Molecular imaging of factor XIIIa activity in thrombosis using a novel, near-infrared fluorescent contrast agent that covalently links to thrombi.
BACKGROUND
Activated factor XIII (FXIIIa) mediates fibrinolytic resistance and is a hallmark of newly formed thrombi. In vivo imaging of FXIIIa activity could further elucidate the role of this molecule in thrombosis and other biological processes and aid in the clinical detection of acute thrombi.


RESULTS
An FXIIIa-sensitive near-infrared fluorescence imaging agent (A15) was engineered by conjugating a near-infrared fluorochrome to a peptide ligand derived from the amino terminus of alpha2-antiplasmin. To evaluate the molecular specificity of A15 for FXIIIa, a control agent (C15) was also synthesized by modifying a single key glutamine residue in A15. Fluorescence imaging experiments with A15 demonstrated stronger thrombosis enhancement in human plasma clots in vitro (P<0.001 versus C15 clots and other controls). A15 was found to be highly specific for the active site of FXIIIa and was covalently bound to fibrin. In vivo murine experiments with A15 demonstrated significant signal enhancement in acute intravascular thrombi (P<0.05 versus C15 group). Minimal A15 enhancement was seen in older aged thrombi (>24 hours), consistent with an expected decline of FXIIIa activity over time. Imaging results were confirmed on correlative histopathology and fluorescence microscopy.


CONCLUSIONS
A15 is a novel optical imaging agent that is specifically crosslinked to fibrin by FXIIIa, permitting detection of FXIIIa activity in experimental thrombi in vivo. This agent should permit assessment of FXIIIa activity in a broad range of biological processes and could aid in the clinical diagnosis of acute thrombi.
Jaffer
Farouc A
FA
Center for Molecular Imaging Research, Massachusetts General Hospital, Harvard Medical School, Charlestown, USA. fjaffer@partners.org
Tung
Ching-Hsuan
CH
Wykrzykowska
Joanna J
JJ
Ho
Nan-Hui
NH
Houng
Aiilyan K
AK
Reed
Guy L
GL
Weissleder
Ralph
R
D000818:Animals; D002712:Chlorides; D003287:Contrast Media / Q000493:pharmacokinetics*; D003432:Cross-Linking Reagents / Q000494:pharmacology; D026122:Factor XIIIa / Q000032:analysis*; D005240:Feasibility Studies; D005290:Ferric Compounds / Q000633:toxicity; D005337:Fibrin / Q000032:analysis / Q000187:drug effects*; D005456:Fluorescent Dyes / Q000493:pharmacokinetics*; D006801:Humans; D051379:Mice; D008807:Mice, Inbred BALB C; D008856:Microscopy, Fluorescence; D010446:Peptide Fragments / Q000493:pharmacokinetics*; D011485:Protein Binding; D019265:Spectroscopy, Near-Infrared; D013927:Thrombosis / Q000139:chemically induced / Q000378:metabolism*; D013997:Time Factors; D000979:alpha-2-Antiplasmin / Q000493:pharmacokinetics*
D002712:Chlorides; D003287:Contrast Media; D003432:Cross-Linking Reagents; D005290:Ferric Compounds; D005456:Fluorescent Dyes; D010446:Peptide Fragments; C494793:acetyl-glycyl-asparagyl-glutaminyl-glutamyl-glutaminyl-valyl-seryl-prolyl-leucyl-threonyl-leucyl-leucyl-lysyl-lysyl-tryptophyl-cysteinyl-Alexa Fluoro 680 C2C2-maleimide conjugate; D000979:alpha-2-Antiplasmin; D005337:Fibrin; D026122:Factor XIIIa; C024555:ferric chloride