Molecular and structural characterization of the L1 virus-like particles that are used as vaccine antigens in Cervarix™, the AS04-adjuvanted HPV-16 and -18 cervical cancer vaccine.
Cervarix™ is a prophylactic human papillomavirus (HPV)-16/18 vaccine developed for the prevention of cervical cancer. The vaccine antigens are HPV-16 and HPV-18 L1 virus-like particles (VLPs) made from baculovirus expression vector system (BEVS)-produced HPV-16 and HPV-18 L1 proteins, respectively. In this study, we demonstrate that truncation of the nuclear targeting and DNA binding signals at the C-terminus of the HPV-16 and HPV-18 L1 proteins prevented intranuclear formation of the VLPs in the host cells and led to cytoplasmic localization of the L1 proteins as shown by in situ immunogold detection and electron microscopy. Following purification, these L1 proteins were able to form VLPs. The characteristics of these HPV-16 and HPV-18 L1 VLPs were studied using various physicochemical and immunological techniques. Amino acid analysis, SDS-PAGE and western blotting demonstrated the high purity of the L1 proteins and batch-to-batch consistency. The structure of the VLPs was shown to be similar to that reported for the native virions, as evaluated by microscopic observations, protein tomography and disc centrifugation experiments. The presence of important conformation-dependent neutralizing epitopes, such as U4, V5 and J4, was confirmed by ELISA and surface plasmon resonance. Structural robustness and consistency among batches was also observed by differential scanning calorimetry and electron microscopy. Moreover, adsorption to aluminum was shown not to impair VLP structure. In conclusion, the BEVS-produced HPV-16 and HPV-18 L1 VLPs display key structural and immunological features, which contribute to the efficacy of Cervarix™ vaccination.
GlaxoSmithKline Biologicals, Rixensart, Belgium.
Michel
M
Deschuyteneer
Abdelatif
A
Elouahabi
Dominique
D
Plainchamp
Michel
M
Plisnier
Dominique
D
Soete
Yvon
Y
Corazza
Laurence
L
Lockman
Sandra
S
Giannini
Marguerite
M
Deschamps
D000596:Amino Acids / Q000032:analysis; D015153:Blotting, Western; D036022:Capsid Proteins / Q000737:chemistry / Q000648:ultrastructure; D002942:Circular Dichroism; D003593:Cytoplasm / Q000737:chemistry / Q000648:ultrastructure; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D000939:Epitopes / Q000276:immunology; D006801:Humans; D046911:Macromolecular Substances / Q000737:chemistry / Q000648:ultrastructure; D016253:Microscopy, Immunoelectron; D009856:Oncogene Proteins, Viral / Q000737:chemistry / Q000648:ultrastructure; D053918:Papillomavirus Vaccines / Q000737:chemistry*; D011487:Protein Conformation; D022721:Vaccines, Virosome / Q000737:chemistry; D022701:Virosomes / Q000737:chemistry* / Q000648:ultrastructure*
D000596:Amino Acids; D036022:Capsid Proteins; D000939:Epitopes; D046911:Macromolecular Substances; D009856:Oncogene Proteins, Viral; D053918:Papillomavirus Vaccines; D022721:Vaccines, Virosome; D022701:Virosomes; C510352:human papillomavirus vaccine, L1 type 16, 18; C063317:L1 protein, Human papillomavirus type 16; C480565:L1 protein, Human papillomavirus type 18