Enhanced Saquinavir Exposure in Human Immunodeficiency Virus Type 1-Infected Patients with Diarrhea and/or Wasting Syndrome

The protease inhibitor saquinavir was administered to 100 humanimmunodeficiency virus type 1 (HIV-1)-infected patients as asingle 600-mg oral dose (hard gelatin capsules) with a standardbreakfast, including 200 ml of grapefruit juice, during an open-labeltrial to assess whether diarrhea and/or wasting syndrome hasconsequences on its pharmacokinetics. Three groups of patientswere enrolled: group 1, asymptomatic patients (n = 30); group2, AIDS symptomatic patients without body weight loss or diarrhea(n = 37); and group 3, AIDS symptomatic patients with severebody weight loss and/or diarrhea (n = 33). Clinical and biologicaldata (covariates) were collected. A population approach wasperformed with three blood samples per patient to estimate themean population pharmacokinetic parameters (clearance [CL]/oralbioavailability [F], V/F, k_a, and lag time) and the derivedones (k_el, C_max, T_max, and area under the curve [AUC]). Therelationships between groups, exposure (i.e., estimated individualpost hoc AUCs), and covariates were explored by using multiplelinear regressions. A significant increase in median AUCs (165,349, and 705 ng · h · ml^-1 for groups 1, 2, and3, respectively [P < 0.0001]) was observed. The enhancementin saquinavir exposure could be due to the destruction of thetransporters in enterocytes and/or to the enlargement of theirtight junctions, allowing a paracellular crossing of saquinaviras the illness spreads. Because of grapefruit juice intake byevery patient, no implication of CYP3A4 could be assessed. Theseresults strongly suggest that, despite its low intrinsic oralbioavailability, saquinavir can be considered as a relevanttreatment for HIV-1-infected patients with diarrhea and/or wastingsyndrome. This must be evaluated in a long-term period.

INTRODUCTION

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Introduction

Intestinal troubles (diarrhea and malabsorption) are commonsymptoms during human immunodeficiency virus (HIV) infectionand are responsible for weight loss throughout the course ofthe illness (32). Several studies have suggested that theseintestinal dysfunctions are linked to HIV disease stages (18,28, 29, 32). Malabsorption, which has nutritional consequences(11), may have great pharmacological effects, particularly froma pharmacokinetic point of view. Moreover, it is usual to considerdiarrhea as a factor decreasing drug absorption, but this hasnot been demonstrated to be fact. Indeed, oral bioavailability(F), i.e., the absorption and first-pass effect-related pharmacokineticparameter, is expected to be disrupted in such clinical situations.However, there has been no alteration in F reported for someantiretrovirals, such as zidovudine, in AIDS patients with smallintestinal disease (SID), i.e., chronic diarrhea with wasting(36). The only change observed in such cases was a delayed absorption(prolonged time to reach the concentration peak [i.e., T_max])with respect to the total exposure of the drug: no significantdifference in areas under the concentration-time curves (AUCs)from time zero to time 6 h between patients with or withoutSID (17). Thus, it is possible that the absorption of drugswith relatively high bioavailability, such as zidovudine (15),is not significantly disrupted by chronic diarrhea with wastingcompared to that of drugs with low bioavailability at the samestage of disease.

The protease inhibitor (PI) saquinavir (SQV), in a hard gelatincapsule (HGC), has a kinetic profile characterized by a reducedabsorption and a high first-pass effect, resulting in the lowestbioavailability (F = 0.04 to 0.06) among the HIV type 1 (HIV-1)PIs available today (3, 27). It has not been pointed out, untilnow, whether patients with diarrhea and malabsorption couldexhibit infratherapeutic SQV concentrations in plasma. Moreover,SQV is known to display a high interindividual variability whenadministered to HIV-1-infected patients (30). This characteristic,associated with troubled absorption, should be taken into accountto either avoid resistance in cases of decreased drug exposureor lower potential side effects in cases of prolonged increasedexposure.

