Past Presentations: American Society of Virology (ASV) Minneapolis, MN July, 2011

Posted on Jul 20, 2011 | Tags: | Comments (0)

Abstracts submitted and approved for presentation at the American Society for Virology Annual Meeting
Minneapolis, MN
July 16-20, 2011

INFLUENZA RESEARCH

Human monoclonal antibody 53 shows unique cross-clade neutralization of influenza

William R. Usinger¹. Scott K. Johnson², Shelly J. Samet², Dan A. Dlugolenski², Jon D. Gabbard², Minha Park¹, Krista McCutcheon¹, Stote L. Ellsworth¹, Lawrence M. Kauvar¹, Ralph A. Tripp¹, and Stephen M. Tompkins²

¹ Trellis Bioscience, South San Francisco, CA 94080, ² Univ. of Georgia, Athens, GA 30602

Monoclonal antibody 53 is an Influenza HA reactive monoclonal antibody discovered by CellSpotTM multiplexing from cultured human PBMC cells. Binding studies demonstrate sub-nanomolar affinity of mAb 53 to HA purified from Group 1 clades including H1N1, H5N1 and H9N2. Surprisingly, mAb 53 also binds strongly to H7N3 HA of Group 2. Plaque neutralization bioassays demonstrate that mAb 53 blocks infection of H1N1, H9N1 and a highly pathogenic strain of H5N1 virus in MDCK target cells at low doses of antibody (0.1-5mg/mL). In initial studies, mice pretreated with graded doses of mAb 53 survive challenge with otherwise lethal titers of H1N1 and H5N1 viruses. Kaplan-Meier survival analysis of these mouse experiments demonstrated 100% and 90% protection against H1 challenge, respectively, (10 and 2MPK, single doses), with significant protection (~50% survival) afforded at the lowest dose examined (O.4MPK). Similar results were obtained for the H5 viral challenge. Additional studies are ongoing to measure therapeutic activity of mAb 53 in mice treated after infection with H1 and H5, and in studies testing mAb 53 efficacy against HA clades not yet examined.

Novel cross-subtype, fully human, influenza hemagglutinin-specific monoclonal antibodies

Stephen M. Tompkins¹. Scott K. Johnson¹, Shelly J. Samet¹, Dan A. Dlugolenski¹, Jon D. Gabbard¹, Minha Park², Krista McCutcheon², Stote L. Ellsworth², Lawrence M. Kauvar², Ralph A. Tripp¹, and William R. Usinger²

¹Univ. of Georgia, Athens, GA 30602, ²Trellis Bioscience, South San Francisco, CA 94080

Seasonal influenza virus infections cause significant morbidity and mortality each year, particularly in the young and elderly populations, and pandemic influenza continues to be a looming threat. While vaccines are generally effective, production time, strain matching, and vaccine failure in at-risk populations continue to be major problems. Licensed anti-viral drugs can reduce disease; however drug-resistance is increasing and may render existing anti-virals useless. Accordingly, new therapeutic strategies are needed. Therapeutic monoclonal antibodies have been shown to be efficacious in humans against a variety of diseases, including viral respiratory infection. While neutralizing antibodies against the hemagglutinin (HA) of influenza A virus are protective, the multiple circulating subtypes and constant antigenic drift of the immunodominant portion of the HA limit the utility of most monoclonal antibodies. That is, neutralizing broadly cross-reactive monoclonal antibodies are exceedingly rare. We have used CellSpot™ technology to identify and isolate these rare B cells from human peripheral blood that produce monoclonal antibodies reactive against multiple subtypes of influenza A, including seasonal influenza, avian influenza (H5, H7, and H9), highly pathogenic avian influenza, and pandemic H1N1 viruses. We show that these antibodies neutralize multiple strains and subtypes of influenza virus in MDCK cells and protected against lethal influenza virus challenge in a murine model of infection.

