Exploring drug delivery for the DOT1L inhibitor pinometostat (EPZ-5676): Subcutaneous administration as an alternative to continuous IV infusion, in the pursuit of an epigenetic target.

Protein methyltransferases are emerging as promising drug targets for therapeutic intervention in human cancers. Pinometostat (EPZ-5676) is a small molecule inhibitor of the DOT1L enzyme, a histone methyltransferase that methylates lysine 79 of histone H3. DOT1L activity is dysregulated in the pathophysiology of rearranged mixed lineage leukemia (MLL-r). Pinometostat is currently in Phase 1 clinical trials in relapsed refractory acute leukemia patients and is administered as a continuous IV infusion (CIV). The studies herein investigated alternatives to CIV administration of pinometostat to improve patient convenience. Various sustained release technologies were considered, and based on the required dose size as well as practical considerations, subcutaneous (SC) bolus administration of a solution formulation was selected for further evaluation in preclinical studies. SC administration offered improved exposure and complete bioavailability of pinometostat relative to CIV and oral administration. These findings warranted further evaluation in rat xenograft models of MLL-r leukemia. SC dosing in xenograft models demonstrated inhibition of MLL-r tumor growth and inhibition of pharmacodynamic markers of DOT1L activity. However, a dosing frequency of thrice daily (t.i.d) was required in these studies to elicit optimal inhibition of DOT1L target genes and tumor growth inhibition. Development of an extended release formulation may prove useful in the further optimization of the SC delivery of pinometostat, moving towards a more convenient dosing paradigm for patients.

Nonclinical pharmacokinetics and metabolism of EPZ-5676, a novel DOT1L histone methyltransferase inhibitor.

(2R,3R,4S,5R)-2-(6-Amino-9H-purin-9-yl)-5-((((1r,3S)-3-(2-(5-(tert-butyl)-1H-benzo[d]imidazol-2-yl)ethyl)cyclobutyl)(isopropyl)amino)methyl)tetrahydrofuran-3,4-diol (EPZ-5676) is a novel DOT1L histone methyltransferase inhibitor currently in clinical development for the treatment of MLL-rearranged leukemias. This report describes the preclinical pharmacokinetics and metabolism of EPZ-5676, an aminonucleoside analog with exquisite target potency and selectivity that has shown robust and durable tumor growth inhibition in preclinical models. The in vivo pharmacokinetics in mouse, rat and dog were characterized following i.v. and p.o. administration; EPZ-5676 had moderate to high clearance, low oral bioavailability with a steady-state volume of distribution 2-3 fold higher than total body water. EPZ-5676 showed biexponential kinetics following i.v. administration, giving rise to a terminal elimination half-life (t1/2 ) of 1.1, 3.7 and 13.6 h in mouse, rat and dog, respectively. The corresponding in vitro ADME parameters were also studied and utilized for in vitro-in vivo extrapolation purposes. There was good agreement between the microsomal clearance and the in vivo clearance implicating hepatic oxidative metabolism as the predominant elimination route in preclinical species. Furthermore, low renal clearance was observed in mouse, approximating to fu -corrected glomerular filtration rate (GFR) and thus passive glomerular filtration. The metabolic pathways across species were studied in liver microsomes in which EPZ-5676 was metabolized to three monohydroxylated metabolites (M1, M3 and M5), one N-dealkylated product (M4) as well as an N-oxide (M6).

Moisture assay of an antifungal by near-infrared diffuse reflectance spectroscopy.

Near-infrared (NIR) diffuse reflectance spectroscopy was employed in the method development and validation of a moisture assay for the novel antifungal caspofungin acetate. Spectra were obtained over the entire spectral region available (950-1650 nm) using an InGaAs photodiode array detector equipped with a diffuse reflectance probe. No sample pre-treatment was required and the analysis time was less than 1 min. Primary reference data were obtained using a Karl Fischer (KF) titration (coulometric, volumetric or both). The investigated range of water content was 2.6-9.9% (w/w) with a standard error of prediction (SEP) of 0.2%. The predictive capabilities of the partial least-squares (PLS) regression calibration model used in the moisture assay were verified using independent test sets. The NIR predicted values of the developed method were equivalent to the reference method sets and the prediction error was equivalent to the reference method error. These results reveal that the predictive model constructed by means of a PLS regression is valid, rugged and could be used to determine moisture levels on-line in caspofungin acetate drug substance.

