Validation of an LC-MS/MS method for simultaneous quantification of abiraterone, enzalutamide and darolutamide in human plasma.

Currently, several oral androgen receptor signalling inhibitors are available for the treatment of advanced prostate cancer. Quantification of plasma concentrations of these drugs is highly relevant for various purposes, such as Therapeutic Drug Monitoring (TDM) in oncology. Here, we report a liquid chromatography/tandem mass spectrometric (LC-MS/MS) method for the simultaneous quantification of abiraterone, enzalutamide, and darolutamide. The validation was performed according to the requirements of the U.S. Food and Drug Administration and European Medicine Agency. We also demonstrate the clinical applicability of the quantification of enzalutamide and darolutamide in patients with metastatic castration-resistant prostate cancer.

Influence of probenecid on endoxifen systemic exposure in breast cancer patients on adjuvant tamoxifen treatment.

Introduction: In breast cancer patients treated with the anti-estrogen tamoxifen, low concentrations of the active metabolite endoxifen are associated with more disease recurrence. We hypothesized that we could increase endoxifen concentrations by induction of its formation and inhibition of its metabolism by co-administration of probenecid. Methods: We conducted a crossover study and measured endoxifen concentrations in patients on steady-state tamoxifen monotherapy and after 14 days of combination treatment with probenecid. Eleven evaluable patients were included. Results: Treatment with tamoxifen and probenecid resulted in a 26% increase of endoxifen area under the plasma concentration-time curve from 0 to 24 h (AUC0-24h) compared to tamoxifen monotherapy (95% confidence interval [CI]: 8-46%; p < 0.01), while the maximum observed endoxifen concentration increased with 24% (95% CI: 7-44%; p < 0.01). The metabolic ratio of endoxifen to tamoxifen increased with 110% (95% CI: 82-143%; p < 0.001) after the addition of probenecid. Conclusion: Probenecid resulted in a clinically relevant increase of endoxifen concentrations in breast cancer patients treated with adjuvant tamoxifen. This combination therapy could provide a solution for patients with a CYP2D6-poor metabolizer phenotype or endoxifen concentrations below the threshold despite earlier tamoxifen dose.

Fasting protects against the side effects of irinotecan treatment but does not affect anti-tumour activity in mice.

BACKGROUND: The main limitation to the use of irinotecan in the treatment of colorectal cancer is the severity of side effects, including neutropaenia and diarrhoea. Here, we explored the effects of 3 days of fasting on irinotecan-induced toxicities, on plasma, liver and tumour pharmacokinetics and on anti-tumour activity in mice. EXPERIMENTAL APPROACH: Male BALB/c mice received C26 colon carcinoma cells subcutaneously. They were randomized 1:1 into equally sized ad libitum fed and fasted groups after which they were treated with irinotecan. Weight and adverse side effects were recorded daily. At the end of the experiment, tumours were resected and weighed, and concentrations of irinotecan and its active metabolite SN-38 were determined in plasma and tumour. KEY RESULTS: Fasting prevented the diarrhoea and visible signs of discomfort induced by irinotecan. Ad libitum fed animals developed leucopenia compared with untreated controls, whereas fasted mice did not. Irinotecan suppressed tumour growth equally in both treated groups, compared with untreated controls. Levels of the active irinotecan metabolite SN-38 9 (calculated as AUC values) were significantly lower in fasted mice in both plasma and liver, but not in tumour tissue. CONCLUSIONS AND IMPLICATIONS: Fasting protected against irinotecan-induced side effects without interfering with its anti-tumour efficacy. Fasting induced a lower systemic exposure to SN-38, which may explain the absence of adverse side effects, while tumour levels of SN-38 remained unchanged. These data offer important new approaches to improve treatment with irinotecan in patients.

Influence of drug formulation on OATP1B-mediated transport of paclitaxel.

Taxane antineoplastic agents are extensively taken up into hepatocytes by OATP1B-type transporters before metabolism and excretion. Because the biodistributional properties imposed upon these agents by different solubilizers drive clinically important pharmacodynamic endpoints, we tested the hypothesis that the in vitro and in vivo interaction of taxanes with OATP1B transporters is affected by the choice of drug delivery system. Transport of paclitaxel, docetaxel, and cabazitaxel was studied in vitro using various cell lines transfected with OATP1B1, OATP1B3, or the rodent equivalent OATP1B2. Pharmacokinetic studies were done in wild-type and OATP1B2-knockout mice in the presence or absence of polysorbate 80 (PS80) or Kolliphor EL (formerly Cremophor EL; CrEL). Paclitaxel and docetaxel, but not cabazitaxel, were transported substrates of OATP1B1, OATP1B3, and OATP1B2, and these in vitro transport processes were strongly reduced in the presence of clinically relevant concentrations of PS80 and CrEL. When paclitaxel was administered without any solubilizers, deficiency of OATP1B2 in mice was associated with a significantly decreased systemic clearance because of a liver distribution defect (P=0.000484). However, this genotype dependence of paclitaxel clearance was masked in the presence of PS80 or CrEL because of significant inhibition of OATP1B2-mediated hepatocellular uptake of the drug (P<0.05). Our findings confirm the importance of OATP1B-type transporters in the hepatic elimination of taxanes and indicate that this process can be inhibited by PS80 and CrEL. These results suggest that the likelihood of drug-drug interactions mediated by these transporters is strongly dependent on the selected taxane solubilizer.

