Quantification by LC-MS(E) of outer membrane vesicle proteins of the Bexsero® vaccine.
Meningococcal disease is a major cause of morbidity and mortality worldwide. Its epidemiology is currently dominated by five capsular serogroups (A, B, C, W, and Y). While effective vaccines already exist for serogroups A, C, W and Y, except for under clonal outbreaks, no vaccine was available against serogroup B. Recently, a four component vaccine, Bexsero(®), designed to prevent infection caused by this serogroup, has been approved in Europe and other Countries for use in individuals from two months of age and older. The active components of this vaccine are three recombinant proteins identified by reverse vaccinology combined with detergent extracted outer membrane vesicles (DOMV) prepared from a New Zealand epidemic strain. Considering their intrinsic complexity, we performed additional characterization of DOMVs on top of the standard quality control testing carried out for batch release. We applied the Hi3 label-free LC-MS(E) methodology to qualitatively and quantitatively characterize the DOMV protein content. We first, successfully investigated the robustness and the accuracy of the methodology for the DOMV characterization and we then applied it to compare six DOMV production lots. Around 100 proteins were quantified from each preparation. When classified according to their predicted cellular localization, about 90% of the total protein amount belongs consistently to the outer membrane compartment. Using nonparametric hypothesis testing and complementary log-log linear regression, the quantifications of a subset of 21 proteins common to all lots and including approximately 90% (85-92%) of the total protein amount quantified in any DOMV lot were found consistent across lots. The relevance of these results is two-fold, showing that the Hi3 quantification methodology is robust for a broad range of proteins and indicating that the manufacturing process currently used for the production of the Bexsero(®) DOMV components is highly reproducible and consistent.
Novartis Vaccines and Diagnostics, Via Fiorentina, 1 53100 Siena, Italy.
Chiara
C
Tani
Novartis Vaccines and Diagnostics, Via Fiorentina, 1 53100 Siena, Italy.
Maria
M
Stella
Novartis Vaccines and Diagnostics, Via Fiorentina, 1 53100 Siena, Italy.
Danilo
D
Donnarumma
Novartis Vaccines and Diagnostics, Via Fiorentina, 1 53100 Siena, Italy.
Massimiliano
M
Biagini
Novartis Vaccines and Diagnostics, Via Fiorentina, 1 53100 Siena, Italy.
Pierino
P
Parente
Novartis Vaccines and Diagnostics, Via Fiorentina, 1 53100 Siena, Italy.
Alessandro
A
Vadi
Novartis Vaccines and Diagnostics, Via Fiorentina, 1 53100 Siena, Italy.
Claudia
C
Magagnoli
Novartis Vaccines and Diagnostics, Via Fiorentina, 1 53100 Siena, Italy.
Paolo
P
Costantino
Novartis Vaccines and Diagnostics, Via Fiorentina, 1 53100 Siena, Italy.
Fabio
F
Rigat
Novartis Vaccines and Diagnostics, Via Fiorentina, 1 53100 Siena, Italy. Electronic address: nathalie.norais@novartis.com.
Nathalie
N
Norais
D001425:Bacterial Outer Membrane Proteins / Q000032:analysis*; D002853:Chromatography, Liquid; D004591:Electrophoresis, Polyacrylamide Gel; D013058:Mass Spectrometry; D022401:Meningococcal Vaccines / Q000032:analysis*; D011786:Quality Control
C570015:4CMenB vaccine; D001425:Bacterial Outer Membrane Proteins; D022401:Meningococcal Vaccines
Bexsero(®); Neisseria meningitidis; Outer membrane vesicles; Quantitative proteomics; Vaccine