Major depressive disorder afflicts ~16 percent of the world population at some point in their lives. Despite a number of available monoaminergic-based antidepressants, most patients require many weeks, if not months, to respond to these treatments, and many patients never attain sustained remission of their symptoms. The non-competitive glutamatergic N-methyl-D-aspartate receptor (NMDAR) antagonist, (R,S)-ketamine (ketamine), exerts rapid and sustained antidepressant effects following a single dose in depressed patients. Here we show that the metabolism of ketamine to (2S,6S;2R,6R)-hydroxynorketamine (HNK) is essential for its antidepressant effects, and that the (2R,6R)-HNK enantiomer exerts behavioural, electroencephalographic, electrophysiological and cellular antidepressant actions in vivo. Notably, we demonstrate that these antidepressant actions are NMDAR inhibition-independent but they involve early and sustained α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA) receptor activation. We also establish that (2R,6R)-HNK lacks ketamine-related side-effects. Our results indicate a novel mechanism underlying ketamine’s unique antidepressant properties, which involves the required activity of a distinct metabolite and is independent of NMDAR inhibition. These findings have relevance for the development of next generation, rapid-acting antidepressants.

Background Ketamine has rapid antidepressant effects lasting as long as 1 week in patients with major depressive disorder (MDD) and bipolar depression (BD). Ketamine is extensively metabolized. This study examined the relationship between ketamine metabolites and response, diagnosis, and psychotomimetic symptoms in MDD and BD patients. Methods Following a 40-minute ketamine infusion (.5 mg/kg), plasma samples were collected at 40, 80, 110, and 230 minutes and day 1 postinfusion in 67 patients currently experiencing a major depressive episode (MDD, n = 45; BD, n = 22). Concentrations of ketamine, norketamine (NK), dehydronorketamine (DHNK), six hydroxynorketamine metabolites (HNK), and hydroxyketamine (HK) were measured. Plasma concentrations were analyzed by diagnostic group and correlated with patients’ depressive, psychotic, and dissociative symptoms. The relationship between cytochrome P450 gene polymorphisms and metabolites, response, and diagnosis was also examined. Results Ketamine, NK, DHNK, four of six HNKs, and HK were present during the first 230 minutes postinfusion. Patients with BD had higher plasma concentrations of DHNK, (2S,6S;2R,6R)-HNK, (2S,6R;2R,6S)-HNK, and (2S,5S;2R,5R)-HNK than patients with MDD, who, in turn, had higher concentrations of (2S,6S;2R,6R)-HK. Higher (2S,5S;2R,5R)-HNK concentrations were associated with nonresponse to ketamine in BD patients. Dehydronorketamine, HNK4c, and HNK4f levels were significantly negatively correlated with psychotic and dissociative symptoms at 40 minutes. No relationship was found between cytochrome P450 genes and any of the parameters examined. Conclusions A diagnostic difference was observed in the metabolism and disposition of ketamine. Concentrations of (2S,5S;2R,5R)-HNK were related to nonresponse to ketamine in BD. Some hydroxylated metabolites of ketamine correlated with psychotic and dissociative symptoms.

The N-methyl-D-aspartate antagonist ketamine has rapid antidepressant effects in patients with treatment-resistant major depression (TRD); these effects have been reported to last for 1 week in some patients. However, the extent and duration of this antidepressant effect over longer periods has not been well characterized under controlled conditions. Riluzole, a glutamatergic modulator with antidepressant and synaptic plasticity-enhancing effects, could conceivably be used to promote the antidepressant effects of ketamine. This study sought to determine the extent and time course of antidepressant improvement to a single-ketamine infusion over 4 weeks, comparing the addition of riluzole vs placebo after the infusion. Forty-two subjects (18-65) with TRD and a Montgomery-Asberg Depression Rating Scale (MADRS) score of ≥ 22 received a single intravenous infusion of ketamine (0.5 mg/kg). Four to six hours post-infusion, subjects were randomized to double-blind treatment with either riluzole (100-200 mg/day; n=21) or placebo (n=21) for 4 weeks. Depressive symptoms were rated daily. A significant improvement (P<0.001) in MADRS scores from baseline was found. The effect size of improvement with ketamine was initially large and remained moderate throughout the 28-day trial. Overall, 27% of ketamine responders had not relapsed by 4 weeks following a single ketamine infusion. The average time to relapse was 13.2 days (SE=2.2). However, the difference between the riluzole and placebo treatment groups was not significant, suggesting that the combination of riluzole with ketamine treatment did not significantly alter the course of antidepressant response to ketamine alone.

