Pharmacokinetic Limitations on Effects of an Alpha7-Nicotinic Receptor Agonist in Schizophrenia: Randomized Trial with an Extended-Release Formulation.

The aim of the trial was to assess whether extending plasma levels of the alpha7-nicotinic acetylcholine receptor (nAChR) agonist 3-(2,4-dimethoxybenzylidene)-anabaseine (DMXB-A) over time enhances its cognitive effects in schizophrenia. Both smoking and non-smoking patients were studied, to determine whether effects differ between these two groups. Forty-three smokers and thirty-seven non-smokers who met DSM-IV criteria for schizophrenia were enrolled in a double-blind, randomized, placebo-controlled 1 month trial. DMXB-A 150 mg was formulated with hypromellose to produce extended release over 4 h and administered four times daily. The primary outcome (the Neurocognitive Composite of the MATRICS Consensus Cognitive Battery) and secondary outcomes (the MATRICS Attention-Vigilance Domain and P50 gating), showed no significant effect. Plasma levels were obtained 2.5 h post administration. In non-smokers, levels were similar to those reached transiently with 75-150 mg DMXB-A immediate-release formulations twice daily, which were earlier shown to be effective doses. However, the extended-release formulation produced no cognitive or clinical effect either in non-smokers or smokers. The 10-fold lower DMXB-A plasma levels in smokers suggest that chronic smoking enhances DMXB-A metabolism. Pro-cognitive effects of DMXB-A may result from transient increases in cell signaling that are limited by receptor tachyphylaxis. Future efforts to improve cognition in schizophrenia by enhancing alpha7 nAChR function may require consideration of these pharmacokinetic limitations.

Application of a sensitive and accurate LC-MS/MS method for determination of dryocrassin ABBA in rat plasma for a bioavailability study.

A sensitive and accurate liquid chromatography-tandem mass spectrometry method was developed and validated for the determination of dryocrassin ABBA, a potential active component isolated from Dryopteris crassirhizoma, in rat plasma. Chromatographic separation was achieved on a Zorbax SB-C18 column (50 × 2.1 mm, 1.8 µm), with elution consisting of eluent (A) 10 mm ammonium acetate in methanol containing 0.1% formic acid and (B) 10 mm ammonium acetate in water containing 0.1% formic acid (A:B = 99:1, v/v) at a flow rate of 0.3 mL/min. Multiple reaction monitoring mode was used to monitor the precursor-product ion transitions of m/z 819.3 → 403.4 for dryocrassin ABBA and m/z 426.2 → 409.2 for internal standard. This assay exhibited a good linearity with a correlation coefficient >0.99 and showed no endogenous interference with the analyte and internal standard. The lower limit of quantification of dryocrassin ABBA was 4 ng/mL in 50 µL of rat plasma. The method was successfully applied in the pharmacokinetic study of dryocrassin ABBA in rats after intravenous (2.35 mg/kg) and oral (23.5 mg/kg) doses of dryocrassin ABBA. The oral bioavailability (F) of dryocrassin ABBA was estimated to be 50.1%. Our study is the first to clarify the pharmacokinetic behaviors of dryocrassin ABBA in animals.

Effects of the α7 nicotinic acetylcholine receptor agonist GTS-21 on the innate immune response in humans.

