Volume 23 Number 6, Part 2 December 1990
Management of patients and side effects during cyelosporine therapy
often unpredictable and variable. A patient with "brittle" psoriasis that tended to relapse rapidly on discontinuation of other previous therapies will often have a similar response to cyclosporine cessation. Such patients may flare within a week after the drug has been stopped. Some
of these patients may get psoriasis in places that they have not had it, such as on the face or periungual area. Other patients, however, may go months with less than 1% body surface involvement after discontinuingcyclosporine.
II
I
Laboratory monitoring of cyclosporine levels: Guidelines for the dermatologist Gary Mockli, MD, Pokar M. Kabra, PhD,I" and Theodore W. Kurtz, MD
San Francisco, California The following guidelines are recommended for laboratory monitoring of circulating levels of cyclosporine in dermatology patients. Measurements should be determined as trough levels in whole blood, not plasma or serum. The measurements should be performed with an assay that is specific for the parent cyclosporine compound (e.g., a high-performance liquid chromatography method or a specific monodonal immunoassay). The results of nonspecific immunoassays that detect cyclosporine as well as its metabolites are difficult to interpret and cannot readily be compared among different studies or laboratories. In psoriasis patients, the circulating concentration of cydosporine does not correlate reliably with the therapeutic response. Some patients may achieve an excellent response with blood levels in the range of 50 ng/ml; others may show little or no response despite blood levels as high as 200 ng/ml. In patients with a poor clinicalresponse, monitoring of cyclosporine levels may be useful to confirm that the drug has been taken and may provide an estimate of the degree of absorption and metabolism of the parent compound. Because an upper limit of safety for the circulating concentration of cyclosporine has not been dearly defined, one should attempt toachieve a therapeutic response with the lowest possible dose. Clinicians must carefully monitor patients for signs of cydosporine toxicity, regardless of the circulating concentration of the drug. Whole blood levels exceeding 250 ng/ml should be avoided. (J AM ACAD DERMATOL 1990;23:1275-9.)
In transplant patients, measuring circulating concentrations of cyclosporine has been widely advocated as a way to ensure adequate immune suppression and to minimize toxicity. 1-3 In this review, we focus on the issue of laboratory monitoring of cyclosporine levels in the management of dermatology patients. From the Department of Laboratory Medicine, University of California, San Francisco, Reprint requests:Theodore W, Kurtz, MD, Department of Laboratory Medicine, University of California, Box 0134, San Francisco, CA 94143-0134. 1"Deceased.This work is dedicated to the memory of Dr. Pokar Kabra, a pioneer in the use of liquid chromatography for therapeutic drug monitoring, 16/0/23798
METHODOLOGIC CONSIDERATIONS To interpret measurements of circulating concentrations appropriately, several problems of methodology must be considered. Type of sample: Whole blood versus plasma A t physiologic temperature, a large fraction of the cyclosporine in blood is distributed in erythrocytes. The partitioning of the drug between erythrocytes and plasma is both temperature and time dependent. For this reason, the same plasma sample can indicate highly variable blood levels, depending on the sample temperature and the time involved in collection and processing. To minimize this variability and to facilitate comparisons among different laboratories, we recommend that circulating concentrations 1275
Journal of the American Academy of Dermatology
1276 M o c k l i et al. of cyclosporine be measured in whole blood, preferably with EDTA used as the anticoagulant. 1,2 Although most laboratories have begun to follow this procedure, a substantial proportion (10% to 20%) may still be measuring cyclosporine in plasma. 4 Measurement .in whole blood instead of plasma also may minimize variation in cyclosporine levels that might be associated with changes in hematocrit. 1 Because the drug levels in whole blood are greater than in plasma, it is important to exercise caution when comparing studies in which different "therapeutic" levels of cyclosporine are reported. There is no universal and reliable method for converting a plasma level into an "equivalent" blood level. Because of the variability associated with cyclosporine measurements in plasma, we will focus on studies in which drug levels were measured in whole blood.
Timing of samples Approximately 10% to 50% of an oral dose of cyclosporine is absorbed through the small intestine, and, typically, peak circulating levels are achieved between 2 and 4 hours after administration. 2 Although extensive variability can be observed in and among patients in steady-state trough levels, even greater variability may be expected in peak levels of the drug. For that reason, it is important to monitor trough levels of cyclosporine.1, 2 Ingestion of food may increase or decrease the absorption of cyclosporine.2 It has also been reported that trough levels may vary substantially, depending on whether they are obtained before a morning or an evening dose. 5 These considerations make it necessary to measure each patient's trough levels on a regular schedule.
Type of assay Cyclosporine levels can be determined by irnmunoassay or high-performance liquid chromatography (HPLC). Because these methods may yield widely differing values in the same blood sample, one must know the technology used when comparing cyclosporine values produced by different studies or laboratories. Depending on the type of technology and laboratory, charges for blood level assays range from $20 to $100 per sample.