New AIDS patients naive to antiretrovirals and with diarrheaand/or wasting syndrome still exist today, particularly in Africaand Asia. Few studies have been published to date that examinethe absorption of major HIV oral medications, such as PIs, whichexhibit very low oral bioavailability in such clinical situations(5). Given the importance of understanding the effect of diarrheaand/or wasting on SQV oral absorption, we undertook a two-centerstudy (the Lariboisière and Paul Brousse hospitals inParis and Villejuif, France, respectively) that compares thepharmacokinetic behaviors of SQV in patients with and withoutdiarrhea and/or wasting syndrome. The goal of the present studyis to evaluate the consequences of diarrhea and/or wasting syndromeon the pharmacokinetics of SQV in HIV-1-infected patients inorder to assess the potential therapeutic relevance of thatPI in these patients.

MATERIALS AND METHODS

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Materials and Methods

Study design. In this open-label trial, which took place from 1996 to 1998,patients enrolled in either of the two centers during the sametime period (half of the patients in each center) and received,in addition to their usual antiretroviral treatment (19, 22),SQV-HGC (Produits Roche, Neuilly-sur-Seine, France) as a singleoral 600-mg dose immediately after a standard breakfast including200 ml of grapefruit juice to optimize SQV absorption. All doseswere checked to have been taken. To lower drug interactionswith SQV, the patients were chosen from an HIV-1-infected populationthat had never before received any PI. The study was approvedby the ethical committee of Paris-Saint-Louis Hospital, Paris,France. A written informed consent was obtained from each patientbefore starting the trial.

To perform the population pharmacokinetics study, three bloodsamples (5 ml each) were collected in vials containing lithiumheparinate from each patient, chosen randomly among 15 samplingtimes, i.e., 0, 0.25, 0.5, 1.0, 1.5, 2.0, 2.5, 3.0, 3.5, 4.0,5.0, 6.0, 8.0, 10.0, and 12.0 h after SQV-HGC administration,according to the following protocol. Each patient had one samplecollected randomly among three periods: 0 to 1.5 h, 2 to 4 h,and 5 to 12 h. Hence, each of the 15 points had six determinationsin each group. Plasma was separated from blood cells by centrifugationimmediately after collection. In accordance with good laboratorypractices, a decontamination procedure was set up by heatingplasma 30 min at 56°C. After they returned to room temperature,plasma samples were frozen at -20°C until analysis. Concentrationsof SQV in plasma were determined by a radioimmunoassay method(Huntington Life Sciences, Ltd., Huntington, Campbridgeshire,England) whose lower limit of quantification was 1 ng/ml (unpublishedmethod).

Patients. HIV-1-infected men and women over 18 years old, whatever theirrisk factors for HIV infection, treated or not treated withantiretroviral therapy (if so, no PI was taken), were enrolledin the present study for 21 months. Neither patients with changesin intestinal absorption as a result of intestinal surgery,severe renal failure (creatinine clearance < 20 ml ·min^-1), or hepatic insufficiency (prothrombin time < 60%)nor patients with signs of clinical or biological pancreatitisor known absorption disorders were included in this clinicaltrial. After patients were clinically examined, three groupswere defined according to the 1993 revised classification ofthe U.S. Centers for Disease Control and Prevention (CDC): group1 (n = 30), asymptomatic subjects (CDC-A or -B); group 2 (n= 37), AIDS symptomatic subjects (CDC-C) without wasting syndromeor diarrhea; and group 3 (n = 33), AIDS symptomatic patients(CDC-C) with severe body weight loss (>10% body weight duringthe past month) and/or chronic diarrhea (more than three dailybowel movements for at least 3 weeks and not related to antiretroviraltherapy). All of the drugs taken by each patient were listedin order to check for drug interactions.

Clinical and biological data. A complete biological profile was defined for each patient,including height and weight to assess the body mass index (BMI),age, total weight of stools per 24 h, viral load, CD4 cell count,creatinine to assess the creatinine clearance (CL_CR) accordingto the Cockroft and Gault formula (8), intestinal permeabilityand absorption tests, plasma albumin, and alkaline phosphatase.

Intestinal absorption and permeability were assessed by theD-xylose and lactulose-mannitol tests, respectively, in which5 g of D-xylose (Pharmacie Centrale des Hôpitaux, Nanterre,France) and 5 g of lactulose (Duphalac; Solvay Pharma, Suresnes,France) plus 1 g of mannitol (Pharmacie Centrale des Hôpitaux,Nanterre, France) were given orally to each patient.

The clinical, intestinal function tests and biological parametersare referred to as covariates below.

