Lopinavir/ritonavirpopulation pharmacokineticsin neonates and infantsSaik Urien,1,2,3 Ghislaine Firtion,4 Suzanne T. Anderson,5,11

Deborah Hirt,1,2,3 Caroline Solas,7 Gilles Peytavin,8 Albert Faye,9

Isabelle Thuret,10 Marthe Leprevost,6 Carole Giraud,1,2,3

Hermione Lyall,11 Saye Khoo,12 Stéphane Blanche1,3,13 &Jean-Marc Tréluyer1,2,3,14

1EA 3620, Université Paris Descartes, 2Unité de Recherche Clinique Paris Centre Descartes, Groupe

Hospitalier Cochi – Saint Vincent de Paul, Assistance Publique Hôpitaux de Paris, 3Premup et CIC 0901,

INSERM, 4Néonatologie, Groupe Hospitalier Cochin – Saint Vincent de Paul, Assistance Publique

Hôpitaux de Paris, Paris, France, 5Department of Medicine, Brighton and Sussex Medical School,

Falmer, Sussex BN1 9PS, 6Northwick Park Hospital, North West London Hospitals NHS Trust, Middlesex,

UK, 7Laboratoire de Pharmacocinétique, Hôpital La Timone, Marseille, 8Service de Pharmacie Clinique,

Hôpital Bichat Claude Bernard, Assistance Publique Hôpitaux de Paris, Paris, 9Pédiatrie Générale,

Hôpital Robert Debré, Assistance Publique Hôpitaux de Paris, Paris, 10Pédiatrie, Hôpital La Timone,

Marseille, France, 11Imperial College Healthcare NHS Trust, London, 12Department of Pharmacology at

the University of Liverpool, Liverpool, UK, 13Immuno-Hématologie, Hôpital Necker Enfants Malades,

Assistance Publique Hôpitaux de Paris and 14Pharmacologie Clinique, Groupe Hospitalier Cochin –

Saint Vincent de Paul, Assistance Publique Hôpitaux de Paris, Paris, France

CorrespondenceDr Saik Urien MD PhD, Unité deRecherche Clinique, Groupe HospitalierCochin Saint Vincent de Paul, HopitalTarnier, 89 rue d’Assas, Paris 75006,France.Tel.: + 33 1 5841 2880Fax: + 33 1 5841 1183E-mail: saik.urien@svp.aphp.fr----------------------------------------------------------------------

Keywordschildren, lopinavir/ritonavir, neonates,population pharmacokinetics----------------------------------------------------------------------

Received25 October 2010

Accepted10 January 2011

WHAT IS ALREADY KNOWN ABOUTTHIS SUBJECT• Lopinavir/ritonavir pharmacokinetics have

been fully investigated in adults and children.

WHAT THIS STUDY ADDS• Lopinavir/ritonavir population

pharmacokinetics in 96 neonates and infantsfrom birth to less than 2 years (1.16 to 10.4 kg)showed that CL/F and V/F were dependent onbody weight on an allometric basis andpost-menstrual age.

AIMSBecause of immature hepatic metabolism, lopinavir could presentspecific pharmacokinetics in the first weeks of life. We aimed atdetermining the optimal dosing regimen in neonates and infantsweighing 1 to 10.5 kg.

METHODSLopinavir/ritonavir (LPV/r) pharmacokinetics were studied in 96 infantsusing a population approach.

RESULTSA one compartment model described LPV/r pharmacokinetics.Normalized to a 70 kg adult using allometry, clearance (CL/F) anddistribution volume (V/F) estimates were 5.87 l h-1 70 kg-1 and91.7 l 70 kg-1. The relative bioavailabilty, F, increased withpost-menstrual age (PMA) and reached 50% of the adult value at 39.7weeks.

CONCLUSIONSSize and PMA explained some CL/F and V/F variability inneonates/infants. Based upon trough concentration limitations,suggested LPV/r dosing regimens were 40 mg 12 h-1, 80 mg 12 h-1 and120 mg 12 h-1 in the 1–2 kg, 2–6 kg and 6–10 kg group, respectively.