The vagus nerve can reflexively attenuate the innate immune response via binding of the vagal neurotransmitter acetylcholine (ACh) to the α7 nicotinic ACh receptor (α7nAChR). We recently reported potent anti-inflammatory effects of the α7nAChR agonist GTS-21 in human leukocytes. In the present work, we investigated the anti-inflammatory effects of GTS-21 on the innate immune response during experimental human endotoxemia. We performed a double-blind placebo-controlled pilot study in 14 healthy nonsmoking male volunteers. Subjects received 150 mg GTS-21 (n = 7) or placebo (n = 7) orally three times per day during 3 days before endotoxin administration and on the day of the human endotoxemia experiment. This GTS-21 dosage scheme is the highest reported to be safe in humans. Subsequently, subjects were i.v. administered 2 ng/kg endotoxin (LPS derived from Escherichia coli O:113). Serial blood withdrawals were performed to determine GTS-21 plasma concentrations and inflammatory mediators. Plasma concentrations of GTS-21 and its active metabolite 4-OH-GTS-21 were highly variable between subjects. LPS administration resulted in a transient inflammatory response. There were no differences in the LPS-induced cytokine response between the GTS-21- and placebo-treated groups. However, within the GTS-21-treated group, higher GTS-21 plasma concentrations correlated with lower levels of TNF-α (r = -0.78, P = 0.03), IL-6 (r = -0.76, P = 0.04), and IL-1RA (r = -0.86, P = 0.01), but not IL-10 (r = -0.35, P = 0.25). In conclusion, although higher GTS-21 plasma concentrations significantly correlated with lower cytokine levels, the highest dose tested to be safe in humans did not result in significant differences in inflammatory mediators between the GTS-21- and placebo-treated groups.

Effects of an alpha 7-nicotinic agonist on default network activity in schizophrenia.

BACKGROUND: 3-(2,4-dimethoxybenzylidene)-anabaseine (DMXB-A) is a partial agonist at α7 nicotinic acetylcholine receptors that has been evaluated clinically for treatment of schizophrenia. This study examined the effects of DMXB-A on default network activity as a biomarker for drug effects on pathologic brain function associated with schizophrenia. METHODS: Placebo and two doses of DMXB-A were administered in a random, double-blind crossover design during a Phase 2 study of DMXB-A. Functional magnetic resonance imaging was performed on 16 nonsmoking patients with schizophrenia while they performed a simple eye movement task. Independent component analysis was used to identify the default network component. Default network changes were evaluated in the context of a polymorphism in CHRNA7, the α7-nicotinic acetylcholine receptor subunit gene, which was previously found to be associated with schizophrenia. RESULTS: Compared with placebo, both 150 and 75 mg twice daily DMXB-A altered default network activity, including a reduction in posterior cingulate, inferior parietal cortex, and medial frontal gyrus activity and an increase in precuneus activity. The most robust difference, posterior cingulate activity reduction, was affected by CHRNA7 genotype. CONCLUSIONS: The observed DMXB-A-related changes are consistent with improved default network function in schizophrenia. Pharmacogenetic analysis indicates mediation of the effect through the α7-nicotinic receptor. These results further implicate nicotinic cholinergic dysfunction in the disease and suggest that default network activity may be a useful indicator of biological effects of novel therapeutic agents.

Continuous administration of a selective alpha7 nicotinic partial agonist, DMXBA, improves sensory inhibition without causing tachyphylaxis or receptor upregulation in DBA/2 mice.

Stimulation of nicotinic receptors, specifically the alpha7 subtype, improves sensory inhibition and cognitive function in receptor deficient humans and rodents. However, stimulation with a full agonist, such as nicotine, produces rapid tachyphylaxis of the P20N40-measured sensory inhibition process. 3-(2,4-dimethoxybenzylidine) anabaseine (DMXBA, also GTS-21) selectively activates the alpha7 nicotinic receptor, and in acute administration studies, has been shown to improve deficient sensory inhibition in both humans and rodents with repeated dosing. Unlike nicotine, this partial agonist acted without inducing tachyphylaxis. Here, we assessed the ability of DMXBA to improve sensory inhibition in DBA/2 mice after 7 days of continuous administration via a subcutaneously implanted osmotic minipump. When assessed on day 8, mice receiving saline showed the characteristic deficient sensory inhibition seen with untreated DBA/2 mice. The 25- and 50-mg/ml infusion concentrations of DMXBA, but not the 100-mg/ml, produced significantly improved sensory inhibition in the mice, exclusively through a decrease in test amplitude. No concentration significantly upregulated hippocampal alpha7 receptor levels. DMXBA levels in the brain were higher than plasma at 2 of the 3 concentrations infused. These data suggest that continuous exposure to DMXBA does not significantly affect the underlying responsiveness of the sensory inhibition pathway to this partial agonist, nor cause receptor upregulation, at these relatively low brain concentrations. The ability of DMXBA to maintain its effectiveness during constant administration conditions may be due to an ability to activate alpha7 receptors at low concentrations, and consequently low fractional occupancy of the five possible binding sites on this homomeric receptor.