Population pharmacokinetic analysis. The blood sampling design was sparse, based on a populationpharmacokinetic approach in order to include a large numberof subjects in a clinical situation by greatly limiting thenumber of samples: three blood samples were drawn per patient,according to the recommendations of Bréant et al. (6).Since only three concentration measurements were available foreach subject, the kinetics profiles in each group were analyzedseparately by using a population approach implemented in theP-PHARM software (InnaPhase Corp., Philadelphia, Pa.). Thissoftware allows general nonlinear mixed-effects models to fitexperimental data, particularly in sparse individual data situations.P-PHARM uses an EM-like algorithm for computing the maximum-likelihoodestimates of the population pharmacokinetic parameters (25).

A one-compartment model with first-order elimination and first-orderabsorption with a lag time was selected. Since no referenceinput route was available (intravenous route), the estimationof the absolute bioavailability (F) was not possible. Hence,clearance and volume of distribution of SQV were evaluated asCL/F and V/F, respectively. All pharmacokinetic parameters wereassumed to be log-normally distributed. Before the algorithmwas applied, initial values of the pharmacokinetic parameters(CL/F, V/F, k_a, and T_lag, the so-called independent parameters)were selected from previously reported values (3, 27). The variancemodel was chosen to display the best fit among constant homoscedasticor heteroscedastic values (1/Y or 1/Y²). The three models wererun until convergence. The model retained was the one that gavethe lowest maximum-likelihood and Akaike criterion values.

The adequacy of the estimated population models was evaluatedby using several standard goodness-of-fit criteria implementedin the P-PHARM software: (i) visual examination of the goodness-of-fitcurves (population fitted model and individual fittings); (ii)visual examination of the scatterplot of observed versus predictedconcentrations of the drugs in plasma after individual Bayesianfitting; (iii) statistical comparison of observed versus predictedconcentrations of the drugs in plasma by a Student paired ttest; (iv) comparison of the mean of the normalized residualsto zero by a Student t test; (v) detection of outliers by usingthe two-sided Student t test; (vi) comparison of the cumulativedistribution of the normalized residuals and individual pharmacokineticparameters to a normal or log-normal distribution, accordingto the initial hypothesis of this dispersal, by a Kolmogorov-Smirnovtest; (vii) visual comparison of the cumulative distributionof the normalized residuals and individual pharmacokinetic parametersto a normal or log-normal distribution; and (viii) checkingthe decrease of the interindividual variability of the pharmacokineticparameters, after the covariables were taken into account, comparedto the variability before their inclusion in the structuralmodel.

From the estimated mean population pharmacokinetic parameters,the individual (post hoc) parameters were computed (i.e., CL/F,V/F, k_a, and T_lag) by the Bayes method implemented in the P-PHARMsoftware. Then, the individual derived parameters, namely, thek_el (elimination rate constant) and AUC, were calculated (13).T_max, the time to reach the maximal concentration, called C_max,and C_max were graphically determined from each individual profile.These four last parameters are called the derived parameters(i.e., k_el, AUC, T_max, and C_max). Since one of the most pertinentparameters to evaluate the total body exposure of a drug isthe AUC, this parameter was used as the primary end point forassessing SQV exposure.

Relationships between groups, exposure, and covariates. The following covariates were studied: gender, age, BMI, meanweight of stools per 24 h, plasma albumin, CL_CR, xylose, lactulose/mannitol(L/M) ratio, alkaline phosphatase levels, and CD4 cell count.The variables that were not normally distributed, i.e., xylose,L/M ratio, alkaline phosphatase levels, and CD4 cell count,were log transformed. Changes in AUC between groups and relationshipsbetween AUC and several biological and clinical parameters,such as intestinal absorption, were both studied. Initially,comparisons between groups of the covariates and the logarithmof all of the individual pharmacokinetic parameters were performedby using univariate analysis of variance (ANOVA) for quantitativevariables and a ${chi}$ ² test for gender. When the results of the ANOVAwere significant, the pairwise comparisons among the three groupswere performed by using a Tukey correction. AUCs were log transformedby using decimal logarithm. Then, univariate relationships betweenthe log(AUC) and the covariates were tested by using a Pearsoncorrelation coefficient for quantitative variables and a Studentt test for gender. A Bonferroni correction was performed toaccount for multiple testing all of the univariate analyses.A P value of <10^-3 was consequently considered statisticallysignificant.

