title: Specific targeting of atherosclerotic plaques in ApoE(-/-) mice using a new Camelid sdAb binding the vulnerable plaque marker LOX-1.
abstract: OBJECTIVE
  Molecular imaging has the potential to provide quantitative information about specific biological aspects of developing atherosclerotic lesions. This requires the generation of reliable, highly specific plaque tracers. This study reports a new camelid single-domain antibody fragment (sdAb) targeting the Lectin-like oxidized low-density lipoprotein receptor (LOX-1), a biomarker for the detection and molecular phenotyping of vulnerable atherosclerotic plaques.
  
  
  METHODS
  A camelid sdAb was generated and selected for high affinity binding to LOX-1. Ex vivo biodistribution and in vivo single photon emission computed tomography (SPECT)/computed tomography (CT) imaging studies were performed in wild-type mice and in fat-fed atherosclerotic apolipoprotein E-deficient mice with (99m)Tc-labeled sdAbs. Gamma-counting and autoradiography analyses were performed on dissected aorta segments with different degrees of plaque burden. The specificity of the LOX-1-targeting sdAb was evaluated by blocking with unlabeled sdAb or by comparison with a nontargeting (99m)Tc-labeled control sdAb.
  
  
  RESULTS
  We generated a sdAb binding LOX-1 with a KD of 280 pM ± 62 pM affinity. After (99m)Tc-labeling, the tracer had radiochemical purity higher then 99 % and retained specificity in in vitro binding studies. Tracer blood clearance was fast with concomitant high kidney retention. At 3 h after injection, uptake in tissues other than plaques was low and not different than background, suggesting a restricted expression pattern of LOX-1. Conversely, uptake in aortic segments increased with plaque content and was due to specific LOX-1 binding. In vivo SPECT/CT imaging 160 min after injection in atherosclerotic mice confirmed specific targeting of LOX-1-expressing aortic plaques.
  
  
  CONCLUSIONS
  The LOX-sdAb specifically targets LOX-1-expressing atherosclerotic plaques within hours after injection. The possibility to image LOX-1 rapidly after administration combined with the favourable biodistribution of a sdAb are beneficial for molecular phenotyping of atherosclerotic plaques and the generation of a future prognostic tracer.
authors.affiliation: Laboratory of Cellular and Molecular Immunology (CMIM), Vrije Universiteit Brussel (VUB), Pleinlaan 2, Brussels, 1050, Belgium, Jens.De.Vos@vub.ac.be.
authors.forename: Jens
authors.initials: J
authors.lastname: De Vos
authors.forename: Iris
authors.initials: I
authors.lastname: Mathijs
authors.forename: Catarina
authors.initials: C
authors.lastname: Xavier
authors.forename: Sam
authors.initials: S
authors.lastname: Massa
authors.forename: Ulrich
authors.initials: U
authors.lastname: Wernery
authors.forename: Luc
authors.initials: L
authors.lastname: Bouwens
authors.forename: Tony
authors.initials: T
authors.lastname: Lahoutte
authors.forename: Serge
authors.initials: S
authors.lastname: Muyldermans
authors.forename: Nick
authors.initials: N
authors.lastname: Devoogdt
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001011:Aorta / Q000378:metabolism / Q000473:pathology; D001057:Apolipoproteins E / Q000172:deficiency*; D015415:Biomarkers / Q000378:metabolism; D016466:CHO Cells; D002162:Camelus; D006224:Cricetinae; D003412:Cricetulus; D005260:Female; D008810:Mice, Inbred C57BL; D008969:Molecular Sequence Data; D058226:Plaque, Atherosclerotic / Q000378:metabolism* / Q000473:pathology; D011485:Protein Binding; D051127:Scavenger Receptors, Class E / Q000378:metabolism*; D061905:Single-Domain Antibodies / Q000737:chemistry / Q000276:immunology*; D013667:Technetium; D014018:Tissue Distribution
chemical_list: D001057:Apolipoproteins E; D015415:Biomarkers; D051127:Scavenger Receptors, Class E; D061905:Single-Domain Antibodies; D013667:Technetium