High-performance liquid chromatography H P L C can provide a specific and precise assay of cyclosporine levels in whole blood?, 6 Its sensitivity
may not be as great as that of some immunoassays, but this is not an important consideration because, for clinical purposes, cyclosporine blood levels are usually well within the sensitivity of most H P L C assays. Most HPLC assays clearly distinguish cyclosporine from its metabolites. Thus the results of such assays reflect the circulating concentration of the parent compound itself and are not affected by changes in metabolite concentrations. As long as appropriate standards are used, it should be possible to compare HPLC measurements of cyclosporine in whole blood across different laboratories and studies. Even so, this method is not widely used in clinical laboratories because it tends to require specialized equipment and expertise. However, in some laboratories, including ours, HPLC is efficiently used to measure cyclosporine levels in large numbers of specimens. In fact, when an automated H P L C injection system is used, a single medical technologist can analyze more than 50 blood samples per day.
lmmunoassay When immunoassay measurements of cyclosporine are interpreted, it is important to know whether they were based on a polyclonal or monoclonal antiserum. A monoclonal antiserum that is specific for cyclosporine is the assay of choice.l, 2, 4 The polyclonal immunoassays detect the parent compound as well as a variety of metabolites. Thus, for a given sample, an assay that uses a polyclonal antibody will yield much higher results than one that uses a specific monoclonal antibody or an H P L C method. Although there is little evidence that cyclosporine metabolites are nephrotoxic, the exact immunosuppressive and toxic nature of the metabolites remains unknown. ~ Furthermore, cyclosporine metabolite profiles may differ from patient to patient. Therefore, it is unclear how the results of polyclonal immunoassays could be used to ensure adequate immune suppression or to minimize toxicity. Several polyclonal assays are available and exhibit variable cross-reactivities with cyclosporine metabolites, l Because these assays may be measuring different metabolites, any comparison of results between studies that use different polyclonal assays can be problematic. Some investigators have proposed that both monoclonal and polyclonal assays be run on the same sample as a means of determining the relative amounts of cyclosporine and its metabolites. 7 However, for now, the clinical utility of
Volume 23 Number 6, Part 2 December 1990
Laboratory monitoring of cyclosporine 1277
such expensive testing is uncertain, and its use should be confined to the research setting. Generally, polyclonal assays use either radioactive labels or fluorescent labels (e.g., the Abbott TDx fluorescence polarization immunoassay). There is no reliable method for estimating the true level of the parent cyclosporine from a result generated with a nonspecific immunoassay. In patients with normal liver function, nonspecific immunoassays usually yield results that are two to four times greater than those of specific immunoassays or HPLC measurements. 2 As might be expected, the results of nonspecific immunoassays correlate poorly with results of HPLC or of monoclonal immunoassays that are specific for cyclosporine, s In contrast, the results of specific monoclonal immunoassays appear to correlate well with those of HPLC. 8' 9 Recently, the accuracy of cyclosporine standards supplied in at least one commercial monoclonal immunoassay kit has been questioned. 1~ Consequently, when selecting a laboratory for measurement of cyclosporine levels, one should determine the type of assay used. How well the laboratory has performed on external proficiency testing surveys is another important piece of information.
Table I. Guidelines for cyclosporine monitoring in psoriasis patients
CLINICAL INTERPRETATION OF CYCLOSPORINE LEVELS
found no evidence of a correlation between clinical response and circulating concentrations. In some patients given 14 mg/kg per day, whole blood levels of approximately 200 ng/ml were associated with complete clearing of severe plaque psoriasis. In oth. ers, much less dramatic clinical responses were observed, although cyclosporine levels were in the range of 250 ng/ml. However, in the preliminary study by Ellis et al., the dose used was well above current recommendations. Consequently, a possible dose-response relationship may have been obscured by the high efficacy of the large dose. Nevertheless, several other investigators have noted that the circulating concentration of this compound does not correlate reliably with the therapeutic response.12"16 In an open trial in psoriasis patients who were given "low"-dose cyclosporine therapy (2 to 4 nag/ kg per day), and in whom whole-blood trough levels of the drug were measured with a nonspecific radioimmunoassay, Griffiths et al) 2 reported that complete clearing of severe lesions could occur with circulating levels as low as 200 ng/ml. Presumably, the nonspecific assay used in this study was detecting cyclosporine metabolites as well as the parent compound. If so, some of the patients may have experi-