Lastly, a multiple linear regression was performed to assessthe association between covariates and log(AUC). Two modelswere built. The first model included all covariates; then, astep-by-step deletion was performed, beginning with the covariateswith the higher P value. The second model included only thevariable "group." A step-by-step backward selection method witha "P = 0.05 criterion" was used. Goodness of fit was assessedaccording to scatter plots and quantile-quantile plots of theresiduals.

RESULTS

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Results

Patients. One hundred patients participated in the present study (19 femalesand 81 males). Thirty, thirty-seven, and thirty-three patientswere enrolled in groups 1, 2, and 3, respectively, as describedabove. The female/male ratios were 6/24 in group 1, 7/30 ingroup 2, and 6/27 in group 3. The antiretroviral treatmentsused were zidovudine, didanosine, zalcitabine, lamivudine, andstavudine. No drug known to affect SQV metabolism was reportedexcept in 12 patients: 2 patients with rifabutin in group 1;3 patients with rifabutin and 1 patient with rifampin in group2; and 5 patients with rifabutin and 1 patient with rifampinin group 3. No antidiarrheal agents were noted. Twenty patientswere naive to antiretrovirals (11, 8, and 1 patients in groups1, 2, and 3, respectively). The main demographic, clinical,and biological characteristics are listed in Table 1. Gender,viral load, and age were not found to differ significantly amongthe three groups. As expected, BMI, CD4 cell count, CL_CR, andalbumin significantly decreased from group 1 to group 3. Thedecrease in BMI is related to an incremental decrease in totalbody weight from group 1 to group 3 since the height of patientswas statistically the same in the three groups (result not shownin Table 1).

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TABLE 1. Mean biological values, intestinal absorption, and permeability test values of the HIV-1-infected patients included in the present study

It should be noted, as already described by Pernet et al. (29),that intestinal absorption and permeability were abnormal inevery patient of our study, regardless the stage of illness,especially for those in group 3. These observations are quiteconsistent with findings published earlier (7, 14, 23). Chronicdiarrhea and/or severe body weight loss, as defined above, permittedus to identify three subgroups in group 3: patients with chronicdiarrhea (n = 18; 54.5%), patients with severe body weight loss(n = 26; 78.8%), and patients with diarrhea and severe bodyweight loss (n = 14; 42.4%).

A significant decrease in D-xylose elimination (group 1 comparedto groups 2 and 3) and a significant increase in L/M ratio (group3 compared to groups 1 and 2) were found. D-Xylose excretionwas decreased in AIDS patients (group 3) with chronic diarrheaand severe body weight loss compared to those with severe bodyweight loss alone. However, L/M ratio did not significantlydiffer among patients in these three subgroups. Moreover, themean weight of stools per 24 h was significantly higher in group3 than in groups 1 and 2.

Population pharmacokinetic analysis. The one-compartment model adequately fit each group with satisfactorygoodness-of-fit plots. The best fits were obtained by usinga homoscedastic variance model for the three groups. The individualconcentrations (solid black circles) and the corresponding meanestimated concentration profiles are displayed in Fig. 1 foreach group. Population parameter mean values, interindividualvariability, and residual errors are given in Table 2. Also,the mean population values of the derived parameters are given(namely, k_el and AUC). The k_el and AUC were calculated by usingCL/V and dose/(CL/F), respectively. C_max and T_max were graphicallydetermined. It should be noted that because the derived populationmean parameters were calculated from estimated mean populationparameters (CL/F, V/F, k_a, and T_lag), no direct estimates ofinterpatient variability of these parameters were available.It is of particular interest that during the evolution of illness(from group 1 to group 3), the total body clearance of SQV expressedas CL/F decreased (Table 2), whereas C_max and AUC increased(Table 2 and Fig. 1). The high interindividual variability inSQV kinetics (30) previously evoked is illustrated in Fig. 1.

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FIG. 1. Individual observed concentrations (indicated by solid black circles [three per patient]) and mean population profile curves after the administration of 600 mg of SQV-HGC as a single oral dose after a standard breakfast that included 200 ml of grapefruit juice.

