Structural basis of activation-dependent binding of ligand-mimetic antibody AL-57 to integrin LFA-1

Zhang, Hongmin; Liu, Jin-huan; Yang, Wei; Springer, Timothy A.; Shimaoka, Motomu; Wang, Jia-Huai

doi:10.1073/pnas.0909301106

Cited by 19 publications

(17 citation statements)

References 27 publications

(37 reference statements)

Supporting

Mentioning

Contrasting

Order By: Relevance

“…The crystal structures (3HI5 and 3HI6) of the Fab fragment of AL-57 in complex with the locked IA I-domain demonstrates that AL-57 coordinates with the MIDAS in a similar geometry as ICAM-1 and that the binding site of AL-57 overlaps the ICAM-1 binding site on the I-domain (34). Additionally, a hydrophobic patch further stabilizes the AL-57/I-domain interface to outcompete ICAM-1 for the MIDAS site (32,34).…”

Section: Discussionmentioning

confidence: 99%

2E8 Binds to the High Affinity I-domain in a Metal Ion-dependent Manner

Carreño¹,

Brown²,

Li³

et al. 2010

Journal of Biological Chemistry

View full text Add to dashboard Cite

The activation of leukocyte function-associated antigen-1 (LFA-1) plays a critical role in regulating immune responses. The metal ion-dependent adhesion site on the I-domain of LFA-1 ␣ L subunit is the key recognition site for ligand binding. Upon activation, conformation changes in the I-domain can lead LFA-1 from the low affinity state to the high affinity (HA) state. Using the purified HA I-domain locked by disulfide bonds for immunization, we developed an mAb, 2E8, that specifically binds to cells expressing the HA LFA-1. The surface plasmon resonance analysis has shown that 2E8 only binds to the HA I-domain and that the dissociation constant (K D ) for HA I-domain is 197 nM. The binding of 2E8 to the HA I-domain is metal ion-dependent, and the affinity decreased as Mn 2؉ was replaced sequentially by Mg 2؉ and Ca 2؉ . Surface plasmon resonance analysis demonstrates that 2E8 inhibits the interaction of HA I-domain and ICAM-1. Furthermore, we found that 2E8 can detect activated LFA-1 on both JY and Jurkat cells using flow cytometry and parallel plate adhesion assay. In addition, 2E8 inhibits JY cell adhesion to human umbilical vein endothelial cells and homotypic aggregation. 2E8 treatment reduces the proliferation of both human CD4 ؉ and CD8 ؉ T cells upon OKT3 stimulation without the impairment of their cytolytic function. Taken together, these data demonstrate that 2E8 is specific for the high affinity form of LFA-1 and that 2E8 inhibits LFA-1/ICAM-1 interactions. As a novel activation-specific monoclonal antibody, 2E8 is a potentially useful reagent for blocking high affinity LFA-1 and modulating T cell activation in research and therapeutics.Leukocyte function-associated antigen-1 (LFA-1, 3 ␣ L ␤ 2 integrin, CD11a/CD18) belongs to the ␤ 2 subclass of integrins and is important for leukocyte adhesion and T cell activation (1, 2). The ligands for LFA-1 are intercellular adhesion molecule-1 (ICAM-1), ICAM-2, and ICAM-3, which are members of the Ig superfamily (1). Although LFA-1 is constitutively expressed on the surface of leukocytes in an inactive state, binding to its ligands requires cellular activation involving both affinity (conformational change within the molecule) and avidity (receptor clustering) enhancement (3-5). The regulation of LFA-1 activation plays a critical role during inflammatory and immune responses in mice and humans (6 -9).The ␣ L subunit of LFA-1 has two prominent structural features, an inserted-domain (I-domain) of about 200 amino acid residues and three EF hand-like repeats containing putative divalent cation binding sites (1). The I-domain of the LFA-1 ␣ L subunit is the ligand binding site and changes conformation upon activation (10). The changes in the I-domain from the low affinity state to the high affinity state lead to an increased affinity for ligand binding (11)(12)(13)(14)(15). The key recognition site for ligand binding is at the metal ion-dependent adhesion site (MIDAS) on the I-domain of the LFA-1 ␣ L subunit (1, 2). Constitutively expressed on the surface of leukoc...

show abstract

Section: Discussionmentioning

confidence: 99%

2E8 Binds to the High Affinity I-domain in a Metal Ion-dependent Manner

Carreño¹,

Brown²,

Li³

et al. 2010

Journal of Biological Chemistry

View full text Add to dashboard Cite

show abstract

“…Unlike AQC2, AL-57 binds to the open form of ␣L (68). It was suggested that a key salt bridge formed with Glu-241 of ␣L by either the native ligand ICAM-1 or AL-57 is necessary for the allosteric rearrangement to occur and that the reason AQC2 binds to the open form of ␣1I is that it does not form a similar bridge with the corresponding residue in ␣1I, Glu-255 (68). Unlike in the ␣L-ICAM interaction, this conserved glutamic acid (Glu-255 in ␣1 or Glu-256 in ␣2) is not critical for collagen binding by integrins (6,67).…”