In transplant patients, it has been suggested that maintenance doses of cyclosporine be adjusted to produce whole-blood trough concentrations between 80 and 150 ng/ml, as measured by HPLC or drugspecific immunoassays.1 These guidelines have been proposed in the hope of ensuring adequate immunosuppression while minimizing toxicity. Although cyclosporine has been used in transplant patients for some time, it must be recognized that these guidelines are based largely on poorly controlled, retrospective studies. It is known that patients may experience rejection or nephrotoxicity, even when drug levels are in the so-called therapeutic range.1 Given the lack of reliable studies of therapeutic monitoring in transplant patients, it is not surprising that even less information is available on measuring cyclosporine in dermatology patients. In dermatology patients, most of the data on monitoring this drug comes from studies in which it has been used to treat psoriasis. In a double-blind study in psoriasis patients in whom whole-blood trough levels of cyclosporine were monitored by specific HPLC, Ellis et al. 11
Preferred sample: Preferred timing: Preferred method: Therapeutic range:
Toxic range:
Whole blood (EDTA or heparin as anticoagulant) Trough level HPLC or immunoassay specific for cyclosporine Unknown--some patients may respond with blood levels as low as 50 ng/rnl; others may not respond even at 200 rig/m1 The level at which toxicity begins to occur is unknown--patients must be monitored carefully for toxicity regardless of the circulating concentration of cyelosporine; however, persistent levels above 250 ng/ml, when measured as above, may be inadvisable!8
1278
Journal of the American Academy of Dermatology
M o c k l i et aL
enced an excellent clinical response, although the true drug levels were as low as 50 ng/ml. Given the uncertain relation between circulating levels of cyclosporine and clinical response to therapy, why measure the drug at all? In contrast to transplant patients in whom the clinician cannot assess therapeutic effectiveness of cyclosporine until it fails and rejection begins to occur, the dermatologist can directly monitor his patients' clinical response to the drug. I f the patient shows no clinical response, one might check a cyclosporine level to confirm that the patient has taken the drug and that at least some amount has been absorbed. However, as has been noted, the minimum level required to achieve a satisfactory clinical response is unknown and might vary widely among patients. If a patient exhibits a satisfactory clinical response, the only reason to measure a cyclosporine blood level would be to minimize the chance of toxicity. It should be noted that with prolonged oral administration of this compound, the bioavailability m a y increase, thereby causing an unanticipated elevation in blood levels. 2 Unfortunately, an upper limit of safety has not been defined. Obviously, it is desirable to achieve therapeutic response with the smallest possible dose of the drug. In renal transplant patients, it has been reported that whole-blood trough levels below 250 ng/ml (measured by H P L C ) have been associated with acceptable renal function) However, in psoriasis patients, even a "moderate" dose of cyclosporine (approximately 4 to 5 m g / k g per day, a dose that should yield circulating blood levels below 250 ng/ml) can be associated with a decrease in glomerular filtration rate.t7 Furthermore, individual patients m a y vary in their susceptibility to cyclosporine toxicity. Given the foregoing considerations, we believe far less emphasis should be placed on monitoring circulating cyelosporine levels than on monitoring the patient for clinical response and early signs of toxicity. 17 It is incumbent on the clinician to monitor patients carefully for signs of toxicity, regardless of the circulating concentration of cyclosporine. Our recommendations for cyclosporine monitoring are summarized in Table I. REFERENCES 1. Shaw LM. Advances in eyclosporine pharmacology, measurement, and therapeutic monitoring, Clin Chem 1989; 35:1299-308. 2. Shaw LM, Bowers L, Demers L, et al. Critical issues in cy-
closporine monitoring: report of the task force on cyclosporine monitoring. Clin Chem 1987;33:1269-88. 3. Moyer TP, Post GR, Sterioff S, et al. Cyclosporine nephrotoxicity is minimized by adjusting dosage on the basis of drug concentration in blood. Mayo Clin Proc 1988;63: 241-7. 4. Shaw L. Cyclosporinemonitoring. Clin Chem 1989;35:5-6. 5. BowersLD, Canafax DM, Singh J, et al. Studies of cyelosporine blood levels: analysis, clinical utility, pharmacokinetics, metabolites, and chronopharmacology. Transplant Proc 1986;18:137-44. 6. Kabra PM, Wall JH, Blanckaert N. Solid phase extraction and liquid chromatography for improved assay of cyclosporine in whole blood and plasma. Clin Chem 1985; 31:1717-20, 7. Trull AK, Tan KKC, Roberts NB, et al. Cyclosporine metabolites and neurotoxicity [Letter]. Lancet 1989;II:448. 8. Subbarao MN, Swanson JR, Mueggler PA. Cyclosporine determinations in heart and kidney transplant patients: comparison of high-performance liquid chromatography, polyclonal, and monoelonal methods. Ther Drug Monit 1989;11:53-6. 9. WolfBA, Daft MC, Koenig JW, et al. Measurement ofcyclospofine concentrations in whole blood: HPLC and radioimmunoassay with a specifie monoclonal antibody and 3H- or 125I-labeled ligand compared. Clin Chem 1989; 35:120-4. 10. MacBride JH, Rodgerson DO, Park SS, et al. Measurement of cyclosporine in plasma from patients with various transplants: HPLC and radioimmunoassay with a specific monoclonal antibody compared. Clin Chem 1989;35: 1726-30. 11. Ellis CN, Gorsulowsky DC, Hamilton TA, et al. Cyclospofine improves psoriasis in a double-blind study. JAMA 1986;256:3110-6. 12. Griffiths CEM, Powles AV, Leonard JN, et al. Clearance of psoriasis with low-dose cyclosporin. Br Med J 1986; 293:731-2. 13. Heule F, Meinardi MM, van Joost T, et al. Low-dose cyclosporineeffectivein severe psoriasis: a double-blind study. Transplant Proe 1988;20(suppl 4):32-41. 14. Meinardi MM, Bos JD. Cyclosporine maintenance therapy in psoriasis. Transplant Proe 1988;20(suppl 4):42-9. 15. Picascia DD, Garden JM, Freinkel RK, etal. Resistant severe psoriasis controlled with systemic cyclosporine therapy. Transplant Proe 1988;20(suppl 4):58-62. 16. Van Joost T, Bos JD, Heule F, et al. Low-dosecyclosporin A in severe psoriasis: a double-blind study. Br J Dermatol 1988;118:183-90. 17. Gilbert S, Emmett M, Menter A, et al. Cyclosporine therapy for psoriasis: serum creatinine measurements are an unreliable predictor of decreased renal function. J AM ACADDERMATOL1989;21:470-4. 18. A consensusreport: cyclosporin A therapy for psoriasis. Br J Dermatol 1990;122(suppl 36):1-3. DISCUSSION Dr. Ellis. Dr. Kurtz, how often do you think you should cheek for cyclosporine trough levels to see if the presumed level of toxicity has been reached? Dr. Kurtz. With a patient who is just starting cyelosporine therapy, it makes sense to assess the blood level once or twice in the beginning and to use 200 to 250 ng/ml by
