Charles R. Ashby

Correlation between plasma levels of glutamate, alanine and serine with severity of depression

The goal of this study was to evaluate the utility of using plasma levels of amino acids as an indicator of the severity of depression. The samples were collected from 23 depressed patients receiving antidepressant medication, and were compared to 31 healthy subjects. The plasma levels of amino acids were determined using HPLC with fluorometric detection. The severity of depression was evaluated using the Hamilton Depression Rating Scale (HAM-D) scores. Plasma levels of glutamate, glutamine, glycine and taurine were significantly increased in the depressed patients compared to the controls. Statistical analysis indicated a positive correlation between glutamate and alanine levels and HAM-D scores and a negative correlation of L-serine with HAM-D scores. The results indicate that plasma level of glutamate, alanine and L-serine could reflect the severity of depression rather than glutamine, glycine and taurine.

Nilotinib (AMN107, Tasigna®) reverses multidrug resistance by inhibiting the activity of the ABCB1/Pgp and ABCG2/BCRP/MXR transporters

Nilotinib, a BCR-Abl tyrosine kinase inhibitor (TKI), was developed to surmount resistance or intolerance to imatinib in patients with Philadelphia positive chronic myelogenous leukemia. Recently, it was shown that several human multidrug resistance (MDR) ATP-binding cassette (ABC) proteins could be modulated by specific TKIs. MDR can produce cancer chemotherapy failure, typically due to overexpression of ABC transporters, which are involved in the extrusion of therapeutic drugs. Here, we report for the first time that nilotinib potentiates the cytotoxicity of widely used therapeutic substrates of ABCG2, such as mitoxantrone, doxorubicin, and ABCB1 substrates including colchicine, vincristine, and paclitaxel. Nilotinib also significantly enhances the accumulation of paclitaxel in cell lines overexpressing ABCB1. Similarly, nilotinib significantly increases the intracellular accumulation of mitoxantrone in cells transfected with ABCG2. Furthermore, nilotinib produces a concentration-dependent inhibition of the ABCG2-mediated transport of methotrexate (MTX), as well as E(2)17betaG a physiological substrate of ABCG2. Uptake studies in membrane vesicles overexpressing ABCG2 have indicated that nilotinib inhibits ABCG2 similar to other established TKIs as well as fumitremorgin C. Nilotinib is a potent competitive inhibitor of MTX transport by ABCG2 with a K(i) value of 0.69+/-0.083 microM as demonstrated by kinetic analysis of nilotinib. Overall, our results indicate that nilotinib could reverse ABCB1- and ABCG2-mediated MDR by blocking the efflux function of these transporters. These findings may be used to guide the design of present and future clinical trials with nilotinib, elucidating potential pharmacokinetic interactions. Also, these findings may be useful in clinical practice for cancer combination therapy with nilotinib.

The selective dopamine D3 receptor antagonist SB-277011-A attenuates ethanol consumption in ethanol preferring (P) and non-preferring (NP) rats.

The mesolimbic dopamine (DA) system plays an important role in mediating addiction to alcohol and other drugs of abuse. Recent evidence points toward the role of the DA D3 receptor (D3R) in drug-induced reward, drug-taking, as well as cue-, drug-, and stress-triggered relapse to drug-seeking behavior. Accordingly, the present study examined the effects of acute selective antagonism of the D3R on ethanol consumption in alcohol Preferring (P) and Non-Preferring (NP) rats. We employed the two-bottle choice paradigm to monitor ethanol consumption in these rats before and after treatment with 3, 10, and 30 mg/kg (i.p.) of the selective D3R antagonist SB-277011-A. Results indicated a significant attenuation in ethanol preference, intake and lick responses in P rats treated with 10 and 30 mg/kg SB-277011-A. A similar, though not as robust effect was observed in ethanol consumption in the NP rats when treated with 30 mg/kg SB-277011-A. Finally, the acute administration of SB-277011-A did not produce extrapyramidal side effects, as indicated by stable lick response-volume ratios and lick response time distributions. These results further support the notion that the D3R is important in mediating the addictive properties of alcohol and suggest that selective blockade of the D3R may constitute a new and useful target for prospective pharmacotherapeutic approaches to alcoholism.

Cepharanthine is a potent reversal agent for MRP7(ABCC10)-mediated multidrug resistance.

