Therapeutic drug monitoring of immunosuppressant drugs in clinical practice

CLINICAL THEFWF’EUTICSVVOL. 24, NO. 3,2002

Therapeutic Drug Monitoring of Immunosuppressant Drugs in Clinical Practice Barry D. Kahan, MD, PhD,l Paul Keown, MD,2 Gary A. Levy, MD,3 and Atholl Johnston, MD 4 ‘Division of Immunology and Organ Transplantation, University of Texas Health Science Center at Houston Medical School, Houston, Texas, 2Vancouver General Hospital and University of British Columbia, Vancouver British Columbia, 3Division of Gastroenterology, Toronto General Hospital, University Health Network, University of Toronto, Toronto, Ontario, Canada, and 4Clinical Pharmacology, St. Bartholomew’s Hospital and the Royal London School of Medicine and Dentistry, London, United Kingdom

ABSTRACT Background: Therapeutic drug monitoring (TDM) is essential to maintain the efficacy of many immunosuppressant drugs while minimizing their toxicity. TDM has become more refined with the development of new monitoring techniques and more specific assays. Objective: This article summarizes current data on TDM of the following immunosuppressant drugs used in organ transplantation: cyclosporine, tacrolimus, sirolimus, everolimus, and mycophenolate mofetil. Methods: Published data were identified by a MEDLINE search of the Englishlanguage literature through March 2001 using the terms therapeutic drug monitoring, cyclosporine, tacrolimus, sirolimus, everolimus, and mycophenolate mofetil. Relevant conference abstracts were also included. Resdts: TDM of cyclosporine has been well studied, and recent findings indicate that monitoring of drug levels 2 hours after dosing is a more sensitive predictor of outcome than trough (C,) monitoring. C, levels are being used more widely in TDM of tacrolimus; however, the relationship between C, and area under the curve has varied widely in clinical trials, with correlations ranging from 0.11 to 0.92. The use of TDM of sirolimus, everolimus, and mycophenolate mofetil is evolving rapidly. Conclusions: TDM of immunosuppressant drugs that have a narrow therapeutic index is an increasingly useful tool for minimizing drug toxicity while maximizing prevention of graft loss and organ rejection. Key words: therapeutic drug monitoring, TDM, immunosuppressant therapy, transplantation, cyclosporine, C,. (Clin Ther 2002;24:330-350) Accepted

for publication

September

18, 2001.

Printed in the USA. Reproduction in whole or part is not permitted.

330

0149-2918/02/$19.00

B.D. KAHAN ET AL.

INTRODUCTION Optimal immunosuppressant drug therapy is essential for maintaining a viable organ allograft. The therapeutic approach has evolved from the use of nonspecific steroids and cytotoxic agents to the use of drugs targeted to modulate T-cell function, the prototype of which was the immunosuppressant cyclosporine. Hariharan et al’ reported a substantial improvement in survival of >90,000 US renal transplant patients and their grafts between 1988 and 1996, the period during which cyclosporine was widely introduced.

IMPORTANCE OF THERAPEUTIC DRUG MONITORING Among the reasons for the improved outcomes seen in transplant recipients is the application of therapeutic drug monitoring (TDM). Cyclosporine and the immunosuppressants tacrolimus, sirolimus, and mycophenolate mofetil (MMF) are critical-dose drugs2a; that is, they have a narrow therapeutic index, exhibiting the desired therapeutic effect with acceptable tolerability only within a narrow range of blood concentrations.* Furthermore, they exhibit a high degree of interindividual and intraindividual pharmacokinetic and pharmacodynamic variability, which increases the possibility of therapeutic failure if these agents are used at uniform doses in all patients.2,3 The correlation between drug concentrations and clinical outcomes is an important factor supporting the use of TDM of immunosuppressant drugs.5-8 With cyclosporine, for example, systemic exposure (measured as the area under the concentrationtime curve [AUC]) correlates with the occurrence of acute rejection episodes and

graft survival in renal allograft recipients.5 Furthermore, intrapatient variability in cyclosporine exposure represents a risk factor for chronic rejection9 and is associated with increased treatment costs.10 Algorithms that use concentrations during the first 4 hours after dosing (C,)” or at the single sampling point of 2 hours after dosing (C,) to estimate AUC have been proposed as correlating with acute rejection episodes during the first 90 days in recipients of renal and hepatic transplants.8 There is mounting evidence that TDM is critical to optimizing immunosuppression and minimizing toxicity with tacrolimus,‘**i3 sirolimus,14-16 and MMF.*,” This review examines current data on the available methods for and clinical applications of TDM of the following immunosuppressant drugs used in organ transplantation: cyclosporine, tacrolimus, sirolimus, everolimus, and MMF. Published data were identified by a MEDLlNE search of the English-language literature through March 2001 using the terms therapeutic drug monitoring, cyclosporine, tacrolimus, sirolimus, everolimus, and mycophenolate mofetil. Relevant conference abstracts were also included.