Volume 23 Number 6, Part 2 December 1990
HPLC or monoclonal immunoassay as an upper limit. With prolonged therapy, the level should be checked again later if there is concern that the bioavailability of the fixed dose might be changing. However, because we do not know for certain what constitutes a safe level, it's difficult to know whether assessing blood levels frequently will provide important information. In some patients, even a level of 200 ng/ml could be toxic. Checking the blood pressure, checking renal function carefully--these are the most critical monitoring steps. Of course, you judge the clinical response by looking at the patient, not at the blood level. Dr. Ellis. With low doses of cyclosporine of 5 mg/kg per day or less, there is not a good correlation between blood levels and either clinical response or change in renal function. I think overall, particularly at higher doses, the data would indicate that specific blood levels greater than 200 ng/ml have a somewhat higher association with elevated creatinine levels. Dr. Kurtz. Although there is a rough correlation between toxicity and blood levels, in practical terms, managing the individual patient through blood level measurements is not feasible. Dr. Eisen. A recent review article on cyclosporine stated that one of the newer monoclonal radioimmunoassays gives more reliable levels than HPLC.I Dr. Kurtz, I'm aware of that review and I thought it was a sketchy presentation of that point of view. I don't have any objection to specific monoclonal radioimmunoassays, and nonisotopic assays are being developed as well. At least one commercial kit has been criticized because the calibrator supplied with it is said to be inaccurate. So it may be difficult to interpret a target level that is recommended as therapeutically appropriate, based on someone else's study using blood levels of cyclosporine that do not correspond to your laboratory's results. Still, I believe these specific monoclonal assays can be made to correlate well with HPLC, provided the calibration issue is resolved. In the future, we may be able to measure free cyclosporine in the blood more readily. Now we are measuring both bound and free cyclosporine. Whether free cyclosporine would have more clinical value is unknown.
Laboratory monitoring of cyclosporine
1279
Dr. Ellis. If a dermatologist were to prescribe cyclosporine for a patient with pyoderma gangrenosum, for example, what test is the local laboratory likely to use in determining cyclosporine levels? Dr. Kurtz. If it's a neighborhood laboratory, the sample probably will be sent out to a reference laboratory. Generally, if it's a large-volume reference laboratory, an immunoassay will be used because most such labs cannot keep up with the workload involved in performing the HPLC. One can only hope that the level is being measured in whole blood, and this can be ascertained from the report or by asking. Then it will be a question of determining whether the test used is a polyclonal immunoassay or a specific monoclonal immunoassay. I think eventually all laboratories will move to the monoolonal tests but, for now, the clinician has to check that. The practicing dermatologist also must check to see how the laboratory has performed on an external proficiency testing survey. What type of results are they obtaining in comparison with the overall mean of all laboratories? It is wise to see if the laboratory participates in such a proficiency survey. It might be using a reasonable assay but generating a poor result, which would go unrecognized without proficiency testing. Dr. Ellis. When reference laboratories send hack the results, they often give the physician a guideline to a therapeutic range. What does that mean, and can the physician use the guideline? Dr. Kurtz. With respect to cyclosporine, it will often be meaningless. The lab probably will have set its values on the basis of a journal article. Moreover, different labs will have read different articles. The therapeutic range may be unreliable, particularly if the laboratory has based its range on the results of a study performed in another laboratory that uses a different methodology. Finally, few if any laboratories will offer a therapeutic range specifically for the dermatology patient. Most such ranges are intended to guide cyclosporine therapy in transplant patients. REFERENCES
1. Kahan BD. Cyclosporine. N Engl J Med 1989;321: 1725-38.
Management of patients and side effects during cyelosporine therapy
often unpredictable and variable. A patient with "brittle" psoriasis that tended to relapse rapidly on discontinuation of other previous therapies will often have a similar response to cyclosporine cessation. Such patients may flare within a week after the drug has been stopped. Some
of these patients may get psoriasis in places that they have not had it, such as on the face or periungual area. Other patients, however, may go months with less than 1% body surface involvement after discontinuingcyclosporine.