Section: Discussionmentioning

confidence: 99%

“…AQC2 can be classified as a pseudo-ligand mimetic antibody, because it binds to ␣1I in a ligand-mimetic fashion via direct interaction of an antibody aspartate residue with the MIDAS metal ion, but it does not induce a change to the open conformation (30,68). The AQC2-␣1I complex was generated in the presence of Mg 2ϩ , and DXMS studies were carried out in a similar manner as described above for the collagen-␣1I complex (Fig.…”

Section: Dxms Profile Of the Complex Between ␣1imentioning

confidence: 99%

Dynamic Structural Changes Are Observed upon Collagen and Metal Ion Binding to the Integrin α1 I Domain

Weinreb

Gao

et al. 2012

Journal of Biological Chemistry

View full text Add to dashboard Cite

Background: Integrin ␣1 I domain undergoes conformational changes upon collagen binding. Results: Deuterium exchange was used to measure the effects of cations, collagen, or an antibody on the ␣1I solution structure. Conclusion: Full-length collagen and metal ions induce changes that differ in key aspects from previously proposed models for ␣1I activation. Significance: These studies support a new model for integrin I domain activation.

show abstract

“…A number of monoclonal antibodies (mAbs) that activate CD11b/CD18 and other β2 integrins or that bind in an activation-sensitive manner (together referred to as “activating mAbs”) have also been previously described in the literature [14–23]. KIM127 is an activation-dependent antibody that also activates human CD11b/CD18 by recognizing sites in the CD18 EGF2 domain that are buried in the inactive integrin conformation [15, 19, 24].…”

Section: Introductionmentioning

confidence: 99%

Small molecule agonists of integrin CD11b/CD18 do not induce global conformational changes and are significantly better than activating antibodies in reducing vascular injury

Faridi

Altintas

Gomez

et al. 2013

Biochimica et Biophysica Acta (BBA) - General Subjects

View full text Add to dashboard Cite

BACKGROUND CD11b/CD18 is a key adhesion receptor that mediates leukocyte adhesion, migration and immune functions. We recently identified novel compounds, leukadherins, that allosterically enhance CD11b/CD18-dependent cell adhesion and reduce inflammation in vivo, suggesting integrin activation to be a novel mechanism of action for the development of anti-inflammatory therapeutics. Since a number of well-characterized anti-CD11b/CD18 activating antibodies are currently available, we wondered if such biological agonists could also become therapeutic leads following this mechanism of action. METHODS We compared the two types of agonists using in vitro cell adhesion and wound-healing assays and using animal model systems. We also studied effects of the two types of agonists on outside-in signaling in treated cells. RESULTS Both types of agonists similarly enhanced integrin-mediated cell adhesion and decreased cell migration. However, unlike leukadherins, the activating antibodies produced significant CD11b/CD18 macro clustering and induced phosphorylation of key proteins involved in outside-in signaling. Studies using conformation reporter antibodies showed that leukadherins did not induce global conformational changes in CD11b/CD18 explaining the reason behind their lack of ligand-mimetic outside-in signaling. In vivo, leukadherins reduced vascular injury in a dose-dependent fashion, but, surprisingly, the anti-CD11b activating antibody ED7 was ineffective. CONCLUSIONS Our results suggest that small molecule allosteric agonists of CD11b/CD18 have clear advantages over the biologic activating antibodies and provide a mechanistic basis for the difference. GENERAL SIGNIFICANCE CD11b/CD18 activation represents a novel strategy for reducing inflammatory injury. Our study establishes small molecule leukadherins as preferred agonists over activating antibodies for future development as novel anti-inflammatory therapeutics.

show abstract

Structural basis of activation-dependent binding of ligand-mimetic antibody AL-57 to integrin LFA-1

Cited by 19 publications

References 27 publications

2E8 Binds to the High Affinity I-domain in a Metal Ion-dependent Manner

2E8 Binds to the High Affinity I-domain in a Metal Ion-dependent Manner

Dynamic Structural Changes Are Observed upon Collagen and Metal Ion Binding to the Integrin α1 I Domain

Small molecule agonists of integrin CD11b/CD18 do not induce global conformational changes and are significantly better than activating antibodies in reducing vascular injury

Contact Info

Product

Resources

About