Oral Abstract Session 01 - Global HIV Vaccine Enterprise

Topic 12: Vaccine Concepts and Design
P12.29
Transforming Epitope-Specific gp120 Monomer-
Based Probes into Immunogens with N-linked Glycan
Masking
G. Chuang 1 , J.C. Boyington 1 , G.J. Nabel 1 , P.D. Kwong 1 ,
I. Georgiev 1
1 National Institutes of Health/NIAID, Bethesda, MD, USA
Background: HIV-1 gp120 monomer-based probes have been
used for the identification of broadly neutralizing antibodies.
Such probes could represent starting points in the design of
HIV-1 immunogens, though efforts must be made to silence
immune responses directed toward non-neutralizing epitopes.
One possible approach would be to mask these epitopes by
introducing N-linked glycan. A potential complication to such an
approach is glycan occupancy: although N-linked glycosylation
generally occurs at N-X-T/S sequons, many such sequons are
not occupied.
Methods: A computational protocol was developed to identify
the putative positions for insertion of N-linked glycan on the
gp120 surface. The first step involves the identification of
residue positions on the gp120 surface where the insertion of
the N-X-T/S sequon is predicted as energetically-tolerable. The
second step involves the application of NGlycPred, a Random
Forest-based predictor, to predict the glycan occupancy at the
inserted sequons.
Results: The glycan occupancy prediction of the protocol is
highly correlated to validated N-X-T sequon insertion designs.
Multiple sequon insertions to gp120 monomer-based probes
were generated based on the protocol.
Conclusion: A computational protocol was implemented to
identify putative sites for insertion of N-X-T/S sequons with
improved likelihood of glycan occupancy. The protocol is
applicable to the design of N-linked glycans for masking nonneutralizing
antibody epitopes on gp120-based probes as well as
other immunogen candidates.
P12.30
AIDS Vaccine 2012
Posters
Development of an Imaging Based Virus Aggregation
Assay for Vaccine Development
D. Stieh 1 , C. Gioia 1 , M. McRaven 1 , G. Cianci 1 , P. Kiser 2 , T. Hope 1
1 Northwestern University, Chicago, IL, USA; 2 University of
Utah, Salt Lake City, UT, USA
Background: Vaccination strategies capable of eliciting
neutralizing antibody responses to HIV remain elusive
despite extensive efforts. Alternative antibody functions offer
opportunities for protection without necessarily achieving
broad neutralization breadth. Viral immune exclusion through
aggregation has been proposed as an alternative protection
pathway, but mechanisms for studying this phenomenon at the
scale necessary for clinical trials have not been explored.
Methods: Concentrated fluorescent virions of two colors,
suspended in hydroxyethylcellulose (HEC) gel, which has been
formulated to simulate the diffusion characteristics of cervical
mucus, are imaged over time. Mean squared displacement and
incidence of colocalized viral particles are determined. The
addition of monoclonal antibodies of various specificities and
isotypes affects these parameters is explored. Immunoglobulin
isolated from HIV-1 positive individuals was examined as
well. Correlative scanning electron micrographs of the
same preparations were performed to confirm the nature of
suspected aggregates.
Results: Multimeric antibodies, rather than monomeric isoforms,
selectively hinder the diffusion characteristics of colocalized
virions, more so than non-aggregated virions. The incidence of
colocalized virions is also increased in a concentration dependent
manner. Excessive antibody or virus concentration is seen to
obviate aggregate formation. These results are seen in both
monoclonal antibody experiments alongside polyclonal patient
IgA isolated from breast milk and IgM from serum.
Conclusion: Virus aggregation has been demonstrated
as a feasible effector function of HIV-1 specific antibody
preparations. This assay platform is amenable to adaptation for
medium to high throughput sample screening and low sample
size. Monoclonal antibodies as well as patient samples are able
to induce similar behaviors. Measurement of viral aggregation
as a corollary of vaccine efficacy is deserving of further
exploration in a clinical setting.
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POSTERS