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British Journal of ClinicalPharmacology

DOI:10.1111/j.1365-2125.2011.03926.x

Br J Clin Pharmacol / ••:•• / ••–•• / 1© 2011 The AuthorsBritish Journal of Clinical Pharmacology © 2011 The British Pharmacological Society

Introduction

The protease inhibitor lopinavir/ritonavir (LPV/r) has beenshown to be effective in children. Because of the availabiltyof the oral formulation, lopinavir is administered to neo-nates having a high risk of perinatal or postnatal mother-to-child transmission of HIV. In infants and children, studieshave shown a fast elimination half-life of LPV/r as comparedwith adults [1–4]. LPV is a substrate of CYP3A4, immature inneonates compared with infants and children,and could beassociated with a specific pharmacokinetic profile in thefirst weeks after birth [5]. Therefore the objective of thisstudy was to assess LPV/r dosing recommendations in neo-nates and infants weighing 1 to 10 kg by developing apopulation pharmacokinetic model, including the investi-gation of weight,gestational age and postnatal age effects.

Methods

Patients from the hospitals and institutions correspondingto authors’ affiliation sites received the liquid formulation ofLPV/r for the prevention of mother-to-child HIV transmis-sion.For the youngest infants or when necessary, LPV/r wasadministered directly in the back of the mouth using a smallsyringe. The antiretroviral therapy was monitored on aroutine basis. For each sampling, the drug dosing regimen,the time from the last dosing, body weight, post-natal age(PNA),gestational age (GA) and combined treatments wererecorded. The assays for lopinavir were performed on fourdifferent sites: site 1 Saint Vincent de Paul Paris; site 2 UKsites;site 3 Hopital Tenon Paris;site 4,Marseille,according tovalidated HPLC methods [4].The quantification limit of themethod was 0.05 mg l-1, the inter-assay precision and biasbeing 7% in the calibration range of 0.1–20 mg l-1.

Data were analyzed using the nonlinear mixed effectmodelling software program Monolix version 31s [6](http://wfn.software.monolix.org). Parameters were esti-mated by computing the maximum likelihood estimator ofthe parameters without any approximation of the model(no linearization) using the stochastic approximationexpectation maximization (SAEM) algorithm combinedwith a MCMC (Markov Chain Monte Carlo) procedure. Thenumber of MCMC chains was fixed to 15 for all estimations.A proportional model was used to describe the residualvariability (e), and the between-subject variabilities (BSV orh) were ascribed to an exponential model. Parametershrinkage was calculated as [1–SD(h)/w], where SD(h) andw are the SD of individual h parameters and the populationmodel estimate of the BSV, respectively. The LikelihoodRatio Test (LRT) including the log-likelihood, the Akaikeinformation criterion (AIC) and the Bayesian informationcriterion (BIC) were used to test different hypothesesregarding the final model, covariate effect(s) on pharma-cokinetic parameter(s), residual variability model (propor-tional vs. proportional plus additive error model), and

structure of the variance-covariance matrix for the BSVparameters.

Main covariates of interest in the population werepostnatal age (PNA), body weight, postmenstrual age(PMA = PNA + GA) and the formulation. Pharmacokineticparameters, V and CL, were standardized (STD) for a meanstandard bodyweight using the ‘1/4 power’ allometricmodels, Vi = VSTD ¥ (BWi/BWSTD)1 and CLi = CLSTD ¥ (BWi/BWSTD)3/4 for the ith patient, Vi. CLi and BWi are the param-eters and bodyweight of the ith individual [7, 8].

Age-related change functions for CL or V as a functionof PNA or post-menstrual age (PMA = GA + PNA) havebeen described in detail [7]. Other covariates were genderand combined drugs including other antiretroviral agents.

The model was validated by simulation techniques, thenormalized prediction distribution errors (npde) metrics[6]. Diagnostic plots and statistics were obtained using theR program [9].

Results

Among the 96 neonates/infants (46 boys, 50 girls), sevenwere treated for neonatal HIV infection. The median post-natal age of these infants was 2 weeks (range 1 day–102 weeks; interquartile range 3 days–14 weeks), themedian gestational age was 38 weeks (range 27.3–41 weeks), the median body weight and body surface areawere 3.3 kg (range 1.16–10.4 kg, interquartile range 2.73–5 kg) and 0.22 m2 (range 0.13–0.48 m2, interquartile range0.2–0.28 m2). A total of 163 LPV concentrations were avail-able. All samples were at steady-state except one (corre-sponding to a 1 day newborn). Sampling was performedon more than one occasion for 34 patients. Patients (n)received LPV/r twice (n = 73), thrice (n = 19) or once a day(n = 4). The mean doses were 39 mg kg-1 24 h-1 (range11–110) and 590 mg m-2 24 h-1 (range 106–1454). Sincethere were only three concentrations below the limit ofquantification, they were fixed at LOQ/2.