II
I
Laboratory monitoring of cyclosporine levels: Guidelines for the dermatologist Gary Mockli, MD, Pokar M. Kabra, PhD,I" and Theodore W. Kurtz, MD
San Francisco, California The following guidelines are recommended for laboratory monitoring of circulating levels of cyclosporine in dermatology patients. Measurements should be determined as trough levels in whole blood, not plasma or serum. The measurements should be performed with an assay that is specific for the parent cyclosporine compound (e.g., a high-performance liquid chromatography method or a specific monodonal immunoassay). The results of nonspecific immunoassays that detect cyclosporine as well as its metabolites are difficult to interpret and cannot readily be compared among different studies or laboratories. In psoriasis patients, the circulating concentration of cydosporine does not correlate reliably with the therapeutic response. Some patients may achieve an excellent response with blood levels in the range of 50 ng/ml; others may show little or no response despite blood levels as high as 200 ng/ml. In patients with a poor clinicalresponse, monitoring of cyclosporine levels may be useful to confirm that the drug has been taken and may provide an estimate of the degree of absorption and metabolism of the parent compound. Because an upper limit of safety for the circulating concentration of cyclosporine has not been dearly defined, one should attempt toachieve a therapeutic response with the lowest possible dose. Clinicians must carefully monitor patients for signs of cydosporine toxicity, regardless of the circulating concentration of the drug. Whole blood levels exceeding 250 ng/ml should be avoided. (J AM ACAD DERMATOL 1990;23:1275-9.)
In transplant patients, measuring circulating concentrations of cyclosporine has been widely advocated as a way to ensure adequate immune suppression and to minimize toxicity. 1-3 In this review, we focus on the issue of laboratory monitoring of cyclosporine levels in the management of dermatology patients. From the Department of Laboratory Medicine, University of California, San Francisco, Reprint requests:Theodore W, Kurtz, MD, Department of Laboratory Medicine, University of California, Box 0134, San Francisco, CA 94143-0134. 1"Deceased.This work is dedicated to the memory of Dr. Pokar Kabra, a pioneer in the use of liquid chromatography for therapeutic drug monitoring, 16/0/23798
METHODOLOGIC CONSIDERATIONS To interpret measurements of circulating concentrations appropriately, several problems of methodology must be considered. Type of sample: Whole blood versus plasma A t physiologic temperature, a large fraction of the cyclosporine in blood is distributed in erythrocytes. The partitioning of the drug between erythrocytes and plasma is both temperature and time dependent. For this reason, the same plasma sample can indicate highly variable blood levels, depending on the sample temperature and the time involved in collection and processing. To minimize this variability and to facilitate comparisons among different laboratories, we recommend that circulating concentrations 1275
Journal of the American Academy of Dermatology
1276 M o c k l i et al. of cyclosporine be measured in whole blood, preferably with EDTA used as the anticoagulant. 1,2 Although most laboratories have begun to follow this procedure, a substantial proportion (10% to 20%) may still be measuring cyclosporine in plasma. 4 Measurement .in whole blood instead of plasma also may minimize variation in cyclosporine levels that might be associated with changes in hematocrit. 1 Because the drug levels in whole blood are greater than in plasma, it is important to exercise caution when comparing studies in which different "therapeutic" levels of cyclosporine are reported. There is no universal and reliable method for converting a plasma level into an "equivalent" blood level. Because of the variability associated with cyclosporine measurements in plasma, we will focus on studies in which drug levels were measured in whole blood.
Timing of samples Approximately 10% to 50% of an oral dose of cyclosporine is absorbed through the small intestine, and, typically, peak circulating levels are achieved between 2 and 4 hours after administration. 2 Although extensive variability can be observed in and among patients in steady-state trough levels, even greater variability may be expected in peak levels of the drug. For that reason, it is important to monitor trough levels of cyclosporine.1, 2 Ingestion of food may increase or decrease the absorption of cyclosporine.2 It has also been reported that trough levels may vary substantially, depending on whether they are obtained before a morning or an evening dose. 5 These considerations make it necessary to measure each patient's trough levels on a regular schedule.
Type of assay Cyclosporine levels can be determined by irnmunoassay or high-performance liquid chromatography (HPLC). Because these methods may yield widely differing values in the same blood sample, one must know the technology used when comparing cyclosporine values produced by different studies or laboratories. Depending on the type of technology and laboratory, charges for blood level assays range from $20 to $100 per sample.