CYCLOSPORINE TDM of cyclosporine has been accepted as an essential tool in the management of transplant recipients for almost 20 years. The availability of simple, sensitive assays that measure amounts of parent drug in blood samples (Table I) permits the use of whole-blood cyclosporine concentrations to individualize dosing regimens according to pharmacokinetic principles.‘* The most widely used index is the predose, or trough, concentration (Co).i8 Although

331

CLINICAL THERAPEUTICS’

Table I. Commercially

available assays for cyclosporine.

5pe

Sensitivity, ng/mL

Cost per Sample, $

TDX

Fluorescence polarization immunoassay

25

5

Abbott Laboratories, Abbott Park, Ill

EMIT

Homogeneous enzyme immunoassay

40

7

Dade Behring Inc, Deerfield, Ill

CYCLO-Trac

Radioimmunoassay

10

9

DiaSorin Inc, Stillwater, Minn

Name

guidelines for the use of target C, levels in the TDM of cyclosporine were developed, it was subsequently recognized that these values did not correlate with the AUC.19*20

AUC Monitoring as a Measure of Drug Exposure Cyclosporine exposure, as represented by AUC, is a more sensitive method than Co for predicting clinical outcome in de novo renal transplant recipients.21 In 160 renal transplant recipients, cyclosporine bioavailability <25%, clearance >325 mL/min, steady-state concentrations ~350 ng/mL during IV infusion, and average concentrations (C,) ~400 ng/mL during oral therapy were all found to be associated with lower rates of graft survival 1 year after transplantation and/or a higher incidence of acute rejection.5 The relationship between exposure and outcome was also seen in a study in 95 de novo renal transplant recipients22; mean cyclosporine AUC values in 39 patients who experienced a rejection episode at 12 months were 3023 ng*h/mL, compared with 10,042 ng*hlmL in 56 patients who did not. Results of another study confirmed that patients with an initial C, 2550 ng/mL had significantly higher 6-year

332

Manufacturer

graft survival rates than patients with an initial C, c550 ng/mL.23 An analysis of cyclosporine pharmacokinetics in a cohort of 204 renal transplant recipients identified intrapatient variability in cyclosporine exposure (% coefficient of variation [CV] = 100 times the quotient of the SD and the mean value over a 5-year period) as a significant risk factor for chronic rejection.‘O Receiveroperating-characteristic curves demonstrated that 27% of the total risk for the occurrence of chronic rejection was attributable to a >28.4% CV for the C,. An economic analysis of total costs over the 5 years after discharge following transplantation showed total mean facility costs and physician costs of $48,789 per patient in the group with less variability in cyclosporine exposure, compared with $60,998 per patient in the group with greater variability. lo Low cyclosporine exposure was independently shown to contribute to an increased risk for chronic rejection in 65 renal transplant patients.24

Cyclosporine Microemulsion and the Use of Sparse-Sampling Techniques The inconvenience and added cost of obtaining multiple blood samples over a

B.D. KAHAN ET AL.

1Zhour dosing period constituted important barriers to the acceptance of cyclosporine AUC monitoring as a routine TDM tool. Sparse-sampling algorithms allow estimation of drug exposure through the use of a limited number of sampling points to calculate AUC. Although Johnston et alz5 introduced the concept of the sparse-sampling algorithm for use with cyclosporine, the large intraindividual variation with the Sandimmune formulation (Novartis Pharmaceuticals Corporation, East Hanover, NJ) meant that the potential of this algorithm as an effective TDM tool was not realized until development of a microemulsion formulation.2c28 This method uses 2 samples: a 2-hour sample for estimation of the drug-absorption rate and a 6-hour sample for estimation of the drug-clearance rate. An alternative sparse-sampling algorithm was used in a multicenter Canadian study that measured only the absorption phase.29,3oA recent review of limited-sampling strategies concluded that many algorithms are suitable for accurate prediction of the AUC.3’ Absorption Profiling Using Absorption-Phase AUC

Current evidence suggests that use of the absorption-phase AUC (AUC,) of the cyclosporine microemulsion formulation allows prediction of early clinical outcomes.7,11An analysis of pharmacokinetic profiles found that the greatest degree of intrapatient and interpatient variability in cyclosporine pharmacokinetics occurred within the first 4 hours after dosing.32 The mean percentage CV was higher in the first 2 hours after dosing (9%-22%) than 4 to 12 hours after dosing (-5%). Thus, a strategy that used 2 sampling times0 and 2 hours after dosing-showed a bet-