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TABLE 2. Population mean estimated pharmacokinetic parameters, median (25th to 75th percentiles) post hoc individual values, and derived parameters for each group^a

Relationships between groups, exposure, and covariates. The median AUC values (interquartile range, 25th to 75th percentiles)for each group were estimated from the individual post hoc estimatedAUCs: these were 165 (range, 90 to 370), 349 (range, 179 to767), and 705 (range, 342 to 1,271) ng · h · ml^-1for groups 1, 2, and 3, respectively. The log(AUC) was foundto increase significantly from group 1 to group 3 (P < 0.0001;Fig. 2). In the univariate analysis, the log(AUC) was foundto be significantly correlated to BMI (r = -0.42, P < 0.0001)and log(xylose) (r = -0.36, P = 0.0007). Each of these variableshad a negative effect on the log(AUC) (Fig. 3). The correlationsremained statistically significant for BMI (r = -0.35 and P= 0.0004) and xylose (r = -0.24 and P = 0.0271) when, for eachanalysis, one outlier with very high BMI or very low xylose(see arrows in Fig. 3) was removed from the analysis. No significantrelationship was found between log(AUC) and sex, age, CL_CR,mean weight of stools per 24 h, and log(alkaline phosphatase)by using the Bonferroni correction.

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FIG. 2. Boxplots of individual estimated AUC in each group. The boxes represent the 25th and 75th percentiles, with the median (solid lines) shown within the boxes. The error bars represent 1.5 times the interquartile range, and the circles indicate extreme values.

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FIG. 3. Correlations between AUC and BMI and log(xylose) for patients from group 1 ( ${circ}$ ), patients from group 2 ( ${square}$ ), and patients from group 3 ( ${triangleup}$ ). Arrows indicate outliers. The correlations remain statistically significant for BMI (r = -0.35 and P = 0.0004) and log(xylose) (r = -0.24 and P = 0.0271) when the outliers are removed from the analysis.

When all covariates, except "group," entered the descendingprocedure of the multivariate analysis, BMI and log(xylose)remained in the final model. This strongly suggests that AUCswere increased from groups 1 to 3 because of two independenteffects: decrease in BMI and decrease in xylose, i.e., decreasein intestinal absorption. When the variable "group" was addedto the model with all covariates, only BMI remained in the model.In that case, groups 2 and 3 had higher values for log(AUC)and BMI had a negative effect on SQV exposure. Goodness-of-fitplots were satisfying for both multivariate models.

DISCUSSION

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Discussion

Homogeneity in the three groups with regard to demographic parametershas been found, whereas the patients differed markedly withrespect to several biological covariates (Table 1). The associationof high viral load (means of 4.44, 4.85, and 4.92 log copies/mlfor groups 1, 2 and 3, respectively) with low CD4 cell counts(means of 248, 118, and 53 cells/mm³ for groups 1, 2, and 3,respectively) reflects relatively advanced stages of the illnessin every group (mainly in groups 2 and 3). The decreases inBMI and CD4 cell counts, in addition to the damaged hepaticfunction (revealed by the decrease in plasma albumin and theincrease in alkaline phosphatase), are clearly a sign of theevolution of the illness from group 1 to group 3.

The population approach we used, limiting the blood sample schedule,was chosen for the comfort of the patients. The population analyseswere performed separately for the three groups of patients,allowing us to adequately estimate the population parametersfor each of them. The large interpatient variability of theobserved concentrations, although a single identical dose ofSQV was given (Fig. 1), was previously described by Regazziet al. (30). A great scatter in the observations was especiallynoted around C_max for groups 2 and 3.

The pharmacokinetics of SQV is often described as a two-compartmentmodel (3). The overall parameters in such a model can be estimatedonly when enough experimental data are available, especiallyin the terminal elimination phase. In the present study, thesample schedule tends to better characterize the absorptionphase up to T_max than the elimination phase. Moreover, sincesome patients prematurely abandoned the study, the later points(i.e., 10 and 12 h after dosing) were not often available, preventingthe algorithm from correctly fitting the elimination phase.Therefore, a one-compartment model with first-order absorptionwith a lag-time and first-order elimination was selected. Possiblemisspecifications on the estimated parameters with respect tothe final elimination phase may have been made as a result.To avoid this main limitation on the pharmacokinetic parameterevaluation, another sample schedule could have been proposedwith more sampling times during the elimination phase. For example,two samples per phase (i.e., absorption, distribution, and elimination)could have been judiciously chosen as we recommended for anotherstudy already published (26). Furthermore, an optimal populationdesign approach using the Fisher information matrix-based methodcould have been implemented to define the best sampling scheduleto consider (24, 31).

