In vitro generation and type-specific neutralization of a human papillomavirus type 16 virion pseudotype

Abstract

We report a system for generating infectious papillomaviruses in vitro that facilitates the analysis of papillomavirus assembly, infectivity, and serologic relatedness. Cultured hamster BPHE-1 cells harboring autonomously replicating bovine papillomavirus type 1 (BPV1) genomes were infected with recombinant Semliki Forest viruses that express the structural proteins of BPV1. When plated on C127 cells, extracts from cells expressing L1 and L2 together induced numerous transformed foci that could be specifically prevented by BPV neutralizing antibodies, demonstrating that BPV infection was responsible for the focal transformation. Extracts from BPHE-1 cells expressing L1 or L2 separately were not infectious. Although Semliki Forest virus-expressed L1 self-assembled into virus-like particles (VLPs), viral DNA was detected in particles only when L2 was coexpressed with L1, indicating that genome encapsidation requires L2. Expression of human papillomavirus type 16 (HPV16) L1 and L2 together in BPHE-1 cells also yielded infectious virus. These pseudotyped virions were neutralized by antiserum to HPV16 VLPs derived from European (114/K) or African (Z-1194) HPV16 variants but not by antisera to BPV VLPs, to a poorly assembling mutant HPV16 L1 protein, or to VLPs of closely related genital HPV types. Extracts from BPHE-1 cells coexpressing BPV L1 and HPV16 L2 or HPV16 L1 and BPV L2 were not infectious. We conclude that (i) mouse C127 cells express the cell surface receptor for HPV16 and are able to uncoat HPV16 capsids; (ii) if a papillomavirus DNA packaging signal exists, then it is conserved between the BPV and HPV16 genomes; (iii) functional L1-L2 interaction exhibits type specificity; and (iv) protection by HPV virus-like particle vaccines is likely to be type specific.

Production of recombinant virus-like particles from human papillomavirus types 6 and 11, and study of serological reactivities between HPV 6, 11, 16 and 45 by ELISA: implications for papillomavirus prevention and detection

“…Antibodies against the 16 L1a peptide, as against VLPs, could be useful for the detection of productive HPV virions and L1 protein in infected tissue [25,26]. The serological results obtained in this study con-¢rm those of Hines et al [27] and Roden et al [7] concerning the presence of type-restricted and typecommon antigen epitopes on VLPs. Moreover, a linear epitope located on the surface of the HPV 16 VLP was identi¢ed, in agreement with the results of Heino et al [25].…”

“…Analysis of the reactivities of anti-VLPs or anti-L1 antibodies to the four VLPs tested con¢rms that HPV 6 and 11 are highly homologous, as reported by Roden et al [7] using a hemagglutination inhibition assay. Our results also con¢rm that, according on capsid antigenicity, HPV 45 is more closely related to HPV 16 than to HPV 6 or 11.…”

“…The L1 protein of papillomavirus was shown to self-assemble, forming empty virus-like particles (VLPs) resembling empty native HPV. Such VLPs were shown to have immunological similarities to HPV virions [5] and to elicit neutralizing antibodies that could be detected using several systems such as the xenograft mouse system [6], neutralization of VLP binding to cells and, more recently, by neutralization of papillomavirus pseudo-types [7,8]. Moreover, immunization with puri¢ed VLPs composed of L1 or L1 plus L2 proteins was shown to protect beagle dogs, cotton-tail rabbits and cattle against homologous viruses [9^11].…”

Gene transfer using human papillomavirus pseudovirions varies according to virus genotype and requires cell surface heparan sulfate

“…The packaged DNA could be transferred to cells as shown by the expression of a reporter gene into mammalian cells [3]. Gene transfer has also been reported for HPV 11, 16, 18 and 33 L1/L2 VLPs [1,2,4,15]. To dispose of a range of vectors that could be used sequentially in vivo, we investigated ¢ve other types of HPVs, namely types 31, 39, 45, 58, 59 and 68, for gene transfer.…”

Immunogenicity of next-generation HPV vaccines in non-human primates: Measles-vectored HPV vaccine versus Pichia pastoris recombinant protein vaccine

“…L1, the major capsid protein of HPV, capable to self-assemble into virus-like particles (VLPs), is a type-specific and highly immunogenic antigen, and can elicit high titres of neutralizing antibodies, conferring protection against HPV infection [4,5].…”