Review. Therapeutic Drug Monitoring of Antiretroviral Therapy

AIDS PATIENT CARE and STDs Volume 18, Number 1, 2004 Mary Ann Liebert, Inc. Review Therapeutic Drug Monitoring of Antiretroviral Therapy NATELLA Y. RAKHMANINA, M.D., 1,4 JOHN N. VAN DEN ANKER, M.D., Ph.D., 2,4 6 and STEVEN J. SOLDIN, Ph.D. 3 5,7 ABSTRACT The concept of managing pharmacotherapy based on plasma drug concentrations has been used for decades in a variety of clinical settings. The interest in therapeutic drug monitoring (TDM) of antiretroviral drugs has grown significantly since highly active antiretroviral therapy (HAART) became a standard of care in clinical practice. A primary characteristic of TDM of antiretroviral drugs is that multiple agents are concomitantly used in HAART regimens. Inadequate drug concentrations may lead to evolution of drug resistance mutations and endanger present and future treatment options. A number of clinical trials have demonstrated that drug serum concentrations are an important factor in response to therapy for HIV, but whether TDM will become a tool for the routine management of HIV infection remains to be determined. This review includes an illustrative case report of measuring concentrations of antiretroviral drugs in a pediatric patient. CONCEPTS OF THERAPEUTIC DRUG MANAGEMENT IN HIV THERAPEUTIC DRUG MONITORING (TDM) permits timely dosage adjustments based on individual plasma concentrations to optimize the therapeutic effect of the medication and prevent exposure to toxic or subtherapeutic concentrations of the drug. Criteria for successful TDM include: well-defined therapeutic range, strong correlation between drug concentration and therapeutic effect, high interpatient and low intrapatient variability in plasma concentrations and failure of the long-term outcome with inadequate target levels. 1 A reliable, cost-effective method should be available for the determination of drug concentrations. 2 While several features of antiretroviral therapy (ART) fit the criteria for TDM, a number of questions need to be answered before largescale introduction of TDM can be justified. For the majority of antiretroviral drugs the therapeutic window is narrow, with toxicities ranging from nausea and vomiting to pancreatitis, nephrolithiasis, and neurologic effects. 2 4 Some of these toxicities might be prevented with the 1 Special Immunology Program, 2 Division of Pediatric Clinical Pharmacology, and 3 Department of Laboratory Medicine, Children s National Medical Center, Washington, D.C. Departments of 4 Pediatrics, 5 Pathology and 6 Pharmacology, The George Washington University School of Medicine, Washington, D.C. 7 Department of Pharmacology, Georgetown University and Georgetown Clinical Research Center, Washington, D.C. 7

8 RAKHMANINA ET AL. appropriate use of TDM. 5,6 Concentrations of antiretroviral agents vary greatly among individual patients who are treated with the same dose, often more than 10-fold. 7,8 The cause of such significant interpatient pharmacokinetic variability is multifactorial and includes differences in metabolism, bioavailability, complex drug-drug and food-drug interactions. Several studies have demonstrated a lack of adequate viral suppression in the absence of therapeutic drug levels. 9 11 The pharmacokinetic (PK) data from the Viradapt study have shown a significant correlation between suboptimal drug concentrations and the risk of virologic failure. 12 This is of particular importance because preliminary studies from centers that offer TDM have been reporting that a substantial proportion of patients may have subtherapeutic levels of antiretroviral agents. 8,13,14 Adverse events, nonadherence, and drug interactions are frequently responsible for subtherapeutic drug levels. A recent study by De Maat et al., 15 however, demonstrated a high number of subtherapeutic drug concentrations without an identifiable cause. Inadequate concentrations of antiretroviral agents may increase the potential of drug resistance and treatment failure. A narrow therapeutic index of many antiretroviral agents suggests the possibility of using TDM for concentration-targeted dosing. Applying TDM to enhance treatment outcome is much more complex and difficult to achieve. There are no studies currently available that have compared peak plasma concentrations (C max ), the area under the serum concentration versus time curves (AUC), and trough plasma concentrations (C min ) as predictors of treatment efficacy. The AUC, a measure of systemic exposure, may be an important target because it is more likely to reflect exposure in body compartments. Studies of virologic efficacy have found a significant correlation both with C min and AUC. The ratio of C min to inhibitory concentration at 50% (IC 50 ), defined as inhibitory quotient (IQ), is used by many researchers and is believed to be the parameter most likely to predict efficacy. TDM of antiretrovirals is also being used to monitor patient adherence to the regimen. 