Coordination and Redox Chemistry of Substituted-Polypyridyl Complexes of Ruthenium.

The complexes [Ru(tpy)(acac)(Cl)], [Ru(tpy)(acac)(H(2)O)](PF(6)) (tpy = 2,2',2"-terpyridine, acacH = 2,4 pentanedione) [Ru(tpy)(C(2)O(4))(H(2)O)] (C(2)O(4)(2)(-) = oxalato dianion), [Ru(tpy)(dppene)(Cl)](PF(6)) (dppene = cis-1,2-bis(diphenylphosphino)ethylene), [Ru(tpy)(dppene)(H(2)O)](PF(6))(2), [Ru(tpy)(C(2)O(4))(py)], [Ru(tpy)(acac)(py)](ClO(4)), [Ru(tpy)(acac)(NO(2))], [Ru(tpy)(acac)(NO)](PF(6))(2), and [Ru(tpy)(PSCS)Cl] (PSCS = 1-pyrrolidinedithiocarbamate anion) have been prepared and characterized by cyclic voltammetry and UV-visible and FTIR spectroscopy. [Ru(tpy)(acac)(NO(2))](+) is stable with respect to oxidation of coordinated NO(2)(-) on the cyclic voltammetric time scale. The nitrosyl [Ru(tpy)(acac)(NO)](2+) falls on an earlier correlation between nu(NO) (1914 cm(-)(1) in KBr) and E(1/2) for the first nitrosyl-based reduction 0.02 V vs SSCE. Oxalate ligand is lost from [Ru(II)(tpy)(C(2)O(4))(H(2)O)] to give [Ru(tpy)(H(2)O)(3)](2+). The Ru(III/II) and Ru(IV/III) couples of the aqua complexes are pH dependent. At pH 7.0, E(1/2) values are 0.43 V vs NHE for [Ru(III)(tpy)(acac)(OH)](+)/[Ru(II)(tpy)(acac)(H(2)O)](+), 0.80 V for [Ru(IV)(tpy)(acac)(O)](+)/[Ru(III)(tpy)(acac)(OH)](+), 0.16 V for [Ru(III)(tpy)(C(2)O(4))(OH)]/[Ru(II)(tpy)(C(2)O(4))(H(2)O)], and 0.45 V for [Ru(IV)(tpy)(C(2)O(4))(O)]/[Ru(III)(tpy)(C(2)O(4))(OH)]. Plots of E(1/2) vs pH define regions of stability for the various oxidation states and the pK(a) values of aqua and hydroxo forms. These measurements reveal that C(2)O(4)(2)(-) and acac(-) are electron donating to Ru(III) relative to bpy. Comparisons with redox potentials for 21 related polypyridyl couples reveal the influence of ligand changes on the potentials of the Ru(IV/III) and Ru(III/II) couples and the difference between them, DeltaE(1/2). The majority of the effect appears in the Ru(III/II) couple. ()A linear correlation exists between DeltaE(1/2) and the sum of a set of ligand parameters defined by Lever et al., SigmaE(i)(L(i)), for the series of complexes, but there is a dramatic change in slope at DeltaE(1/2) approximately -0.11 V and SigmaE(i)(L(i)) = 1.06 V. Extrapolation of the plot of DeltaE(1/2) vs SigmaE(i)(L(i)) suggests that there may be ligand environments in which Ru(III) is unstable with respect to disproportionation into Ru(IV) and Ru(II). This would make the two-electron Ru(IV)O/Ru(II)OH(2) couple more strongly oxidizing than the one-electron Ru(IV)O/Ru(III)OH couple.