High-performance liquid chromatography H P L C can provide a specific and precise assay of cyclosporine levels in whole blood?, 6 Its sensitivity
may not be as great as that of some immunoassays, but this is not an important consideration because, for clinical purposes, cyclosporine blood levels are usually well within the sensitivity of most H P L C assays. Most HPLC assays clearly distinguish cyclosporine from its metabolites. Thus the results of such assays reflect the circulating concentration of the parent compound itself and are not affected by changes in metabolite concentrations. As long as appropriate standards are used, it should be possible to compare HPLC measurements of cyclosporine in whole blood across different laboratories and studies. Even so, this method is not widely used in clinical laboratories because it tends to require specialized equipment and expertise. However, in some laboratories, including ours, HPLC is efficiently used to measure cyclosporine levels in large numbers of specimens. In fact, when an automated H P L C injection system is used, a single medical technologist can analyze more than 50 blood samples per day.
lmmunoassay When immunoassay measurements of cyclosporine are interpreted, it is important to know whether they were based on a polyclonal or monoclonal antiserum. A monoclonal antiserum that is specific for cyclosporine is the assay of choice.l, 2, 4 The polyclonal immunoassays detect the parent compound as well as a variety of metabolites. Thus, for a given sample, an assay that uses a polyclonal antibody will yield much higher results than one that uses a specific monoclonal antibody or an H P L C method. Although there is little evidence that cyclosporine metabolites are nephrotoxic, the exact immunosuppressive and toxic nature of the metabolites remains unknown. ~ Furthermore, cyclosporine metabolite profiles may differ from patient to patient. Therefore, it is unclear how the results of polyclonal immunoassays could be used to ensure adequate immune suppression or to minimize toxicity. Several polyclonal assays are available and exhibit variable cross-reactivities with cyclosporine metabolites, l Because these assays may be measuring different metabolites, any comparison of results between studies that use different polyclonal assays can be problematic. Some investigators have proposed that both monoclonal and polyclonal assays be run on the same sample as a means of determining the relative amounts of cyclosporine and its metabolites. 7 However, for now, the clinical utility of
Volume 23 Number 6, Part 2 December 1990
Laboratory monitoring of cyclosporine 1277
such expensive testing is uncertain, and its use should be confined to the research setting. Generally, polyclonal assays use either radioactive labels or fluorescent labels (e.g., the Abbott TDx fluorescence polarization immunoassay). There is no reliable method for estimating the true level of the parent cyclosporine from a result generated with a nonspecific immunoassay. In patients with normal liver function, nonspecific immunoassays usually yield results that are two to four times greater than those of specific immunoassays or HPLC measurements. 2 As might be expected, the results of nonspecific immunoassays correlate poorly with results of HPLC or of monoclonal immunoassays that are specific for cyclosporine, s In contrast, the results of specific monoclonal immunoassays appear to correlate well with those of HPLC. 8' 9 Recently, the accuracy of cyclosporine standards supplied in at least one commercial monoclonal immunoassay kit has been questioned. 1~ Consequently, when selecting a laboratory for measurement of cyclosporine levels, one should determine the type of assay used. How well the laboratory has performed on external proficiency testing surveys is another important piece of information.
Table I. Guidelines for cyclosporine monitoring in psoriasis patients
CLINICAL INTERPRETATION OF CYCLOSPORINE LEVELS
found no evidence of a correlation between clinical response and circulating concentrations. In some patients given 14 mg/kg per day, whole blood levels of approximately 200 ng/ml were associated with complete clearing of severe plaque psoriasis. In oth. ers, much less dramatic clinical responses were observed, although cyclosporine levels were in the range of 250 ng/ml. However, in the preliminary study by Ellis et al., the dose used was well above current recommendations. Consequently, a possible dose-response relationship may have been obscured by the high efficacy of the large dose. Nevertheless, several other investigators have noted that the circulating concentration of this compound does not correlate reliably with the therapeutic response.12"16 In an open trial in psoriasis patients who were given "low"-dose cyclosporine therapy (2 to 4 nag/ kg per day), and in whom whole-blood trough levels of the drug were measured with a nonspecific radioimmunoassay, Griffiths et al) 2 reported that complete clearing of severe lesions could occur with circulating levels as low as 200 ng/ml. Presumably, the nonspecific assay used in this study was detecting cyclosporine metabolites as well as the parent compound. If so, some of the patients may have experi-