The present study mainly displayed a significant increase inSQV exposure in HIV-1-infected patients when disease worsens.This is brought to the fore by a steady increase in AUCs, observedfrom group 1 (asymptomatic HIV-1-infected patients) to group3 (AIDS symptomatic patients with severe body weight loss and/ordiarrhea) (Fig. 2 and Table 2). The multivariate analysis ofthe individual post hoc AUC estimates showed that this increasewas mainly correlated to two independent effects: (i) a decreasein intestinal absorption and (ii) an increase of the dose ofSQV expressed in milligrams per kilogram (because of the decreasein total body weight).

The D-xylose test assesses intestinal absorption. D-Xylose isa monosaccharide absorbed unchanged by the duodenum and thejejunum due to a facilitated transport after oral administration.It is poorly metabolized. A decrease in its urinary excretionsignals a decrease in the mucosal functional area of the jejunum.In HIV-1-infected patients, small-bowel histological modificationssuch as villous atrophy, which leads to a decreased D-xyloseabsorption, are already well known (9, 18, 21). In the presentstudy, this test displayed a significant incremental urinaryD-xylose decrease as the illness spreads from group 1 to 3.This argues for possible disabled absorption transporters whoseconsequence should be a decrease in SQV absorption. Furthermore,it is now well known that peptides and peptidomimetics, suchas the HIV PIs, are substrates of P-glycoprotein and/or CYP3A4(2, 4, 12, 20, 33, 34). P-glycoprotein, an ATP-dependent drugefflux pump, typically associated with multidrug resistancein cancer chemotherapy, has been shown to be present in theenterocyte brush border and to closely interact with the majorphase I drug-metabolizing enzyme in humans, CYP3A4 (35), furtherlimiting the bioavailability of many drugs. In addition to itshepatic biotransformation (see below), it has been shown thatSQV undergoes another metabolism in the enterocyte that potentiallycontributes to its high first-pass metabolism (12). Thus, itmay be that in normal situations (without intestinal disease)P-glycoprotein, together with CYP3A4, acts as a concerted barrierto SQV absorption, which is responsible for its fairly low bioavailability.As the HIV infection spreads, i.e., as wasting syndrome developsand diarrhea appears, the histological integrity of the villusis challenged. Hence, the destruction, at least in part, ofthe efflux, together with the metabolism mechanisms, permitslarger drug quantities to be passively absorbed by the body,causing an increase in its AUCs. However, in our study, allpatients used grapefruit juice, which acts as a blocker of CYP3A4(10, 22). Indeed, when the present study started (at the endof the 1990s), it was usual practice to coadminister grapefruitjuice in order to increase SQV absorption (22) because of thehigh cost of that treatment when prescribed in doses high enoughto be effective. Therefore, the design of this protocol correspondsto the routinely prescribed method at that time. As a consequence,the mechanism involving P-glycoprotein should be consideredthe only one in our study that could explain the increase inAUCs from group 1 to group 3. Furthermore, since grapefruitjuice was given to every patient at the same dose, it allowedcomparison between the groups. Finally, some of our patientswere treated with rifabutin and/or rifampin. Both substancesstrongly induce the CYP3A4 PI metabolism, thus decreasing theirconcentrations. The many patients with rifabutin and/or rifampinin group 1 could have explained the lower disposition of SQVobserved in this group compared to that of group 3 (AUC₁ <AUC₃, P < 0.0001). However, most patients receiving rifabutinand/or rifampin were in group 3 (2 of 30 in group 1 [6.7%],4 of 37 in group 2 [10.8%], and 6 of 33 in group 3 [18.2%]).However, an increase in SQV disposition from group 1 to group3 was observed despite these drug interactions. Hence, the presenceof rifabutin and/or rifampin in some of our patients does notinvalidate our overall findings.

