Abstract
A simple, sensitive and rapid assay method has been developed and validated for the estimation of enzalutamide, N-desmethylenzalutamide (active metabolite of enzalutamide), darolutamide and ORM-15341 (active metabolite of darolutamide) on mice dried blood spots (DBS) using liquid chromatography coupled to tandem mass spectrometry with electro spray ionization in the positive-ion mode. The method utilizes liquid extraction of enzalutamide, N-desmethylenzalutamide, darolutamide and ORM-15341 from 3 mm punched disks from DBS cards (spiked or study samples). The extracted sample was chromatographed using an isocratic mobile phase (0.2 % formic acid : acetonitrile; 30:70, v/v) on an Atlantis dC18 column. The total run time was 2.5 min. The MS/MS ion transitions monitored were m/z 465 → m/z 209, m/z 451 → m/z 195, m/z 399 → m/z 178, m/z 397 → m/z 194 and m/z 481 → m/z 453 for enzalutamide, N-desmethylenzalutamide, darolutamide, ORM-15341 and the IS (apalutamide-d3), respectively. Method validation was performed as per regulatory guideline. The assay had a good linearity over the range of 0.93-2000 ng/mL. The intra- and inter-batch accuracy and precision (%RE & RSD) across quality controls met the acceptance criteria for all the analytes. Stability studies showed that all the analytes were stable on DBS cards for one month. This novel method has been applied to analyze the DBS samples of enzalutamide, N-desmethylenzalutamide, darolutamide and ORM-15341 obtained from a pharmacokinetic study in mice.
Highlights
Despite therapeutic advances, prostate adenocarcinoma is the highly prevalent cancer in men and responsible for 20 % of cancer related deaths in the Western world [1]
The results have shown that the precision and accuracy for analyzed samples were within acceptance range (Table 2)
We developed and validated an analytical method for the determination of enzalutamide, Ndesmethylenzalutamide, darolutamide and ORM-15341 in mice dried blood spots (DBS) samples for these second generation non-steroidal anti-androgens
Summary
Prostate adenocarcinoma is the highly prevalent cancer in men and responsible for 20 % of cancer related deaths in the Western world [1]. Despite an initial response, most patients develop a most aggressive form of disease called metastatic castration-resistant prostate cancer (mCRPC) that is associated with tumor progression and survival less than 18-24 months [3]. LC-MS/MS determination of enzalutamide/darolutamide with metabolites on DBS become available like targeting androgen receptor signalling such as CYP17A1 inhibitor (abiraterone acetate) [4] and a second-generation androgen receptor antagonist (enzalutamide, apalutamide and darolutamide) [5,6,7]. Treatment with these drugs showed improved overall survival and quality of life in mCRPC patients
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