ter correlation with 12-hour AUC than did the single value, Co (12= 0.945 vs 0.53).30 A Canadian multicenter study30 comparing the oil-based and microemulsion formulations of cyclosporine in a subset of 421 renal transplant recipients demonstrated that AUC, was a sensitive surrogate for the full AUC of the cyclosporine microemulsion and displayed less variability than Co over a 6-month period. The point of greatest pharmacodynamic effect (calcineurin inhibition) has also been shown to occur within the first 2 hours after dosing, with peak cyclosporine levels of 800 to 2285 ng/mL producing 70% to %% calcineurin inhibition at 1 to 2 hours.33 Furthermore, Sindhi et al34 reported that maximum suppression of interleukin-2 release from T cells occurred 2 hours after dosing. Therefore, the AUC, may be a sensitive indicator of pharmacokinetic and pharmacodynamic differences between patients. Results of a retrospective analysis correlating pharmacokinetics with clinical outcomes in 156 de novo renal transplant recipients suggested that compared with Co, AUC, was a superior predictor of the incidence of acute rejection episodes within the first 3 months after transplantation.” Mean AUC, values were significantly lower in patients with acute rejection episodes than in patients who did not experience acute rejection (mean f SEM, 3934 + 306 vs 4802 f 166 ng*h/mL; P = 0.006). In contrast, mean Co values between the 2 groups showed no significant difference (mean f SEM, 293 f 21 vs 294 * 11 ng/rnL). In another study,” AUC, values between 4400 and 5500 ngh/mL appeared to be associated with a lower incidence of drug-induced renal dysfunction. In a subsequent prospective study in 89 de novo renal transplant re333

CLINICALTHERAPEUTICS”

cipients,’ those who achieved the target AUC, by the third day after the introduction of cyclosporine microemulsion had a 4% rate of acute rejection episodes, compared with 41% in patients who did not achieve this threshold value (P c 0.001). Selection of the C, Data Point for TDM

In a study in 188 de novo hepatic transplant recipients randomized to receive either the oil-based or microemulsion formulation of cyclosporine,35 it was only in the latter cohort that a high correlation was observed between C, concentrations at days 5 and 10 after transplantation and AUC from 0 to 6 hours after dosing (3 = 0.93), as well as a low incidence of acute rejection episodes. A series of studies have shown that C, usefully predicts AUC,. An open-label, randomized, parallel-group study in 204 de novo renal transplant recipients at 20 transplant centers found that C, correlated with AUC, (3 = 0.85), whereas C, and C, showed inferior rates of prediction (3 = 0.12 and 0.70, respectively).36 Among 9 de novo pediatric hepatic transplant recipients, C, showed a better correlation with AUC, than did C, (9 = 0.89 vs 0.03, respectively).37 In 27 stable (at least 3 months after transplantation) pediatric hepatic transplant recipients, C, was also a better predictor than C, (3 = 0.93 vs 0.53, respectively).37 In a subsequent single-center study in 164 renal transplant recipients whose condition was stable 3 months after transplantation,38 the 28 patients whose cyclosporine dose was reduced because of elevated C, levels subsequently showed improved renal function (mean [&SD] serum creatinine, from 164 + 45 to 125 f 20 pmol/L; P c 0.01) with no evidence of 334

acute rejection episodes. Thus, C, monitoring seems to represent a sensitive and practical tool for TDM of cyclosporine after transplantation. Correlation Between C, and Clinical Outcomes

In a Canadian study,39 the 26 renal transplant recipients who had no episodes of rejection 1 month after transplantation had a mean (*SD) cyclosporine C, of 1.9 -c 0.5 kg/L at day 7, compared with 1.1 + 0.2 pg/L for the 10 patients who experienced rejection by day 28 (P c 0.001). Patients who achieved a C, of >1.5 pg/L by day 7 after transplantation had no rejection episodes, compared with a 58% incidence of rejection in those who did not achieve such concentrations (P < 0.001). In the open-label, randomized, parallelgroup study in 204 de novo renal transplant recipients, 36Cox regression analysis showed a trend toward a correlation between cyclosporine C, levels and probable freedom from rejection at 90 days.36,39 Those patients who absorbed the drug poorly and were thus considered at higher risk for inadequate immunosuppression but had C, levels > 1.7 p,g/L at day 3 had a 20% rejection rate within 3 months after transplantation, compared with a 40% incidence of rejection in those with a C, of
B.D. KAHAN ET AL.

C, arm had an -25% lower incidence of acute rejection episodes compared with the C, group (23.6% vs 31.0%, respectively; P = NS) and a significantly lower incidence of moderate and severe (grades II and III) acute rejection episodes (P = 0.01). Tolerability and safety profiles were similar in both groups. A recent study found that early acute rejection episodes occurred in 2 of 30 patients (7% incidence) who achieved target C, values 2 1.O p,g/rnL within 3 to 5 days after transplantation.41 Both of these episodes were mild and were reversed by a single course of high-dose steroids. In summary, monitoring of cyclosporine C, values in hepatic and renal transplant recipients represents a more sensitive predictor of susceptibility to acute rejection episodes than does monitoring of C, values.

