A pharmacokinetic and pharmacogenetic study of efavirenz in children: dosing guidelines can result in subtherapeutic concentrations.

BACKGROUND: Our main objectives were to study the population pharmacokinetics of efavirenz and to explore the adequacy of dosing guidelines. METHODS: A total of 33 HIV-1-infected patients were recruited from the Emma Children's Hospital (Amsterdam, the Netherlands). Gender, age, drug formulation, the presence of the c.516G>T polymorphism in the CYP2B6 gene and the quantitation of liver enzymes alanine aminotransferase and aspartate aminotransferase at baseline were collected. A non-linear mixed effect pharmacokinetic model was developed. RESULTS: CYP2B6 genotype and drug formulation significantly influenced efavirenz pharmacokinetics. Clearance was 29.7% lower in children carrying the CYP2B6-516-G/T genotype compared with children carrying the G/G genotype. Relative bioavailiability of the oral liquid compared with tablets or capsules was 46.6%. Children carrying the CYP2B6-516-G/G genotype had a 50-70% probability of developing a subtherapeutic trough level of efavirenz and only 1-3% probability of developing a trough level >4 mg/l. To reduce the probability of developing a subtherapeutic trough concentration, we propose to give an adult efavirenz dose to children weighing > or =25 kg and to allometrically scale doses for other weight levels a priori. The dose of the oral solution should be twice the dose of capsules. CONCLUSIONS: Population pharmacokinetics of efavirenz in children were adequately described. Current dosing guidelines can result in subtherapeutic concentrations in children carrying the CYP2B6-516-G/G genotype and with the liquid formulation. A priori dose adaptations in the paediatric population seem feasible and need prospective validation.

Antiviral activity of HIV type 1 protease inhibitors nelfinavir and indinavir in vivo is not influenced by P-glycoprotein activity on CD4+ T cells.

P-glycoprotein (P-gp) can compromise the antiretroviral effect of a protease inhibitor (PI)-containing regimen for HIV-1, but can also reduce HIV-1 replication. We studied the net effect of P-gp on the intracellular HIV-1 RNA and DNA load in vivo. CD4(+) T cells were isolated from 27 HIV-1 patients (13 without and 14 with a PI-containing regimen) and subsequently sorted in CD45RO(-) (naive) and CD45RO(+) (memory) subsets with either high (P-gp(high)) or low (P-gp(low)) P-gp activity. Unspliced HIV-1 RNA and HIV-1 DNA load were determined. For each patient P-gp(high) and P-gp(low) subsets were compared. In patients on a PI-containing regimen, intracellular unspliced HIV-1 RNA was significantly lower in P-gp(high)-naive CD4(+) cells compared to P-gp(low)-naive CD4(+) cells (p = 0.04). The same trend was seen in naive CD4(+) cells of treatment naive patients. In both treated and untreated patients HIV-1 DNA levels were significantly lower in P-gp(high) than in P-gp(low) memory CD4(+) cells (p = 0.02 and p = 0.04). High cellular P-gp activity coincided with a reduced intracellular HIV-1 load in vivo, both in therapy-naive and in PI-treated patients. Therefore we conclude that the potential efflux function of P-gp on PIs may be clinically less relevant than the effect of P-gp on intracellular HIV-1 replication.

Population pharmacokinetics and pharmacodynamics of nelfinavir and its active metabolite M8 in HIV-1-infected children.

BACKGROUND: The objectives of this study are to develop and validate a population pharmacokinetic model that adequately describes the pharmacokinetics of nelfinavir and its active metabolite M8 in HIV-1-infected children; to define factors involved in the pharmacokinetic variability, which could aid in defining dosing strategies; and to correlate the pharmacokinetics to the treatment response. METHODS: Protease inhibitor-naive, HIV-1-infected children were included. A population pharmacokinetic model of nelfinavir and M8 was developed using NONMEM. Bayesian analysis was used to estimate pharmacokinetic values. A pharmacokinetic-pharmacodynamic analysis was performed to study relationships between these values and the virologic response to therapy. RESULTS: From 38 children, 724 nelfinavir and 636 M8 plasma concentrations were available. The pharmacokinetics of both compounds were described simultaneously with a one-compartment model with first-order elimination. Clearance (CL/F) and volume of distribution (V/F) were 32.6 L/h (interindividual variability [IIV]: 31.6%) and 281 L/h (IIV: 29.7%) for nelfinavir and 86.2 L/h (IIV: 43.1%) and 42.3 L/h for M8. No factors could be defined that affected the pharmacokinetics of nelfinavir or M8. The overall virologic response was 78% (HIV-1 RNA <500 copies/mL, on-treatment analysis). No differences in exposure to nelfinavir and M8 were observed between responders and nonresponders. The only factor distinguishing the two groups was a higher baseline HIV-1 RNA concentration in nonresponders. CONCLUSION: A model was developed and validated that adequately described the population pharmacokinetics of nelfinavir and M8 in a childhood population. No factors affecting dosing strategies were identified, and no correlation could be demonstrated between the exposure to nelfinavir and M8 and the virologic treatment response.

