Humanization of an anti-ICAM-1 antibody with over 50-fold affinity and functional improvement

Luo, Guang X.; Kohlstaedt, Lori A.; Charles, Catherine H.; Gorfain, Elliott; Morantte, Ianessa; Williams, Jacqueline; Fang, Fang

doi:10.1016/s0022-1759(02)00542-2

Cited by 24 publications

(22 citation statements)

References 27 publications

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“…In light of our present findings, the trial of humanized ICAM-1 or VCAM-1 Abs (for example, Ref. 42) in formats that specifically target the lymphatics is worthy of renewed consideration.…”

Section: Hivanp Inhibits Cd8mentioning

confidence: 83%

Blocking Development of a CD8+ T Cell Response by Targeting Lymphatic Recruitment of APC

Teoh

Johnson

Hanke

et al. 2009

The Journal of Immunology

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Generating a protective immune response to viral infection is known to depend upon the priming and clonal expansion of virus-specific CD8؉ T cells by Ag-loaded dendritic cells (DC) within secondary lymphoid tissue. However, the actual initiation of the response involves critical upstream events that control the recruitment of mature Ag-charged DC from the periphery via afferent lymphatics, events that are still only partly understood. Recent evidence has revealed that transmigration of lymphatic endothelium by DC is regulated by the adhesion molecules ICAM-1 and VCAM-1 both in vitro and in vivo. These findings imply that lymphatic entry may be an important rate-limiting step in primary immunity and a possible target for immune intervention. In this study, we have explored such possibilities using an F 5 TCR-transgenic mouse model to assess the contribution of lymphatic cell adhesion molecules in the CD8 ؉ T cell response to influenza virus nucleoprotein (NP). We show for the first time that immunization with ICAM-1-and VCAM-1-blocking mAbs can impair the T cell response in lymph node-draining sites of dermally administered nucleoprotein vaccine (MVA.HIVA.NP) by targeting lymphatic uptake of Ag-loaded DC ahead of other cell adhesion molecule-dependent events. These results reveal lymphatic entry as an important step that may be rate limiting in the development of immunity and reconfirm its potential as a target for localized immunotherapy in inflammation and tissue rejection.

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“…In light of our present findings, the trial of humanized ICAM-1 or VCAM-1 Abs (for example, Ref. 42) in formats that specifically target the lymphatics is worthy of renewed consideration.…”

Section: Hivanp Inhibits Cd8mentioning

confidence: 83%

Blocking Development of a CD8+ T Cell Response by Targeting Lymphatic Recruitment of APC

Teoh

Johnson

Hanke

et al. 2009

The Journal of Immunology

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“…Such ligands include high-affinity monoclonal ICAM antibodies (anti-ICAM) or their F(abЈ) 2 , Fab, and single chain Fv fragments, some of which undergo clinical testing (Takei et al, 1996;Luo et al, 2003;. Numerous studies have reported targeting to ECs in cell cultures, perfused organs, and in vivo using anti-ICAM-conjugated isotopes, ultrasound and magnetic resonance imaging contrast probes (Villanueva et al, 1998;Weiner et al, 2001;Weller et al, 2002), therapeutic agents (Murciano et al, 2003;Muro et al, , 2006, and drug carriers including liposomes, PNCs, and micron-size polymer particles (Bloemen et al, 1995;Sakhalkar et al, 2003).…”

mentioning

confidence: 99%

Endothelial Targeting of High-Affinity Multivalent Polymer Nanocarriers Directed to Intercellular Adhesion Molecule 1

Muro

Dziubla²,

Qiu³

et al. 2006

J Pharmacol Exp Ther

178

278

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Targeting of diagnostic and therapeutic agents to endothelial cells (ECs) provides an avenue to improve treatment of many maladies. For example, intercellular adhesion molecule 1 (ICAM-1), a constitutive endothelial cell adhesion molecule upregulated in many diseases, is a good determinant for endothelial targeting of therapeutic enzymes and polymer nanocarriers (PNCs) conjugated with anti-ICAM (anti-ICAM/PNCs). However, intrinsic and extrinsic factors that control targeting of anti-ICAM/PNCs to ECs (e.g., anti-ICAM affinity and PNC valency and flow) have not been defined. In this study we tested in vitro and in vivo parameters of targeting to ECs of anti-ICAM/ PNCs consisting of either prototype polystyrene or biodegradable poly(lactic-coglycolic) acid polymers (ϳ200 nm diameter spheres carrying ϳ200 anti-ICAM molecules). Anti-ICAM/ PNCs, but not control IgG/PNCs 1) rapidly (t 1/2 ϳ5 min) and specifically bound to tumor necrosis factor-activated ECs in a dose-dependent manner (B max ϳ350 PNC/cell) at both static and physiological shear stress conditions and 2) bound to ECs and accumulated in the pulmonary vasculature after i.v. injection in mice. Anti-ICAM/PNCs displayed markedly higher EC affinity versus naked anti-ICAM (K d ϳ80 pM versus ϳ8 nM) in cell culture and, probably because of this factor, higher value (185.3 Ϯ 24.2 versus 50.5 Ϯ 1.5% injected dose/g) and selectivity (lung/blood ratio 81.0 Ϯ 10.9 versus 2.1 Ϯ 0.02, in part due to faster blood clearance) of pulmonary targeting. These results 1) show that reformatting monomolecular anti-ICAM into high-affinity multivalent PNCs boosts their vascular immunotargeting, which withstands physiological hydrodynamics and 2) support potential anti-ICAM/PNCs utility for medical applications.