Need for C2 Conversion Factors for Different Cyclosporine Assays Cyclosporine assays differ in their ability to accurately measure concentrations of parent drug in whole blood (Table I).42-44 These differences are the result of

systematic errors in calibration and variable cross-reactivities of the reference antibodies, as well as of their assay systems, to specific metabolites. This raises questions of whether the differences between the available immunoassays are important in clinical practice and whether it is necessary to compensate for these betweenassay differences by adjusting the target therapeutic ranges used for cyclosporine TDM-in other words, are the cyclosporine target ranges assay specific? Results of the initial comparisons between cyclosporine assays based on measurements of C, cannot be applied to measurements of C,, because the latter concentrations contain a smaller fraction of metabolites and thus demonstrate less metabolite interference and less divergence between assays. Table II compares C, and C, values obtained with 3 assay methods in an analysis of 108 paired blood samples from renal transplant recipients and confirms that C, target ranges are less assay specific: the mean variation was 89% to 104% for C,, compared with 94% to 122% for C,. 45 Similarly, among 165 samples from hepatic transplant recipients, C, values were relatively assay in-

Table II. Comparison of results obtained with 3 methods for assaying cyclosporine concentrations at C, (predose, or trough) and C, (2 hours after dosing) in 108 paired blood samples from renal transplant recipients.45 Method TDx EMIT HPLCIMS/MS

C,, Mean % (95% CI)

C,, Mean % (95% CI)

122 (1 W-136) 106 (95-l 17) 94 (85-105)

104 (94-l 15) 102 (92-l 13) 89 (80-99)

Results are expressed as a percentage ratio to the radioimmunoassay. TDx = a fluorescence polarization immunoassay (Abbott Laboratories, Abbott Park, Ill); EMIT = a homogeneous enzyme immunoassay (Dade Behring Inc, Deeriield, Ill); HPLC/MS/MS = high-performance liquid chromatography with tandem mass-spectrometric detection.

335

CLINICAL THERAPEUTICS”

dependent (Figure 1A) compared with C, values (Figure 1B). 45 Therefore, no conversion factor appears to be necessary to adjust for differences in the assays used to determine cyclosporine C, concentrations. Recommendations for C, Monitoring A whole-blood sample has been shown to display a value within 10% of the true concentration within 10 minutes of the 2-hour time point following the morning dose of cyclosporine microemulsion (Figure 2). Furthermore, it is currently proposed that the dose of cyclosporine microemulsion be individualized using linear extrapolation“‘j to achieve concentrations within the target range (Table III).38 Although use of this guideline seemed to reduce the incidence of acute rejection episodes in the initial trial~,~*~’ their impact on long-term efficacy and toxicity is not yet clear. Physicians should use prudence in tailoring cyclosporine regimens to individual patients.

TACROLIMUS Like cyclosporine, tacrolimus has a narrow therapeutic index; its absorption is erratic and incomplete, resulting in high intrapatient and interpatient variability.47 These characteristics suggest the usefulness of monitoring tacrolimus blood concentrations to optimize immunosuppression.48 The lack of a sensitive commercial assay has been an obstacle to exploration of tacrolimus TDM. Pro-Trac I (DiaSorin Inc, Stillwater, Minn), until recently the only commercially available assay, had a modest sensitivity of 5 ngAnL, representing the lower therapeutic limit of the drug when used as primary therapy.4g Table IV lists the current commercially available assays.

336

Assays such as the Pro-Trac II (DiaSorin) enzyme-linked immunosorbent assay (ELISA) and the IMx Tacrolimus II (Abbott Laboratories, Abbott Park, Ill) microparticle enzyme immunoassay (MEIA) are now widely available. These newer assays have lowered the range of sensitivity to ~5 ng/mL, the lower limit of the drug’s range for primary therapy.50 Pharmacokinetics The oral bioavailability of tacrolimus averages 25% (range 4%-89%),47 with peak steady-state blood levels occurring -1.5 hours after dosing.13 As with cyclosporine, tacrolimus is metabolized by the cytochrome P450 (CYP) 3A4 isozyme, and its potential for drug-drug interactions seems similar to that of cyclosporine. One exception is that tacrolimus does not seem to inhibit MMF absorption as cyclosporine does,13 leading to greater mycophenolic acid (MPA) exposure when tacrolimus and MMF are used in combination. Available data on the pharmacokinetic variability in tacrolimus concentrations in the early posttransplantation period are both limited and inconsistent, possibly because of interassay differences. Ihara et a14g reported a 38.8% intrapatient variability and 85.8% interpatient variability in AUC among renal transplant recipients. Another study in renal transplant recipients receiving a fixed dose of tacrolimus reported a 50% interpatient variability in Co whole-blood concentrations.48 When the tacrolimus AUC was estimated based on 17 sequential samples from 1 patient,49 intrapatient variability was -45% for Co and -39% for AUC,,, when normalized for dose. In addition, a substantial degree of variability in the pharmacokinetic prop-

B.D. KAHAN ET AL.

