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TM208 and TM209, dithiocarbamate derivatives with potential anti-cancer effects, were evaluated in reversible and time-dependent cytochrome P450 (CYP) 3A inhibition assays in rat liver microsomes using testosterone as probe substrate. Both compounds were found to be weak reversible inhibitors and moderate mechanism-based inhibitors of rat CYP3A. For reversible inhibition on rat CYP3A, the Ki values of competitive inhibition model were 12.10±1.75 and 13.94±1.31 μM, respectively. For time-dependent inhibition, the inactivation constants (Kl) were 31.93±12.64 and 32.91±15.58 μM, respectively, and the maximum inactivation rates (kinact) were 0.03497±0.0069 and 0.07259±0.0172 min−1 respectively. These findings would provide useful in vitro information for future in vivo DDI studies on TM208 or TM209.

Highlights: [•] Highlight of evaluation of TM208 toxic effects by metabonomics. [•] The biomarkers were identified with metabonomics analytical software and online database. [•] Mechanism of TM208 toxicity was related to tricarboxylic acid (TCA) cycle and lysine metabolism. [ABSTRACT FROM AUTHOR]

Abstract: A rapid, simple, and sensitive on-line solid-phase extraction HPLC–DAD method for simultaneous evaluation of the activity of five CYP450 isoforms (CYP1A2, CYP2C19, CYP2D6, CYP2E1 and CYP3A4) in vivo has been developed and validated. The five specific probe substrates include caffeine (1A2), metoprolol (2D6), dapsone (3A4), omeprazole (2C19) and chlorzoxazone (2E1). Automated pre-purification of plasma and enrichment of analytes were performed using a C18 on-line solid-phase extraction cartridge. After being eluted from the cartridge, the analytes and the internal standard antipyrine were separated on a C18 RP analytical column and analyzed by DAD. The method was validated to quantify the concentration ranges of 0.05–50.0μg/ml for dapsone and omeprazole, 0.1–50.0μg/ml for caffeine and 0.2–50.0μg/ml for metoprolol and chlorzoxazone. The linearity (R 2) for all analytes tested was exceeded 0.99. The intra-day precision ranged from 0.29 to 13% and the inter-day precision ranged from 5.0 to 15%, respectively. The intra-day and inter-day accuracy were between 86.7% and 113.6%. The extraction recoveries were in the range 82.8–109.9% for all the analytes and internal standard antipyrine. This method was successfully applied to evaluate the effects of TM208 on rat five CYP450 isoforms. [Copyright &y& Elsevier]

ABSTRACT 4-Methyl-piperazine-1-carbodithioc acid 3-cyano-3, 3-diphenylpropyl ester hydrochloride (TM208), a newly synthesized anticancer compound, was quantified using liquid chromatography-tandem mass spectrometry (LC-MS/MS) for the first time. A simple, rapid and sensitive assay method using propranolol as internal standard (IS) after one-step precipitation with acetonitrile was developed and validated to determine TM208 in rat plasma. Separation was achieved on a reverse-phase C18 column with a mobile phase composed of methanol-water (pH4.0) containing 5 m m ammonium acetate in gradient elution mode. A triple quadrupole tandem mass spectrometer with electrospray ionization source was used as detector and operated by multiple reaction monitoring in the positive ion mode. Calibration curves were linear ( r > 0.99) between 0.2 and 500 ng/mL. The quantitative limit was 0.2 ng/mL; reliable precision and accuracy were validated by relative standard deviation values in the range 3.44-13.15% and relative error values between −0.58 and −9.78%. The method was successfully applied to preclinical pharmacokinetic studies of TM208. Copyright © 2011 John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

Abstract: TM208 and TM209, dithiocarbamate derivatives with potential anti-cancer effects, were evaluated in reversible and time-dependent cytochrome P450 (CYP) 3A inhibition assays in rat liver microsomes using testosterone as probe substrate. Both compounds were found to be weak reversible inhibitors and moderate mechanism-based inhibitors of rat CYP3A. For reversible inhibition on rat CYP3A, the K i values of competitive inhibition model were 12.10±1.75 and 13.94±1.31μM, respectively. For time-dependent inhibition, the inactivation constants (K l) were 31.93±12.64 and 32.91±15.58μM, respectively, and the maximum inactivation rates (k inact) were 0.03497±0.0069 and 0.07259±0.0172min−1 respectively. These findings would provide useful in vitro information for future in vivo DDI studies on TM208 or TM209. [Copyright &y& Elsevier]

A specific, simple, and fast online-solid-phase extraction-high performance liquid chromatography-diode array detector ( SPE- HPLC- DAD) method was developed and validated to quantify 4-methylpiperaine-1-carbodithioc acid 3-cyano-3,3-diphenylpropyl ester hydrochloride ( TM208) in small volume samples of rats' plasma for the first time. In this method, the 50-μL plasma sample was taken to perform protein precipitation with 75 μL methanol, and then 50 μL supernatant containing the target analytes was injected and concentrated automatically in a C18 solid-phase extraction ( SPE) cartridge. After that the sample was separated on a C18 RP analytical column and analyzed by DAD. The run cycle time is 6.0 min for each sample, and the calibration curve over the range of 0.03 to 25.00 μg/mL has a good linear relationship ( r > 0.9998). The recoveries of the quality control samples were all greater than 90%. The limit of detection and the lowest limit of quantification were 0.01 and 0.03 μg/mL, respectively. Finally, this method was successfully applied to a pharmacokinetic study of TM208 in rats. [ABSTRACT FROM AUTHOR]