Multidrug resistance protein 2 (MRP2) transports HIV protease inhibitors, and transport can be enhanced by other drugs.

BACKGROUND: Various drug transporters of the ATP-binding cassette (ABC) family restrict the oral bioavailability and cellular, brain, testis, cerebrospinal fluid and fetal penetration of substrate drugs. MDRI P-glycoprotein (P-gp) has been demonstrated to transport most HIV protease inhibitors (HPI) and to reduce their oral bioavailability and lymphocyte, brain, testis and fetal penetration, possibly resulting in major limiting effects on the therapeutic efficacy of these drugs. OBJECTIVES: To investigate whether the ABC transporters MRP1, MRP2, MRP3, MRP5 and breast cancer resistance protein 1 (Bcrp1) are efficient transporters of the HPI saquinavir, ritonavir and indinavir. METHODS: Polarized epithelial non-human (canine) cell lines transduced with human or murine complementary DNA (cDNA) for each of the transporters were used to study transepithelial transport of the HPI. RESULTS: MRP2 efficiently transported saquinavir, ritonavir and indinavir and this transport could be enhanced by probenecid. Sulfinpyrazone was also able to enhance MRP2-mediated saquinavir transport. In contrast, MRP1, MRP3, MRP5, or Bcrp1 did not efficiently transport the HPI tested. CONCLUSIONS: Human MRP2 actively transports several HPI and could, based on its known and assumed tissue distribution, therefore reduce HPI oral bioavailability. It may also limit brain and fetal penetration of these drugs and increase their hepatobiliary, intestinal and renal clearance. MRP2 function and enhancement of its activity could adversely affect the therapeutic efficacy, including the pharmacological sanctuary penetration, of HPI. In vivo inhibition of MRP2 function might, therefore, improve HIV/AIDS therapy.

Drug-drug interaction between itraconazole and the antiretroviral drug lopinavir/ritonavir in an HIV-1-infected patient with disseminated histoplasmosis.

We describe a drug-drug interaction between coformulated lopinavir/ritonavir and itraconazole in a patient infected with human immunodeficiency virus type 1 who had disseminated histoplasmosis. Coadministration of these agents led to a strong increase in itraconazole concentrations and a decrease in concentrations of its metabolite, hydroxyitraconazole, which is equally active pharmacologically. The dosage of itraconazole was reduced when it was used in combination with lopinavir/ritonavir.

The plasma and intracellular steady-state pharmacokinetics of lopinavir/ritonavir in HIV-1-infected patients.

Therapeutic drug monitoring of protease inhibitors (PIs) is usually performed on plasma samples although their antiretroviral effect takes place inside cells. Little is known, however, about the intracellular accumulation and related plasma pharmacokinetics of PIs such as lopinavir/ritonavir (LPV/RTV). Therefore, we studied the plasma and intracellular (cell-associated) steady-state pharmacokinetics of this PI combination in a dosage of 400/100 mg twice daily in a non-randomized cohort of HIV-1-infected individuals. Plasma (0-12 h) and peripheral blood mononuclear cell (PBMC; 0-8 h) samples were drawn during a 12-h dosing interval in 11 subjects. The plasma concentrations versus time curves of LPV and RTV were characterized by an irregular absorption phase showing double-peaks (Cmax) in most subjects and single-peaks in the remaining patients between 1 and 3 h after drug intake. Pre-dose concentrations of both agents in plasma were significantly higher than the concentrations at the end of the dosing interval indicating the presence of a circadian rhythm in their pharmacokinetics. The course of the intracellular concentrations versus time curves appeared to be similar to the plasma concentration curves, with the highest intracellular concentration measured 3 h after drug intake. The intracellular RTV concentrations were higher than reported in vitro EC50 values and might therefore contribute to the antiretroviral effect of LPV/RTV. The median intracellular-to-plasma concentration ratios (interquartile range) were 1.18 (0.74-2.06) and 4.59 (3.20-7.70) for LPV and RTV, respectively. In conclusion, both LPV and RTV accumulate to potential therapeutic concentrations in PBMCs. Irregular absorption and circadian plasma clearance patterns were observed for the PI combination LPV/RTV.

Practical guidelines to interpret plasma concentrations of antiretroviral drugs.