The lactulose-mannitol test assesses intestinal permeability.In this test, the two sugars are simultaneously given orally,and their urinary recovery is determined. The intestinal permeabilitymeasures the ability of compounds to passively cross the intestinalmucosa through paracellular tight junction areas (16). The urinaryrecovery of the two sugars is expressed as the L/M ratio, whichis known to be a reproducible measure of the mucosal integrityof the small intestine. Many studies in the literature utilizedthat test. Some of these studies demonstrated that childrenwith small-intestine mucosal damage, determined by jejunal biopsy,had a significantly higher L/M urinary excretion ratio thancontrol groups (1). In these situations, the sugar permeabilitytest has shown good correlation with jejunal biopsy results.The enhanced L/M ratio detected in our study as the sicknessworsened (Table 1) probably reveals an SQV-increased intestinalpermeability from group 1 to group 3 that could, at least inpart, explain and argue for the increased SQV disposition. Althoughthe L/M ratio significantly increased from group 1 to group3 (P < 0.01), the multivariate analysis did not correlateit with log(AUC). A higher number of patients in every groupcould probably have revealed such a correlation. Nevertheless,the increase in L/M ratio from group 1 to group 3 was probablyrelated to an enlargement of the tight junctions, allowing apassive paracellular crossing of SQV. This latter mechanismcould take over from the transporter-mediated absorption thattakes place in normal situations (patients in group 1 withoutdigestive disease).

The total body clearance of SQV (expressed as CL/F = dose/AUC)decreased as the illness spread (Table 2). This effect can beexplained by a decrease in the dose of SQV, the increase inAUC being a consequence of the decrease in CL/F. If one expressesthe SQV dosing in milligrams per kilogram, the doses are 8.6± 1.5, 9.3 ± 1.5, and 11.1 ± 1.8 mg/kgfor groups 1, 2, and 3, respectively. The mean dose of SQV was1.3-fold greater in group 3 than in group 1. This is relatedto an incremental decrease in the total body weights of ourpatients from group 1 to group 3, their heights being not significantlydifferent in the three groups (Table 1, decrease in BMI). Thus,the increase in SQV dosing, expressed in milligrams per kilogram,should not be the only explanation for the CL/F decrease, whosemedian post hoc individual value was 4.3 times greater in group3 than in group 1. The changes observed in CL/F could then berelated to a decrease in CL and/or an increase in F. As alreadystated above, CL could not be evaluated because the intravenousdata were not available. If bioavailability was implicated (increasein F), this could be due to an increase in the "absorption"(from a pharmacokinetic point of view) or to a decrease in thefirst-pass metabolism effect. Even if the liver function wasdisrupted (decrease in plasma albumin and increase in alkalinephosphatase), the ANOVA did not show any significant relationshipbetween plasma albumin or alkaline phosphatase and log(AUC).Moreover, since grapefruit juice was given to all patients,the argument involving CYP3A4 in the enterocytes cannot be putforward to explain the possible increase in F. This increasein SQV bioavailability could then be explained, at least inpart, by the increase of the intestinal permeability observedin the present study.

The present study demonstrated that SQV, despite its low intrinsicoral bioavailability, remains a relevant treatment for HIV-1-infectedpatients with diarrhea and/or wasting syndrome, mainly in developingcountries where these digestive troubles still persist. Theresults support a major role of the efflux protein, P-glycoprotein,together with the integrity of the gut barrier, in determiningthe level of the bioavailability of SQV. The role of the metabolismenzyme CYP3A4 in the enterocyte, although reduced because ofthe coadministration of grapefruit juice to our patients, isnot to be neglected. Further investigations are needed to determinethe clinical efficacy of SQV for HIV-1-infected patients withdiarrhea and/or wasting syndrome over a long-term period.

ACKNOWLEDGMENTS

We thank Produits Roche and the Agence Française du Médicamentfor financial support.

We also thank to Huntington Life Sciences Ltd., Huntington,Campbridgeshire, England, for SQV assays; Bertrand Diquet forreviewing the manuscript; the patients for their kind contributionin that trial; and Martine Mallet, Maguy Parrinello, LaurenceBonhomme-Faivre, and the nurses of the Departments of InfectiousDiseases of Lariboisière and Paul Brousse Hospitals,who actively participated in this study.

FOOTNOTES

* Corresponding author. Mailing address: Laboratoire de Toxicologie et Pharmacologie Clinique, Service de Pharmacie, Hôpital Lariboisière, 2 rue Ambroise Paré, 75475 Paris Cedex 10, France. Phone: 33 (1) 49956463. Fax: 33 (1) 49956470. E-mail: herve.trout{at}lrb.ap-hop-paris.fr.

REFERENCES

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