16 Poor adherence has been identified as a leading cause of treatment failure by many authors. 17,18 Nonadherence is a serious problem and is reported in 33% to 69% of this patient population. 4,19 Common reasons for missing doses include clinical toxicity, forgetfulness, sleeping through the time of prescribed dose, and being away from home. TDM can help identify nonadherence, although a drug concentration only reflects the last few drug doses taken by the patient. It is crucial that any TDM results are incorporated with careful adherence assessment to facilitate interpretation of the results and adherence counseling should be established as part of clinical care in the centers practicing TDM. TDM OF DIFFERENT ANTIRETROVIRAL AGENTS Adequate drug concentrations may not reach the virus because of poor compliance, poor absorption, increased metabolism and/or elimination, and drug-drug and drug-food interactions. The most important and most common pharmacokinetic drug interactions involve inhibition of metabolism, induction of metabolism, altered drug absorption, inhibition of renal secretion, and displacement from plasma protein and/or binding sites. The nucleoside reverse transcriptase inhibitors (NRTI) are active as intracellular triphosphates, and there is little evidence to suggest that their measurement would be helpful other than to assess adherence to the drug regimen. Several studies have established a relationship between plasma concentrations of NRTI and virologic and immunologic outcomes. 20,21 The usefulness of NRTI plasma concentration in predicting the intracellular levels of their triphosphate metabolites remains unclear. The relationship between the intracellular drug levels and outcome parameters for zidovudine and lamivudine has been shown to be significant, 20 but the methodology of intracellular triphosphate metabolites is expensive and labor-intensive and is limited to highly specialized centers. For the protease inhibitors (PIs) and to lesser extent for non-nucleoside reverse transcriptase inhibitors (NNRTIs), the relationship between

THERAPEUTIC DRUG MONITORING 9 plasma drug concentrations and their efficacy and toxicity has been identified. 9,11,12,22 24 Large interindividual and intraindividual differences in drug disposition have been observed with the use of many antiretroviral agents, which makes dose based prediction of plasma concentrations unreliable. 7,25,26 The patients taking NNRTIs are at the greatest risk for developing resistance and treatment-limiting toxicity. These drugs are metabolized by CYP 450 2B6, 1A2, 2A6, 2C and 2D6 (efavirenz, nevirapine, nelfinavir, and ritonavir) and by 3A, primarily 3A4 (indinavir, delavirdine, ritonavir, nevirapine, saquinavir, and nelfinavir) creating a solid base for multiple drug-drug interactions. 27,28 Many drugs used in the treatment of infections associated with HIV-1 disease (ketoconazole, fluconazole, rifampin, rifabutin, methadone) as well as a herbal supplement (St. John s wort) have shown significant interactions with antiretroviral drugs leading to toxicity-related complications and subtherapeutic concentrations. The interactions between PIs and NNRTIs, and among different PIs, especially in salvage regimens, have become clinical issues with the growing resistance pattern of the HIV. The application of TDM in the patients who are using two and more P-450 interacting agents seems therefore indicated. The PIs are strongly protein bound, and especially to a-1-acid glycoprotein (AAG), an acute phase reactant. 29,30 In general there is a lack of data on the effect of varying concentrations of the binding protein on the free drug concentrations and its antiviral activity. This is particularly true for pediatric patients, in whom concentrations of albumin and/or AAG may vary substantially. Currently, studies are being conducted on the value of free drug concentration measurements and its relationship with outcome measurements. Recently, randomized, prospective clinical trials evaluating the role of TDM in the management of HIV-1 infected patients have shown promising results. 4 The ATHENA study conducted in The Netherlands included two clinical trials using TDM in treatment-naïve patients who started indinavir- or nelfinavir-based regimens. TDM of nelfinavir improved viral load effects but did not reduce toxicity, while dose adjustment for indinavir reduced toxicity but did not improve antiviral effect. 13,14 TDM did prevent either virologic failure (presumably by preventing development of resistance) or treatment discontinuation because of concentrationrelated toxicities. These findings are even more impressive, taking into consideration that only 20% of the physicians responded to the dosing recommendations of the intervention group. This protocol, however, did not incorporate resistance data in the dosage adjustment calculation. Another study conducted in France (PharmAdapt) failed to find a significant benefit of TDM versus standard care. 31 Application of routine TDM in other patients groups (treatment-experienced patients) or for different drugs (NNRTIs, other PIs, combination of PIs) has still to be proven. 