PT  - JOURNAL ARTICLE
AU  - Remy L. Brim
AU  - Mark R. Nance
AU  - Daniel W. Youngstrom
AU  - Diwahar Narasimhan
AU  - Chang-Guo Zhan
AU  - John J. G. Tesmer
AU  - Roger K. Sunahara
AU  - James H. Woods
TI  - A Thermally Stable Form of Bacterial Cocaine Esterase: A Potential Therapeutic Agent for Treatment of Cocaine Abuse
AID  - 10.1124/mol.109.060806
DP  - 2010 Jan 19
TA  - Molecular Pharmacology
PG  - mol.109.060806
4099  - http://molpharm.aspetjournals.org/content/early/2010/01/19/mol.109.060806.short
4100  - http://molpharm.aspetjournals.org/content/early/2010/01/19/mol.109.060806.full
AB  - Rhodococcal cocaine esterase (CocE) is an attractive potential treatment for both cocaine overdose and cocaine addiction. CocE directly degrades cocaine into inactive products, whereas traditional small-molecule approaches require blockade of the inhibitory-action of cocaine on a diverse array of monoamine transporters and ion channels. The usefulness of wild-type (wt) cocaine esterase, however, is hampered by its inactivation at 37°C. Herein we characterize the most thermostable form of this enzyme to date, CocE-L169K/G173Q. In vitro kinetic analyses reveal that CocE-L169K/G173Q displays a half-life of 2.9 days at 37°C, representing a 340-fold improvement over wt and 15-fold greater than previously reported mutants. Crystallographic analyses of CocE-L169K/G173Q, determined at 1.6 Å resolution, suggest that stabilization involves enhanced domain-domain interactions involving van der Waals interactions and hydrogen bonding. In vivo rodent studies reveal that intravenous pre-treatment with CocE-L169K/G173Q in rats provides protection from cocaine-induced lethality for longer time periods prior to cocaine administration than wt CocE. Furthermore, intravenous administration (pre-treatment) of CocE-L169K/G173Q prevents self-administration of cocaine in a time-dependent manner. Termination of its in vivo effects appears to be dependent on, but not proportional to, its clearance from plasma as its half-life is approximately 2.3 hours and similar to that of wt CocE (2.2 hrs). Taken together these data suggest that CocE-L169K/G173Q possesses many of the properties of a biological therapeutic for treating cocaine abuse, but requires additional development to improve its serum half-life.The American Society for Pharmacology and Experimental Therapeutics