Simultaneous Determination of Ripretinib and Its Desmethyl Metabolite in Human Plasma Using LC-MS/MS.

BACKGROUND: Ripretinib, a recently developed tyrosine kinase inhibitor with switch-control abilities, can inhibit both primary and secondary activation of KIT(KIT proto-oncogene receptor tyrosine kinase) and platelet-derived growth factor receptor alpha (PDGFRA) mutants, which contribute to gastrointestinal stromal tumor progression. METHODS: In this study, a high-performance liquid chromatography-tandem mass spectrometry method to measure the concentrations of ripretinib and its active desmethyl metabolite DP-5439 in human plasma was developed and validated. Plasma samples were extracted and recovered by precipitation with acetonitrile containing the internal standard and diluted with acetonitrile before analysis. Ripretinib and DP-5439 were separated using chromatography on a Waters ACQUITY UPLC HSS T3 column (2.1 mm × 50 mm, 1.8 µm) with gradient elution using 0.1% formic acid and 5 mM ammonium formate in water as mobile phase A and acetonitrile as mobile phase B. The mobile phase was set to a flow rate of 0.5 mL/min. RESULTS: The calibration curves were linear across the following concentration range: 7.5 to 3000 ng/mL for ripretinib and 10 to 4000 ng/mL for DP-5439. The intraday and interday precisions were approximately 15% for all analytes in the quality control samples. The relative matrix effects in extracted plasma samples (90.3%-108.8% at different levels) were considered acceptable. CONCLUSIONS: This method will be a useful tool in oncology to facilitate the further clinical development of ripretinib.

LC-MS/MS methods for determination of venetoclax in human plasma and cerebrospinal fluid.

We developed and validated sensitive MS/MS methods for the determination of venetoclax, an oral selective B-cell lymphoma-2 inhibitor, in human plasma and cerebrospinal fluid (CSF). Acetonitrile was used as protein precipitant. The mobile phase was 10 mM ammonium formate consisting of 0.1% formic acid and acetonitrile (40:60, v/v). The analytes were separated on an ACQUITY UPLC HSS T3 column (2.1 × 50 mm, 1.8 µm) in 5 min. An API 4000 mass spectrometer was selected to quantify venetoclax and internal standard using m/z 868.3 → 636.3 and 876.3 → 644.3 under multiple response monitoring mode. In plasma, the calibration curve exhibited good linearity ranging from 20.0 to 5000 ng/mL, whereas in the CSF, the linear range was 0.500-100 ng/mL. The matrix effect of venetoclax and internal standard (venetoclax-d8) was not obvious in both plasma and CSF. The inter- and intra-run accuracy was within ±11.9%, and the inter- and intra-run precision was below 13.6%. Both methods had no carryover, and the recovery was close to 100%. The validated methods were employed to quantify the concentrations of venetoclax in the plasma and CSF of patients diagnosed with chronic lymphocytic leukemia or acute myelogenous leukemia.

Simultaneous Determination of Orelabrutinib, Zanubrutinib, Ibrutinib and Its Active Metabolite in Human Plasma Using LC-MS/MS.

Ibrutinib, orelabrutinib, and zanubrutinib are all Bruton's tyrosine kinase inhibitors, which have greatly improved the treatment of B-cell malignancies. In this study, an LC-MS/MS method was developed and validated for the determination of orelabrutinib, zanubrutinib, ibrutinib, and its active metabolite dihydrodiol ibrutinib in human plasma. The Ibrutinib-d5 was used as the internal standard. Pretreatment was performed using a simple protein precipitation step using acetonitrile. The ACQUITY UPLC HSS T3 column (2.1×50 mm, 1.8 µm) was used to separate the analytes, and the run time was 6.5 min. The mobile phase consisted of acetonitrile and 10 mM of ammonium formate, which contained 0.1% formic acid. The multiple reactions' monitoring transitions were selected at m/z 428.1→411.2, 472.2→455.2, 441.1→304.2, 475.2→304.2 and 446.2→309.2 respectively for orelabrutinib, zanubrutinib, ibrutinib, dihydrodiol ibrutinib and ibrutinib-d5 using positive ion electrospray ionization. The standard curves were linear, from 0.400 to 200 ng/mL for ibrutinib and dihydrodiol ibrutinib, 1.00-500 ng/mL for orelabrutinib, and 2.00-1000 ng/mL for zanubrutinib. Selectivity, the lower limit of quantitation, precision, accuracy, matrix effect, recovery, stability, and dilution integrity all met the acceptance criteria of FDA guidance. This method was used to quantify the plasma levels of orelabrutinib, zanubrutinib, ibrutinib, and dihydrodiol ibrutinib in clinical patients.