A one-compartment model described the data. Esti-mates for Ka and its associated BSV were inaccurate (>100%relative standard error, %rse), so Ka and hKa were fixed to0.564 h-1 and zero without any detrimental effect on thequality of the fit. The covariance term between hCL/F andhV/F was significant, the correlation coefficient was 0.84(rse 20%). Because multiple samples on a same occasionwere available for only four patients (four, four, four andtwo samples per patient), between-occasion variabilitywas also investigated. However, the BOV parameters on Ka

or V or CL were not accurate and dramatically increasedthe BIC value.

Weight and age were the main influential covariates.The weight effects on CL/F and V/F were then normalizedfor a 70 kg body weight adult. This size effect was signifi-cant (LRT test P = 0.0002), and the BSVs for CL/F and V/F,hCL/F and hV/F, decreased from 0.54 to 0.49 and from 0.66 to

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0.55, respectively. Further investigation of age on CL/F andV/F showed a similar decrease of CL/F and V/F as a functionof age, PMA or PNA (not shown). Given the similarity of thedecrease and the inflated CL/F70 and V/F70 values (relativeto expected values in a 70 kg adult) at the intercept, thisage effect was thought to influence the relative bioavail-ability, F, in these very young patients

F F= +( )ADULTPMA PMA PMA50

where FADULT is the reference bioavailability in adults, fixedto 1, and PMA50 is the PMA corresponding to F = 0.5. AICand BIC decreased by 8 and 3 units, respectively, and hCL/F

and hV/F decreased from 0.49 to 0.46 and from 0.55 to 0.46,respectively. The final model was then

CL l h kg BW− −( ) = × × ( )1 1 3 470 5 87 70. F

V Fl kg BW70 92 701 1−( ) = × × ( )

with

F = +( )PMA PMA 39 7.

where the CL, V and PMA50 typical values (% rse) are5.87 l h-1 70 kg-1 (23%), 92 l 70 kg-1 (25%) and 39.7 weeks(48%). The corresponding hCL/F and hV/F estimates (%rse)[shrinkage] were 0.46 (11%) [0.20] and 0.45 (41%) [0.34].The proportional residual variability estimate was 0.37 (rse18%).

Figure 1 shows the diagnostic plots for the final model.The mean, variance and distribution of the npde metrics(500 Monte Carlo simulations) were not significantly differ-ent from zero (Wilcoxon signed rank test, P = 0.92),1 (Fishervariance test, P = 0.74) and a normal distribution (Shapiro-Wilk test, P = 0.52). The parameters were well estimatedwith acceptable relative standard errors. Only V/F hadnotable shrinkage.

A dosage regimen was derived from the predictedtrough concentrations, Ctrough. The distribution of Ctrough

following 20 to 160 mg 12 h-1 or 40–320 mg 24 h-1 was

DV

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Figure 1Diagnostic plots for the final population pharmacokinetic model: Observed LPV/r concentrations (DV) vs. model-predicted concentrations (Pop_mean) andindividual predictions (Pred_ind_cond_mode) in mg l-1 and the corresponding normalized prediction distribution errors (npde) metrics vs. predictions orvs. time after dose. The dotted lines join the data from same individuals. The lines indicate the lines of unity (left, middle) and the y = 0 line (right). npdestatistics, mean and variance were not significantly different from zero (Wilcoxon signed rank test, P = 0.92) and 1 (Fisher variance test, P = 0.74) and thedistribution was not significantly different from a normal distribution (Shapiro-Wilk test of normality, P = 0.52)

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Short report

Br J Clin Pharmacol / ••:•• / 3

studied by 500 Monte Carlo simulations of the final popu-lation model. Figure 2 shows the probability of observingCTROUGH values between 1 mg l-1 and 8 mg l-1, therapeuticrange, and above 8 mg l-1, toxic range, in different bodyweight groups.

Discussion

In this neonate/infant cohort, weighing 1–10.5 kg, LPV/rpharmacokinetics were satisfactorily described by a onecompartment model, including size effects on pharmaco-kinetic parameters. The allometric body weight scaling ofCL and V plus the effect of PMA on the relative bioavail-ability decreased some variability associated with CL.However, the physiological circumstances in very younginfants and neonates probably alter the relationshipbetween body weight and clearance and volume of dis-tribution, explaining the moderate decrease of clearanceBSV. The variabilities, BSV and residual, remained at highlevels, as previously observed in other childhood studies[1, 2, 4]. In adults, the residual variability, approximatedto 0.21 (from the additive 1.15 mg l-1 and proportional0.075 components reported, 0.21 = square root ((1.15/6)2 + 0.0752) was lower and BSVs were estimated via a full