Functional magnetic resonance imaging of effects of a nicotinic agonist in schizophrenia.

3-(2,4-Dimethoxybenzylidene)-anabaseine (DMXB-A) is a partial agonist at alpha7-nicotinic acetylcholine receptors and is now in early clinical development for treatment of deficits in neurocognition and sensory gating in schizophrenia. During its initial phase 2 test, functional magnetic resonance imaging (fMRI) studies were conducted to determine whether the drug had its intended effect on hippocampal inhibitory interneurons. Increased hemodynamic activity in the hippocampus in schizophrenia is found during many tasks, including smooth pursuit eye movements, and may reflect inhibitory dysfunction. Placebo and two doses of drug were administered in a random, double-blind crossover design. After the morning drug/placebo ingestion, subjects underwent fMRI while performing a smooth pursuit eye movement task. Data were analyzed from 16 nonsmoking patients, including 7 women and 9 men. The 150-mg dose of DMXB-A, compared with placebo, diminished the activity of the hippocampus during pursuit eye movements, but the 75-mg dose was ineffective. The effect at the 150-mg dose was negatively correlated with plasma drug levels. The findings are consistent with the previously established function of alpha7-nicotinic receptors on inhibitory interneurons in the hippocampus and with genetic evidence for deficits in these receptors in schizophrenia. Imaging of drug response is useful in planning future clinical tests of this compound and other nicotinic agonists for schizophrenia.

A phase I pharmacokinetic study of HMN-214, a novel oral stilbene derivative with polo-like kinase-1-interacting properties, in patients with advanced solid tumors.

PURPOSE: HMN-214 is an oral prodrug of HMN-176, a stilbene derivative that interferes with the subcellular spatial location of polo-like kinase-1, a serine/threonine kinase that regulates critical mitotic events. We conducted a dose escalation study of HMN-214 in patients with advanced cancer to assess the safety profile and pharmacokinetics of HMN-214 and to establish the maximum tolerated dose. EXPERIMENTAL DESIGN: Thirty-three patients were enrolled onto four dosing cohorts of HMN-214 from 3 to 9.9 mg/m2/d using a continuous 21-day dosing schedule every 28 days, with pharmacokinetic sampling during cycle 1. RESULTS: A severe myalgia/bone pain syndrome and hyperglycemia were dose-limiting toxicities at 9.9 mg/m2/d. A dose reduction and separate enrollment by pretreatment status (lightly versus heavily pretreated) was undertaken, with one dose-limiting toxicity (grade 3 bone pain) at 8 mg/m2/d. The maximum tolerated dose was defined as 8 mg/m2/d for both treatment cohorts. Dose-proportional increases were observed in AUC but not Cmax. There was no accumulation of HMN-176, the metabolite of HMN-214, with repeated dosing. Seven of 29 patients had stable disease as best tumor response, including 6-month stable disease in a heavily pretreated breast cancer patient. A transient decline in carcinoembryonic antigen in a patient with colorectal cancer was noted. CONCLUSIONS: The maximum tolerated dose and recommended phase II dose of HMN-214 when administered on this schedule was 8 mg/m2/d regardless of pretreatment status. Further development of HMN-214 will focus on patient populations for which high expression of polo-like kinase-1 is seen (i.e., prostate and pancreatic cancer patients).

Prediction of plasma concentration of GTS-21 in hairless rats following monolithic transdermal delivery.

