Belén Valenzuela

Simultaneous online SPE–HPLC–MS/MS analysis of docetaxel, temsirolimus and sirolimus in whole blood and human plasma

Docetaxel and temsirolimus are some of the most used drugs in a wide range of solid tumors. In preclinical studies, mTOR inhibitors such as temsirolimus have demonstrated synergistic cytotoxic effects with taxanes providing the rationale for combination studies. These anticancer agents exhibit a narrow therapeutic concentration range and due to their high inter- and intra-individual pharmacokinetic variability, therapeutic dose monitoring by highly sensitive methods as LC-MS/MS are important for clinical research. Therefore, the aim of this study was to develop and validate a sensitive, fast and convenient method for the simultaneous identification and quantification of docetaxel, temsirolimus and its main metabolite, sirolimus, using paclitaxel, another anticancer drug, as the internal standard. These analytes were quantified by an integrated online solid phase extraction-high performance liquid chromatography-tandem mass spectrometry (SPE-HPLC-MS/MS) system. Separation was performed on a Zorbax eclipse XDB-C8 (150mm×4.6mm, 5μm) column. The mass spectrometer tandem quadruple detector was equipped with jet stream electrospray ionization, monitored in multiple reactions monitoring (MRM) and operated in positive mode. A combination of protein precipitation with methanol/zinc sulphate (70:30) (v/v) and online SPE using a Zorbax eclipse plus C8 (12.5mm×4.6mm, 5μm) cartridge was used to extract the compounds. This method allows the use of the same reagents, sample treatment and analytical technique independently of whether the samples are whole blood or plasma. The method has been successfully validated and applied to real samples. It is a suitable method for dose adjustment and for evaluating potential drug interactions during combined treatments.

Development and validation of an HPLC-UV method for sorafenib quantification in human plasma and application to patients with cancer in routine clinical practice.

Background: Several factors such as low therapeutic index, large interindividual variability in systemic exposure, and the relationships between exposure and toxicity for sorafenib could justify its therapeutic drug monitoring (TDM). To support TDM, a selective and precise high-performance liquid chromatography with ultraviolet detection (HPLC-UV) method was developed and validated for the determination of sorafenib in human plasma. Methods: After protein precipitation with acetonitrile, sorafenib and lapatinib (internal standard) were separated using isocratic elution on a Kromasil C18 column using a mobile phase of acetonitrile and 20 mmol/L ammonium acetate in a proportion 53:47 (vol/vol) pumped at a constant flow rate of 1.2 mL/min. Quantification was performed at 260 nm. Validation experiments were carried out after the guidelines for Bioanalytical Method Validation published by the Food and Drug Administration and the European Medicines Agency. Results: Calibration curves were linear over the range 0.1–20 mcg/mL. Inter- and intra-day coefficients of variation were <3%. The limit of detection and the lower limit of quantification were 0.06 and 0.1 mcg/mL, respectively. Recoveries of sorafenib from plasma were >99% in all cases. Conclusions: This method was successfully applied to the determination of the drug in the plasma of 2 patients with cancer receiving sorafenib 200 and 400 mg orally twice daily, respectively, and could be useful for TDM of sorafenib in routine clinical practice.

Development and validation of a high-performance liquid chromatography ultraviolet method for lapatinib quantification in human plasma.

Abstract: A selective and precise high-performance liquid chromatography ultraviolet method was developed and validated for the determination of lapatinib in human plasma. After protein precipitation with acetonitrile, lapatinib and sorafenib were separated using isocratic elution (on a C18 Ultrabase column using a mobile phase of acetonitrile/20 mM ammonium acetate in a proportion 53:47 (v/v) pumped at a constant flow rate of 1.2 mL/min). Quantification was performed at 260 nm. Calibration curves were linear over the range 0.2–10 µg/mL. Inter- and intraday coefficients of variation were less than 7%. The limit of detection and the lower limit of quantification were 0.1 and 0.2 µg/mL, respectively. Recoveries of lapatinib from plasma were higher than 86.7% in all cases. The assay was applied to the determination of the drug in the plasma of 2 cancer patients receiving lapatinib, 1000 and 1250 mg orally, and could be useful for therapeutic drug monitoring of lapatinib in routine clinical practice.

Effect of grapefruit juice on the pharmacokinetics of docetaxel in cancer patients: a case report

Docetaxel is a taxane antineoplasic agent that acts by inducing microtubular stability and disrupting the dynamics of the microtubular network. It is approved for the adjuvant treatment of patients with breast cancer, non-small cell lung cancer (NSCLC), hormone refractory prostate cancer and gastric cancer (http://www.taxotere.com). Moreover, docetaxel is active against different types of solid tumours, including oesophageal squamous cell carcinoma and advanced squamous cell carcinoma of the head and neck [1]. Its dose limiting toxicity is neutropenia, peripheral neurotoxicity and oedema [2].