A

0

T I

I I CEDIA+

I EMIT

I

I TDx

HPLChlS

Assay Method

B 150

140 1

80

1

I CEDIA+

I

I

I EMIT

HPLC/MS

I

I

TDx

Assay Method

Figure 1. Comparison of results obtained with various assay methods for cyclosporine at (A) C, (2 hours after dosing) and (B) C, (predose, or trough) in 165 paired blood samples from hepatic transplant recipients. RIA = radioimmunoassay; CEDIA+ = cloned enzyme donor immunoassay; EMIT = a homogeneous enzyme immunoassay (Dade Behring Inc, Deerfield, Ill); HPLCYMS = highperformance liquid chromatography with mass-spectrometric detection; TDx = a fluorescence polarization immunoassay (Abbott Laboratories, Abbott Park, Ill).

331

CLINICAL THERAF’EUTICS”

60 70 1

0

6o/h 0

50 40 30 20

10

1

0

+

+

I

1

h

III

35

40

45

I

50

I

55

-

I

2h

I

I

I

I

I

I

5

10

15

20

25

2h 30 min

30 min

Time After Dosing

Figure 2. Timing accuracy error from 1 hour 30 minutes to 2 hours 30 minutes after the morning dose of cyclosporine microemulsion. erties of tacrolimus is suggested by the wide range of oral doses (l-44 mg/d) used to maintain Cc values between 5 and 20 ng/mL in clinically stable hepatic and renal transplant patients.47 No clear relationship has been observed between tacrolimus dose and C, values.47

Guidelines for COMonitoring With a single exception,51 the results of clinical studies suggest that C, levels of tacrolimus are to some extent predictive of efficacy and toxicity.52*53The manufacturer recommends using C, values for

Table III. Target C, levels (concentrations 2 hours after dosing) for cyclosporine microemulsion therapy.38 Patient Population

Target C, Levels, pg/mL

Hepatic transplant recipients Time after transplantation, mo O-3 3-6 6+ Renal transplant recipients Time after transplantation, mo 1 2 3 4-6 7-12 12+

338

1.0

0.8 0.6

1.7 1.5 1.3 1.1 0.9 0.8

B.D. KAHAN ET AL.

Table IV. Commercially

available assays for tacrolimus. Sensitivity, 5pe

ng/mL

Cost per Sample, $

Pro-Trac II

Enzyme-linked immunosorbent assay

0.18

12.50

DiaSorin Inc, Stillwater, Minn

EMIT 2000 Tacrolimus

Homogeneous enzyme immunoassay

1.5

13.50

Dade Behring Inc. Deerfield, Ill

IMx Tacrolimus II

Microparticle enzyme immunoassay

1.5

13.50

Abbott Laboratories, Abbott Park, Ill

Name

monitoring adult renal transplant recipients (months l-3: 7-20 ng/rnL; months 4-12: 5-15 ng/mL), adult hepatic transplant recipients (months 1-12: 5-20 ng!mL), and pediatric hepatic transplant recipients (months 1-12: 5-20 ng/mL).50 Three studies53-55 have reported that Co concentrations of tacrolimus in whole blood correlate with both effectiveness and toxicity in renal and hepatic transplant recipients. Tacrolimus Co levels >25 ng/mL (measured by ELISA) were associated with a significantly greater risk of nephrotoxicity in renal transplant recipients.54g55 Statistically significant relationships were observed between tacrolimus Co and the incidence of acute rejection episodes (P = 0.02) and toxicity (P = 0.01) in 92 renal transplant recipients.12 In >700 hepatic transplant recipients, Co levels showed a significant correlation with drug-induced toxicity (P c 0.01) but not with efficacy. l2 Results of a retrospective analysis in de novo renal transplant recipients suggested that a reduced risk of rejection episodes was correlated with a Co concentration of 210 ng/mL achieved by day 2 or 3 after transplantation.53 However, another study observed no useful relationship between tacrolimus Co and clinical outcomes.51

Manufacturer

As with cyclosporine, tacrolimus Co may not be a good predictor of outcomes in hepatic12 and renal transplant recipients.51 In an open-label, concentrationranging trial in 120 renal transplant recipients with low, medium, or high tacrolimus Co values, there was no significant relationship between any of the Co subgroups and the incidence of biopsyproven rejection in the first month after transplantation. 51 However, when the data were reexamined using logistic regression analysis of maximum and minimum Co levels and outcomes, a significant relationship was identified between Co and episodes of acute rejection. In a multicenter study in 270 hepatic transplant recipients using MEIA (lower limit of detection, 0.5 ng/mL in whole blood),13 no significant relationship was observed between tacrolimus blood levels and selected adverse events or serum creatinine levels in the 6 months after transplantation.

