Cocaine Esterase Prevents Cocaine-Induced Toxicity and the Ongoing Intravenous Self-Administration of Cocaine in Rats

Abstract

Cocaine esterase (CocE) is a naturally occurring bacterial enzyme, is a very efficient protein catalyst for the hydrolysis of cocaine, and has previously been shown to protect rodents from the lethal effects of cocaine. The current studies were aimed at evaluating the capacity of a longer acting mutant form (CocE T172R/G173Q; DM CocE) of CocE to protect against the lethal effects of cocaine, and alter ongoing intravenous cocaine self-administration in rats. A dose-response analysis revealed a dose-dependent suppression of cocaine-reinforced responding with 1.0 mg of CocE T172R/G173Q producing saline-like rates of responding. The effects of 1.0 mg of CocE T172R/ G173Q on cocaine-reinforced responding were then compared with responding when saline was available for injection, whereas the selectivity of CocE T172R/G173Q's effects was assessed by evaluating the effects of 1.0 mg of CocE T172R/ G173Q on (Ϫ)-2␤-carbomethoxy-3␤-phenyltropane (WIN-35065-2)-and food-reinforced responding. Although 1.0 mg of CocE T172R/G173Q suppressed responding maintained by 0.1 mg/kg/injection cocaine, a significant increase in responding was observed when responding was maintained by 1.0 mg/kg/ injection cocaine, resulting in a 10-fold rightward shift in the dose-response curve for cocaine self-administration at a dose that did not significantly alter responding maintained by either WIN-35065-2 or food. These findings demonstrate that a longacting form of CocE is effective at abruptly reducing the ongoing self-administration of low doses of cocaine, and provides a robust antagonism of cocaine's reinforcing effects. Furthermore, these studies provide strong evidence for the potential usefulness of a suitable, stable, and long-acting form of CocE as a pharmacotherapy for cocaine abuse in humans.

Long-Lasting Effects of a PEGylated Mutant Cocaine Esterase (CocE) on the Reinforcing and Discriminative Stimulus Effects of Cocaine in Rats

“…Previous studies with mutant bacterial CocEs (Brim et al, 2010;Collins et al, 2009) or mutant human BChEs (Carroll et al, 2011) have demonstrated the capacity of highly efficient cocaine-specific hydrolase to inhibit the reinforcing effects of cocaine; however, in each of these studies the inhibitory effects of the enzymes were short-lived, lasting from minutes to hours. The results of the current studies with PEG-CCRQ CocE not only confirm this effect of cocaine-specific esterases, but they also provide convergent evidence that PEG-CCRQ CocE represents a significant improvement over previously described mutant bacterial and human cocaine hydrolases with respect to both the magnitude and duration of its antagonism of the reinforcing effects of cocaine.…”

“…Nevertheless, just as the prolonged duration of PEG-CCRQ CocE's protective effects differentiated it from DM CocE in rodent models of acute cocaine toxicity (Collins et al, 2011b;Narasimhan et al, 2011) differences between these enzymes began to emerge when PEG-CCRQ CocE was evaluated against cocaine self-administration using a multiple dose procedure that allowed for the longitudinal evaluation of an animal's sensitivity to the reinforcing effects of cocaine. Unlike the effects of B3 mg/ kg DM CocE, which had completely dissipated by 24 h (Collins et al, 2009), pretreatment with 3.2 mg/kg PEG-CCRQ CocE produced a longer lasting change in cocaine self-administration with a three-fold rightward shift in the dose-response curve observed 24 h later, and baseline-like rates of responding not observed until 48 h after administration. Interestingly, in addition to producing larger Figure 7 Effects of 32.0 mg/kg; IV PEG-CCRQ CocE on the cocaine-like interoceptive effects of methylphenidate, d-amphetamine, and cocaine.…”

“…In an extension of previous findings with DM CocE (T172R/G173Q CocE or RQ CocE; Collins et al, 2009Collins et al, , 2011aGao et al, 2009;Narasimhan et al, 2010), the current studies evaluated impact of introducing two cysteine residues at the dimer interface, in combination with the PEGylation of this crosslinked CocE dimer (PEG-CCRQ CocE; Narasimhan et al, 2011) on duration and effectiveness of CocE to alter the behavioral effects of cocaine. Importantly, just as these changes did not alter the enzymatic activity of PEG-CCRQ CocE in vitro , the DM (RQ) and PEG-CCRQ forms of CocE appeared to be equally effective at eliminating cocaine in vivo as both enzymes produced at least 10-fold rightward shifts in the reinforcing effects of cocaine when comparable doses of B3 mg/kg were administered immediately before the start of cocaine self-administration sessions (Collins et al, 2009).…”

“…Importantly, just as these changes did not alter the enzymatic activity of PEG-CCRQ CocE in vitro , the DM (RQ) and PEG-CCRQ forms of CocE appeared to be equally effective at eliminating cocaine in vivo as both enzymes produced at least 10-fold rightward shifts in the reinforcing effects of cocaine when comparable doses of B3 mg/kg were administered immediately before the start of cocaine self-administration sessions (Collins et al, 2009). Nevertheless, just as the prolonged duration of PEG-CCRQ CocE's protective effects differentiated it from DM CocE in rodent models of acute cocaine toxicity (Collins et al, 2011b;Narasimhan et al, 2011) differences between these enzymes began to emerge when PEG-CCRQ CocE was evaluated against cocaine self-administration using a multiple dose procedure that allowed for the longitudinal evaluation of an animal's sensitivity to the reinforcing effects of cocaine.…”

“…Sitedirected mutagenesis studies aimed at improving the thermostability of CocE have identified an equally efficient mutant CocE (T172R/G173Q CocE, RQ CocE, DM CocE) with an in vivo half-life of B4.5 h in mice (Gao et al, 2009;Narasimhan et al, 2010), and a high degree of pharmacologic specificity (Brim et al, 2011). In addition to dose dependently inhibiting the cardiovascular, convulsant, lethal, and reinforcing effects of cocaine in rats (Collins et al, 2009(Collins et al, , 2011b, DM CocE has also been shown to effectively reverse the cardiovascular effects associated with large intravenous (IV) doses of cocaine in rhesus monkeys (Collins et al, 2011a). While these findings suggest that DM CocE should provide an effective treatment option for acute cocaine toxicity in humans, its in vivo duration of action is likely too short to provide a realistic option for the treatment of cocaine abuse.…”