Radiolabeled Constructs for Evaluation of the Asialoglycoprotein Receptor Status and Hepatic Functional Reserves

University of Nebraska at Omaha, Omaha, Nebraska, United States
Bioconjugate Chemistry (Impact Factor: 4.51). 08/2003; 14(5):997-1006. DOI: 10.1021/bc034081a
Source: PubMed


Transplantation of isolated hepatocytes may eventually replace a whole liver transplantation for the treatment of selected liver metabolic disorders and acute hepatic failure. To understand the behavior of transplanted hepatocytes, methods for longitudinal assessment of functional activity and survival of hepatocyte transplants must be developed. Targeting of asialoglycoprotein receptor (ASGPr) with various radiolabeled or Gd-labeled constructs of asialofetuin (AF) is expected to allow noninvasive and quantitative assessments of the ASGPr status in functioning hepatocytes before and after the transplant. Six new constructs of (125)I-, (99m)Tc-, (153)Gd-, and (111)In-radiolabeled AF with distinct stabilities and clearance rates were prepared and evaluated in vitro in mice, rat, porcine, and human hepatocytes, and in vivo in mice and rats. The blood and organ clearance rates, as well as liver and spleen uptake, were measured. Even extensive chemical modifications of AF with poly-l-lysine and various chelating agents do not appear to diminish AF's binding to ASGPr. Binding to isolated hepatocytes and the in vivo liver uptake studies indicate unimpaired functional activity of AF as evidenced by the rapid (<10 min) and nearly complete hepatic extraction of AF constructs from the systemic circulation. The catabolic processing and elimination of AF constructs from liver depend on the chemical modification used in the preparation of a given reagent. Radioiodinated AF has by far the shortest postabsorption (5.1 min +/- 0.05 min) and elimination half-lives (2.8 +/- 0.06 h) in liver. In comparison, the AF construct prepared by conjugation of DTPA- and 2-iminothiolane-substituted p-Lys with N-sulfosuccinimidyl 4-(p-maleimidophenyl)butyrate (SMPB)-modified AF (AF-SMPB-Traut-p-Lys-((111)In-DTPA)(20)(-)(30)) has a hepatic postabsorption time of 9.1 +/- 0.1 min and an elimination half-life of 44.3 +/- 3.08 h, whereas [(99m)Tc]technetium-labeled AF appears to be permanently retained in liver. These differences in rates of liver uptake and clearance of catabolized radiolabeled AF can be used to determine functional activity of liver and transplanted hepatocytes.

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    • "An example of this approach is the ability to determine and quantitate the risks of intrapulmonary shunting of transplanted cells via portosystemic collaterals. Moreover, transplanted cells may be localized in extrahepatic sites by imaging of hepatic receptor function, e.g., asialoglycoprotein receptor as has been shown in mice and rats [69]. For longer-term tracking of transplanted cells, molecular imaging methods have also been developed. "
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    ABSTRACT: Work over several decades has laid solid foundations for the advancement of liver cell therapy. To date liver cell therapy in people has taken the form of hepatocyte transplantation for metabolic disorders with a hepatic basis, and for acute or chronic liver failure. Although clinical trials using various types of autologous cells have been implemented to promote liver regeneration or reduce liver fibrosis, clear evidence of therapeutic benefits have so far been lacking. Cell types that have shown efficacy in preclinical models include hepatocytes, liver sinusoidal endothelial cells, mesenchymal stem cells, endothelial progenitor cells, and macrophages. However, positive results in animal models have not always translated through to successful clinical therapies and more realistic preclinical models need to be developed. Studies defining the optimal repopulation by transplanted cells, including routes of cell transplantation, superior engraftment and proliferation of transplanted cells, as well as optimal immunosuppression regimens are required. Tissue engineering approaches to transplant cells in extrahepatic locations have also been proposed. The derivation of hepatocytes from pluripotent or reprogramed cells raises hope that donor organ and cell shortages could be overcome in the future. Critical hurdles to be overcome include the production of hepatocytes from pluripotent cells with equal functional capacity to primary hepatocytes and long-term phenotypic stability in vivo. Copyright © 2015 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.
    Journal of Hepatology 04/2015; 62(1S):S157-S169. DOI:10.1016/j.jhep.2015.02.040 · 11.34 Impact Factor
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    • "Natural ASGPR ligands have been shown to specifically accumulate in liver in vivo following, for example, intravenous administration of radio-iodinated asialorosomucoid in rats [30],111In labelled asialofetuin in mice and rats [31] and liposomes decorated with asialofetuin in mice [32]. Here we show, using microSPECT/CT imaging and biodistribution studies, striking differences between the distribution of the ASGPR specific dAb DOM26h-196-61 and the VHD2 isotype control dAb following radio labelling with 111In, with up to 20% of the injected dose of DOM26h-196-61 distributing to the liver 3 hours after intravenous administration. "
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    ABSTRACT: Interferon alpha (IFNα) is used for the treatment of hepatitis C infection and whilst efficacious it is associated with multiple adverse events including reduced leukocyte, erythrocyte, and platelet counts, fatigue, and depression. These events are most likely caused by systemic exposure to interferon. We therefore hypothesise that targeting the therapeutic directly to the intended site of action in the liver would reduce exposure in blood and peripheral tissue and hence improve the safety and tolerability of IFNα therapy. We genetically fused IFN to a domain antibody (dAb) specific to a hepatocyte restricted antigen, asialoglycoprotein receptor (ASGPR). Our results show that the murine IFNα2 homolog (mIFNα2) fused to an ASGPR specific dAb, termed DOM26h-196-61, could be expressed in mammalian tissue culture systems and retains the desirable biophysical properties and activity of both fusion partners when measured . Furthermore a clear increase in targeting of the liver by mIFNα2-ASGPR dAb fusion protein, compared to that observed with either unfused mIFNα2 or mIFNα2 fused to an isotype control dAb VD2 (which does not bind ASGPR) was demonstrated using microSPECT imaging. We suggest that these findings may be applicable in the development of a liver-targeted human IFN molecule with improved safety and patient compliance in comparison to the current standard of care, which could ultimately be used as a treatment for human hepatitis virus infections.
    PLoS ONE 02/2013; 8(2):e57263. DOI:10.1371/journal.pone.0057263 · 3.23 Impact Factor
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