IN-VIVO TARGETING OF ACUTE MYOCARDIAL-INFARCTION WITH NEGATIVE-CHARGE, POLYMER-MODIFIED ANTIMYOSIN ANTIBODY - USE OF DIFFERENT CROSS-LINKERS

Background. Cell surfaces and intercellular matrixes contain acidic re sidues, making them negatively charged. Antibodies are basic, positive ly charged glycoproteins. Therefore the potential for nonspecific ioni c interaction exists, which could increase the background activity. Mo dification 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-l inkers used covalently to link the chelating negatively charge-modifie d polylysine to antimyosin Fab (AM-Fab). The cross-linking was perform ed through peptide (AM-I) or thioether (AM-II) bonds. The in vitro eva luation of the immunointegrity and the in vivo assessment were perform ed 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 on ly partially negatively charge modified [AM-IIns]) was also assessed. Methods and Results. All polymer-modified preparations (AM-I, AM-IIs, and AM-IIns) retained the immunoreactivities relative to the unmodifie d or conventional diethylenetriaminepentaacetic acid-coupled AM-Fab as assessed by radioimmunoassay or enzyme-linked immunosorbent assay. Th ese polymer-modified preparations labeled with In-111 were assessed in 13 rabbits with acute experimental myocardial infarction. Acute infar cts were produced by 40 minutes of left anterior descending coronary a rtery occlusion followed by reperfusion. At between 10 and 30 minutes of reperfusion, 10.4 +/- 1.8 mBq In-111-AM-I (10 to 20 mu g; n = 7) or 11.4 +/- 2.3 mBq In-111-AM-II (n or ns) (20 to 25 mu g; n = 6) was ad ministered intravenously. Gamma imaging was performed in the left late ral position and arterial blood samples were withdrawn serially for th e next 3 hours. At the end of the final imaging session, AM-I uptake w as determined to be 1.09% +/- 0.11% (mean percent injected dose per gr am myocardium +/- SFM) in 20 infarcted myocardial segments from seven rabbits, compared with 0.031% +/- 0.003% in 20 normal myocardial segme nts (infarct/normal myocardial ratio 53.9 +/- 18.41). The mean percent injected dose of In-111-labeled thioether-linked AM-Fab preparations in nine infarcted myocardial segments from each group was 0.067% a 0.0 08% (infarct/normal myocardial ratio 9.0 +/- 1.5) and 0.144% +/- 0.011 % (infarct/normal myocardial ratio 10.2 +/- 1.9) with AM-IIs in = 3) a nd AM-IIns (n = 3), respectively (p < 0.0001). The non-target organ di stribution of the AM-I and AM-IIs was similar. AM-IIns preparation res ulted in high non-target organ activities. Conclusions. This study sho ws that the charge of the antibody can be manipulated favorably by cro ss-linking with negatively charged polymers, which results in the redu ced in vivo non-target organ activities. Charge modification does not adversely affect the apparent affinity of the antibody. However, the t ype of cross-linkers used may significantly influence the in vivo stab ility of the modified antibody preparations for target organ visualiza tion. These data may find potential application in future clinical ima ging protocols.