Measurement of neutralizing serum antibodies of patients vaccinated with human papillomavirus L1 or L2-based immunogens using furin-cleaved HPV Pseudovirions.

Chenguang Wang,Joshua W Wang,Warner K Huh,Henry C Kitchener,Richard B S Roden,Peter L Stern,Susana Pang,Subhashini Jagu,Patricia M Day,Sai Daayana

doi:10.1371/journal.pone.0101576

Abstract

Antibodies specific for neutralizing epitopes in either Human papillomavirus (HPV) capsid protein L1 or L2 can mediate protection from viral challenge and thus their accurate and sensitive measurement at high throughput is likely informative for monitoring response to prophylactic vaccination. Here we compare measurement of L1 and L2-specific neutralizing antibodies in human sera using the standard Pseudovirion-Based Neutralization Assay (L1-PBNA) with the newer Furin-Cleaved Pseudovirion-Based Neutralization Assay (FC-PBNA), a modification of the L1-PBNA intended to improve sensitivity towards L2-specific neutralizing antibodies without compromising assay of L1-specific responses. For detection of L1-specific neutralizing antibodies in human sera, the FC- PBNA and L1-PBNA assays showed similar sensitivity and a high level of correlation using WHO standard sera (n = 2), and sera from patients vaccinated with Gardasil (n = 30) or an experimental human papillomavirus type 16 (HPV16) L1 VLP vaccine (n = 70). The detection of L1-specific cross-neutralizing antibodies in these sera using pseudovirions of types phylogenetically-related to those targeted by the L1 virus-like particle (VLP) vaccines was also consistent between the two assays. However, for sera from patients (n = 17) vaccinated with an L2-based immunogen (TA-CIN), the FC-PBNA was more sensitive than the L1-PBNA in detecting L2-specific neutralizing antibodies. Further, the neutralizing antibody titers measured with the FC-PBNA correlated with those determined with the L2-PBNA, another modification of the L1-PBNA that spacio-temporally separates primary and secondary receptor engagement, as well as the protective titers measured using passive transfer studies in the murine genital-challenge model. In sum, the FC-PBNA provided sensitive measurement for both L1 VLP and L2-specific neutralizing antibody in human sera. Vaccination with TA-CIN elicits weak cross-protective antibody in a subset of patients, suggesting the need for an adjuvant.

Highlights

The seminal discovery by zur Hausen that certain oncogenic genotypes of Human papillomaviruses (HPV) typified by human papillomavirus type 16 (HPV16) are the etiologic agents of cervical cancer has led to the commercial development of two preventive vaccines, Gardasil and Cervarix [1]
Our results showed the L1-PBNA and FCPBNA estimated titers of 120.6 and 146.8 respectively for the HPV16 (05/134) serum
This EC50 value was within the range of titers (80–1350) detected by 13 laboratories which previously tested the HPV18 standard. Both standards to 16 and 18 were tested against different HPV types such as HPV 31 and 45 which resulted in no detectable neutralization at 1:50 dilution (Data not shown). This indicates that the L1and FurinCleaved Pseudovirion-Based Neutralization Assay (FC-PBNA) are sensitive in detecting the HPV16 and HPV18 L1 virus-like particles (VLP)-specific antibody in the WHO standards which were derived by pooling sera from naturally infected individuals

Summary

Introduction

The seminal discovery by zur Hausen that certain oncogenic genotypes of Human papillomaviruses (HPV) typified by HPV16 are the etiologic agents of cervical cancer has led to the commercial development of two preventive vaccines, Gardasil and Cervarix [1]. In line with preclinical studies, vaccination of patients with HPV16 L1 VLP induces type-restricted neutralizing antibodies, suggesting the need for multivalent formulation [5] As a result, both licensed vaccines contain L1 VLPs derived from HPV16 and HPV18, the oncogenic genotypes that respectively cause circa 50% and 20% of all cervical cancer cases. Gardasil contains L1 VLP of benign genotypes HPV6 and HPV11, which are the most common cause of genital warts These L1 VLP vaccines were proven safe, highly immunogenic, and protective against infection and anogenital neoplasia associated with the vaccinal genotypes [6,7,8,9,10]. A nonavalent prophylactic VLP vaccine being developed by Merck is intended to broaden protection against the remaining oncogenic HPV types, but this complex formulation may be costly to produce, limiting access for low resource settings [14]

Methods

Results

Conclusion