In transplant patients, it has been suggested that maintenance doses of cyclosporine be adjusted to produce whole-blood trough concentrations between 80 and 150 ng/ml, as measured by HPLC or drugspecific immunoassays.1 These guidelines have been proposed in the hope of ensuring adequate immunosuppression while minimizing toxicity. Although cyclosporine has been used in transplant patients for some time, it must be recognized that these guidelines are based largely on poorly controlled, retrospective studies. It is known that patients may experience rejection or nephrotoxicity, even when drug levels are in the so-called therapeutic range.1 Given the lack of reliable studies of therapeutic monitoring in transplant patients, it is not surprising that even less information is available on measuring cyclosporine in dermatology patients. In dermatology patients, most of the data on monitoring this drug comes from studies in which it has been used to treat psoriasis. In a double-blind study in psoriasis patients in whom whole-blood trough levels of cyclosporine were monitored by specific HPLC, Ellis et al. 11
Preferred sample: Preferred timing: Preferred method: Therapeutic range:
Toxic range:
Whole blood (EDTA or heparin as anticoagulant) Trough level HPLC or immunoassay specific for cyclosporine Unknown--some patients may respond with blood levels as low as 50 ng/rnl; others may not respond even at 200 rig/m1 The level at which toxicity begins to occur is unknown--patients must be monitored carefully for toxicity regardless of the circulating concentration of cyelosporine; however, persistent levels above 250 ng/ml, when measured as above, may be inadvisable!8
1278
Journal of the American Academy of Dermatology
M o c k l i et aL
enced an excellent clinical response, although the true drug levels were as low as 50 ng/ml. Given the uncertain relation between circulating levels of cyclosporine and clinical response to therapy, why measure the drug at all? In contrast to transplant patients in whom the clinician cannot assess therapeutic effectiveness of cyclosporine until it fails and rejection begins to occur, the dermatologist can directly monitor his patients' clinical response to the drug. I f the patient shows no clinical response, one might check a cyclosporine level to confirm that the patient has taken the drug and that at least some amount has been absorbed. However, as has been noted, the minimum level required to achieve a satisfactory clinical response is unknown and might vary widely among patients. If a patient exhibits a satisfactory clinical response, the only reason to measure a cyclosporine blood level would be to minimize the chance of toxicity. It should be noted that with prolonged oral administration of this compound, the bioavailability m a y increase, thereby causing an unanticipated elevation in blood levels. 2 Unfortunately, an upper limit of safety has not been defined. Obviously, it is desirable to achieve therapeutic response with the smallest possible dose of the drug. In renal transplant patients, it has been reported that whole-blood trough levels below 250 ng/ml (measured by H P L C ) have been associated with acceptable renal function) However, in psoriasis patients, even a "moderate" dose of cyclosporine (approximately 4 to 5 m g / k g per day, a dose that should yield circulating blood levels below 250 ng/ml) can be associated with a decrease in glomerular filtration rate.t7 Furthermore, individual patients m a y vary in their susceptibility to cyclosporine toxicity. Given the foregoing considerations, we believe far less emphasis should be placed on monitoring circulating cyelosporine levels than on monitoring the patient for clinical response and early signs of toxicity. 17 It is incumbent on the clinician to monitor patients carefully for signs of toxicity, regardless of the circulating concentration of cyclosporine. Our recommendations for cyclosporine monitoring are summarized in Table I. REFERENCES 1. Shaw LM. Advances in eyclosporine pharmacology, measurement, and therapeutic monitoring, Clin Chem 1989; 35:1299-308. 2. Shaw LM, Bowers L, Demers L, et al. Critical issues in cy-
closporine monitoring: report of the task force on cyclosporine monitoring. Clin Chem 1987;33:1269-88. 3. Moyer TP, Post GR, Sterioff S, et al. Cyclosporine nephrotoxicity is minimized by adjusting dosage on the basis of drug concentration in blood. Mayo Clin Proc 1988;63: 241-7. 4. Shaw L. Cyclosporinemonitoring. Clin Chem 1989;35:5-6. 5. BowersLD, Canafax DM, Singh J, et al. Studies of cyelosporine blood levels: analysis, clinical utility, pharmacokinetics, metabolites, and chronopharmacology. Transplant Proc 1986;18:137-44. 6. Kabra PM, Wall JH, Blanckaert N. Solid phase extraction and liquid chromatography for improved assay of cyclosporine in whole blood and plasma. Clin Chem 1985; 31:1717-20, 7. Trull AK, Tan KKC, Roberts NB, et al. Cyclosporine metabolites and neurotoxicity [Letter]. Lancet 1989;II:448. 8. Subbarao MN, Swanson JR, Mueggler PA. Cyclosporine determinations in heart and kidney transplant patients: comparison of high-performance liquid chromatography, polyclonal, and monoelonal methods. Ther Drug Monit 1989;11:53-6. 9. WolfBA, Daft MC, Koenig JW, et al. Measurement ofcyclospofine concentrations in whole blood: HPLC and radioimmunoassay with a specifie monoclonal antibody and 3H- or 125I-labeled ligand compared. Clin Chem 1989; 35:120-4. 10. MacBride JH, Rodgerson DO, Park SS, et al. Measurement of cyclosporine in plasma from patients with various transplants: HPLC and radioimmunoassay with a specific monoclonal antibody compared. Clin Chem 1989;35: 1726-30. 11. Ellis CN, Gorsulowsky DC, Hamilton TA, et al. Cyclospofine improves psoriasis in a double-blind study. JAMA 1986;256:3110-6. 12. Griffiths CEM, Powles AV, Leonard JN, et al. Clearance of psoriasis with low-dose cyclosporin. Br Med J 1986; 293:731-2. 