Protection from the 2009 H1N1 Pandemic Influenza by an Antibody from Combinatorial Survivor-Based Libraries

Influenza viruses elude immune responses and antiviral chemotherapeutics through genetic drift and reassortment. As a result, the development of new strategies that attack a highly conserved viral function to prevent and/or treat influenza infection is being pursued. Such novel broadly acting antiviral therapies would be less susceptible to virus escape and provide a long lasting solution to the evolving virus challenge. Here we report the in vitro and in vivo activity of a human monoclonal antibody (A06) against two isolates of the 2009 H1N1 pandemic influenza virus. This antibody, which was obtained from a combinatorial library derived from a survivor of highly pathogenic H5N1 infection, neutralizes H5N1, seasonal H1N1 and 2009 “Swine” H1N1 pandemic influenza in vitro with similar potency and is capable of preventing and treating 2009 H1N1 influenza infection in murine models of disease. These results demonstrate broad activity of the A06 antibody and its utility as an anti-influenza treatment option, even against newly evolved influenza strains to which there is limited immunity in the general population

Antibody A06 therapy protects Balb/C mice from death by 2009 pandemic H1N1 influenza infection.

<p>Balb/C (n = 10, except groups 4dpi and PBS where n = 9) were infected with 25MLD50 A/California/04/2009. A single administration of 15 mg/kg per group was given 1–6 days post-infection. Open blue squares- 1 day post infection, open red circles- 2 days post- infection, open green triangles- 3 days post-infection, filled triangles- 4 days post- infection, filled diamonds- 5 days post-infection, open black circles- 6 days post- infection, and open black squares- PBS vehicle 1 day post-infection. Survival (left panel) and weight (right panel) were monitored for 17 days after infection.</p

Antibody A06 therapy protects DBA/2 mice from death by 2009 pandemic H1N1 influenza infection.

<p>DBA/2 mice were infected with 3.3MLD50 of A/Netherlands/602/2009 and treated with a single administration of antibody A06 1 day (A) or 2 days (B) post-infection. Three different A06 concentrations were tested along with vehicle (PBS) and non-specific IgG controls. Animals were monitored for survival (left panels) and weight (right panels) over a 14 day period. Treatment groups (n = 5) are labeled as in <a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1000990#ppat-1000990-g001" target="_blank">Figure1B</a> and <a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1000990#ppat-1000990-g002" target="_blank">2C</a>. Open blue squares- 25 mg/kg antibody A06, open red circles- 10 mg/kg antibody A06, open green triangles- 2.5 mg/kg antibody A06, black filled circles- PBS vehicle control, open orange diamonds- 25 mg/kg human IgG control.</p

The A-helix epitope targeted by the A06 antibody is highly conserved across numerous types of influenza.

<p>A-helix epitope sequences from novel H1N1, current and past seasonal isolates of H1N1, H5N1 and avian H9N2 hemagglutinin proteins were aligned. Positions are labeled according to HA2 numbering. Amino acids at positions 19-21, 41, 42, 45, 46, 49, 52, 53 and 56 are antibody contact points <a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1000990#ppat.1000990-Sui1" target="_blank">[13]</a>, <a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1000990#ppat.1000990-Ekiert1" target="_blank">[17]</a>. The column on the far right indicates <i>in vitro</i> microneutralization minimal inhibitory concentrations (MIC) of antibody A06 in µg/ml for the isolates tested (ND = not done, * = previously reported data <a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1000990#ppat.1000990-Cheng1" target="_blank">[8]</a>).</p

Sequence analysis of novel H1N1 HA isolates shows limited variation in the predicted neutralization epitope.

<p>Alignment of hemagglutinin protein from novel H1N1 isolates deposited in the NCBI Influenza Virus Resource was performed using the multiple sequence alignment application within the database. 1000 full length HA sequences contained in the database on November 11, 2009 were analyzed for variation in HA2 residues previously shown to be contacted by neutralizing antibodies binding to the stem region <a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1000990#ppat.1000990-Sui1" target="_blank">[13]</a>, <a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1000990#ppat.1000990-Ekiert1" target="_blank">[17]</a>.</p