CYP2C9 Polymorphism Influence in PK/PD Model of Naproxen and 6-O-Desmethylnaproxen in Oral Fluid

Abstract

Polymorphisms in CYP2C9 can significantly interfere with the pharmacokinetic (PK) and pharmacodynamic (PD) parameters of nonsteroidal anti-inflammatory drugs (NSAIDs), including naproxen. The present research aimed to study the PK/PD parameters of naproxen and its metabolite, 6-O-desmethylnaproxen, associated with allelic variations of CYP2C9. In our study, a rapid, selective, and sensitive Liquid Chromatography-Tandem Mass Spectrometry (LC-MS/MS) method was developed and validated for the determination of naproxen and its main metabolite, 6-O-desmethylnaproxen, in oral fluid. Naproxen and its main metabolite were separated using a Shim-Pack XR-ODS 75L × 2.0 column and C18 pre-column at 40 °C using a mixture of methanol and 10 mM ammonium acetate (70:30, v/v), with an injection flow of 0.3 mL/min. The total analytical run time was 3 min. The volunteers, previously genotyped for CYP2C9 (16 ancestral—CYP2C9 *1 and 12 with the presence of polymorphism—CYP2C9 *2 or *3), had their oral fluids collected sequentially before and after taking a naproxen tablet (500 mg) at the following times: 0.25, 0.5, 0.75, 1, 1.5, 2, 3, 4, 5, 6 8, 11, 24, 48, 72 and 96 h. Significant differences in the PK parameters (* p < 0.05) of naproxen in the oral fluid were: Vd/F (L): 98.86 (55.58–322.07) and 380.22 (261.84–1097.99); Kel (1/h): 0.84 (0.69–1.34) and 1.86 (1.09–4.06), in ancestral and mutated CYP2C9 *2 and/or *3, respectively. For 6-O-desmethylnaproxen, no PK parameters were significantly different between groups. The analysis of prostaglandin E2 (PGE2) proved to be effective and sensitive for PD parameters analysis and showed higher levels in the mutated group (p < 0.05). Both naproxen and its main metabolite, 6-O-desmethylnaproxen, and PGE2 in oral fluid can be effectively quantified using LC-MS/MS after a 500 mg oral dose of naproxen. Our method proved to be effective and sensitive to determine the lower limit of quantification of naproxen and its metabolite, 6-O-desmethylnaproxen, in oral fluid (2.4 ng/mL). All validation data, such as accuracy, precision, and repeatability intra- and inter-assay, were less than 15%. Allelic variations of CYP2C9 may be considered relevant in the PK of naproxen and its main metabolite, 6-O-desmethylnaproxen.

Liquid Chromatography-Tandem Mass Spectrometry Method for Detection and Quantification of Meloxicam and 5′-Carboxymeloxicam in Oral Fluid Samples

“…The MEPS methodology was selected for the extraction of analytes from oral fluid samples and calibration curves. MEPS consists of the miniaturization of conventional SPE with a conditioned needle incorporated directly into an injector syringe (Trajan Scientific Australia Pty Ltd) which is an alternative to other extraction methodologies in different matrixes [ 36 ], thus demonstrating its specificity and reduction in matrix effects [ 37 , 38 ], including in oral fluid samples [ 16 , 21 , 27 , 39 , 40 ].…”

“…Finally, the syringe and BIN were washed (100 µL—5× MeOH) between each patient sample. The same process was performed for the calibration curve [ 21 , 27 , 38 ].…”

“…The extraction steps with the MEPS device, which were standardized in previous works by our group [ 21 , 27 ], are described in Table 1 .…”

“…Due to the numerous sample collections required for a pharmacokinetic study (up to 96 h), the use of alternative samples, such as oral fluid, has shown promise, even for NSAIDs [ 26 , 27 , 28 , 29 , 30 , 31 ]. Some analytical methodologies were developed in human plasma for the detection of meloxicam after the ingestion of a 7.5 mg tablet [ 32 ], 15 mg tablet [ 33 ] and transdermal application [ 34 ], but studies about the detection and determination of this drug in oral fluid samples have not been reported.…”

“…MEPS is a miniaturization of solid-phase extraction (SPE) with a packed bed (1–2 mg). Although its applicability in oral fluid is recent, it has shown to be promising with some drugs [ 21 , 27 , 37 , 38 ].…”