Evidence of Different Propofol Pharmacokinetics under Short and Prolonged Infusion Times in Rabbits

Propofol is an anaesthetic widely used in both human beings and animals. However, the characterization of propofol pharmacokinetics (PK) is not well understood when long‐term infusions are used. The main objective of this study was to explore the PK behaviour of propofol in a rabbit model during short and prolonged propofol infusions and to develop an internally validated PK model, for propofol dose individualization in the rabbit for future use. Population 1 (P1) was constituted by seven New Zealand rabbits and was used to characterize the PK profile of propofol at short infusions. Animals were anaesthetized with a bolus of 20 mg/kg, followed by an infusion rate of 50 mg/kg/hr of propofol at 1%, which was then maintained for 30 min. A second rabbit population (P2, n = 7) was sedated according to reflexes responses and Index of Consciousness values, for 20 consecutive hours using propofol 2% aiming at characterizing propofol behaviour at long‐term infusions. Clinical data and blood samples were collected at specific time‐points in both populations. Propofol plasma concentrations were determined by gas chromatography/ion trap mass spectrometry. The NONMEM VII software was used to evaluate the relationships between dose and plasma concentrations. A linear two‐compartment model with different central compartment volume and plasma clearance (separately modelled in the two populations) was the one that best described propofol concentrations. The time course of propofol plasma concentrations was well characterized by the PK model developed, which simultaneously accounts for propofol short‐ and long‐term infusions and can be used to optimize future PK studies in rabbits.

Population pharmacokinetics applied to optimising cisplatin doses in cancer patients

OBJECTIVE To develop and internally validate a population pharmacokinetics model for cisplatin and assess its prediction capacity for personalising doses in cancer patients. METHOD Cisplatin plasma concentrations in forty-six cancer patients were used to determine the pharmacokinetic parameters of a two-compartment pharmacokinetic model implemented in NONMEN VI software. Pharmacokinetic parameter identification capacity was assessed using the parametric bootstrap method and the model was validated using the nonparametric bootstrap method and standardised visual and numerical predictive checks. The final models prediction capacity was evaluated in terms of accuracy and precision during the first (a priori) and second (a posteriori) chemotherapy cycles. RESULTS Mean population cisplatin clearance is 1.03 L/h with an interpatient variability of 78.0%. Estimated distribution volume at steady state was 48.3 L, with inter- and intrapatient variabilities of 31,3% and 11,7%, respectively. Internal validation confirmed that the population pharmacokinetics model is appropriate to describe changes over time in cisplatin plasma concentrations, as well as its variability in the study population. The accuracy and precision of a posteriori prediction of cisplatin concentrations improved by 21% and 54% compared to a priori prediction. CONCLUSION The population pharmacokinetic model developed adequately described the changes in cisplatin plasma concentrations in cancer patients and can be used to optimise cisplatin dosing regimes accurately and precisely.

Estudio de estabilidad de oxaliplatino y doxorrubicina para su administración intraperitoneal con hipertermia

Objective: To evaluate the in vitro physicochemical stability of oxaliplatin anddoxorubicin when the in vivo hyperthermic intraperitoneal conditions arereproduced.Methods: Three solutions were prepared, A (oxaliplatin 200 mg/L), B(doxorubicin 15 mg/L) and C (oxaliplatin 200 mg/L with doxorubicin 15mg/L) in glucose 5%. The three solutions were subjected to the maximumtemperature reached in vivo (49° C) for two hours. Physical stability wasfocused on visual control of particles or precipitates in solutions, dischargeof gases, odor and color. Samples were taken every 15 minutes and thechemical stability was evaluated by determining the concentration of oxaliplatinand doxorubicin remaining in the samples. Oxaliplatin concentrationswere determined by atomic absorption graphite chamber whiledoxorubicin was determined by high performance liquid chromatography.The chemical stability criteria selected was the one described by the AmericanPharmacopoeia, which sets a permissible variation range betweenthe 90-110% of the initial concentration.Results: During the assay there was no appearance of particles, precipitatesin the samples, discharge of gases, nor colour changes in the solutions. Thesamples showed a remaining concentration of oxaliplatin and doxorubicinwithin the 90-110% limit. The stability of the samples that follow to twocycles of freeze-thaw after hyperthermia was also found within the specifiedlimits.Conclusion: A, B and c solutions in 5% glucose, are physically and chemicallystable at 49° C for two hours. Under these conditions, these solutionscould be used with guarantees of stability in patients with peritoneal carcinomatosissubsidiary of intraperitoneal hyperthermic chemotherapybased in these antineoplastic agents.

