Elsevier

Journal of Chromatography B

Volume 809, Issue 1, 25 September 2004, Pages 81-86
Journal of Chromatography B

Determination of the cyclic depsipeptide FK228, a histone deacetylase inhibitor, by liquid chromatography–mass spectrometry

https://doi.org/10.1016/j.jchromb.2004.06.007Get rights and content

Abstract

An analytical method was developed for the quantitative determination of the novel histone deacetylase inhibitor, depsipeptide FK228 (formerly FR901228; NSC 630176), in human plasma. Calibration curves were constructed in the range of 0.5–100 ng/ml, and were analyzed using a weight factor proportional to the nominal concentration. Sample pretreatment involved a liquid–liquid extraction with ethyl acetate using 500 μl aliquots of plasma. The analyte was separated on a column (50 mm × 4.6 mm i.d.) packed with 3.5 μm C8 material, and eluted with methanol—10 mM ammonium formate (55:45; v/v; pH 8). The column effluent was monitored by mass spectrometry with electrospray ionization. The values for precision and accuracy were always ≤7.88% and <3.33% relative error, respectively. The method was successfully applied to examine the pharmacokinetics of FK228 in a cancer patient.

Introduction

Acetylation and deacetylation of histones plays a major role in the regulation of gene transcription and in the modulation of chromatin structure [1]. The state of acetylation of these nucleosome core proteins is determined by two classes of enzymes with opposing activity, which are referred to as histone acetyl transferases and histone deacetylases (HDACs). During the last decade, various agents have been identified that inhibit HDAC activity and induce cell growth arrest, differentiation and/or apoptotic cell death [2]. These agents belong to diverse structural classes and include short-chain fatty acids, hydroxamic acids, synthetic benzamides, and certain cyclic tetrapeptides. In the latter group, cyclic depsipeptide FK228 (formerly FR901228, NSC 630176; (E)-(1S,4S,10S,21R)-7-[(Z)-ethylidene]4,21-diisopropyl-2-oxa-12,13-dithia-5,8,20,23-tetraazabicyclo[8,7,6]tricos-16-ene-3,6,9,19,22-pentanone; Fig. 1) is a novel, highly potent histone deacetylase inhibitor [3], which was first isolated from the fermentation broth of Chromobacterium violaceum [4]. This agent induces expression of the cyclin-dependent kinase inhibitor p21WAF1 [5], [6], [7] through ataxia telangiectasia-mutated-related protein kinase [8], and has also shown antiproliferative activity in various in vitro and in vivo models for human solid tumors [9], [10] and chronic lymphocytic leukemia [11], [12].

Clinical trials of FK228 in patients with refractory solid tumors and hematological malignancies are currently ongoing [13], [14]. Preliminary data indicate that FK228 is a potentially effective agent for the treatment of peripheral and cutaneous T-cell lymphoma [15] as well as renal cell carcinoma [14]. It has been suggested that, in vivo, FK228 may act as a prodrug that requires glutathione-mediated intracellular activation [16], [17]. As part of a project to further assess the disposition and the pharmacodynamic profile of FK228, we report here on the development and validation of an analytical method that allows the determination of the drug at low concentrations in human plasma samples.

Section snippets

Chemicals and materials

Depsipeptide (HPLC purity, 98.56%) was supplied as a crystalline white powder by the Pharmaceutical Management Branch, Cancer Therapy Evaluation Program, National Cancer Institute (Bethesda, MD, USA). HPLC-grade methanol was obtained from J.T. Baker (Phillipsburg, NJ, USA). Ammonium formate and ammonium hydroxide were purchased from Sigma (St. Louis, MO, USA). Deionized water was generated with a hydro-reverse osmosis system (Durham, NC, USA) connected to a Milli-Q UV Plus purifying system

Results and discussion

In recent years, several analytical methods based on reversed-phase HPLC have been reported for the quantitative determination of FK228 in human plasma either based on UV detection [18] or triple-quadrupole MS detection [19], [20]. These methods seem to either lack in sensitivity to support clinical pharmacological investigations or require specialized equipment that is currently unavailable in most laboratories for routine analyses, such as HPLC linked to tandem mass spectrometry. The present

Conclusion

A novel assay for the measurement of a novel histone deacetylase inhibitor, the cyclic depsipeptide FK228, was developed. The method was validated according to the US Food and Drug Administration bioanalytical guidance, and met the pre-defined acceptance criteria for precision and accuracy [21]. The described method permits the analysis of patient samples to concentrations of FK228 as low as 0.5 ng/ml, which is sufficiently sensitive to allow pharmacokinetic monitoring after intravenous

References (21)

  • Y Sasakawa et al.

    Biochem. Pharmacol.

    (2002)
  • Y Sasakawa et al.

    Biochem. Pharmacol.

    (2003)
  • J.L Aron et al.

    Blood

    (2003)
  • R.L Piekarz et al.

    Blood

    (2001)
  • C Chassaing et al.

    J. Chromatogr. B.: Biomed. Sci. Appl.

    (1998)
  • Z Li et al.

    J. Pharm. Biomed. Anal.

    (2000)
  • C Hartmann et al.

    J. Pharm. Biomed. Anal.

    (1998)
  • R.R Rosato et al.

    Expert Opin. Invest. Drugs

    (2004)
  • G Kouraklis et al.

    Curr. Med. Chem. Anti-Cancer Agents

    (2002)
  • T.A Miller et al.

    J. Med. Chem.

    (2003)
There are more references available in the full text version of this article.

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    After fermentation resins and cell debris of each strain were collected by centrifugation and lypholized to dryness. Ten ml of ethyl acetate was used to extract the dried mass and 20 μl of such organic extract was analyzed with an Agilent 1100 series LC/MSD Trap mass spectrometer (Agilent, Santa Clara, CA) for the detection and quantification of FK228 production by relating the peak area of ion signals to that of FK228 standard, as described elsewhere (Cheng et al., 2007; Hwang et al., 2004). A larger quantity of FK228 was purified from 15 L fermentation culture of wild-type C. violaceum according to a previously published procedure (Ueda et al., 1994), and saved as amorphous powder at −20°C until use.

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