Integrins are ?/? heterodimeric adhesion receptors that play essential roles in kidney formation during development and in regeneration of the adult kidney following inflammatory or vascular injury. Abnormal integrin function can lead, however, to inflammatory, autoimmune and thrombotic disorders affecting the kidney and the vasculature, thus making integrins important therapeutic targets. Existing ligand-mimetic monoclonal antibodies (mAbs) and small molecules (fashioned based on structures of natural ligands) exert agonist-like properties, by engaging the integrin ligand binding site, like the parent physiologic ligand, thus inducing cell adhesion responses these drugs were designed to prevent, and underscoring the need for novel approaches targeting integrins in disease. Integrins comprise two subgroups, depending on the presence or absence of a von Willebrand's Factor Type A-domain (vWFA) (?A- or I domain) found in one-half of the integrin ?-subunits. ?A mediates Mg2+- dependent binding of an aspartate (Asp) or glutamate (Glu) residue from integrin ligands (or ligand-mimetic drugs) at a metal-ion-dependent-adhesion site (MIDAS) in ?A. ?A-lacking integrins bind ligands through an ?A-like domain (?A or I-like) present in their ?-subunits. The ligand Asp/Gu binds the Mg2+ at MIDAS monodentately, completing an octahedral coordination sphere around the metal ion. Formation of the Mg2+- Asp (or Glu) bond triggers activating conformational changes in the integrin leading to cell adhesion, accounting for the agonist-like properties of existing integrin antagonists. In solving the crystal structure of a ligand-mimetic mAb, which binds at MIDAS in the ?A domain but prevents conformational switching in the integrin (i.e. acting as a pure antagonist), we found that a Ca2+ is heptacoordinated at MIDAS, the result of the unusual symmetric bidentate ligation by the mAb-derived Asp. Stable heptacoordination of Ca2+ blocked the conformational switching normally induced by ligand occupancy of MIDAS. We hypothesize that it should therefore be feasible to design structure-based cyclized peptide antagonists to the ?A domain based on our crystal structure, and that switching ligand Asp coordination from the monodentate to the bidentate binding mode in the ?A MIDAS, thus stabilizing a Ca2+ there, will also achieve a similar outcome in the ?A-lacking integrins. The present proposal represents a multidisciplinary approach with participation by cell-, structural biology and medicinal chemistry groups. An outcome from these studies will be the development of new probes for assessing structure-activity relationships of these dynamic receptors and could provide new therapeutic strategies to treat a wide-spectrum of disease states affecting the kidney and the vasculature.

Public Health Relevance

Improper activation of integrins plays critical roles in a diverse range of diseases including inflammatory, autoimmune diseases of the kidney and the vasculature as well as cancer metastasis. Current drugs targeting these receptors have contributed to adverse autoimmune reactions and to increased mortality, the result of inadvertent activation of integrins. Identifying drugs that maintain the integrin in the inactive state is highly desirable and is the subject of this proposal.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Exploratory/Developmental Grants (R21)
Project #
5R21DK096334-02
Application #
8549221
Study Section
Special Emphasis Panel (KMBD)
Program Officer
Hoshizaki, Deborah K
Project Start
2012-09-22
Project End
2014-08-31
Budget Start
2013-09-01
Budget End
2014-08-31
Support Year
2
Fiscal Year
2013
Total Cost
$253,051
Indirect Cost
$71,410
Name
Massachusetts General Hospital
Department
Type
DUNS #
073130411
City
Boston
State
MA
Country
United States
Zip Code
02199
Rui, Xianliang; Mehrbod, Mehrdad; Van Agthoven, Johannes F et al. (2014) The ?-subunit regulates stability of the metal ion at the ligand-associated metal ion-binding site in ?3 integrins. J Biol Chem 289:23256-63
Mahalingam, Bhuvaneshwari; Van Agthoven, Johannes F; Xiong, Jian-Ping et al. (2014) Atomic basis for the species-specific inhibition of ?V integrins by monoclonal antibody 17E6 is revealed by the crystal structure of ?V?3 ectodomain-17E6 Fab complex. J Biol Chem 289:13801-9
Van Agthoven, Johannes F; Xiong, Jian-Ping; Alonso, José Luis et al. (2014) Structural basis for pure antagonism of integrin ?V?3 by a high-affinity form of fibronectin. Nat Struct Mol Biol 21:383-8