Discovery of broadly cross-reactive antibodies against influenza A virus from circulating human B cells

Minha Park, Stote L. Ellsworth, Keyi Liu, Krista M. McCutcheon, Neal DeChene, Evelene G. Lomongsod, Natalie Y. Chen, Julia Gray, Lawrence M. Kauvar, William R. Usinger

Trellis Bioscience, South San Francisco, CA 94080

Memory B cells from people vaccinated against or infected with circulating strains of influenza virus provide a natural repertoire of antibodies that have been selected and affinity matured to provide effective immunity to future infection. Antibodies obtained from healthy humans have been selected to be safe in the host and may exhibit lower levels of adverse immunogenicity as compared to humanized antibodies, or antibodies called “fully human” but in fact not obtained directly from humans. However, demands for high affinity and rare specificity force screening methods to have high throughput and render circulating human memory B cells as a challenging source for antibody discovery. The Trellis Cellspot™ technology is a high-throughput, multiplex platform that enables rapid, concurrent screening of B cells on multiple antigens. We have used our platform to screen over 10 million IgG-secreting B cells from various donors to identify rare B cells expressing antibodies that have broad reactivity against multiple subtypes of the influenza A hemagglutinin glycoprotein. The heavy and light chain Ig variable regions cloned from single B cells, when recombinantly expressed as a whole IgG1, recapitulate the binding properties seen in CellSpot™. Strikingly, some antibody clones display high-affinity reactivity to multiple subtypes, including H5, H7 and H9, despite no evidence of prior exposure to these viruses by PBMC donors, suggesting broadly reactive antibody responses may be primed by immunization or infection with only one or two influenza virus subtypes. These data demonstrate both the breadth and quality of the human B cell repertoire in circulating PBMCs. Moreover, they suggest that a fully human monoclonal antibody with reactivity against all influenza A subtypes could be rescued from an individual using CellSpot™ technology and developed for human therapeutic use.

CMV RESEARCH

High affinity native human monoclonal antibodies to CMV gB glycoprotein

Minha Park¹, Keyi Liu¹, Neal DeChene¹, Michael McVoy², Stuart Adler², Lenore Pereira³, Stote Ellsworth¹, Lawrence Kauvar¹, William Usinger¹

¹Trellis Bioscience, South San Francisco, CA 94080, ²Virginia Commonwealth University School of Medicine, Richmond, VA 23298, ³Univ California San Francisco (UCSF), San Francisco, CA 94143

About 40,000 infants born in the US each year shed cytomegalovirus (CMV). Of these, 8000 are born with neurological symptoms of varying severity and up to 8000 more will later develop symptoms, notably progressive hearing loss. Only about half of pregnant mothers have adequate native immunity to CMV. To date, none of the multiple vaccine efforts have yielded a product with better than 50% reduction in transmission to the fetus (Pass, RF, et al., N Engl J Med, 360 (12):1191 (2009)), a frustrating result in light of clinical evidence that passive immunization with CMV-specific gamma globulin is very effective (Nigro, G., et al., N Engl J Med 353 (13):1350 (2005)). The vaccine setbacks may represent a failure to induce antibodies with sufficiently high affinity, as it has been established that weak affinity antibodies to CMV actually promote transmission across the human placenta (Nozawa, N., et al. J Clin Virol. (2009)). Antibodies to the viral surface protein gB are known to neutralize CMV infectivity and thus gB has been a target for vaccine construction. Using our proprietary CellSpot™ multiplexing platform for monoclonal antibody (mAb) discovery from naturally immune people, we have discovered several fully human antibodies with sub-nanomolar affinity to the gB protein AD-2 epitope. Infection of HFF target cells can be blocked using <0.5 ug/mL mAb, an IC50 superior to CH177, a potent murine antibody against the gB-AD-2 epitope used as a comparator in our neutralization studies. Neutralization studies against additional CMV isolates is ongoing with the hope of developing an effective passive immunotherapy for pregnant women and their newborns at risk for CMV-induced pathology.

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