Establishment of a pig influenza challenge model for evaluation of monoclonal antibody delivery platforms

Abstract

31Monoclonal antibodies are a possible adjunct to vaccination and drugs in treatment of 32 influenza virus infection. However questions remain whether small animal models 33 accurately predict efficacy in humans. We have established the pig, a large natural 34 host animal for influenza, with many physiological similarities to humans, as a robust 35 model for testing monoclonal antibodies. We show that a strongly neutralizing 36 monoclonal antibody (2-12C) against the hemagglutinin head administered 37prophylactically at 15 mg/kg reduced viral load and lung pathology after pandemic 38 H1N1 influenza challenge. A lower dose of 1 mg/kg of 2-12C or a DNA plasmid 39 encoded version of 2-12C, reduced pathology and viral load in the lungs, but not viral 40 shedding in nasal swabs. We propose that the pig influenza model will be useful for 41 testing candidate monoclonal antibodies and emerging delivery platforms prior to 42 human trials. 43 therapeutic administration to reduce viral load after infection has been established. 94We went on to evaluate the potential of an in vivo produced 2-12C using synthetic 95 dMAb technology in support of the translation of this technology to humans as an 96 immunoprophylactic. 97 98 Results: 99Establishment of a prophylactic pig influenza challenge model with recombinant 100 2-12C mAb. To establish a positive control protective mAb and delivery method in the 101 pig influenza model we tested the strongly neutralizing human IgG1 anti-head 2-12C 102 mAb 31 in a prophylactic experiment. We administered 15 mg/kg of 2-12C intravenously 103 and the control groups received an isotype matched IgG1 mAb or the diluent. Twenty-104 four hours later the pigs were challenged with pandemic swine H1N1 isolate, 105 A/swine/England/1353/2009 (pH1N1) and 4 days later culled to assess viral load and 106 pathology (Fig. 1a). 2-12C significantly decreased viral load in nasals swabs on each 107 day over the course of infection, although the decrease was less on days 2 and 3 than 108 days 1 and 4 (Fig. 1b). The total viral load in nasal swabs over the 4 days in animals 109 treated with 2-12C was also significantly less as determined by the area under the 110 curve (AUC) compared to diluent and isotype controls (p=0.0267 and p=0.021). Viral 111 load in the 2-12C group was significantly reduced in the BAL and no virus was detected 112 in the lungs at 4 days post infection (DPI). Overall 2-12C had a clear effect on viral 113 load in nasal swabs, BAL and lung. 114The isotype and diluent control groups showed the most severe gross 115 pathology and histopathology scores (Fig. 2a). The red tan areas of pulmonary 116 consolidation were present mainly in the cranial and middle lung lobes of animals from 117 the isotype and diluent groups, but no lesions were found in the 2-12C group (Fig. 2b). 118 6 Similarly, necrotizing bronchiolitis and bronchial and alveolar exudation, and mild 119 thickening of the alveolar septa was shown in the animals from both control groups. 120

Simultaneous Aerosol and Intramuscular Immunization with Influenza Vaccine Induces Powerful Protective Local T Cell and Systemic Antibody Immune Responses in Pigs

“…Because no difference was found in analyses of the samples challenged on different days, the results are presented together. Gross and histopathological analyses were performed as previously described (11,12,16). Briefly, the lungs were removed, and digital photographs were taken of the dorsal and ventral aspects.…”

“…ELISA was performed using recombinant HA (from A/England/195/2009) containing a C-terminal thrombin cleavage site, a trimerization sequence, a hexahistidine tag, and a BirA recognition sequence as previously described (20). Microneutralization (MN) was performed using standard procedures as described previously (9,16).…”

Magnitude and Kinetics of T Cell and Antibody Responses During H1N1pdm09 Infection in Inbred Babraham Pigs and Outbred Pigs

“…Hemagglutination inhibition (HAI) Ab titers were determined using 0.5% chicken red blood cells and H1N1pdm09 at a concentration of 4 HA units/ml. Microneutralization (MN) was performed using standard procedures as described previously ( 15 , 31 ).…”

“…Pigs exhibit similar clinical manifestations and pathogenesis when infected with IAV making them an excellent model to study immunity to influenza. Furthermore, we have defined the dynamics of H1N1pdm09 influenza virus transmission in pigs and demonstrated the utility of the pig model to test therapeutic antibody delivery platforms and vaccines ( 14 , 15 ).…”

Respiratory and intramuscular immunization with ChAdOx2 NPM1-NA induces distinct immune responses in H1N1pdm09 pre-exposed pigs

“…Endpoint titer ELISAs and microneutralization (MN) assays for pH1N1 and H3N2 viruses were performed as described before 48 . In addition, Abs against recombinant N2 protein were measured in serum.…”