32 Additional clinical trials with improved design are needed to investigate if routine TDM as standard of care for the treatment of HIV infection is warranted. Several important limitations to the application of TDM for ART should be recognized, including uncertainty about the best PK predictor of response and insufficient validation of target concentrations for individual PIs and NNRTIs. The availability of a large TDM database may facilitate the establishment of expected concentration ranges for a variety of antiretroviral drugs. The concept of inhibitory quotient, which integrates drug concentrations and resistance testing, shows promise in a number of retrospective analyses. 13,14,33 A logistical problem remains with regard to its feasibility, and theoretical issues such as protein binding, variability, and the appropriate time of sampling continue to be debated. Available technology allows to set target inhibitory concentrations for a particular virus isolate based on genotypic and phenotypic sensitivity, however it is expensive and requires highly specialized assistance. 34 Ongoing research in the field of the HIV pharmacology is necessary for the future improvement of ART. CURRENT APPLICATION OF TDM IN CLINICAL PRACTICE The major reasons for TDM include individualizing therapy, preventing drug toxicity, and

verifying patient compliance. Poor adherence and pharmacokinetic factors may contribute to highly active antiretroviral therapy (HAART) failure. 34 Other factors such as degree of underlying resistance, drug tolerability and treatment history also play a role in determining virologic outcome. Today TDM has been proven to be most useful in selected patients groups: children, pregnant women, patients with renal and hepatic dysfunction and for the measurement of drug-drug interactions. 8,32,34 39,42,43 The recommendations of antiretroviral drugs for the treatment of HIV-infected pregnant women are subject to special considerations, including potential changes in dosing requirement and potential effects on the fetus or newborn. The pharmacokinetics of antiretroviral agents have not been adequately studied in pregnant women. The pharmacokinetic disposition of antiretroviral drugs may be profoundly altered during pregnancy as a result of dramatic physiologic, hormonal, and metabolic changes, which may lead to subtherapeutic or toxic systemic and intracellular drug levels in pregnant women. Available information is limited to the compounds currently used to decrease the risk of vertical transmission of HIV infection to the fetus. 35 37 Patients with renal or hepatic insufficiency are at increased risk of toxic concentrations for antiretroviral medications. The interindividual variability in plasma concentration has been reported to be higher in patients with liver dysfunction and examples of the application of TDM for HAART drugs in patients with liver impairment have been published. 38,39 Rapid changes in drug metabolism and distribution among pediatric patients, especially infants, can significantly alter their pharmacokinetic profiles. As with many other medications the extrapolation of the adult PK data and its application in pediatrics has been proven to have poor predictive value. 40 42 As children grow, drug pharmacokinetics are affected by changes in absorption, liver metabolizing capacity, plasma protein concentrations, body compartments, and renal function. Brundgade et al. 43 have shown that the dose based on pharmacokinetic data in older children may not be applicable to younger children. Most researchers agree that HIV-infected children may 10 be a target population for TDM application due to the unpredictability of plasma concentrations based on the administered dose. 34 Many clinicians feel drug concentrations may be of greatest utility shortly after initiating any new regimen, to ensure concentrations are adequate. Others see TDM only in the settings of previously failed therapy and in conjunction with resistance testing. 44 TDM for antiretroviral agents in HIV is available in a number of large reference centers in the United States and in Europe. The data from United Kingdom reported that the most common reasons for using TDM in HIV were: the administration of nonrecommended doses, suspected treatment failure, pharmacologic enhancement, suspected drug-drug or drug-food interaction, clinical symptoms of toxicity, change in therapy, and pediatric treatment. 8 The panel of experts in the field of pharmacology of antiretroviral agents held in Perugia in 2000 has agreed that TDM may represent a practical tool to improve the outcome of patients receiving HAART. The current limitations of routine clinical use of TDM include the insufficiency of data, lack of the assays standardization, and need for an expert data interpretation. The panel recommended further the conduct of large randomized trials to assess the clinical utility of TDM in the management of HIV-1 infection, assay standardization and educational programs for the caregivers and patients. 