The effect of the (R,S)-ketamine metabolites (R,S)-norketamine, (R,S)-dehydronorketamine, (2S,6S)-hydroxynorketamine and (2R,6R)- hydroxynorketamine on the activity of α7 and α3β4 neuronal nicotinic acetylcholine receptors was investigated using patch-clamp techniques. The data indicated that (R,S)-dehydronorketamine inhibited acetylcholine-evoked currents in α7-nicotinic acetylcholine receptor, IC50 = 55 ± 6 nM, and that (2S,6S)-hydroxynorketamine, (2R,6R)-hydroxynorketamine and (R,S)-norketamine also inhibited α7-nicotinic acetylcholine receptor function at concentrations ≤1μM, while (R,S)-ketamine was inactive at these concentrations. The inhibitory effect of (R,S)-dehydronorketamine was voltage-independent and the compound did not competitively displace selective α7-nicotinic acetylcholine receptor ligands [125I]-α-bungarotoxin and [3H]-epibatidine indicating that (R,S)-dehydronorketamine is a negative allosteric modulator of the α7-nicotinic acetylcholine receptor. (R,S)-Ketamine and (R,S)-norketamine inhibited (S)-nicotine-induced whole-cell currents in cells expressing α3β4-nicotinic acetylcholine receptor, IC50 3.1 and 9.1μM, respectively, while (R,S)-dehydronorketamine, (2S,6S)-hydroxynorketamine and (2R,6R)-hydroxynorketamine were weak inhibitors, IC50 >100μM. The binding affinities of (R,S)-dehydronorketamine, (2S,6S)-hydroxynorketamine and (2R,6R)-hydroxynorketamine at the NMDA receptor were also determined using rat brain membranes and the selective NMDA receptor antagonist [3H]-MK-801. The calculated Ki values were 38.95 μM for (S)-dehydronorketamine, 21.19 μM for (2S,6S)-hydroxynorketamine and > 100 μM for (2R,6R)-hydroxynorketamine. The results suggest that the inhibitory activity of ketamine metabolites at the α7-nicotinic acetylcholine receptor may contribute to the clinical effect of the drug.

WHAT IS ALREADY KNOWN ABOUT THIS SUBJECT • (R,S)‐ketamine is a phencyclidine derivative that was initially developed as an anaesthetic agent and which is currently being studied in the treatment of pain and depression. After administration, the drug is extensively N‐demethylated to (R,S)‐norketamine. The pharmacokinetics of ketamine and norketamine have been extensively studied in volunteers and patients after the administration of anaesthetic and sub‐anaesthetic doses. However, ketamine and norketamine are extensively transformed into a series of diastereomeric hydroxyketamines and hydroxynorketamines and (R,S)‐dehydronorketamine metabolites. The plasma kinetics of these metabolites have not been elucidated. WHAT THIS STUDY ADDS • The current study expands the characterization of the disposition kinetics of (R,S)‐ketamine and (R,S)‐norketamine and presents a population pharmacokinetic analysis of (R)‐ketamine, (S)‐ketamine, (R)‐norketamine, (S)‐norketamine, (R)‐dehydronorketamine, (S)‐ dehydronorketamine and (2S,6S;2R,6R)‐hydroxynorketamine and the serum concentration–time profiles of multiple ketamine metabolites observed in the plasma of patients after a single 40 min infusion of a sub‐anaesthetic dose of the drug. The data demonstrate that while norketamine is an initial metabolite, it is not the major circulating metabolite and suggest that the determination of the downstream metabolites of ketamine may play a role in the pharmacological effects of the drug. AIM To construct a population pharmacokinetic (popPK) model for ketamine (Ket), norketamine (norKet), dehydronorketamine (DHNK), hydroxynorketamine (2S,6S;2R,6R)‐HNK) and hydroxyketamine (HK) in patients with treatment‐resistant bipolar depression. METHOD Plasma samples were collected at 40, 80, 110, 230 min on day 1, 2 and 3 in nine patients following a 40 min infusion of (R,S)‐Ket (0.5 mg kg−1) and analyzed for Ket, norKet and DHNK enantiomers and (2S,6S;2R,6R)‐HNK, (2S,6S;2R,6R)‐HK and (2S,6R;2R,6S)‐HK. A compartmental popPK model was constructed that included all quantified analytes, and unknown parameters were estimated with an iterative two‐stage algorithm in ADAPT5. RESULTS Ket, norKet, DHNK and (2S,6S;2R,6R)‐HNK were present during the first 230 min post infusion and significant concentrations (>5 ng ml−1) were observed on day 1. Plasma concentrations of (2S,6S;2R,6R)‐HK and (2S,6R;2R,6S)‐HK were below the limit of quantification. The average (S) : (R) plasma concentrations for Ket and DHNK were <1.0 while no significant enantioselectivity was observed for norKet. There were large inter‐patient variations in terminal half‐lives and relative metabolite concentrations; at 230 min (R,S)‐DHNK was the major metabolite in four out of nine patients, (R,S)‐norKet in three out of nine patients and (2S,6S;2R,6R)‐HNK in two out of nine patients. The final PK model included three compartments for (R,S)‐Ket, two compartments for (R,S)‐norKet and single compartments for DHNK and HNK. All PK profiles were well described, and parameters for (R,S)‐Ket and (R,S)‐n...

