Abstract
There are concerns that effectiveness and consistency of biopharmaceutical formulations, including vaccines, may be compromised by differences in size, concentration and shape of particles in suspension. Thus, a simple method that can help monitor and characterize these features is needed. Here, nanoparticle tracking analysis (NTA) was used to characterize particle concentration and size distribution of a highly-purified rabies vaccine (RABV), produced in Vero cells without raw materials of animal origin (RMAO). The NTA technique was qualified for characterization of RABV particles by assessing the stability profile of vaccine particles over 5–55 °C. Antigenicity of the viral particle was also monitored with the enzyme-linked immunosorbent assay (ELISA) and NTA. RABV particle size diameters were 100–250 nm (mean:150 nm), similar to sizes obtained when labelled with rabies anti-G D1–25 monoclonal antibody, suggesting mainly antigenic virus-like particles, also confirmed by transmission electron microscopy. Thermal stress at 55 °C decreased the concentration of anti-G D1–25-labelled particles from 144 hours, coherent with conformational changes leading to loss of G protein antigenicity without impacting aggregation. Results from RABV antigenicity assessment during the 24 months monitoring of stability showed good correlation between NTA and ELISA. NTA is a suitable approach for the characterization of biopharmaceutical suspensions.
Highlights
Rabies is a fatal zoonotic encephalitis caused by lyssaviruses from the Rhabdoviridae family
The discrepancy in particle concentration between drug product (DP) and drug substance (DS) batches is due to differences in the formulation process which is based on enzyme-linked immunosorbent assay (ELISA)
The concentration was unaffected by anti-G D1–25 monoclonal antibody labelling, suggesting that the particles were mainly comprised of antigenic virus particles
Summary
Rabies is a fatal zoonotic encephalitis caused by lyssaviruses from the Rhabdoviridae family. Since the first anti-rabies vaccination conducted by Louis Pasteur and Emile Rouxin 18853–5, a number of different vaccines for use in humans have been developed, including those prepared in human diploid cells, Vero cells, and purified chick embryo cells[6]. It is vitally important to characterize vaccines during the production process in order to develop analytical protocols to ensure that potency and lot-to-lot consistency of the final product is monitored. The technique has shown applicability in characterizing the size, concentration, and stability of a number of viruses including adenoviruses, influenza A, vesicular stomatitis virus, rabies (VRVg), hepatitis E virus-like particles preparations and therapeutic antibodies[21,22,23,24]. NTA was used to characterize Verorab and PVRV-NG2 (VRVg2.0, RABV) by particle quantification and size estimation. The NTA method was qualified to indicate if it was suitable for the characterization of viral particles in vaccines in terms of specificity, linearity, accuracy and precision, with no matrix interference
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