Several relationships have been reported between antiretroviral drug concentrations and the efficacy of treatment, and toxicity. Therefore, therapeutic drug monitoring (TDM) may be a valuable tool in improving the treatment of HIV-1-infected patients in daily practice. In this regard, several measures of exposure have been studied, e.g. trough and maximum concentrations, concentration ratios and the inhibitory quotient. However, it has not been unambiguously established which pharmacokinetic parameter should be monitored to maintain optimal viral suppression. Each pharmacokinetic parameter has its pros and cons. Many factors can affect the pharmacokinetics of antiretroviral agents, resulting in variability in plasma concentrations between and within patients. Therefore, plasma concentrations should be considered on several occasions. In addition, the interpretation of the drug concentration of a patient should be performed on an individual basis, taking into account the clinical condition of the patient. Important factors herewith are viral load, immunology, occurrence of adverse events, resistance pattern and comedication. In spite of the described constraints, the aim of this review is to provide a practical guide for TDM of antiretroviral agents. This article outlines pharmacokinetic target values for the HIV protease inhibitors amprenavir, atazanavir, indinavir, lopinavir, nelfinavir, ritonavir and saquinavir, and the non-nucleoside reverse transcriptase inhibitors efavirenz and nevirapine. Detailed advice is provided on how to interpret the results of TDM of these drugs.

Effect of mycophenolate mofetil on the pharmacokinetics of antiretroviral drugs and on intracellular nucleoside triphosphate pools.

OBJECTIVE: To study the effect of mycophenolate mofetil therapy on the pharmacokinetic parameters of a number of antiretroviral drugs, on intracellular pools of deoxycytidine triphosphate (dCTP) and deoxyguanosine triphosphate (dGTP), and on intracellular concentrations of the triphosphate of lamivudine (3TCTP). DESIGN: Randomised pharmacokinetic study. PARTICIPANTS: Nineteen HIV-1-infected patients. METHODS: Antiretroviral-naive men starting treatment with didanosine 400 mg once daily, lamivudine 150 mg twice daily, abacavir 300 mg twice daily, indinavir 800 mg twice daily, ritonavir 100 mg twice daily and nevirapine 200 mg twice daily were randomised to a group with or without mycophenolate mofetil 500 mg twice daily. After 8 weeks of therapy, the plasma pharmacokinetic profiles of mycophenolic acid (the active metabolite of mycophenolate mofetil), abacavir, indinavir and nevirapine, and triphosphate concentrations (dCTP, dGTP and 3TCTP) in peripheral blood mononuclear cells, were determined. RESULTS: Nine of the 19 patients received mycophenolate mofetil. There was no difference in plasma clearance of indinavir or abacavir between the two groups. The clearance of nevirapine was higher in patients using mycophenolate mofetil (p = 0.04). In 12 patients, of whom five also received mycophenolate mofetil, intracellular triphosphates were measured. There was no significant difference in intracellular dCTP, dGTP or 3TCTP concentrations between the two groups. CONCLUSION: In this small cohort of patients, mycophenolate mofetil therapy reduced the plasma concentration of nevirapine but had no effect on plasma concentrations of indinavir and abacavir. There were no consistent effects of mycophenolic acid on the intracellular concentrations of dCTP, dGTP or 3TCTP.

Liquid chromatographic assay for the antiviral nucleotide analogue tenofovir in plasma using derivatization with chloroacetaldehyde.

A sensitive and selective reversed-phase liquid chromatographic assay for tenofovir in human plasma has been developed and validated. Tenofovir was isolated from a 200 microl plasma sample using protein precipitation with trichloroacetic acid. The fluorescent 1,N(6)-etheno derivative is formed at 98 degrees C in the buffered extract with chloroacetaldehyde. This derivative was analysed using gradient ion-pair liquid chromatography and fluorescence detection at 254 nm for excitation and 425 nm for emission. In the evaluated concentration range (20-1000 ng/ml), the intra-day precision was 4% and the inter-day precision was 5-6%. An accuracy of between 97 and 110% was determined. The lower limit of quantification was 20 ng/ml with an inter-day precision of 11%, an intra-day precision of 12% and an accuracy of 103%. The assay is subject to interference from co-administered abacavir. The usefulness of the assay was demonstrated for samples obtained from an HIV-infected patient treated with tenofovir.

Selective high-performance liquid chromatographic assay for itraconazole and hydroxyitraconazole in plasma from human immunodeficiency virus-infected patients.