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“…Fab19 and Fab48 are derived from two versions of humanized anti-ICAM-1 MAb 1A6 (R. J. Colonno, J. H. Condra, J. E. Tomassini, and V. V. Sardana, European patent application 0459577A2), called HscB and HscE, respectively (11). The expression vector produces protein from a bicistronic operon that directs the expression of both lightchain and composite heavy-chain polypeptides.…”

Section: Resultsmentioning

confidence: 99%

“…The gene segment encoding the light chain was synthesized by PCR with two overlapping templates, the V L fragment derived from two versions of humanized scFv MAb 1A6 (HscB for Fab19 and HscE for Fab48 [11]) and the C L fragment derived from the human 1 light-chain constant region (14). The PCR product was cloned into the pCR 2.1 TOPO cloning vector (Invitrogen).…”

Section: Methodsmentioning

confidence: 99%

“…The vector includes a mechanism for generating a variety of multimeric Fab molecules through insertion of polymerization domain sequences between a unique SacI site just before the stop codon in the C H domain and a unique EcoRI site just after the stop codon. The expression vector pTexK was derived from pBR322 and contains the isopropyl-␤-D-thiogalactopyranoside (IPTG)-inducible tac promoter and a kanamycin resistance gene.The gene segment encoding the light chain was synthesized by PCR with two overlapping templates, the V L fragment derived from two versions of humanized scFv MAb 1A6 (HscB for Fab19 and HscE for Fab48 [11]) and the C L fragment derived from the human 1 light-chain constant region (14). The PCR product was cloned into the pCR 2.1 TOPO cloning vector (Invitrogen).…”

mentioning

confidence: 99%

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Prevention of Human Rhinovirus Infection by Multivalent Fab Molecules Directed against ICAM-1

Charles

Luo

Kohlstaedt

et al. 2003

Antimicrob Agents Chemother

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We have developed a technology for improving avidity by making bivalent, trivalent, or tetravalent recombinant polypeptides. We designed tripartite proteins consisting of the Fab fragment of an antibody fused with a hinge derived from human immunoglobulin D that was further linked to polymerization domains derived from human coiled-coil proteins. We report here on the application of this method with a Fab domain directed against the major human rhinovirus receptor, intercellular adhesion molecule 1 (ICAM-1). Multivalent anti-ICAM-1 molecules were produced in bacteria and purified as soluble preassembled homogeneous proteins at high yield. These proteins successfully blocked rhinovirus infection in vitro, with the efficiency increasing from monomer to dimer, trimer, and tetramer. The diminished dissociation rate of these multivalent antibodies and their improved efficacy in preventing rhinovirus infection provide a foundation for producing prophylactic and therapeutic molecules against human rhinovirus, the causative agent of the majority of common colds.Human rhinoviruses (HRVs) are the major cause of the common cold (23,24) and are among the most frequently occurring human pathogens. These viruses infect cells of the nasal epithelium by binding to cell surface receptors. On the basis of their cellular receptor specificities, the more than 100 HRV serotypes can be divided into two groups. The major group contains about 90% of all serotypes and uses intercellular adhesion molecule 1 (ICAM-1) as its receptor (25). A receptor-blocking approach has shown that anti-ICAM-1 monoclonal antibody (MAb) 1A6 prevents HRV infection of cells in vitro (3). In human clinical trials, the antibody diminished cold symptoms but failed to prevent onset of the disease (8). The limited efficacy of MAb 1A6 is most likely due to its low functional affinity (or avidity) for ICAM-1 compared to that of the multivalent HRV particles. Consistent with this interpretation was a study in which several MAbs against ICAM-1 were shown to dissociate from ICAM-1 at the same rate as HRV itself (i.e., they had equal dissociation rate constants [k off s]) (2).For over a decade, science has tried to improve on nature by using multivalency to increase the avidities of antibodies. The majority of these proteins were engineered by fusing the antigen-binding scFv fragment of an antibody with a polymerization domain, such as streptavidin (10), the yeast protein GCN4 leucine zipper domain (7, 18), or the tetramerization domain of p53 (18). Fab fragments of antibodies, however, generally exhibit significantly higher binding affinities than their corresponding scFv fragments. Because of the greater complexity of assembling multichain proteins, little work has been done on making trimeric and tetrameric Fab molecules. Our method involves the generation of tripartite proteins consisting of a humanized Fab, a linker sequence derived from the human immunoglobulin D (IgD) hinge (13), and a multimerization domain derived from either of the human transcription factors...

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Humanization of an anti-ICAM-1 antibody with over 50-fold affinity and functional improvement

Cited by 24 publications

References 27 publications

Blocking Development of a CD8+ T Cell Response by Targeting Lymphatic Recruitment of APC

Blocking Development of a CD8+ T Cell Response by Targeting Lymphatic Recruitment of APC

Endothelial Targeting of High-Affinity Multivalent Polymer Nanocarriers Directed to Intercellular Adhesion Molecule 1

Prevention of Human Rhinovirus Infection by Multivalent Fab Molecules Directed against ICAM-1

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