The transdermal therapeutic systems (TTS) usually achieve constant plasma concentration for an extended period of time. This is because a sufficient drug stored in the device can keep the constant concentration on the surface of the stratum corneum during the system application. When the drug molecules are not enough to provide the constant surface concentration, the rate of drug penetration decreases with time because of decreased supply of the drug molecules from the delivery device. This paper has proposed an empirical simple approach to predict the plasma concentration for such a TTS. A novel compound, GTS-21, for Alzheimers' disease currently under development was used as a model drug. In vivo and in vitro experiments were carried out in hairless rats. The in vivo plasma concentration-time profile in hairless rats following the application of TTS well agreed with the predicted profile based on the skin pharmacokinetic model together with the model parameters determined from the in vitro experiment.

Intragastric DMXB-A, an alpha7 nicotinic agonist, improves deficient sensory inhibition in DBA/2 mice.

BACKGROUND: Abnormal sensory inhibition is observed in the majority of schizophrenic patients. DBA/2 mice spontaneously exhibit a similar deficit in sensory inhibition and thus provide a model for drug development targeted to this physiologic abnormality. The impaired sensory inhibition is characterized by diminished response of the hippocampal evoked potential to the second of closely paired auditory stimuli (500-m/sec interstimulus interval). Subnormal levels of hippocampal alpha7 nicotinic cholinergic receptors are associated with the deficient sensory inhibition in both DBA/2 mice and people with schizophrenia. METHODS: Our study examined the inhibition of the P20-N40 auditory evoked potential in DBA/2 mice after intragastric administration of DMXB-A (3-2,4-dimethoxybenzylidine anabaseine), an alpha7 nicotinic receptor partial agonist. After presentation of auditory stimuli, electroencephalographic responses were recorded and measured to monitor the effects of the DMXB-A, alone and in combination with selective nicotinic antagonists. RESULTS: Gastric administration of DMXB-A (10 mg/kg) improved sensory inhibition in DBA/2 mice. This improvement was blocked by alpha-bungarotoxin, but not mecamylamine, indicating that DMXB-A exerts its effects through the alpha7 nicotinic receptor. CONCLUSIONS: Intragastrically administered DMXB-A improves deficient sensory inhibition in DBA/2 mice through stimulation of alpha7 nicotinic receptors. These studies agree with results from previous studies with subcutaneously administered DMXB-A.

First-pass of GTS-21 on canine gut wall and liver determined by portal-systemic concentration difference.

To clarify the cause of the canine individual variability of plasma concentration after oral administration of GTS-21 [(E)-3-(2,4-dimethoxybenzylidene)-3,4,5,6-tetra-hydro-2,3'-bipyridine dihydrochloride], we evaluated the absorption ratio (F(A)), intestinal availability (F(G)), and hepatic availability (F(H)). The bioavailability (F) was evaluated from the ratio of the area under the plasma concentration versus time curves after oral and intravenous administration. Three isoflurane anaesthetised dogs were fitted with an electromagnetic flow probe attached to the portal vein and cannulated through the portal and the femoral veins. After intraduodenal administration of GTS-21, both plasma concentrations were determined simultaneously. F(A) x F(G) was calculated from the portal-systemic concentration difference taking into consideration the blood-plasma partition ratio. F(A) was calculated from the residual drug contents of the small intestine. F(H) was calculated by dividing F by F(A) x F(G). The F values were 0.072, 0.021, and 0.037, indicating an individual variability of ca. threefold. The F(A) values were close to 1, and the F(G) values ranged from 0.449 to 0.461. Accordingly, the F(H) values were estimated at 0.170, 0.047, and 0.083. GTS-21 was completely absorbed but lost by first-pass effects of passage through the gut wall and liver. The first-pass effect of liver is larger than that of the gut wall, and dominates the individual variability in plasma concentration.

Pharmacokinetics and urinary excretion of DMXBA (GTS-21), a compound enhancing cognition.