Relationship Between Tacrolimus Trough Concentrations and AUC Studies of the relationship between tacrolimus Co and AUC have reported a wide range of correlations, from ? = 0.1 156 to 0.92.47 In contrast, concentrations mea-

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CLINICAL THERAF’EUTICS”

sured precisely at the end of the dosing interval-C,,-have been shown to provide a more robust correlation than C,, with an r-zof 0.89 in de novo hepatic transplant recipients. 57 In a study in 67 recipients of renal, hepatic, or bone marrow transplants, a correlation of 0.76 was reported between steady-state AUC and C,,.58 The correlation between C, and AUC was 0.92 in a limited cohort of small bowel and renal transplant recipients, and 0.86 in de novo renal transplant recipients.59 These differences, although their cause is unclear, may have profound implications for the use of C, levels for TDM of tacrolimus. Indeed, a recent pharmacokinetic study using the sensitive IMx Tacrolimus II immunoassay (lower limit of detection, 1.5 ng/mL) in 18 stable renal transplant recipients observed a low correlation between AUC and C, (9 = 0.1 l), whereas the best single sampling point was C, (? = 0.81).56Based on use of a sparse-sampling method, the authors concluded that tacrolimus concentrations at 2 and 4 hours after dosing provided the best estimate of tacrolimus exposure. A more recent study used logistic regression analysis to examine the relationships between tacrolimus whole-blood concentrations and acute rejection, nephrotoxicity, and adverse reactions requiring reductions in the tacrolimus dose in 111 hepatic transplant recipients.60 In the 0- to 7-day window in which primary end points were examined, a significant relationship was seen between tacrolimus whole-blood levels, a decreasing risk of acute rejection (P = 0.047), and an increasing risk of nephrotoxicity (P c 0.001). Unfortunately, there are no prospective studies of the relationship between tacrolimus exposure as estimated by AUC and clinical outcomes that would provide an 340

index of the optimal benefit to be obtained from TDM. SIROLIMUS Sirolimus is a potent immunosuppressive agent with a narrow therapeutic index. Although the randomized, placebocontrolled nature of pivotal trials means that uniform doses were used to maintain blinding, postapproval use has been based on the application of TDM principles. Table V lists selected analytic methods for the measurement of sirolimus in blood samples.61~62Neither the MEIA nor the immunophilin-binding assay mentioned there is currently available. All other methods measure parent compound exclusively and have been validated against each other. High-performance liquid chromatography with ultraviolet detection (HPLC-UV) methods of analysis are highly precise and accurate and have been used in a number of clinical trials,” but their high cost means that few institutions are able to use HPLC-UV for routine patient management. Nonetheless, the continued unavailability of a commercial assay for sirolimus means that more institutions may adopt HPLC techniques. Phamacokinetics The pharmacokinetic properties of sirolimus have been well described and cannot be predicted based on patients’ demographic characteristics.14*15s63Indeed, its pharmacokinetic parameters have been reported to be similar in African American and white transplant recipients.@ The oral absorption of sirolimus is -14%, with a time to peak absorption ranging from 0.7 to 3 hours after dosing.‘j5Analysis of data in a recent review suggests that sirolimus

B.D. KAHAN ET AL.

Table V. Selected assays for sirolimus.61y62

Assay 5pe

Sensitivity,

Calibration Range,

@mL

@mL

Specificity

MEIA

3

IBA

2.5

2.540

Weak metabolite cross-reactivity

HPLC-UV I II III

2 2.5 6.5

2-50 2.5-25 6.5-356

Specific for sirolimus* Specific for sirolimus* Specific for sirolimus*

HPLC/MS/MS

0.2

0.2-100

Specific for sirolimus

3-30

Metabolite cross-reactivity

MEL4 = microparticle enzyme immunoassay; IBA = immunophilin-binding assay; HPLC-W liquid chromatography with ultraviolet detection; HPLCIMSIMS with tandem mass-spectrometric detection. *Avoidance of nonspecific interference is necessary.

exhibits much the same pharmacokinetic variability as cyclosporine and tacrolimus.16 In a 4-year study in 150 renal transplant recipients, l6 both intrapatient and interpatient variability were high (dose-corrected AUC, 63.8% and 59.5%, respectively). Sirolimus is metabolized by the CYP3A4 isozyme, and its potential for drug-drug interactions is similar to that of cyclosporine, which competes for the same enzyme substrate.‘j6 Because concurrent administration of sirolimus with high doses of cyclosporine microemulsion (but not the oil-based formulation) results in an 80% increase in sirolimus AUC,67 it was advocated that administration of these 2 agents be separated by 4 hours in the pivotal blinded triak6* However, when using low doses of either cyclosporine or tacrolimus, spacing of drug delivery is neither important nor necessary as long as sirolimus TDM is used. TDM of Sirolimus Preliminary data using HPLC-tandem mass-spectrometric detection showed that

= high-performance

= high-performance liquid chromatography

sirolimus C, concentrations correlate with steady-state AUC exposure in stable transplant recipients. 69 Because a single-center experience documented a high correlation between sirolimus C, and AUC, using an HPLC analytic method in both de novo and stable renal transplant recipients (? = 0.946),‘O current strategies are based on Co values. Analysis of 150 patients treated at a single center indicated that sirolimus Co levels correlated with acute rejection episodes. l6 Using an assay selective for parent compound, the investigators found that patients with acute rejection episodes had mean sirolimus Co values 16 ng/mL, whereas patients who did not experience rejection episodes had Co levels of 210 ng/mL. A retrospective review of patients in the pivotal trials who received sirolimus in combination with regular doses of cyclosporine sufficient to produce usual target Co values suggested that effective prophylaxis against acute rejection episodes occurred within the Co range of 5 to 15 ng/mL using whole-blood MEIA analyses, which yield concentrations almost

