title: Multifunctional nanoagent for thrombus-targeted fibrinolytic therapy.
abstract: BACKGROUND
  Current thrombolytic therapies utilize exogenous plasminogen activators (PAs) to effectively lyse clots, restoring blood flow, and preventing tissue and organ death. These PAs may also impair normal hemostasis, leading to life-threatening bleeding, including intracerebral hemorrhage.
  
  
  OBJECTIVE
  This study aims to develop new thrombus-targeted fibrinolytic agents that harness the multifunctional theranostic capabilities of nanomaterials, potentially allowing for the generation of efficacious thrombolytics while minimizing deleterious side effects.
  
  
  METHODS
  A thrombus-targeted nano-fibrinolytic agent was synthesized using a magnetofluorescent crosslinked dextran-coated iron oxide nanoparticle platform that was conjugated to recombinant tissue PA (tPA). Thrombus-targeting was achieved by derivatizing the nanoparticle with an activated factor XIII (FXIIIa)-sensitive peptide. Human plasma clot binding ability of the targeted and control agents was assessed by fluorescence reflectance imaging. Next, the in vitro enzymatic activity of the agents was assessed by S2288-based amidolytic activity, and an ELISA D-dimer assay for fibrinolysis. In vivo targeting of the nanoagent was next examined by intravital fluorescence microscopy of murine arterial and venous thrombosis. The fibrinolytic activity of the targeted nanoagent compared to free tPA was then evaluated in vivo in murine pulmonary embolism.
  
  
  RESULTS
  In vitro, the targeted thrombolytic nanoagent demonstrated superior binding to fresh-frozen plasma clots compared to control nanoagents (analysis of variance, p < 0.05). When normalized by S2288-based amidolytic activity, targeted, control and free tPA samples demonstrated equivalent in vitro fibrinolytic activity against human plasma clots, as determined by ELISA D-dimer assays. The FXIIIa targeted fibrinolytic nanoagent efficiently bound the margin of intravascular thrombi as detected by intravital fluorescence microscopy. In in vivo fibrinolysis studies the FXIIIa-targeted agent lysed pulmonary emboli with similar efficacy as free tPA (p > 0.05).
  
  
  CONCLUSIONS
  The applicability of a FXIIIa-targeted thrombolytic nanoagent in the treatment of thromboembolism was demonstrated in vitro and in vivo. Future studies are planned to investigate the safety profile and overall efficacy of this class of nanoagents.
authors.affiliation: Center for Systems Biology, Harvard Medical School & Massachusetts General Hospital, 149 13th Street, Charlestown, MA 02129, USA. jason_mccarthy@hms.harvard.edu
authors.forename: Jason R
authors.initials: JR
authors.lastname: McCarthy
authors.forename: Irina Y
authors.initials: IY
authors.lastname: Sazonova
authors.forename: S Sibel
authors.initials: SS
authors.lastname: Erdem
authors.forename: Tetsuya
authors.initials: T
authors.lastname: Hara
authors.forename: Brian D
authors.initials: BD
authors.lastname: Thompson
authors.forename: Purvish
authors.initials: P
authors.lastname: Patel
authors.forename: Ion
authors.initials: I
authors.lastname: Botnaru
authors.forename: Charles P
authors.initials: CP
authors.lastname: Lin
authors.forename: Guy L
authors.initials: GL
authors.lastname: Reed
authors.forename: Ralph
authors.initials: R
authors.lastname: Weissleder
authors.forename: Farouc A
authors.initials: FA
authors.lastname: Jaffer
mesh_terms: D000818:Animals; D003911:Dextrans / Q000737:chemistry; D016503:Drug Delivery Systems / Q000379:methods*; D005260:Female; D005290:Ferric Compounds / Q000737:chemistry; D006801:Humans; D008168:Lung / Q000098:blood supply / Q000187:drug effects / Q000473:pathology; D051379:Mice; D008810:Mice, Inbred C57BL; D053758:Nanoparticles / Q000737:chemistry*; D010960:Plasminogen Activators / Q000008:administration & dosage* / Q000627:therapeutic use*; D011651:Pulmonary Artery / Q000187:drug effects / Q000473:pathology; D011655:Pulmonary Embolism / Q000188:drug therapy / Q000473:pathology; D011994:Recombinant Proteins / Q000008:administration & dosage / Q000627:therapeutic use; D015912:Thrombolytic Therapy / Q000379:methods*; D013927:Thrombosis / Q000188:drug therapy* / Q000473:pathology; D014680:Veins / Q000187:drug effects / Q000473:pathology; D020246:Venous Thrombosis / Q000188:drug therapy / Q000473:pathology
chemical_list: D003911:Dextrans; D005290:Ferric Compounds; D011994:Recombinant Proteins; C000499:ferric oxide; D010960:Plasminogen Activators