for HPV 6,11,16,18,31,33,45,52, or 58 that were greater than the predefined seropositive threshold values for one or more HPV type (indicative of previous exposure to the respective vaccine HPV types) were detected in 12.0% (132 of 1102), 38.0% (119 of 313), and 33.2% (365 of 1098) of HM, MSM, and women, respectively. Table 3A displays the month 7 HPV cLIA GMTs in HM, MSM and women for each vaccine HPV type in the PPI population.…”

“…Each vaccine component was analyzed separately. To be included in the primary immunogenicity analysis for the HPV 6 and HPV 11 components, subjects had to be seronegative to both HPV 6 and 11 at day 1 (because of extensive cross-reactivity due to the high amino acid sequence identity [92%] between HPV 6 and HPV 11 L1 proteins) [31]. To be included in the primary immunogenicity analysis for the other vaccine HPV types, subjects were required to be seronegative at day 1 only for the HPV type being analyzed.…”

“…Samples were tested for anti-HPV 6, anti-HPV 11, anti-HPV 16, anti-HPV 18, anti-HPV 31, anti-HPV 33, anti-HPV 45, anti-HPV 52, and anti-HPV 58 by HPV competitive Luminex immunoassay (HPV-9 cLIA) [30]. A subject was defined to be anti- HPV 6,11,16,18,31,33,45,52, or 58 positive if the anti-HPV serum level was ≥30, ≥16, ≥20, ≥24, ≥10, ≥8, ≥8, ≥8, or ≥8 mMU/mL, respectively. Since the cLIA antibody titers to each HPV type were determined using typespecific monoclonal antibodies, it was not possible to make a direct comparison of assay results across HPV types.…”

Development of a human papillomavirus competitive luminex immunoassay for 9 HPV types

“…The generation and characterization of the panels of mAbs to these HPV types, as well as the selection of the pairs of antibodies that were of high affinity, type-specific, and recognized conformationdependent neutralizing epitopes, has been previously described. 10 To determine the specificity of the mAbs used in the HPV-9 cLIA, each of the mAb-PEs was tested at 3 different concentrations: 10 μg/mL, 1.0 μg/mL, and 0.1 μg/mL, against the set of VLP coupled microspheres for all 9 HPV types. The mAb-PE concentrations employed for clinical testing in the HPV-9 cLIA ranged between 0.05 and 0.3 μg/mL.…”