341

CLINICAL THERAPEUTICS@

twice those of the selective assays.67 This review also suggested that dose was not a sensitive predictor of C, levels, indicating that individualization of doses is necessary. Mean (*SD) C, values in patients who did and did not experience rejection within the first 75 days after transplantation were 8.7 + 7.8 and 12.8 f 7.7 ng/mL (MEIA; P c O.OOl), respectively. However, there was also a significant difference in mean (&SD) cyclosporine C, levels between the 2 groups: 302 f 160 and 346 f 139 ng/mL, respectively (P< 0.004). Therefore, isolating the therapeutic effects of cyclosporine from those of sirolimus was not possible in this study analysis. However, a subsequent article using these data in a median-effect analysis showed direct relationships between drug concentrations and efficacy versus toxicity, as well as demonstrating a synergistic interaction between the 2 drugs.16 TDM of sirolimus was performed in a study of islet transplantation in 7 patients with type 1 diabetes mellitus.‘l Patients received a loading dose of sirolimus 0.2 mg/kg, followed by a daily dose of 0.1 mg/kg, and blood levels were monitored to maintain a target range of 12 to 15 ng/mL for the first 3 months and 7 to 10 ng/mL after 3 months. Tacrolimus was given concurrently at an initial dose of 1 mg BID, titrated to a 12-hour trough concentration (IMx Tacrolimus II immunoassay). Daclizumab was given at 1 mgkg every 14 days for a total of 5 doses. All patients achieved insulin independence and continued to be independent of insulin through the time of last follow-up (median time, 11.9 months), indicating that sirolimus and tacrolimus TDM was effective in this patient population. A single-center study examined a combination of sirolimus and low-dose tacro-

342

limus in 42 hepatic transplant recipients.72 The dose was titrated to target C, levels of 7 ng/mL for sirolimus and 5 ng/mL for tacrolimus. Patient (92.9%) and graft survival (90%) were good after a mean of 14 months. Larger studies are ongoing. Future Research on Sirolimus TDM Although there appears to be a relationship between sirolimus C, and clinical outcomes, the value of sirolimus TDM needs to be tested prospectively in the clinical setting using a rigorous study design that compensates for the impact of differences in cyclosporine exposure. Despite the early stage of development of sirolimus, available pharmacokinetic and clinical data suggest that sirolimus has good efficacy at target values of 5 to 10 ng/mL for cyclosporine exposures producing 30% reduction in usual target C, or AUC values, 10 to 15 ng/mL for cyclosporine exposures producing 60% reduction, and 220 ng/mL for no cyclosporine exposure.

EVEROLIMUS Everolimus (SDZ RAD) is a new rapamycin analogue currently in Phase III clinical trials for use as a complementary immunosuppressant to cyclosporine-based protocols in transplant recipients.73-75 Its pharmacokinetics have been examined in a multicenter, randomized, double-blind study in 101 de novo renal transplant patients who received everolimus 0.5, 1, or 2 mg BID in combination with cyclosporine and prednisone.76 The steadystate maximum concentration and AUC were both dose proportional throughout the 12-month study. The intraindividual variability in everolimus concentrations was 40.8%,“j lower than the reported vari-

B.D. KAHAN ET AL.

ability in sirolimus concentrations (63.8%) in another trial.16 Everolimus demonstrated a 40.8% intrapatient variability. There will not be an opportunity to develop a clinically relevant method of TDM for everolimus until data from the Phase III trials become available and allow thorough analysis of blood levels and clinical outcomes in renal transplant recipients. The limited information available so far, however, suggests that everolimus TDM may be of benefit.

MYCOPHENOLATE MOFETIL MMF blocks the generation of guanosine nucleotides in the de novo pathway of DNA synthesis in the T cell.” It is a prodrug that after absorption, releases the active drug MPA. Hence, pharmacokinetic and TDM research measure MPA as well as its primary glucuronide metabolite. As summarized in 2 recent reviews,2T’6 TDM of MMF is still at an investigative stage, but the data to date support the use of TDM to optimize immunosuppression. One randomized study analyzed outcomes in patients stratified to receive increasing exposure to MPA based on target AUC values (low-, intermediate-, and high-AUC groups of 16.1, 32.2, and 60.6 pg*h/mL, respectively). Patients in the high-AUC group had the lowest incidence of biopsy-proven acute rejection episodes compared with the low- and intermediate-AUC groups (1 lS%, 27.5%, and 14.9%, respectively) but also had the greatest number of premature withdrawals due to adverse events compared with the other 2 groups (44.2%, 7.8%, and 23.4%, respectively).78 In a single-center study in 29 renal transplant recipients,79 monitoring of MMF AUC was conducted by measuring