4 METHODOLOGY RAKHMANINA ET AL. The analytical criteria for TDM require a drug assay with high specificity, sensitivity, small sample volume, reasonable cost, and rapid turnaround time. 45 Methods for antiretroviral drug quantitation include high-performance chromatography and tandem-mass spectrometry (MS)/MS. 2,46 49 The tandem- MS/MS procedure developed at our laboratory is simple and allows for the quantitation of 16 HIV medications simultaneously in less than 5 minutes. 50 This procedure provides a better sensitivity and specificity than the high-performance liquid chromatography methods, which also require larger sample volumes, and are far more labor intensive. 51

THERAPEUTIC DRUG MONITORING 11 The ideal sample is drawn at steady state just before the next dose (steady-state trough sample). Steady state is reached after an interval of five drug half-lives have elapsed, which for most of these drugs is less than 48 hours. Exceptions to this are nevirapine and saquinavir, both of which have longer half-lives and should not be monitored until 4 days after initiation of the drug regimen. Most of the PIs have in vitro 95% inhibitory concentrations of approximately 100 ng/ml. At the Children s National Medical Center (CNMC) the lower limit for the therapeutic range for PIs has been 150 ng/ml. For the upper limit we recommend concentrations less than 6000 ng/ml, except for lopinavir, which we recommend should be maintained at concentrations less than 1200 ng/ml. These values were derived from the literature and are tentative therapeutic ranges. 7,9,11,12,25, 52,53 For the NNRTIs we recommend tentative therapeutic ranges between 1200 and 7000 ng/ml. External proficiency testing for the AIDS drugs is available from the International Quality Control Program for Therapeutic drug Monitoring in HIV infection (University Medical Center Nijmegen, The Netherlands). Currently, more than 65 laboratories worldwide are enrolled in this program. Drug standards can be obtained from the National Institutes of Health (NIH) AIDS Reagent Reference Program. CASE REPORT The following is a short illustration of the usefulness of TDM in pediatric practice at CNMC during a complex multiple drug regimen treatment of one of our patients. The patient was an African American girl, 9 years of age, with HIV/AIDS infection in category C3 per Centers for Disease Control (CDC) classification of pediatric HIV infection, a history of poor adherence with medications, encephalopathy, and severe failure to thrive with wasting. Her CD4 counts have dropped down to 3% (22 cells/mm 3 ) and her viral load was at 47,871 copies/ml. Because of profound immunosuppression she has developed disseminated Mycobacterium avium infection (MAI) with significant gastrointestinal disease, confirmed by small bowel biopsy. Her gut involvement was so severe that she was unable to tolerate any enteral intake including HAART therapy during a period of 9 weeks. Her MAI therapy included rifabutin, ethambutol, and amikacin via the intravenous route along with total parenteral nutrition (TPN), trimethoprim-sulphamethoxazole, metoclopramide, ranitidine, ondansetrone and methadone. Unfortunately, despite amikacin serum concentrations in the therapeutic range, she developed moderate sensorineural hearing loss bilaterally. Her MAI regimen was changed to a combination of rifabutin, ethambutol, and azithromycin. After a 9-week interruption of her HIV treatment, she was able to tolerate medications through her gastrojejunal tube and was restarted on HAART regimen consisting of didanosine, zidovudine, and kaletra (lopinavir/ ritonavir). Her new HAART regimen was chosen based on the results of the resistance profile by phenotype (PhenoSense, ViroLogic, South San Francisco, CA), which showed susceptibility to all antiretroviral drugs, except for nevirapine and nelfinavir. Because of known drug-drug interaction between lopinavir/ ritonavir and rifabutin (with lopinavir and ritonavir induced suppression of rifabutin metabolism and increase of rifabutin levels by 570% with lopinavir and by 350% with ritonavir administration) 34 the dose of rifabutin was decreased accordingly. There was also a concern of her gastrointestinal tract ability to absorb antiretroviral agents in view of severe MAI infiltration of the gastric and intestinal walls. To confirm adequate drug concentrations of her antiretroviral agents and to avoid unnecessary toxicity in combination therapy with drug-drug interactions the levels of her antiretroviral medications were obtained. It is important to mention that TDM was conducted in an inpatient setting where the administration of HAART and other medication was performed by medical personnel and assured 100% adherence with the regimen. The levels of the medications were determined at our laboratory using liquid chromatography tandem MS/MS. The trough levels were 18 ng/ml for ddi; 50 ng/ml for zidovudine; 189 ng/ml for ritonavir (reference range of 150 6000 ng/ml) and 10,700 ng/ml for lopinavir (with reference