variance-covariance matrix, explaining in part a lowerBSV(CL) value, 0.17 [3]. The high correlation betweenCL/F and V/F BSVs (r = 0.86) also supports the importanceof bioavailability to explain these high variabilities. Drugintake, with/without concomitant formula milk in infants,as well as the amount of simultaneous milk intake maywidely vary. The model predicts that F will be in theorder (80%) of the adult value at 160 weeks PMA or2.3 years PNA. Although the PMA effect on F decreasedmarginally the BSVs and significantly the AIC and BICcriteria, the final CL/F70 and V/F70 values extrapolatedto a 70 kg adult with F = 1 are similar to the adult esti-mates, 5.73 l h–1 and 61.6 l, respectively [3], whicht sup-ports this population pharmacokinetic model. Higherapparent CL and V values in the youngest infants of agroup have also been observed in other LPV/r childhoodstudies [1, 2].

Given the available LPV/r formulations, the highestprobabilities of Ctrough in the normal 1–8 mg l-1 range wereobtained with 40 mg 12 h-1 in the 1–2 kg group,80 mg 12 h-1 in the 2–6 kg group and 120 mg 12 h-1 in the6–10 kg group (note that in Kaletra*, 40 mg 12 h-1 of lopi-navir correspond to 10 mg 12 h-1 of ritonavir). The once aday regimens produced high probabilities, >40%, ofobserving Cmin <1 mg l-1.

Pro

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4 to 6 kg 6 to 10 kg

2 to 3 kg1 to 2 kg 3 to 4 kg

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20 40 80 120 16020 40 80 120 160

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Figure 2Probability of observing trough lopinavir concentrations, Ctrough in the therapeutic range, between 1–8 mg l-1 (solid lines), below 1 mg l-1 (dashed lines) orabove 8 mg l-1 (toxic range, dotted lines) following administration of 20 to 160 mg 12 h-1 LPV/r in paediatric patients weighing from 1 to 10 kg. Predictedtrough lopinavir concentrations were obtained from 500 Monte Carlo simulations of the final model for each dosage

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Competing Interests

There are no competing interests to declare.The authors thank Abbott Laboratories, which partially

supported this study.

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Rodman JH, Chen J, Palumbo P, Serchuck L, Smith E,Hughes M. Pharmacokinetics, safety and efficacy ofLopinavir/ritonavir in infants less than 6 months of age: 24weeks results. AIDS 2008; 22: 249–55.

2 Verweel G, Burger DM, Sheehan NL, Bergshoeff AS, Warris A,van der Knaap LC, Driessen G, de Groot R, Hartwig NG. Plasmaconcentrations of the HIV-protease inhibitor lopinavir aresuboptimal in children aged 2 years and below. Antivir Ther2007; 12: 453–8.

3 Crommentuyn KM, Kappelhoff BS, Mulder JW, Mairuhu AT,van Gorp EC, Meenhorst PL, Huitema AD, Beijnen JH.Population pharmacokinetics of lopinavir in combinationwith ritonavir in HIV-1-infected patients. Br J Clin Pharmacol2005; 60: 378–89.

4 Jullien V, Urien S, Hirt D, Delaugerre C, Rey E, Teglas JP, Vaz P,Rouzioux C, Chaix ML, Macassa E, Firtion G, Pons G, Blanche S,Treluyer JM. Population analysis of weight-, age-, andsex-related differences in the pharmacokinetics of lopinavir inchildren from birth to 18 years. Antimicrob AgentsChemother 2006; 50: 3548–55.

5 Lacroix D, Sonnier M, Moncion A, Cheron G, Cresteil T.Expression of CYP3A in the human liver – evidence that theshift between CYP3A7 and CYP3A4 occurs immediately afterbirth. Eur J Biochem 1997; 247: 625–34.

6 Kuhn E, Lavielle M. Maximum likelihood estimation innonlinear mixed effects models. Comput Stat Data Anal 2005;49: 1020–30.

7 Anderson BJ, Holford NH. Mechanism-based concepts of sizeand maturity in pharmacokinetics. Annu Rev PharmacolToxicol 2008; 48: 303–32.

8 Comets E, Brendel K, Mentre F. Computing normalisedprediction distribution errors to evaluate nonlinearmixed-effect models: the npde add-on package for R. ComputMethods ••; ••: ••–••.

9 Ihaka R, Gentleman RR. A language for data analysis andgraphics. J Comput Graphic Stat 1996; 5: 299–314.

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