Rapid discovery and optimization of therapeutic antibodies against emerging infectious diseases

Abstract

Using a comprehensive set of discovery and optimization tools, antibodies were produced with the ability to neutralize SARS coronavirus (SARS-CoV) infection in Vero E6 cells and in animal models. These anti-SARS antibodies were discovered using a novel DNA display method, which can identify new antibodies within days. Once neutralizing antibodies were identified, a comprehensive and effective means of converting the mouse sequences to human frameworks was accomplished using HuFR™ (human framework reassembly) technology. The best variant (61G4) from this screen showed a 3.5–4-fold improvement in neutralization of SARS-CoV infection in vitro. Finally, using a complete site-saturation mutagenesis methodology focused on the CDR (complementarity determining regions), a single point mutation (51E7) was identified that improved the 80% plaque reduction neutralization of the virus by greater than 8-fold. These discovery and evolution strategies can be applied to any emerging pathogen or toxin where a causative agent is known.

Recent Developments in Anti-Severe Acute Respiratory Syndrome Coronavirus Chemotherapy

“…Another chemotherapy strategy that has been developed is affinity-selected neutralizing antibodies as potential drugs [78]. For many virus infections, the development of neutralizing antibody is responsible for combating a virus infection, and this appears to be no different for infections caused by SARS-CoV [79].…”

“…Viral entry S protein-activated cellular factor TNF-a-converting enzyme, promotes viral entry [72] N-(R)-(2-[hydroxyaminocarbonyl]methyl)-4-methylpentanoyll-t-butyl-glycine-l-alanine 2-aminoethyl amide 200 nM (reduction in viral RNA) [76] Virus-neutralizing epitopes Virus infectivity 2978/10 humanized antibody 80% neutralization at 1.56 µg/ml [78] Viral entry…”

“…Ferrets (Mustela furo) also support the replication of SARS-CoV, with peak viral lung titers being detected at day 4 post-virus exposure and reaching levels as high as 10 6 median tissue culture infective dose/ml [97,98]. The lung pathology is characterized by multifocal pulmonary lesions only involving a small percentage of the surface future science group 10 mg/kg/mouse [104] Mouse BALB/c Senescent mouse model Humanized mouse antibody 2978/10~15,000-fold reduction in day-3 viral lung titers [78] Mouse BALB/c Senescent mouse model Equine anti-SARS-CoV F(ab')(2) Reduced viral lung titers by ~1000-fold and ~1000-fold fewer copies of N protein mRNA were detected [100] Mouse [108] Hamster Golden Syrian (LVG) Virus lung replication model, golden Syrian hamster mAb 201 2.4-3.9 log 10 drop in viral lung titers, reduced interstitial pneumonitis and lung consolidation [115] Hamster Chinese Virus lung replication model Equine neutralizing antibody F(ab') (2) 4-log 10 drop in viral lung titers, amelioration of lung pathology [114] Ferret Unknown Virus lung replication model CR3014 (human IgG1 neutralizing mAb)…”

“…Mice A number of compounds have been evaluated in both mouse lung replication models and in lethal models of the BALB/c mouse using two similar mouse-adapted SARS coronaviruses. Several studies have evaluated the therapeutic efficacy of antibody therapy [78,100]. In an aerosol infection model using aged mice, one group of mice was treated intraperitoneally with 15 mg/kg of a humanized mouse antibody (2978/10) selected by human framework reassembly technology and optimized for neutralization and specificity [78].…”

Development of Therapeutic Interventions for Emerging Diseases

“…One major factor that makes this an attractive technology is that antibody "factories" can be rapidly turned around to produce a stock of antibodies for therapeutic use. This approach was recently described by Rogers et al (2008) where they developed a panel of neutralising antibodies against the SARS coronavirus, a recently emerged pathogen, using a novel DNA display method. They describe their approach which involved panning the library using whole SARS virus rather than just the spike protein (which had been used by others, being a primary virulence factor for cell binding and entry).…”