DMXBA (3-(2,4-dimethoxybenzylidene)-anabaseine, also known as GTS-21) is currently being tested as a possible pharmacological treatment of cognitive dysfunction in Alzheimer's disease. In this study, plasma and brain pharmacokinetics as well as urinary excretion of this compound have been evaluated in adult rats. DMXBA concentrations were determined by HPLC. Following a 5 mg kg-1 iv dose, DMXBA plasma concentration declined bi-exponentially with mean (+/- SE) absorption and elimination half-lives of 0.71 +/- 0.28 and 3.71 +/- 1.12 h, respectively. The apparent steady state volume of distribution was 2150 +/- 433 mL kg-1, total body clearance was 1480 +/- 273 mL h-1 kg-1, and AUC0-infinity was 3790 +/- 630 ng h mL-1. Orally administered DMXBA was rapidly absorbed. After oral administration of 10 mg kg-1, a peak plasma concentration of 1010 +/- 212 ng mL-1 was observed at 10 min after dosing. Elimination half-life was 1.740 +/- 0.34 h, and AUC0-infinity was 1440 +/- 358 ng h mL-1. DMXBA peak brain concentration after oral administration was 664 +/- 103 ng g-1 tissue, with an essentially constant brain-plasma concentration ratio of 2.61 +/- 0.34, which indicates that the drug readily passes across the blood-brain barrier. Serum protein binding was 80.3 +/- 1.1%. Apparent oral bioavailability was 19%. Renal clearance (21.8 mL h-1 kg-1) was less than 2% of the total clearance (1480 +/- 273 mL h-1 kg-1); urinary excretion of unchanged DMXBA over a 96 h period accounted for only 0.28 +/- 0.03% of the total orally administered dose. Our data indicates that DMXBA oral bioavailability is primarily limited by hepatic metabolism.

Effect of physiological fluids on radical intensity of sodium ascorbate and sodium 5,6-benzylidene-L-ascorbate.

The effect of various physiological fluids on the radical intensity of sodium ascorbate and sodium 5,6-benzylidene-L-ascorbate (SBA) was investigated using ESR spectroscopy. Blood from various animals did not significantly affect the radical intensity of both ascorbates, whereas the corresponding plasma fractions significantly enhanced the radical intensity. This suggests that some populations of blood cells might modify the interaction between plasma components and ascorbates. Saliva contained labile substance(s) which effectively reduced the ascorbate radical intensity. HPLC demonstrated the presence of endogenous ascorbate in rat liver and brain homogenates. When sodium ascorbate or SBA was incubated with any of these homogenates, their radical intensity was synergistically enhanced, but abruptly declined without any apparent ascorbate degradation. Incubation with homogenates elevated the radical intensity of SBA up to the level significantly higher than that of sodium ascorbate. The present data suggest that antitumor action of SBA might be mediated via the accelerated production of ascorbate radical in the target organ.

Simultaneous determination of GTS-21 and its metabolite in rat plasma by high-performance liquid chromatography using solid-phase extraction.

It has been suggested that GTS-21 can improve the learning deficits and inhibit the neuro-degeneration in patients with Alzheimer's disease. This paper describes a reversed-phase high-performance liquid chromatographic assay with visible detection at 405 nm for determination of GTS-21 and its metabolite, 4-hydroxy-GTS-21 in rat plasma. The method uses solid-phase extraction with a Bond Elut C18 column. A quantitation limit of 1.0 ng/ml was achieved using 0.5 ml of rat plasma. In the validation study, the coefficients of variation and the relative errors of each compound were less than 10%. Also freeze-thaw and storage stability were confirmed. This method has proved to be applicable to the pharmacokinetic study of GTS-21 in rats.

Interaction of sulindac and metabolite with human serum albumin.

The binding of the newly developed nonsteroidal anti-inflammatory agent sulindac and its principal active metabolite, sunlindac sulfide, to human serum albumin was investigated. With the methods of dialysis, fluorescence quenching, and difference spectrophotometry, it was found that both agents were extensively bound to albumin. The binding affinity of the metabolite was considerably higher than that of sunlindac and this effect may be related to its prolonged plasma half-life versus the parent drug. Sulindac binding was albumin concentration dependent, which gave rise to an unfamilar Scatchard analysis of the dialysis data.