A sensitive and selective reversed-phase liquid chromatographic assay for itraconazole and hydroxyitraconazole in human plasma has been developed and validated. Itraconazole and hydroxyitraconazole were extracted from the matrix using solid-phase extraction on a strong cation-exchange sorbent. All compounds were detected using fluorescence at 265 and 363 nm for excitation and emission, respectively. The assay has been validated over the range 10-1,000 ng/ml for both compounds, 10 ng/ml being the lower limit of quantification. Accuracies ranged from 104 to 113% for itraconazole and from 91 to 103% for hydroxyitraconazole. The intra-assay precisions were all below 9% for itraconazole and below 8% for hydroxyitraconazole. The selectivity has been evaluated with respect to all registered anti-human immunodeficiency virus (HIV) drugs and other potential co-medications and a few of their metabolites, commonly used by HIV-infected individuals. Both itraconazole and hydroxyitraconazole were stable under relevant conditions for HIV-inactivation and storage of samples. The applicability of the assay was demonstrated for samples collected from a treated HIV-infected patient.

Liquid chromatographic assay for the protease inhibitor atazanavir in plasma.

Atazanavir is the most recently introduced protease inhibitor for the suppression of the anti-human immunodeficiency virus. A sensitive and selective reversed-phase liquid chromatographic assay for this drug in human plasma has been developed and validated. Atazanavir was isolated from a 500 microL plasma sample using liquid-liquid extraction with dichloromethane. After evaporation and reconstitution of the extract the sample was analysed using liquid chromatography and ultraviolet detection at 280 nm. In the evaluated concentration range (44-4395 ng/mL atazanavir), intra-day precisions were < or =7% and inter-day precisions were < or =14%. Accuracies between 96 and 106% were found. The lower limit of quantification was 44 ng/mL with an intra-day precision of 7%, an inter-day precision of 14% and an accuracy of 87%. There was no interference from 32 tested potentially co-administrated drugs and metabolites. The usefulness of the assay was demonstrated for samples obtained from an HIV-infected patient treated with atazanavir.

Liquid chromatographic assay for the non-peptidic protease inhibitor tipranavir in plasma.

Tipranavir is the most recently introduced protease inhibitor for the suppression of the human immunodeficiency virus (HIV). A selective reversed-phase liquid chromatographic assay, previously developed for atazanavir, has been extended and validated for tipranavir in plasma. Compounds were isolated from a 500 microL plasma sample using liquid-liquid extraction with dichloromethane. After evaporation and reconstitution of the extract the sample was analysed using reversed-phase liquid chromatography and ultra violet detection at 280 nm. In the evaluated concentration range (0.2-50 microg/mL tipranavir), intra-day precisions were

Development and validation of a population pharmacokinetic model for ritonavir used as a booster or as an antiviral agent in HIV-1-infected patients.

AIMS: The aim of this study was to develop and validate a population pharmacokinetic model of ritonavir, used as an antiviral agent or as a booster, in a large patient population and to identify factors influencing its pharmacokinetics. METHODS: Ambulatory HIV-1-infected patients from the outpatient clinic of the Slotervaart Hospital, Amsterdam, the Netherlands, who were being treated with a ritonavir-containing regimen were included. During regular visits, blood samples were collected for the determination of ritonavir plasma concentrations and several clinical chemistry parameters. Furthermore, complete pharmacokinetic curves were available in some patients. Single and multiple compartment models with zero-order and first-order absorption, with and without absorption lag-time, with linear and nonlinear elimination were tested, using nonlinear mixed effect modelling (NONMEM). Pharmacokinetic parameters and interindividual, interoccasion and residual variability were estimated. In addition, the influence of several factors (e.g. patient characteristics, comedication) on the pharmacokinetics of ritonavir was explored. RESULTS: From 186 patients 505 ritonavir plasma concentrations at a single time-point and 55 full pharmacokinetic profiles were available, resulting in a database of 1228 plasma ritonavir concentrations. In total 62% of the patients used ritonavir as a booster of their protease inhibitor containing antiretroviral regimen. First order absorption in combination with one-compartment disposition best described the pharmacokinetics of ritonavir. Clearance, volume of distribution and absorption rate constant were 10.5 l h(-1) (95% prediction interval (95% PI) 9.38-11.7), 96.6 l (95% PI 67.2-121) and 0.871 h(-1) (95% PI 0.429-1.47), respectively, with 38.3%, 80.0% and 169% interindividual variability, respectively. The interoccasion variability in the apparent bioavailability was 59.1%. The concomitant use of lopinavir resulted in a 2.7-fold increase in the clearance of ritonavir (P value < 0.001). No patients characteristics influenced the pharmacokinetics of ritonavir. CONCLUSIONS: The pharmacokinetic parameters of ritonavir were adequately described by our population pharmacokinetic model. Concomitant use of the protease inhibitor lopinavir strongly influenced the pharmacokinetics of ritonavir. The model has been validated and can be used for further investigation of the interaction between ritonavir and other protease inhibitors.