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Journal of Virology, December 2008, p. 12069-12081, Vol. 82, No. 24
0022-538X/08/$08.00+0 doi:10.1128/JVI.01379-08
Copyright © 2008, American Society for Microbiology. All Rights Reserved.
Llama Antibody Fragments with Cross-Subtype Human Immunodeficiency Virus Type 1 (HIV-1)-Neutralizing Properties and High Affinity for HIV-1 gp120
,
Anna Forsman,1
Els Beirnaert,2
Marlén M. I. Aasa-Chapman,1
Bart Hoorelbeke,2,
Karolin Hijazi,3
Willie Koh,1
Vanessa Tack,2
Agnieszka Szynol,4
Charles Kelly,3
Áine McKnight,1,
Theo Verrips,4
Hans de Haard,2,4 and
Robin A. Weiss1*
MRC/UCL Centre for Medical Molecular Virology, Division of Infection and Immunity, University College London, 46 Cleveland Street, London W1T 4JF, United Kingdom,1 Ablynx NV, Technologiepark 4, B-9052 Ghent, Belgium,2 King's College London, Dental Institute, Oral Immunology, Tower Wing, Guy's Hospital, London SE1 9RT, United Kingdom,3 Department of Molecular and Cellular Biology, Cellular Architecture & Dynamics, University of Utrecht, H.R. Kruytgebouw, Padualaan 8, 3584 CH Utrecht, The Netherlands4
Received 2 July 2008/ Accepted 26 September 2008
Members of the Camelidae family produce immunoglobulins devoid of light chains. We have characterized variable domains of these heavy chain antibodies, the VHH, from llamas immunized with human immunodeficiency virus type 1 (HIV-1) envelope protein gp120 in order to identify VHH that can inhibit HIV-1 infection. To increase the chances of isolating neutralizing VHH, we employed a functional selection approach, involving panning of phage libraries expressing the VHH repertoire on recombinant gp120, followed by a competitive elution with soluble CD4. By immunizing with gp120 derived from an HIV-1 subtype B'/C primary isolate, followed by panning on gp120 from HIV-1 isolates of subtypes A, B, and C, we could select for VHH with cross-subtype neutralizing activity. Three VHH able to neutralize HIV-1 primary isolates of subtypes B and C were characterized. These bound to recombinant gp120 with affinities close to the suggested affinity ceiling for in vivo-maturated antibodies and competed with soluble CD4 for this binding, indicating that their mechanism of neutralization involves interacting with the functional envelope spike prior to binding to CD4. The most potent VHH in terms of low 50% inhibitory concentration (IC50) and IC90 values and cross-subtype reactivity was A12. These results indicate that camelid VHH can be potent HIV-1 entry inhibitors. Since VHH are stable and can be produced at a relatively low cost, they may be considered for applications such as HIV-1 microbicide development. Antienvelope VHH might also prove useful in defining neutralizing and nonneutralizing epitopes on HIV-1 envelope proteins, with implications for HIV-1 vaccine design.
* Corresponding author. Mailing address: University College London, Wohl Virion Centre, 46 Cleveland St., London W1T 4JF, United Kingdom. Phone and fax: 44207 6799554. E-mail: r.weiss{at}ucl.ac.uk
Published ahead of print on 8 October 2008.
Supplemental material for this article may be found at http://jvi.asm.org/.
Present address: Virology and Chemotherapy Section, Catholic University of Leuven, Minderbroedersstraat 10 Blok x, Bus 1030, 3000 Leuven, Belgium.
Present address: Centre for Infectious Disease, Institute of Cell and Molecular Science, Barts and The London School of Medicine and Dentistry, The Blizard Building, 4 Newark Street, London E1 2AT, United Kingdom.
Journal of Virology, December 2008, p. 12069-12081, Vol. 82, No. 24
0022-538X/08/$08.00+0 doi:10.1128/JVI.01379-08
Copyright © 2008, American Society for Microbiology. All Rights Reserved.
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