title: In vivo targeting of acute myocardial infarction with negative-charge, polymer-modified antimyosin antibody: use of different cross-linkers.
abstract: BACKGROUND
  Cell surfaces and intercellular matrixes contain acidic residues, making them negatively charged. Antibodies are basic, positively charged glycoproteins. Therefore the potential for nonspecific ionic interaction exists, which could increase the background activity. Modification of antibodies with negatively charge-modified polymers have been shown to reduce this nonspecific background activity. This study was performed to investigate the appropriateness of different cross-linkers used covalently to link the chelating negatively charge-modified polylysine to antimyosin Fab (AM-Fab). The cross-linking was performed through peptide (AM-I) or thioether (AM-II) bonds. The in vitro evaluation of the immunointegrity and the in vivo assessment were performed to investigate the potential for reduction of nontarget background activity. Furthermore, the role of the charge of the polymers (whether completely negatively charge modified by succinylation [AM-IIs] or only partially negatively charge modified [AM-IIns]) was also assessed.
  
  
  RESULTS
  All polymer-modified preparations (AM-I, AM-IIs, and AM-IIns) retained the immunoreactivities relative to the unmodified or conventional diethylenetriaminepentaacetic acid-coupled AM-Fab as assessed by radioimmunoassay or enzyme-linked immunosorbent assay. These polymer-modified preparations labeled with 111In were assessed in 13 rabbits with acute experimental myocardial infarction. Acute infarcts were produced by 40 minutes of left anterior descending coronary artery occlusion followed by reperfusion. At between 10 and 30 minutes of reperfusion, 10.4 +/- 1.8 mBq 111In-AM-I (10 to 20 micrograms; n = 7) or 11.4 +/- 2.3 mBq 111In-AM-II (n or ns) (20 to 25 micrograms; n = 6) was administered intravenously. Gamma imaging was performed in the left lateral position and arterial blood samples were withdrawn serially for the next 3 hours. At the end of the final imaging session, AM-I uptake was determined to be 1.09% +/- 0.11% (mean percent injected dose per gram myocardium +/- SEM) in 20 infarcted myocardial segments from seven rabbits, compared with 0.031% +/- 0.003% in 20 normal myocardial segments (infarct/normal myocardial ratio 53.9 +/- 18.41). The mean percent injected dose of 111In-labeled thioether-linked AM-Fab preparations in nine infarcted myocardial segments from each group was 0.067% +/- 0.008% (infarct/normal myocardial ratio 9.0 +/- 1.5) and 0.144% +/- 0.011% (infarct/normal myocardial ratio 10.2 +/- 1.9) with AM-IIs (n = 3) and AM-IIns (n = 3), respectively (p < 0.0001). The non-target organ distribution of the AM-I and AM-IIs was similar. AM-IIns preparation resulted in high non-target organ activities.
  
  
  CONCLUSIONS
  This study shows that the charge of the antibody can be manipulated favorably by cross-linking with negatively charged polymers, which results in the reduced in vivo non-target organ activities. Charge modification does not adversely affect the apparent affinity of the antibody. However, the type of cross-linkers used may significantly influence the in vivo stability of the modified antibody preparations for target organ visualization. These data may find potential application in future clinical imaging protocols.
authors.affiliation: Northeastern University, Boston, MA 02115, USA.
authors.forename: J
authors.initials: J
authors.lastname: Narula
authors.forename: V P
authors.initials: VP
authors.lastname: Torchilin
authors.forename: A
authors.initials: A
authors.lastname: Petrov
authors.forename: S
authors.initials: S
authors.lastname: Khaw
authors.forename: V S
authors.initials: VS
authors.lastname: Trubetskoy
authors.forename: S M
authors.initials: SM
authors.lastname: O'Donnell
authors.forename: N D
authors.initials: ND
authors.lastname: Nossiff
authors.forename: B A
authors.initials: BA
authors.lastname: Khaw
mesh_terms: D000818:Animals; D007140:Immunoglobulin Fab Fragments; D007205:Indium Radioisotopes; D008297:Male; D009203:Myocardial Infarction / Q000000981:diagnostic imaging*; D009218:Myosins / Q000276:immunology*; D011817:Rabbits; D016719:Radioimmunodetection; D014018:Tissue Distribution
chemical_list: D007140:Immunoglobulin Fab Fragments; D007205:Indium Radioisotopes; D009218:Myosins