Stereoisomers of fenoterol and six fenoterol derivatives have been synthesized and their binding affinities for the beta2 adrenergic receptor (Kibeta2-AR), the subtype selectivity relative to the beta1-AR (Kibeta1-AR/Kibeta2-AR) and their functional activities were determined. Of the 26 compounds synthesized in the study, submicromolar binding affinities were observed for (R,R)-fenoterol, the (R,R)-isomer of the p-methoxy, and (R,R)- and (R,S)-isomers of 1-naphthyl derivatives and all of these compounds were active at submicromolar concentrations in cardiomyocyte contractility tests. The Kibeta1-AR/Kibeta2-AR ratios were >40 for (R,R)-fenoterol and the (R,R)-p-methoxy and (R,S)-1-naphthyl derivatives and 14 for the (R,R)-1-napthyl derivative. The binding data was analyzed using comparative molecular field analysis (CoMFA), and the resulting model indicated that the fenoterol derivatives interacted with two separate binding sites and one steric restricted site on the pseudo-receptor and that the chirality of the second stereogenic center affected Kibeta2 and subtype selectivity.

The objective was to determine the cytochrome P450s (CYPs) responsible for the stereoselective and regiospecific hydroxylation of ketamine ((R,S)-Ket) to diastereomeric hydroxyketamines, (2S,6S;2R,6R)-HK (5a) and (2S,6R;2R,6S)-HK (5b) and norketamine ((R,S)-norKet) to hydroxynorketamines, (2S,6S;2R,6R)-HNK (4a), (2S,6R;2R,6S)-HNK (4b), (2S,5S;2R,5R)-HNK (4c), (2S,4S;2R,4R)-HNK (4d), (2S,4R;2R,4S)-HNK (4e), (2S,5R;2R,5S)-HNK (4f).The enantiomers of Ket and norKet were incubated with HLMs and expressed CYPs. Metabolites were identified and quantified using LC/MS/MS and apparent kinetic constants estimated using single-site Michaelis-Menten, Hill or substrate inhibition equation.5a was predominantly formed from (S)-Ket by CYP2A6 and N-demethylated to 4a by CYP2B6. 5b was formed from (R)- and (S)-Ket by CYP3A4/3A5 and N-demethylated to 4b by multiple enzymes. norKet incubation produced 4a, 4c and 4f and minor amounts of 4d and 4e. CYP2A6 and CYP2B6 were the major enzymes responsible for the formation of 4a, 4d and 4f, and CYP3A4/3A5 for the formation of 4e. The 4b metabolite was not detected in the norKet incubates.5a and 4b were detected in plasma samples from patients receiving (R,S)-Ket, indicating that 5a and 5b are significant Ket metabolites. Large variations in HNK concentrations were observed suggesting that pharmacogenetics and/or metabolic drug interactions may play a role in therapeutic response.

Depsipeptide (FR901228) is a bicyclic peptide isolated from Chromobacterium violaceum that has demonstrated potent in vitro cytotoxic activity against human tumor cell lines and in vivo efficacy against human tumor xenografts. The primary mechanism of action is through inhibition of histone deacetylase. Initial development was halted due to significant cardiac toxicity. Subsequent studies performed at the National Cancer Institute demonstrated administration without cardiotoxicity was possible by varying the schedule of administration. A phase I trial was designed to determine the maximum tolerated dose and toxicity profile when administered as a 4-hour infusion weekly x 3 with one week rest. 33 Patients with advanced, incurable cancers were enrolled into this trial and treated with doses of Depsipeptide ranging from 1 mg/m2 to 17.7 mg/m2. At doses above 5 mg/m2, we observed common symptoms of nausea, vomiting, fatigue, and anorexia. Subtle changes in ECGs were seen in several patients. However, no cardiac enzyme abnormalities or reduction in ejection fraction were observed. The MTD was defined as 13.3 mg/m2 with dose limiting toxicities being grade 3 thrombocytopenia and fatigue. Depsipeptide can be safely administered when given as a 4-hour infusion and further clinical trials are warranted.