Mimicry of cocaine by anti-idiotypic antibodies: Molecular basis and antagonist drug design

Drug development for treatment of cocaine addiction is greatly hindered by the extreme difficulty in designing a selective cocaine antagonist.; Peptides, generated from the antigen-combining regions of anti-idiotypic (anti-Id) monoclonal antibodies (mAbs) and selected for their ability to inhibit cocaine binding to the dopamine transporter (DAT) without interfering with dopamine uptake, should function as cocaine antagonists. To test this hypothesis, six specific aims were pursued: (1) identification and characterization of anti-Id mAbs that mimic the configuration of cocaine, (2) stable expression of human DAT (hDAT) in mouse neuroblastoma cells, (3) characterization of selected anti-Id mAbs with respect to their binding affinity and specificity for hDAT, (4) generation of single chain (antibody) variable fragments (scFv), (5) investigation of the ability of these scFv to block cocaine binding to hDAT without interfering with dopamine uptake, and (6) sequence and structure (molecular modeling) analysis.; Mouse neuronal cell lines (N1E-115 and Neuro2A) stably expressing a fully functional hDAT were successfully constructed. When tested in this system, some anti-Id mAbs mimicked the configuration of cocaine and bound DAT with excellent specificity. One scFv engineered from a selected anti-Id mAb (K2-3f) bound DAT and its binding affinity was stronger than that of cocaine. More importantly, scFv K2-3f completely blocked the binding of cocaine at a low molar concentration (50 nM) while allowing equimolar dopamine uptake. Sequence analysis and molecular models of anti-Id mAbs indicated the amino acid residues involved in antigen mimicry. These findings provide a novel approach to develop cocaine antagonists. The intriguing application of anti-Id antibodies in rational design of drugs could be widely applied to potential ligand-receptor interactions in the treatment of other diseases.; [M. Ho was supported by National Research Service Award Predoctoral Fellowship F31-DA14484 from the National Institutes of Health, University of Illinois Fellowship (1-1-10742) and On-Campus Dissertation Research Grant (1-2-16449). This work was supported by NIDA research grant R15-DA10367 and University of Illinois Research Board 1-2-68205.]...