13. Heule F, Meinardi MM, van Joost T, et al. Low-dose cyclosporineeffectivein severe psoriasis: a double-blind study. Transplant Proe 1988;20(suppl 4):32-41. 14. Meinardi MM, Bos JD. Cyclosporine maintenance therapy in psoriasis. Transplant Proe 1988;20(suppl 4):42-9. 15. Picascia DD, Garden JM, Freinkel RK, etal. Resistant severe psoriasis controlled with systemic cyclosporine therapy. Transplant Proe 1988;20(suppl 4):58-62. 16. Van Joost T, Bos JD, Heule F, et al. Low-dosecyclosporin A in severe psoriasis: a double-blind study. Br J Dermatol 1988;118:183-90. 17. Gilbert S, Emmett M, Menter A, et al. Cyclosporine therapy for psoriasis: serum creatinine measurements are an unreliable predictor of decreased renal function. J AM ACADDERMATOL1989;21:470-4. 18. A consensusreport: cyclosporin A therapy for psoriasis. Br J Dermatol 1990;122(suppl 36):1-3. DISCUSSION Dr. Ellis. Dr. Kurtz, how often do you think you should cheek for cyclosporine trough levels to see if the presumed level of toxicity has been reached? Dr. Kurtz. With a patient who is just starting cyelosporine therapy, it makes sense to assess the blood level once or twice in the beginning and to use 200 to 250 ng/ml by
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HPLC or monoclonal immunoassay as an upper limit. With prolonged therapy, the level should be checked again later if there is concern that the bioavailability of the fixed dose might be changing. However, because we do not know for certain what constitutes a safe level, it's difficult to know whether assessing blood levels frequently will provide important information. In some patients, even a level of 200 ng/ml could be toxic. Checking the blood pressure, checking renal function carefully--these are the most critical monitoring steps. Of course, you judge the clinical response by looking at the patient, not at the blood level. Dr. Ellis. With low doses of cyclosporine of 5 mg/kg per day or less, there is not a good correlation between blood levels and either clinical response or change in renal function. I think overall, particularly at higher doses, the data would indicate that specific blood levels greater than 200 ng/ml have a somewhat higher association with elevated creatinine levels. Dr. Kurtz. Although there is a rough correlation between toxicity and blood levels, in practical terms, managing the individual patient through blood level measurements is not feasible. Dr. Eisen. A recent review article on cyclosporine stated that one of the newer monoclonal radioimmunoassays gives more reliable levels than HPLC.I Dr. Kurtz, I'm aware of that review and I thought it was a sketchy presentation of that point of view. I don't have any objection to specific monoclonal radioimmunoassays, and nonisotopic assays are being developed as well. At least one commercial kit has been criticized because the calibrator supplied with it is said to be inaccurate. So it may be difficult to interpret a target level that is recommended as therapeutically appropriate, based on someone else's study using blood levels of cyclosporine that do not correspond to your laboratory's results. Still, I believe these specific monoclonal assays can be made to correlate well with HPLC, provided the calibration issue is resolved. In the future, we may be able to measure free cyclosporine in the blood more readily. Now we are measuring both bound and free cyclosporine. Whether free cyclosporine would have more clinical value is unknown.
Laboratory monitoring of cyclosporine
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Dr. Ellis. If a dermatologist were to prescribe cyclosporine for a patient with pyoderma gangrenosum, for example, what test is the local laboratory likely to use in determining cyclosporine levels? Dr. Kurtz. If it's a neighborhood laboratory, the sample probably will be sent out to a reference laboratory. Generally, if it's a large-volume reference laboratory, an immunoassay will be used because most such labs cannot keep up with the workload involved in performing the HPLC. One can only hope that the level is being measured in whole blood, and this can be ascertained from the report or by asking. Then it will be a question of determining whether the test used is a polyclonal immunoassay or a specific monoclonal immunoassay. I think eventually all laboratories will move to the monoolonal tests but, for now, the clinician has to check that. The practicing dermatologist also must check to see how the laboratory has performed on an external proficiency testing survey. What type of results are they obtaining in comparison with the overall mean of all laboratories? It is wise to see if the laboratory participates in such a proficiency survey. It might be using a reasonable assay but generating a poor result, which would go unrecognized without proficiency testing. Dr. Ellis. When reference laboratories send hack the results, they often give the physician a guideline to a therapeutic range. What does that mean, and can the physician use the guideline? Dr. Kurtz. With respect to cyclosporine, it will often be meaningless. The lab probably will have set its values on the basis of a journal article. Moreover, different labs will have read different articles. The therapeutic range may be unreliable, particularly if the laboratory has based its range on the results of a study performed in another laboratory that uses a different methodology. Finally, few if any laboratories will offer a therapeutic range specifically for the dermatology patient. Most such ranges are intended to guide cyclosporine therapy in transplant patients. REFERENCES
1. Kahan BD. Cyclosporine. N Engl J Med 1989;321: 1725-38.
