Pandemrix

Anesthetized mice [0.1mg/10g: cocktail of Zoletil Forte (250mg/ml; Virbac France), Rompun (20mg/ml; Bayer Animal Health GmbH), and fentanyl (50µg/ml; Actavis, Germany)] were vaccinated intra muscularly (i.m.) with 25µg DNA (αMHCII-HA) into each quadriceps femoris, immediately followed by electroporation over the injection site (Elgen; Inovio Biomedical Co., Blue Bell, PA). The αMHCII-HA plasmid encodes HA from influenza A/California/07/2009 (H1N1), aa 18-541, linked to the MHCII-specific scFv via a dimerization unit consisting of the C_H3 domain of human IgG3 (22 (link)). All DNA vaccines were purified by using an EndoFree Plasmid Mega kit (catalog no. 12381; Qiagen, Hilden, Germany) and dissolved in sterile injection fluid (0.9% NaCl). Alternatively, anaesthetized mice were vaccinated i.m. with 1/10 human dose of Pandemrix with AS03 adjuvant (GlaxoSmithKline, Belgium), or a non-adjuvanted trivalent inactivated seasonal influenza vaccine [strains: A/Michigan/45/2015 (H1N1)pdm09-like virus, A/Singapore/INFIMH-16-0019/2016(H3N2)-like virus B/Colorado/06/2017-like virus (B/Victoria/2/87 lineage)].

Sera were obtained from 42 participants before and 3 weeks after 2 doses of an AS03-H1N1 adjuvanted split-virion vaccine (Pandemrix, GlaxoSmithKline, Rixenstart, Belgium). These samples had originally been obtained to investigate H1N1 seroconversion rates following vaccination [19 (link)].

All the following vaccines, except Pandemrix, were obtained from Sanofi-Pasteur MSD (Lyon, France). Mutagrip (0.5 ml/dose) contains Hemagglutinin (HA) and Neuraminidase (N) proteins from the following three influenza strains (A/Brisbane/59/2007 [H1N1]-like, A/Brisbane/10/2007 [H3N2], B/Brisbane/60/2008-like). Each dose includes 15 μg of the various HA proteins but no Thimerosal. Panenza, in its multidose format (10 doses), contains for each dose 15 μg of HA derived from the A/California/7/2009 [H1N1]- like strain and 45 μg of Thimerosal.
Pandemrix from GlaxoSmithKline (Marly-le-Roi, France) contains for each dose 3.5 μg of HA derived from the A/California/7/2009 [H1N1]-like strain, the AS03 adjuvant and 5 μg of Thimerosal.
PepTivator-CMV pp65, PepTivator-CMV IE1, PepTivator-EBV EBNA-1 and PepTivator-EBV BZLF (Miltenyi Biotec SAS, Paris, France) were used at 0.25 μg/ml, EBV, Tetanus toxoid (TT) and tuberculin PPD (Statens Serum Institut, Copenhagen, Denmark) were used at 5 μg/ml. As positive control, PBMC were stimulated with plate-bound anti-CD3 (1 μg/ml) and anti-CD28- mAbs (2 μg/ml) (Miltenyi Biotec SAS, Paris, France) during 1 to 3 days, according to the experiments.
Thimerosal was purchased from Sigma-Aldrich (St Quentin-Fallavier, France) and diluted in sterile water to obtain a 1 g/ml stock solution.

The 2-component vaccine Pandemrix, containing inactivated influenza A/California/7/2009 (H1N1) split virus (3.75 μg per 0.5 ml emulsion), and AS03 adjuvant (10.69 mg squalene, 11.86 mg DL-alpha-tocopherol and 4.86 mg polysorbate 80 per 0.5 ml emulsion), was obtained from GlaxoSmithKline Biologicals, Rixensart, Belgium.

The vaccine included the A/California/07/2009 (H1N1) virus strain. The vaccine brands were Focetria® (Novartis), adjuvanted with MF59, recommended to children and the elderly, Pandemrix® (Glaxo SmithKline), adjuvanted with ASO3, recommended for adults, and Panenza® (Sanofi Pasteur), unadjuvanted, recommended for pregnancy. Actual exposure involved administration of 970,468 doses in the second half of November 2009, 641,829 in December 2009, 114,220 in January 2010, coming to an end in late February 2010, making a total of 1.7 million doses, less than 5 % of which were administered to persons under 18 years of age. In all, 12 % of health care workers, 9 % of workers in essential public services, 15 % of people from 18 to 60 years old with chronic conditions, and 28 % of people 60 years old and over received the vaccine (Division of Vaccination Programmes, Spain’s Ministry of Health, Social Services, and Equality, unpublished report).

Vaccination dates and seasonal vaccine type were derived from Vaccinera. Three different trivalent inactivated vaccines, Vaxigrip (Sanofi Pasteur MSD, Lyon, France), Fluarix (GSK, Brentford, United Kingdom), and Inflexal V (Crucell, Janssen Vaccines, Leiden, The Netherlands), were used during the seasons covered. No high-dose or adjuvanted vaccines were available in Sweden during the four seasons. Individuals with influenza infection before vaccination, or up to 13 days post-vaccination, were considered to be unvaccinated as were those who did not receive the seasonal vaccine. Those with influenza infection ≥ 14 days post vaccination were considered to be vaccinated. Pandemic influenza (Pandemrix, GSK) vaccination status from 2009/10 was included as a covariate as was pneumococcal vaccination (in the current season or previous seasons since 2009). Vaccination against seasonal influenza in the previous season was also included as a covariate.

The vaccination schedule comprised two doses of the inactivated, split-virion, AS03-adjuvanted H1N1 vaccine (Pandemrix; GlaxoSmithKline, Brentford, Middlesex, UK). Pandemrix was derived from the A/California/7/2009(H1N1) v-like strain antigen (New York Medical College x-179A). The doses were administered at a 28-day interval. The vaccinations were given into the deltoid muscle. Children below the age of 13 years received 0·25 ml per dose, whereas older children received the adult dose of 0·5 ml in keeping with national guidelines issued by the Health Protection Surveillance centre. Participants were requested to report any adverse effects to the vaccination by telephone and were questioned on possible adverse effects at follow-up visits.