Chiral Resolution of the Enantiomers of the Slow-Onset Dopamine Reuptake Inhibitor CTDP-32476 and Their Activities.

An improved synthesis was developed for CDTP-32476, a potent slow-onset dopamine reuptake blocker that may have utility as a treatment for cocaine abuse. The enantiomers of the compound were separated by fractional crystallization with N-acetylleucine enantiomers. An X-ray crystal structure was obtained of the RR enantiomer paired with N-acetyl-d-leucine. Chiral chromatography showed that the resolved enantiomers were pure with little contamination by the other enantiomer. The enantiomers were tested for their ability to block the reuptake of monoamines at their respective transporters and to stimulate locomotor activity in mice. Both enantiomers potently blocked the reuptake of dopamine and stimulated locomotor activity in mice. The RR enantiomer that corresponds to the active RR enantiomer of methylphenidate was slightly more potent at the dopamine reuptake site. The RR enantiomer also was found to be about twice as selective for the dopamine transporter relative to the norepinephrine transporter, which may have clinical implications. A method for designing slow-onset stimulants is proposed since there is increasing evidence that such activity is an important factor in stimulants that may have limited abuse potential.

The macamide N-3-methoxybenzyl-linoleamide is a time-dependent fatty acid amide hydrolase (FAAH) inhibitor.

The Peruvian plant Lepidium meyenii (Maca) has been shown to possess neuroprotective activity both in vitro and in vivo. Previous studies have also demonstrated the activity of the pentane extract and its macamides, the most representative lipophilic constituents of Maca, in the endocannabinoid system as fatty acid amide hydrolase (FAAH) inhibitors. One of the most active macamides, N-3-methoxybenzyl-linoleamide, was studied to determine its mechanism of interaction with FAAH and whether it has inhibitory activity on mono-acyl glycerol lipase (MAGL), the second enzyme responsible for endocannabinoid degradation. Macamide concentrations from 1 to 100 µM were tested using FAAH and MAGL inhibitor assay methods and showed no effect on MAGL. Tests with other conditions were performed in order to characterize the inhibitory mechanism of FAAH inhibition. N-3-methoxybenzyl-linoleamide displayed significant time-dependent and dose-dependent FAAH inhibitory activity. The mechanism of inhibition was most likely irreversible or slowly reversible. These results suggest the potential application of macamides isolated from Maca as FAAH inhibitors, as they might act on the central nervous system to provide analgesic, anti-inflammatory, or neuroprotective effects, by modulating the release of neurotransmitters.

Macamides and their synthetic analogs: evaluation of in vitro FAAH inhibition.

Maca (Lepidium meyenii), a traditional food crop of the Peruvian Andes is now widely touted as a dietary supplement. Among the various chemical constituents isolated from the plant are a unique series of non-polar, long-chain fatty acid N-benzylamides known as macamides. We have synthesized 11 of the 19 reported macamides and have tested each as potential inhibitors of the human enzyme, fatty acid amide hydrolase (FAAH). The five most potent macamides were FAAH inhibitors (IC50=10-17µM). These amides were derivatives of oleic, linoleic and linolenic acids and benzylamine or 3-methoxybenzylamine. Of the three compounds evaluated in a pre-incubation time study, two macamides were not irreversible inhibitors of FAAH. The third, a carbamate structurally related to macamides, was shown to be an irreversible inhibitor of FAAH (IC50=0.153µM).

Design, synthesis and biological evaluation of a series of thioamides as non-nucleoside reverse transcriptase inhibitors.

A series of thioamides were designed as bio-isosteres to the non-nucleoside reverse transcriptase inhibitor trovirdine by replacement of the thiourea NH groups with methylene groups. Eight thioamides were synthesized and in vitro tested for inhibitory effects on the activity of HIV-1 reverse transcriptase wild and mutant types. Three of the 8-thioamides exhibited enzyme inhibitory activities with IC(50) values below 100 microM. While compound (2) exhibited activity against the mutant strain L100I with IC(50) of 70.1 microM, compound (4) showed activity against the mutant strain K103N with IC(50) of 92.7 microM, and compound (8) with activity against the wild type enzyme with IC(50) of 8.9 microM. Each of the three thioamides could serve as a lead compound for further activity optimization.

Slow-onset, long-duration, alkyl analogues of methylphenidate with enhanced selectivity for the dopamine transporter.

