Abstract
A highly sensitive and specific LC-MS/MS method for the quantitation of largazole thiol, the active species of the marine-derived preclinical histone deacetylase inhibitor, largazole (prodrug), was developed and validated. Largazole thiol was extracted with ethyl acetate from human or rat plasma along with the internal standard, harmine. Samples were separated on an Onyx Monolithic C18 column by a stepwise gradient elution with 0.1% formic acid in methanol and 0.1% aqueous formic acid employing multiple reaction monitoring (MRM) detection. Linear calibration curves were obtained in the range of 12.5–400 ng/mL with 200 µL of human plasma. The overall intra-day precision was from 3.87% to 12.6%, and the inter-day precision was from 7.12% to 9.8%. The accuracy at low, medium and high concentrations ranged from 101.55% to 105.84%. Plasma protein bindings of largazole thiol in human and rat plasma as determined by an ultrafiltration method were 90.13% and 77.14%, respectively. Plasma drug concentrations were measured by this LC-MS/MS method. The pharmacokinetics of largazole thiol in rats was studied following i.v. administration at 10 mg/kg and found to follow a two-compartment model. Largazole thiol was rapidly eliminated from systemic circulation within 2 h. The established LC-MS/MS method is suitable for the analysis of largazole thiol in human plasma, as well.
Highlights
The cyclic depsipeptide, largazole, discovered from a marine cyanobacterium, was first described in 2008 as a potential anticancer agent with novel chemical scaffolding and selectivity for cancer cells over non-transformed cells [1]
The electrospray mass spectrum of largazole thiol and harmine are shown in Figures 2 and 3
histone deacetylase (HDAC) inhibitors are already available on the market, but one main concern is the metabolic stability, which affects the in vivo activity [16]
Summary
The cyclic depsipeptide, largazole, discovered from a marine cyanobacterium, was first described in 2008 as a potential anticancer agent with novel chemical scaffolding and selectivity for cancer cells over non-transformed cells [1]. This activity combined with the unusual structural features has triggered the synthesis of this compound and rigorous biological evaluation. Molecular docking studies using an HDAC1 homology model and the HDAC8-largazole thiol X-ray co-crystal structure indicated that the side chain containing the thiol group chelates Zn2+ in the active site of HDACs, and the macrocycle interacts with divergent regions of HDACs to give pronounced selectivity for Class I isoforms, rather than broadly inhibiting the eleven Zn2+-dependent isoforms [2,15]
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