blood levels at 0, 1.5, and 6 hours after administration of MMF using a homogeneous enzyme immunoassay (EMIT, Dade Behring Inc, Deerfield, Ill). Large interindividual variability was observed, and the investigators concluded that dose adjustment of MMF based on MPA AUC was appropriate. A dual-arm study in 45 cardiac transplant recipients compared a group that received MMF at a fixed dose of 2 g/d with a TDM group in which target blood concentrations were from 2.5 to 4.5 p&&80 Both groups received concurrent tacrolimus (target concentrations of 10-15 ng/mL) and corticosteroids. Patients in the fixed-dose MMF group had a 66.7% rate of acute rejection episodes, whereas the TDM group had a 10.0% rate. Although no statistical analysis of betweengroup differences was reported, the results indicate that TDM of MMF may be beneficial. Another study in 38 cardiac transplant recipients measured MPA AUC using a 2hour abbreviated AUC, with measurements at 0,20,40,75, and 120 minutes.81 Free MPA and MPA trough values were also determined. Both MPA AUC and free MPA values, but not MPA trough levels, were significantly lower in patients with grade 2 or 3 rejection compared with grade 0 or 1 rejection (P < 0.05). A subanalysis of the pharmacokinetic data was conducted to determine whether a single time point correlated with MPA AUC.82 There was a fair correlation between the 2-hour postdose level and AUC (r = 0.82; P c 0.001) but a poor correlation at 20 (r = 0.1 l), 40 (r = 0.36), and 75 minutes (r = 0.60), and at trough (r = 0.40). Future trials may examine clinical end points using the 2-hour level as a surrogate for the full AUC.

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The correlation between MPA concentrations and side effects was examined in a trial in 31 renal transplant recipients receiving MMF 1 g BID along with cyclosporine and corticosteroids.83 Patients who reported side effects had elevated mean AUC values (62.1 pgh/mL) compared with patients who did not report side effects (39.8 pgh/rnL; P c O.OOl), but there was no significant betweengroup difference in mean minimum concentrations (C,,) (2.29 vs 1.63 l.&nL, respectively). Another analysis by the same authors studied the relationship between MPA concentrations and side effects in 51 renal transplant recipients treated with MMF 0.5 g BID along with tacrolimus and corticosteroids.84 There was a significant difference in C,, at 30 minutes and 1 hour, AUC, and dose-normalized AUC between patients who reported side effects and those who did not (P c 0.01). An enteric-coated formulation of mycophenolate sodium is being developed and is currently in Phase III clinical trial~.~~Mycophenolate sodium is converted directly to MPA, and 720 mg of entericcoated mycophenolate sodium is equivalent to 1000 mg of MMF. This formulation of MPA also has the potential to benefit from TDM. It has been suggested that because of the mounting clinical evidence for a relationship between MPA AUC and rates of acute rejection, as well as intraindividual differences in MPA pharmacokinetics due to concomitant disease states and medications, TMD of MPA will become the standard of care in the future.77 CONCLUSIONS

Many immunosuppressive drugs have a narrow therapeutic index and exhibit vari344

able pharmacokinetics. Effective therapy requires individualization of dosing based on TDM, and effective TDM depends on knowledge of a drug’s pharmacokinetics and correlations with clinical outcomes. Cyclosporine is the most extensively studied immunosuppressive agent in transplantation, and a large body of data supports the value of using TDM surrogates for clinical effects of this drug. Recent findings suggest that monitoring of C, levels is a sensitive and practical technique for cyclosporine microemulsion therapy in renal and hepatic transplant recipients, as sampling at this time point has been shown to correlate with both pharmacokinetic and pharmacodynamic effects. Similarly, abundant data support the use of C, measurements of sirolimus to optimize clinical outcomes in combination with cyclosporine. It is expected that similar findings will emerge as everolimus achieves more widespread clinical use. TDM seems to be useful in tacrolimus regimens, and C, values have become the standard for optimization of therapy. However, a lack of pharmacokinetic and clinical TDM research has hampered the exploration of more sensitive monitoring tools. This deficiency may be due both to the unavailability of a sensitive commercial assay and to the conflicting data that have emerged from clinical studies of the relationship between blood levels and clinical outcomes. As the clinical community becomes more experienced with the management of tacrolimus therapy, the role of TDM should be clarified. Finally, the preliminary data on MMF suggest that TDM may be useful in individualizing immunosuppressive regimens and minimizing dose-related adverse events in transplant recipients.

B.D. KAHAN ET AL.

Further research into the pharmacokinetics of these agents and their relationship to outcomes is needed before art effective TDM approach can be developed and its importance in transplantation established.

ACKNOWLEDGMENT

This article was funded by an unrestricted educational grant from Novartis Pharma AG, Basel, Switzerland.

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Address correspondence to: Barry D. Kahan, MD, PhD, Professor

of Surgery and Director, Division of Immunology and Organ Transplantation, University of Texas Health Science Center at Houston Medical School, 643 1 Farmin, Suite 6.240, Houston, TX 77030. E-mail: [email protected]

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