Methylphenidate analogues, in which the carbomethoxy has been replaced by an alkyl group and with different phenyl substituents, have been synthesized and tested in monoamine transporter assays. As predicted from a pharmacophore model, most of the RR/SS diastereomers showed high potency as dopamine reuptake inhibitors. Analogues with a 4-chlorophenyl group and an unbranched initial alkyl atom had consistently enhanced selectivity for the dopamine transporter. The most potent compounds were those with a three- or four-carbon chain. The "inactive" RS/SR diastereomers showed substantial activity when the phenyl substituent was 3,4-dichloro. On a locomotor assay, one compound was found to have a slow onset and a long duration of action. The activity of these compounds provides additional evidence for a conformational/superposition model of methylphenidate with cocaine-like structures. A ketone analogue, obtained by hydrogenating a previously described vinylogous amide, had activity similar to that of methylphenidate.

Vinylogous amide analogs of methylphenidate.

In an effort to produce compounds with longer durations of action, we attempted to synthesize ketone analogs of methylphenidate which, however, appear to be highly unstable due to a highly acidic proton alpha to the ketone and phenyl groups. Nevertheless, vinylogous amide by products have been synthesized and tested for activity at dopamine, norepinephrine, and serotonin transporters. The compounds were found to be weak inhibitors of monoamine reuptake despite rigid three dimensional structures that are quite similar to the global minimum of threo-(R,R)-methylphenidate. The structures were confirmed by X-ray crystallography.

Antimalarial activity of lactucin and lactucopicrin: sesquiterpene lactones isolated from Cichorium intybus L.

Folklore reports from Afghanistan prior to the wars described the use of aqueous root extracts of Cichorium intybus (L.) as a light-sensitive plant remedy for malaria. Preparative isolation and bioassay against HB3 clone of strain Honduras-1 of Plasmodium falciparum identified the previously known light-sensitive sesquiterpene lactones Lactucin and Lactucopicrin to be antimalarial compounds.

Time course of recovery of cytochrome p450 3A function after single doses of grapefruit juice.

BACKGROUND: Components of grapefruit juice may impair the activity of intestinal cytochrome P450 (CYP) 3A enzymes, sometimes resulting in clinically important drug interactions. The time course of recovery from CYP3A inhibition after a single exposure to grapefruit juice is not clearly established. METHODS: Healthy volunteer subjects (N = 25) received a single 6-mg oral dose of the CYP3A substrate midazolam in the control condition without exposure to grapefruit juice. Two days later, midazolam was administered 2 hours after 300 mL of regular-strength grapefruit juice. Subjects were then randomly assigned to 3 different groups, receiving a third midazolam challenge at 26, 50, or 74 hours after exposure to grapefruit juice. The capacity of 6'7'-dihydroxybergamottin and bergamottin to inhibit human CYP3A was studied in vitro using human liver microsomes. RESULTS: The area under the plasma concentration curve (AUC) for midazolam increased by a factor of 1.65 (ratio compared with control) when midazolam was given 2 hours after grapefruit juice. At 26, 50, and 74 hours after grapefruit juice, the AUC ratios (mean AUC value at the indicated time divided by the mean control AUC on day 1) were 1.29, 1.29, and 1.06, respectively. The relationship of time after grapefruit juice exposure versus AUC increase over control indicated a recovery half-life estimated at 23 hours. The midazolam elimination half-life did not change significantly from the control value at any time after grapefruit juice exposure. 6'7'-Dihydroxybergamottin inhibited midazolam alpha-hydroxylation in vitro, with a mean 50% inhibitory concentration of 4.7 micro mol/L; preincubation of microsomes with 6'7'-dihydroxybergamottin greatly reduced the 50% inhibitory concentration to 0.31 micro mol/L, consistent with mechanism-based inhibition. Bergamottin itself had much weaker inhibitory potency compared to 6'7'-dihydroxybergamottin. CONCLUSIONS: A usual single exposure to grapefruit juice appears to impair the enteric, but not the hepatic, component of presystemic extraction of oral midazolam. Recovery is largely complete within 3 days, consistent with enzyme regeneration after mechanism-based inhibition. 6'7'-Dihydroxybergamottin was verified as a potent mechanism-based inhibitor of midazolam alpha-hydroxylation by CYP3A in vitro.