range 150 12,000 ng/ml). The conclusion was made that the gastrointestinal absorption of HAART in our patient was adequate and the level of medications were in therapeutic and not in the toxic range, and no adjustments of the medications doses and regimen were warranted. Within 16 weeks of restarting HAART regimen she has increased her CD4 counts to 20% (179 cells/mm 3 ) and her viral load was undetectable at less than 400 copies per milliliter. This clinical case illustrates that TDM for antiretrovirals can be useful to monitor the therapeutic window of antiretroviral therapy with the complex treatment involving multiple drugs and toxicity could be avoided. 12 CONCLUSION Optimal care in HIV requires individualized management and ongoing attention to relevant scientific and clinical information in the field. 54 A growing body of literature supports the concept of TDM in HIV, but it is important that it is incorporated with other interventions such as resistance testing, adherence monitoring, and patient counseling to be an effective tool in patient management. Prospective randomized and double-blinded clinical trials are necessary to establish the role of concentrationtargeted therapy of HIV in routine clinical care. Properly applied TDM program can already benefit many HIV-infected patients on antiretroviral therapy, especially those at risk for subtherapeutic or toxic drug concentrations. REFERENCES 1. Acosta EP, Gerber JG. Adult Pharmacology Committee of the AIDS Clinical Trials Group. Position paper on therapeutic drug monitoring of antiretroviral agents. AIDS Res Hum Retroviruses 2002;18:825 834. 2. Soldin OP, Elin RJ, Soldin SJ. Therapeutic drug monitoring in human immunodeficiency virus/acquired immunodeficiency syndrome. Quo vadis. Arch Pathol Lab Med 2003;127:102 104. 3. Back DJ, Khoo SH, Gibbons SE, et al. Therapeutic drug monitoring of antiretrovirals in human immunodeficiency virus infection. Ther Drug Monit 2000;22: 122 126. 4. van Heeswijk RPG. Critical issues in therapeutic drug monitoring of antiretroviral drugs. Ther Drug Monit 2002;30:313 318. RAKHMANINA ET AL. 5. Casado JL, MorenoA, Sabido R, et al. A clinical study of the combination of 100 mg ritonavir plus 800 mg indinavir as salvage therapy: Influence of increased plasma drug levels in the rate of response. HIV Clin Trials 2000;1:13 19. 6. Lamotte C, Peytavin G, Perre P, et al. Increasing adverse events with indinavir dosages and plasma concentrations in four different ritonavir-indinavir containing regimens in HIV-infected patients [Abstract 738]. Eighth Conference on Retroviruses and Opportunistic Infections. Chicago, 2001. 7. Marzolini C, Telenti, Decosterd LA, et al. Efavirenz plasma levels can predict treatment failure and central nervous system side effects in HIV-infected patients. AIDS 2001;15:71 75. 8. Gibbons ES, Reynolds HE, TIJA JF et al. Therapeutic drug monitoring in the management of subjects on the protease inhibitors nelfinavir and saquinavir: results of the Roche UK TDM service [Abstract P259]. 5th International Congress on Drug therapy in HIV infection. Glasgow, 2000. 9. Hoetelmans RMW, Reijers MHE, Weverling GJ, et al. The effect of plasma drug concentrations on HIV-1 clearance rate during quadruple drug therapy. AIDS 1998;12:F111 F115. 10. Lebergeber B, Egger M, Opravil M, et al. Clinical progression and virologic failure of highly active antiretroviral therapy in HIV-1 patients: A prospective cohort stuffy. Lancet 1999;353:863 868. 11. Veldkamp AI, Weverling GJ, Lange JM, et al. High exposure to nevirapine in plasma is associated with improved virological response in HIV-1 infected individuals. AIDS 2001;15:1089 1095. 12. Durant J, Clevengergh P, Garaffo R, et al. Importance of protease inhibitors plasma levels in HIV-infected patients treated with genotypic-guided therapy: pharmacologic data from the Viradept study. AIDS 2000; 14:1333 1339. 13. Burger DM, Hugen PWH, Droste J, et al. Therapeutic drug monitoring of indinavir in treatment-naïve patients improves therapeutic outcome after one year: results from ATHENA [Abstract 6.2a]. 2nd International Workshop on Clinical Pharmacology of HIV Therapy. Noordwijk, 2001. 14. Burger DM, Hugen PWH, Droste J, et al. Therapeutic drug monitoring of nelfinavir 1250 BID in treatmentnaïve patients improves therapeutic outcome after one year: Results from ATHENA [Abstract 6.2b]. 2nd International Workshop on Clinical Pharmacology of HIV Therapy. Noordwijk, 2001. 15. De Maat MM, Huitema AD, Mulder JW, et al. Subtherapeutic antiretroviral plasma concentrations in routine clinical outpatient HIV care. Ther Drug Monit 2003;25:367 373. 16. Hugen PW, Burger DM, Aarnouste RE, Baede PA, et al. Therapeutic drug monitoring of HIV-protease inhibitors to assess non-compliance. Ther Drug Monit 2002;24:579 587. 17. Descamps D, Flandre P, Calvez V, et al. Mechanisms of virologic failure in previously untreated HIV-in-

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