Quantification of tamoxifen and three of its phase-I metabolites in human plasma by liquid chromatography/triple-quadrupole mass spectrometry.

In view of future pharmacokinetic studies, a highly sensitive ultra performance liquid chromatography/tandem mass spectrometry (UPLC-MS/MS) method has been developed for the simultaneous quantification of tamoxifen and three of its main phase I metabolites in human lithium heparinized plasma. The analytical method has been thoroughly validated in agreement with FDA recommendations. Plasma samples of 200 µl were purified by liquid-liquid extraction with 1 ml n-hexane/isopropanol, after deproteination through addition of 50 µl acetone and 50 µl deuterated internal standards in acetonitrile. Tamoxifen, N-desmethyl-tamoxifen, 4-hydroxy-tamoxifen and endoxifen were chromatographically separated on an Acquity UPLC(®) BEH C18 1.7 µm 2.1 mm×100 mm column eluted at a flow-rate of 0.300 ml/min on a gradient of 0.2mM ammonium formate and acetonitrile, both acidified with 0.1% formic acid. The overall run time of the method was 10 min, with elution times of 2.9, 3.0, 4.1 and 4.2 min for endoxifen, 4-hydroxy-tamoxifen, N-desmethyl-tamoxifen and tamoxifen, respectively. Tamoxifen and its metabolites were quantified by triple-quadrupole mass spectrometry in the positive ion electrospray ionization mode. The multiple reaction monitoring transitions were set at 372>72 (m/z) for tamoxifen, 358>58 (m/z) for N-desmethyl-tamoxifen, 388>72 (m/z) for 4-hydroxy-tamoxifen and 374>58 (m/z) for endoxifen. The analytical method was highly sensitive with the lower limit of quantification validated at 5.00 nM for tamoxifen and N-desmethyl-tamoxifen and 0.500 nM for 4-hydroxy-tamoxifen and endoxifen, which is equivalent to 1.86, 1.78, 0.194 and 0.187 ng/ml for tamoxifen, N-desmethyl-tamoxifen, 4-hydroxy-tamoxifen and endoxifen, respectively. The method was also precise and accurate, with within-run and between-run precisions within 12.0% and accuracy ranging from 89.5 to 105.3%. The method has been applied to samples from a clinical study and cross-validated with a validated LC-MS/MS method in serum.

Increasing tumoral 5-fluorouracil concentrations during a 5-day continuous infusion: a microdialysis study.

PURPOSE: Response to anticancer therapy is believed to be directly related to the concentration of the anticancer drug in the tumor itself. Assessment of intra-tumor drug pharmacokinetics can be helpful to gain more insight into mechanisms involved in the (in)sensitivity of tumors to anticancer therapy. We explored the pharmacokinetics of 5-fluorouracil in both plasma and tumor tissue during a 5-day continuous infusion of 5-fluorouracil in patients with cancer. Sampling for measurement of 5-fluorouracil in tumor tissue was performed using microdialysis. EXPERIMENTAL DESIGN: In seven patients with an accessible (sub)cutaneous tumor treated with a continuous 5-fluorouracil infusion, plasma and microdialysate samples from tumor and normal adipose tissue were collected over a period of 5 days. RESULTS: For six patients, drug concentrations in both tumor tissue and plasma were available. Concentration-time curves of unbound 5-fluorouracil were lower in tumor tissue compared to the curves in plasma, but exposure ratios of tumor tissue versus plasma increased during the 5-day infusion period. The presence of circadian rhythmicity of 5-fluorouracil pharmacokinetics in the tumor itself was demonstrated as 5-fluorouracil concentrations in tumor extracellular fluid were higher during the night than during daytime. CONCLUSION: Microdialysis was successfully employed in patients with cancer during a continuous 5-day 5-fluorouracil infusion. Plasma and tumor pharmacokinetics of 5-fluorouracil differed substantially with increasing 5-fluorouracil concentrations in tumor over time, possibly resulting from a lowered interstitial fluid pressure by 5-fluorouracil itself. This microdialysis 5-fluorouracil model might be useful to monitor the effect of drug delivery modulating strategies in future studies.