Stability study of oxaliplatin and doxorubicin for intraperitoneal administration with hyperthermia

OBJECTIVE To evaluate the in vitro physicochemical stability of oxaliplatin and doxorubicin when the in vivo hyperthermic intraperitoneal conditions are reproduced. METHODS Three solutions were prepared, A (oxaliplatin 200 mg/L), B(doxorubicin 15 mg/L) and C (oxaliplatin 200 mg/L with doxorubicin 15mg/L) in glucose 5%. The three solutions were subjected to the maximum temperature reached in vivo (49° C) for two hours. Physical stability was focused on visual control of particles or precipitates in solutions, discharge of gases, odor and color. Samples were taken every 15 minutes and the chemical stability was evaluated by determining the concentration of oxaliplatin and doxorubicin remaining in the samples. Oxaliplatin concentrations were determined by atomic absorption graphite chamber while doxorubicin was determined by high performance liquid chromatography.The chemical stability criteria selected was the one described by the American Pharmacopoeia, which sets a permissible variation range between the 90-110% of the initial concentration. RESULTS During the assay there was no appearance of particles, precipitates in the samples, discharge of gases, nor colour changes in the solutions. The samples showed a remaining concentration of oxaliplatin and doxorubicin within the 90-110% limit. The stability of the samples that follow to two cycles of freeze-thaw after hyperthermia was also found within the specified limits. CONCLUSION A, B and c solutions in 5% glucose, are physically and chemically stable at 49° C for two hours. Under these conditions, these solutions could be used with guarantees of stability in patients with peritoneal carcinomatosis subsidiary of intraperitoneal hyperthermic chemotherapy based in these antineoplastic agents.

Populational pharmacokinetics of doxorubicin applied to personalised its dosing in cancer patients

OBJECTIVE To develop and internally validate a population pharmacokinetic model for doxorubicin and to evaluate its predictive performance for dose individualization in cancer patients. METHODS Doxorubicin plasma concentrations were determined in thirty-three cancer patients treated with intravenous doxorubicin. A three-compartment pharmacokinetic model was implemented in the NONMEN VI programme to determine the doxorubicin pharmacokinetic parameters. The identifiability of the parameters was assessed by parametric bootstrap and model validation was performed using nonparametric bootstrap, visual predictive check, and numerical predictive check. The final models predictive performance was evaluated in terms of accuracy and precision of plasma concentration predictions during the first and second cycles of chemotherapy. RESULTS Doxorubicin clearance was 58.8 L/h, with interpatient variability of 29.2% and intrapatient variability of 18.9%. The estimated volume of distribution at steady state was 2294 L, with inter-and intrapatient variability of 7.3% and 26.1%, respectively. Internal validation confirmed that the population pharmacokinetic model is appropriate to describe the time course of the doxorubicin plasma concentrations and its variability in this population. The accuracy and precision of an a posteriori prediction of doxorubicin plasma concentrations improved by 63% and 41% compared to the a priori prediction. CONCLUSION The Bayesian population pharmacokinetic model characterised the time course of doxorubicine plasma concentrations and can be accurately and precisely used to optimise doxorubicine dosing regimens in cancer patients.

Epidermal growth factor receptor (EGFR) mutations in a series of non-small-cell lung cancer (NSCLC) patients and response rate to EGFR-specific tyrosine kinase inhibitors (TKIs).

IntroductionEpidermal growth factor receptor (EGFR) mutation related to tyrosine kinase inhibitors’ (TKIs) responsiveness in non-small cell lung cancer (NSCLC) has become an important issue for therapeutic decision-making in NSCLC patients.Material and methodsSixty-nine Caucasian NSCLC patients were screened for mutations in the tyrosine kinase (TK) domain of EGFR by direct sequencing from December 2005 to September 2010.ResultsActivating mutations in the EGFR TK domain were found in 8 of 69 (11.6%) (7 deletions in exon 19 and one L858R mutation in exon 21). Seven of those mutations were found in adenocarcinoma and one mutation in bronchiolo-alveolar carcinoma; five of them in females (one smoker) and three of them in males (one smoker). All patients carrying activating mutations in the TK domain of EGFR were treated with TKIs. Ten patients not carrying an activating mutation in EGFR, who progressed after chemotherapy, were also treated with compassionate use of EGFR-specific TKIs (gefitinib or erlotinib). An objective response (partial response) was observed in all patients carrying an activating mutation in EGFR that received TKIs. Median overall survival for these patients has not been reached, however mean survival has been estimated at 39.5 months (95% CI, 22–57).ConclusionsAs previously reported, EGFR TK mutational analysis was a predictive test for response to targeted therapy with EGFR TKIs. The early identification of these patients consistently attains disease response and clearly improves outcomes.