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Identification of Metabolites of ¹¹¹In-Diethylenetriaminepentaacetic Acid-Monoclonal Antibodies and Antibody Fragments in Vivo¹

The Mallinckrodt Institute of Radiology [B. E. R., F. N. F., J. R. D., W. B. E., C. J. A., M. J. W.] and Department of Surgery [J. M. C.], Washington University, School of Medicine, St. Louis, Missouri 63110

The in vivo fate of various ¹¹¹In-labeled polypeptides has been the subject of many investigations. Intracellular metabolism has been studied through the use of ¹¹¹In-labeled glycoproteins that are concentrated in the lysosome by receptor-mediated endocytosis. These studies have indicated that the main lysosomal metabolite is ¹¹¹In-chelate--lysine, both in vitro and in vivo (Y. Arano et al., J. Nucl. Med., 35: 890–898, 1994; F. N. Franano et al., Nucl. Med. Biol., 21: 1023–1034, 1994). Since the vast majority of radiolabeled antibodies do not localize within the target tissue, an understanding of the metabolism of ¹¹¹In-labeled antibodies in nontarget tissues is important for the rational design of future radiolabeled antibodies.

We investigated the in vivo metabolism of ¹¹¹In-DTPA³-conjugated antibody in female Sprague-Dawley rats using the anticolorectal carcinoma monoclonal antibody (MAb) 1A3 and MAb 1A3-F(ab')₂. Livers and kidneys were harvested from rats injected with either intact MAb or MAb fragments and analyzed by gel filtration chromatography. Thirty-five % of the radioactivity from ¹¹¹In-DTPA-1A3 MAb present in the liver was in the form of a low molecular weight species at 1 through 5 days. In contrast, ¹¹¹In-DTPA-1A3-F(ab')₂ was >98% degraded to a low molecular weight species in the kidney after 1 day. In each case, the low molecular weight metabolites were collected and further analyzed by silica gel thin-layer chromatography, reversed phase high-performance liquid chromatography, and ion-exchange chromatography and compared to ¹¹¹In-DTPA and ¹¹¹In-DTPA--lysine standards. In each system, the major metabolite co-eluted with ¹¹¹In-DTPA--lysine, similar to the results obtained with ¹¹¹In-labeled glycoproteins that are delivered to lysosomes by receptor-mediated endocytosis. A minor metabolite that was more highly charged than ¹¹¹In-DTPA was also observed. Analysis of urine and feces demonstrated that the main excretory product of both ¹¹¹In-labeled intact 1A3 and 1A3-F(ab')₂ was ¹¹¹In-DTPA--lysine. Based on this data, we propose that ¹¹¹In-DTPA-antibodies are degraded within lysosomes of nontarget organs such as the liver and kidneys.