Document Detail

Chemically self-assembled antibody nanostructures as potential drug carriers.
MedLine Citation:
PMID:  23013206     Owner:  NLM     Status:  MEDLINE    
Chemically self-assembled antibody nanorings (CSANs) displaying multiple copies of single-chain variable fragments can be prepared from dihydrofolate reductase (DHFR) fusion proteins and bis-methotrexate (bisMTX). We have designed and synthesized a bisMTX chemical dimerizer (bisMTX-NH(2)) that contains a third linker arm that can be conjugated to fluorophores, radiolabels, and drugs. Monovalent, divalent, and higher-order AntiCD3 CSANs were assembled with a fluorescein isothiocyanate (FITC)-labeled bis-methotrexate ligand (bisMTX-FITC) and found to undergo rapid internalization and trafficking by HPB-MLT, a CD3+ T-leukemia cell line, to the early and late endosome and lysosome. Because the fluorescence of bisMTX-FITC when incorporated into CSANs was found to be significantly greater than that of the free ligand, the stability of the endocytosed AntiCD3 CSANs could be monitored. The internalized CSANs were found to be stable for several hours, while treatment with the nontoxic DHFR inhibitor trimethoprim resulted in a rapid loss (>80%) of cellular fluorescence within minutes, consistent with efficient intracellular disassembly of the nanorings. Over longer time periods (24 h), cellular fluorescence decreased by 75-90%, regardless of whether cells had been treated with DMSO or trimethoprim. Although bisMTX is a potent inhibitor of DHFR, it was found to be nontoxic (GI(50) > 20 μM) to HPB-MLT cells. In contrast, AntiCD3 CSANs prepared with bisMTX were found to be at least 13-fold more cytotoxic (GI(50) = 0.5-1.5 μM) than bisMTX at 72 h. Consistent with our findings from CSAN stability studies, no increase in cytotoxicity was observed upon treatment with trimethoprim. Taken together, our results suggest that cell receptor targeting CSANs prepared with trifunctional bisMTX could be used as potential tissue selective drug carriers.
Adrian Fegan; Sidath C Kumarapperuma; Carston R Wagner
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Publication Detail:
Type:  Journal Article; Research Support, N.I.H., Extramural     Date:  2012-10-16
Journal Detail:
Title:  Molecular pharmaceutics     Volume:  9     ISSN:  1543-8392     ISO Abbreviation:  Mol. Pharm.     Publication Date:  2012 Nov 
Date Detail:
Created Date:  2012-11-05     Completed Date:  2013-04-30     Revised Date:  2013-11-07    
Medline Journal Info:
Nlm Unique ID:  101197791     Medline TA:  Mol Pharm     Country:  United States    
Other Details:
Languages:  eng     Pagination:  3218-27     Citation Subset:  IM    
Department of Medicinal Chemistry, University of Minnesota, Minneapolis, Minnesota 55455, United States.
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MeSH Terms
Antibodies, Monoclonal / chemistry*,  immunology
Antigens, CD3 / immunology*,  metabolism
Cell Proliferation
Chromatography, Gel
Drug Carriers*
Flow Cytometry
Leukemia, T-Cell / drug therapy*
Methotrexate / pharmacology*
Microscopy, Confocal
Microscopy, Fluorescence
Recombinant Fusion Proteins / immunology,  metabolism
Single-Chain Antibodies / immunology,  metabolism
Tetrahydrofolate Dehydrogenase / immunology*,  metabolism
Grant Support
Reg. No./Substance:
0/Antibodies, Monoclonal; 0/Antigens, CD3; 0/Drug Carriers; 0/Recombinant Fusion Proteins; 0/Single-Chain Antibodies; 59-05-2/Methotrexate; EC Dehydrogenase

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