Review of HPLC and LC-MS/MS assays for the determination of various nonsteroidal anti-androgens used in the treatment of prostate cancer.

Prostate cancer is the most common cancer and one of the leading causes of cancer deaths in men. One of the commonly used approaches to treat metastatic prostate cancer was via first-generation nonsteroidal anti-androgens (NSAAs), namely flutamide, nilutamide, bicalutamide and topilutamide. Most prostate cancer patients who are initially responsive develop the most aggressive form of disease called castration-resistant prostate cancer. Second-generation NSAA receptor antagonists (enzalutamide, apalutamide and darolutamide) are emerging as additional new options to treat castration-resistant prostate cancer. The objective of this work was to review the literature on the bioanalytical methods for the quantification of first- and second-generation NSAA inhibitors in clinical (human plasma) and preclinical (mouse plasma, rat plasma, urine and tissue homogenates etc.) studies along with relevant case studies for some chosen drugs. Based on the review, it was concluded that the published methodologies using either HPLC or LC-MS/MS are well suited for the quantification of NSAA inhibitors in various biological fluids to delineate pharmacokinetic data.

Selective androgen receptor modulators: in vitro and in vivo metabolism and analysis.

For future targeted screening in National Residue Control Programmes, the metabolism of seven SARMs, from the arylpropionamide and the quinolinone classes, was studied in vitro using S9 bovine liver enzymes. Metabolites were detected and identified with ultra-performance liquid chromatography (UPLC) coupled to time-of-flight mass spectrometry (ToF-MS) and triple quadrupole mass spectrometry (QqQ-MS). Several metabolites were identified and results were compared with literature data on metabolism using a human cell line. Monohydroxylation, nitro-reduction, dephenylation and demethylation were the main S9 in vitro metabolic routes established. Next, an in vivo study was performed by oral administration of the arylpropionamide ostarine to a male calf and urine samples were analysed with UPLC-QToF-MS. Apart from two metabolites resulting from hydroxylation and dephenylation that were also observed in the in vitro study, the bovine in vivo metabolites of ostarine resulted in glucuronidation, sulfation and carboxylation, combined with either a hydroxylation or a dephenylation step. As the intact mother compounds of all SARMs tested are the main compounds present after in vitro incubations, and ostarine is still clearly present in the urine after the in vivo metabolism study in veal calves, the intact mother molecules were selected as the indicator to reveal treatment. The analytical UPLC-QqQ-MS/MS procedure was validated for three commercially available arylpropionamides according to European Union criteria (Commission Decision 2002/657/EC), and resulted in decision limits ranging from 0.025 to 0.05 µg l⁻¹ and a detection capability of 0.025 µg l⁻¹ in all cases. Adequate precision and intra-laboratory reproducibility (relative standard deviation below 20%) were obtained for all SARMs and the linearity was 0.999 for all compounds. This newly developed method is sensitive and robust, and therefore useful for confirmation and quantification of SARMs in bovine urine samples for residue control programmes and research purposes.