Multidrug resistance protein 7 (MRP7; ABCC10) is an ABC transporter that confers resistance to anticancer agents such as the taxanes. We previously reported that several inhibitors of P-gp and MRP1 were able to inhibit the in vitro transport of E(2)17betaG by MRP7 in membrane vesicles transport assays. However, compounds that are able to reverse MRP7-mediated cellular resistance have not been identified. In this study, we examined the effects of cepharanthine (6,12-dimethoxy-2,2-dimethyl-6,7-[methylenebis(oxy)]oxyacanthan), an herbal extract isolated from Stephania cepharantha Hayata, to reverse paclitaxel resistance in MRP7-transfected HEK293 cells. Cepharanthine, at 2microM, completely reversed paclitaxel resistance in MRP7-transfected cells. In contrast, the effect of cepharanthine on the parental transfected cells was significantly less than that on the MRP7-transfected cells. In addition, cepharanthine significantly increased the accumulation of paclitaxel in MRP7-transfected cells almost to the level of control cells in the absence of cepharanthine. The efflux of paclitaxel from MRP7-transfected cells was also significantly inhibited by cepharanthine. The ability of cepharanthine to inhibit MRP7 was analyzed in membrane vesicle assays using E(2)17betaG, an established substrate of MRP7, as a probe. E(2)17betaG transport was competitively inhibited by cepharanthine with a K(i) value of 4.86microM. These findings indicate that cepharanthine reverses MRP7-mediated resistance to paclitaxel in a competitive manner.

The effect of the acute administration of various selective 5-HT receptor antagonists on focal hippocampal seizures in freely-moving rats

In this study, we assessed the effects of the acute administration of various 5-HT receptor antagonists on hippocampal partial seizures generated by low-frequency electrical stimulation in male Wistar rats. The seizure threshold and severity were determined by measuring the pulse number threshold and primary and secondary afterdischarges, respectively, and the latency of secondary discharge was also determined. The administration of either the selective 5-HT(1A) receptor antagonist N-[2-[4-(2-methoxyphenyl)-1-piperazineyl]ethyl]-N-(pyridinyl)-c yclohe xanecarboximimde 3 HCl (WAY 100635, 0.1-1 mg/kg i.p.), the selective 5-HT(3) receptor antagonist granisetron (0.3-3 mg/kg i.p.), the selective 5-HT(2A) receptor antagonist R-(+)-a-(2, 3-dimethoxyphenyl)-1-[2-(4-fluorophenyl) ethyl]-4-piperidine-methanol (MDL 100907, 0.3-3 mg/kg i.p.) or the 5-HT(2B,C) receptor antagonist antagonist N-(1-methyl-5-indolyl)-N-(3-pyridyl) urea HCl (SKB 200646A, 5-50 mg/kg i.p.) did not alter the pulse number threshold compared to vehicle-treated animals. However, the acute administration of WAY 100635 (0.3 mg/kg) and M100907 (1 mg/kg) significantly increased, whereas granisetron (1 mg/kg) decreased, the primary afterdischarge duration compared to vehicle-treated animals. The latency of secondary after discharge was significantly decreased by WAY 100635 (1 mg/kg) and granisetron (3 mg/kg) compared to vehicle-treated animals. These results suggest that in this model, the antagonism of 5-HT(1A), 5-HT(2A), 5-HT(3) or 5-HT(2B,C) receptors do not lower or raise seizure threshold. However, the antagonism of 5-HT(1A) receptors may increase or augment seizure severity.

The Effect of the Repeated Administration of the Compound 3,4-Methylenedioxymethamphetamineon the Response of Rats to the5-HT2A,C Receptor Agonist (±)-1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane (DOI)

In this study, we examined the effect of the 5-HT2A,C receptor agonist (±)-1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane (DOI) on locomotor activity and the head twitch response in rats by the repeated (twice daily for 4 days) subcutaneous administration of 0.9% saline (1 ml/kg) or (±)-3,4-Methylenedioxymethamphetamine (MDMA) (20 mg/kg), a compound that selectively destroys serotonergic nerve terminals. There was no difference between the MDMA- and saline-treated rats regarding the (±)-DOI-induced (0.625, 1.25 or 2.5 mg/kg, i.p.) increase and decrease in the head twitch response and horizontal locomotor activity, respectively. Overall, repeated MDMA does not alter the behavioral response of rats to (±)-DOI.

Effect of the repeated administration of (+/-)-3,4-methylenedioxymethamphetamine on the behavioral response of rats to the 5-HT1A receptor agonist (+/-)-8-hydroxy-(di-n-propylamino)tetralin.

