Development of an UPLC-MS/MS Method for the Quantitative Analysis of Upadacitinib in Beagle Dog Plasma and Pharmacokinetics Study.

BACKGROUND: Upadacitinib, a novel selective Janus kinase 1 (JAK1) inhibitor, has been recently approved by the US FDA for the treatment of adult patients with moderately to severely active rheumatoid arthritis (RA). An ultra-performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) method for the quantitative analysis of upadacitinib in beagle dog plasma was developed and validated. METHODS: Upadacitinib and fedratinib (internal standard, IS) were extracted with ethyl acetate under alkaline condition and then separated and detected. The chromatographic column was Waters Acquity UPLC BEH C18 column (2.1 mm × 50 mm, 1.7 µm), the mobile phase was acetonitrile and 0.1% formic acid in water with gradient elution procedure, and the flow rate was 0.40 mL/min. Under the positive ion mode, upadacitinib and IS were monitored by multiple reaction monitoring (MRM) as the following mass transition pairs: m/z 447.00 → 361.94 for upadacitinib and m/z 529.82 → 141.01 for IS. RESULTS: In the concentration range of 1-500 ng/mL, upadacitinib had good linearity, and the lower limit of quantification (LLOQ) was 1 ng/mL. The RSD of the intra- and inter-day precision was less than 10.03%, and the RE of accuracy was -3.79% to 2.58%. The extraction recovery of upadacitinib was more than 80%, the matrix effect was around 100%, and upadacitinib was found to be stable. CONCLUSION: The novel optimized UPLC-MS/MS assay was an effective tool for the determination of upadacitinib and had been successfully applied to the pharmacokinetic study of upadacitinib in beagle dogs, and this method would also be used to study DDIs.

FXR1 regulates transcription and is required for growth of human cancer cells with TP53/FXR2 homozygous deletion.

Tumor suppressor p53 prevents cell transformation by inducing apoptosis and other responses. Homozygous TP53 deletion occurs in various types of human cancers for which no therapeutic strategies have yet been reported. TCGA database analysis shows that the TP53 homozygous deletion locus mostly exhibits co-deletion of the neighboring gene FXR2, which belongs to the Fragile X gene family. Here, we demonstrate that inhibition of the remaining family member FXR1 selectively blocks cell proliferation in human cancer cells containing homozygous deletion of both TP53 and FXR2 in a collateral lethality manner. Mechanistically, in addition to its RNA-binding function, FXR1 recruits transcription factor STAT1 or STAT3 to gene promoters at the chromatin interface and regulates transcription thus, at least partially, mediating cell proliferation. Our study anticipates that inhibition of FXR1 is a potential therapeutic approach to targeting human cancers harboring TP53 homozygous deletion.