In this study, we examined the effect of the subcutaneous administration (twice daily for 4 consecutive days) of 3,4-methylenedioxymethamphetamine (MDMA, 20 mg/kg) or saline (1 ml/kg) on the response of rats to the behavioral effects of the 5-HT1A agonist (±)-8-hydroxy-(di-n-propylamino)tetralin (8-OH-DPAT) 30 days after the last saline or MDMA treatment. The reciprocal forepaw treading elicited by the 0.25-mg/kg dose of 8-OH-DPAT was significantly lower in animals pretreated with MDMA compared to vehicle-treated animals. However, there were no significant differences between the MDMA- and vehicle-treated animals in flat body posture, locomotor activity and rectal temperature measured after the systemic administration of 8-OH-DPAT. Overall, our results suggest that the depletion of 5-HT levels by the repeated administration of MDMA does not produce a supersensitivity of central 5-HT1A receptors in the rat as determined via our approach.

The synthesis and SAR study of phenylalanine-derived (Z)-5-arylmethylidene rhodanines as anti-methicillin-resistant Staphylococcus aureus (MRSA) compounds

A focused library of rhodanine compounds containing novel substituents at the C5-position was synthesized and tested in vitro against a panel of clinically relevant MRSA strains. The present SAR study was based on our lead compound 1 (MIC=1.95 μg/mL), with a focus on identifying optimal C5-arylidene substituents. In order to obtain this objective, we condensed several unique aromatic aldehydes with phenylalanine-derived rhodanine intermediates to obtain C5-substituted target rhodanine compounds for evaluation as anti-MRSA compounds. These efforts produced three compounds with significant efficacy: 23, 32 and 44, with MIC values ranging from 0.98 to 1.95 μg/mL against all tested MRSA strains as compared to the reference antibiotics penicillin G (MIC=15.60-250.0 μg/mL) and ciprofloxacin (MIC=7.80-62.50 μg/mL) and comparable to that of vancomycin (MIC=0.48 μg/mL). In addition, compounds 24, 28, 37, 41, 46 and 48 (MIC=1.95-3.90 μg/mL) were efficacious against all MRSA strains. The majority of the synthesized compounds had bactericidal activity at concentrations only two to fourfold higher than their MIC. Overall, the results suggest that compounds 23, 32 and 44 may be of potential use in the treatment of MRSA infections.

The dopamine D3 receptor antagonists PG01037, NGB2904, SB277011A, and U99194 reverse ABCG2 transporter-mediated drug resistance in cancer cell lines

The ATP - binding cassette (ABC) family G2 (ABCG2) transporters are known to produce multidrug resistance (MDR) in cancer, thereby limiting the clinical response to chemotherapy. Molecular modeling data indicated that certain dopamine (DA) D3 receptor antagonists had a significant binding affinity for ABCG2 transporter. Therefore, in this inxa0vitro study, we determined the effect of the D3 receptor antagonists PG01037, NGB2904, SB277011A, and U99194 on MDR resulting from the overexpression of ABCG2 transporters. The D3 receptor antagonists, at concentrations >100xa0μM, did not significantly affect the viability of H460-MX20, S1M1-80, A549-MX10 or wild type ABCG2 overexpressing (HEK293-R2) cells. However, at concentrations ranging from 0.01 to 10xa0μM, the D3 receptor antagonists PG01037, NGB2904, SB-277011A, and U99194 significantly increased the efficacy of the anticancer drugs mitoxantrone and doxorubicin in ABCG2-overexpressing MDR cells. Efflux studies indicated that both PG01037 and NGB2904, at a concentration of 5xa0μM, significantly decreased the efflux of rhodamine 123 from H460-MX20 cells. Interestingly, 5xa0μM of PG01037 or NGB2904 significantly decreased the expression levels of the ABCG2 protein, suggesting that these compounds inhibit both the function and expression of ABCG2 transporters at non-toxic concentrations.

The effects of dentate granule cell destruction on behavioral activity and Fos protein expression induced by systemic MDMA in rats.

In this study, we examined the effect of the s.c. administration of (+/-) 3,4-methylenedioxymethamphetamine (MDMA) or saline on locomotor activity and Fos expression following the bilateral destruction of hippocampal dentate granule cells by colchicine in rats. The lesioned animals, when administered s.c. saline, showed a significantly greater increase in locomotor activity compared to the intact animals, and revealed a marginally significant level of increased locomotor activity compared to the sham-lesioned animals. In addition, when the lesioned animals were given s.c. saline or MDMA, there was a significant increase in Fos expression in the nucleus accumbens core, but not in the medial prefrontal cortex, dorsolateral prefrontal cortex, anterior cingulate cortex, piriform cortex, dorsal striatum, or nucleus accumbens shell, compared to the intact and sham-lesioned animals. Overall, these results suggest that the nucleus accumbens core may be involved in the enhancement of locomotor activity induced by the injection of saline alone (stress loading) or MDMA following bilateral destruction of hippocampal dentate granule cells by colchicine.