Burckart, GJ

Iwatsuki, S

Ptachcinski, RJ

Sanghvi, A

Seltman, H

Shaw, BW

Starzl, TE

Van Thiel, DH

Venkataramanan, R

Yang, S

English

PubMed

Name not available

Biliary excretion of cyclosporine in liver transplant patients

D-Scholarship@Pitt

Biliary Excretion of Cyclosporine in Liver Transplant Patients R. Venkataramanan, T.E. Starzl, S. Yang, G.J. Burckart, R.J. Ptachcinski, B.W. Shaw, S. lwatsuki, D.H. Van Thiel, A. Sanghvi, and H. Seltman CYCLOSPORINE (CyA) is a novel cyclic polypeptide with potent immunosup-pressive properties.1.2 Cy A is presently used with steroids to prevent the rejection of trans-planted kidneys,3 livers,4 and heartsY This drug is primarily eliminated from the body by hepatic metabolism, and biliary excretion is the major route of CyA elimination in ani-mals. 7 In humans, enterohepatic recycling of CyA has been suggested because of the pres-ence of a second peak in CyT (CyA plus . metabolites measured by radioimmunoassay [RIA)) plasma concentrations and the appearance of the drug in the ileostomy drain-age of one patient.8 We have observed increases in CyA blood levels (parent com-pound measured by high-pressure liquid chro-matography [HPLC]) following T-tube clamping in liver transplant patients, which is also indicative of possible enterohepatic recy-cling of Cy A. In order for Cy A to undergo any significant enterohepatic recirculation, large amounts of CyA must be excreted in the bile. In the present study, we quantitated the bil-iary excretion of CyA and CyT in adult liver transplant patients in order to determine whether CyA undergoes significant enterohe-patic recycling. MATERIALS AND METHODS Eighteen biliary excretion studies were performed in 13 patients (12 liver transplant recipients and one liver disease patient) with aT-tube in their common bile ducts. From The Clinical Pharmacokinetics Laboratorv, School of Pharmacy. Departments of Medicine. Surge;y. and Pathology. School of Medicine. University of Pitts-burgh. Address reprint requests to Dr R. Venkataramanan. Clinical Pharmacokinetics Laboratory. 807 Salk Hall. School of Pharmacy. University of PitMburgh. Pitts-burgh.PA 15261. © 1985 by Grune & Stratton. Inc. 0041-1345/85/1701-0100$03.00/0 286 CyA was administered either as an intravenous (IV) infusion (130 to 150 mg; n - 7), as a combination of IV infusion and oral dose (70 to 150 mg I V, 300 to 700 mg orally; n = 3). or as an oral dose (400 to 600 mg; n - 8). Prior to drug administration. the T-tube was drained and the total bile output over the study period of eight to 24 hours was collected at one- to four-hour intervals. The total bile volume in each collection was measured. Ali-quots of the bile samples were kept frozen at - 20 °C until being analyzed for CyA and CyT. HPLC was used to analyze CyA in samples from all of the patients. Samples from four patients were·also analyzed for CyT by RIA. Bile samples (0.1 to 1.0 mL) were extracted and assayed following minor modification of the HPLC proce-dure of Sawchuk and Cartier.9 RIA was performed using a kit provided by Sandoz, Inc (East Hanover, NJ) accord-ing to a procedure described by Donatsch et ai, 10 following I :20 and 1 :50 dilution of the bile samples. Standards were prepared for both methods in blank bile. Biliary excretion studies were peiformed in patients with varying degrees of liver function, as indicated by total serum bilirubin concentrations. Patients with a total bilirubin concentration of 1 mgj dL or less were consid-ered to have normal liver function. Patients with a total serum bilirubin concentration of greater than 1 mgjdL were considered to have poor liver function. Three studies were performed in patients with normal liver function, and the remaining studies were carried out in patients whose liver function was poor. Bile output, bile CyA and CyT concentrations, and the total amounts of CyA and CyT excreted in bile was compared between patient groups with normal and poor liver function, using a Student's I test, assuming a level of significance of P c:s .05. RESULTS Analysis of blank bile by HPLC confirmed the absence of any interfering materials in the bile. The standard curve for HPLC analysis of CyA in bile was run from 25 to 4,000 ng/mL. The coefficient of variation (n = 9) at 887 ng/mL was 6.5%. For the RIA analysis of CyT, the standard curve was linear from 78 to 2,500 ng/mL. The coefficient of variation (n = 6) at 611 ng/mL was 17.8%. The bile samples were dilu ted to ensure that the results of the analysis were in the linear range of the standard curve for both HPLC and RIA. TransplantBtion Proceedings. Vol XVII, No 1 (February), 1985 CYCLOSPORINE BILIARY EXCRETION 287 Table 1. Biliary Excretion of Cyclosporine Total Amount of Dose (mgl Total Duration of Total Cyclosporine in Bile (mgl Study Bilirubin Bile Collection Bile Volume No. IV Oral (mg/dLi 1 150 25.8 2 150 3 150 4 130 24.6 5 140 13.7 6 150 4.3 7 150 300 24.0 8 100 700 14.8 9 70 300 2.5 10 600 2.5 11 400 7.9 12 550 5.8 13 600 2.5 14 500 0.5 15 500 1.0 16 400 1.0 17* 730 18.4 18* 144 18.4 'Pretransplant, liver disease patient. The total volume of bile collected over one dosing interval ranged from 13.5 to 417.5 ml (Table 1). Patients with normal liver function had higher bile output than patients with poor liver function (24.1 mL/h v 7.6 mL/h; P < .05). The maximum bile concentration of CyA ranged from 32 ng/mL in a patient with severe liver disease to 5,212 ng/mL in a liver transplant recipient with normal hepatic func-tion. The total amount of CyA excreted in the bile over a dosing interval ranged from 0.003 to 0.814 mg.· There was a positive linear correlation between the bile output and the total amount of cyclosporine excreted in the bile (r = .686; P < .0 I). The amount of (hi (mLi HPLC (CyAI RIA (CyTI 12 17.0 0.031 12 13.5 0.025 12 23.0 0.028 8 53.5 0.208 8 54.5 0.100 2.990 13 59.0 0.097 8 14.5 0.013 12 50.5 0.035 0.980 12 143.0 0.177 6.900 12 280.0 0.222 13 40.2 0.053 1.200 13 239.0 0.092 12 168.0 0.195 12 417.5 1.052 12 350.5 0.501 12 97.7 0.814 24 129.0 0.003 23 83.0 0.005 cyclosporine excreted in the bile in patients with normal liver function was greater than that excreted in patients whose liver function was poor at the time of the study (789 J.Lg v 98 .ug; P < .01). The patient with severe liver disease who was studied pretransplant had the lowest maximum bile concentration of CyA and excreted the smallest amount of Cy A (3.7 .ug) in the bile. A negative linear correlation was observed between serum bilirubin and the total amount of CyA excreted in the bile (r = -.597; P < .01). The total amount of CyA excreted in the bile over one dosing interval represents less than 1 '1(, of the administered dose in all of the patients studied (Table 2). The concentration of CyT in bile ranged Table 2. Effect of Liver FunctIon and Route of Administration on Cyclosporine Biliary Excretion No. Route of Bile Drainage of CyA mL/Dosing Status of Subjects Studies· Administration Interval Poor liver function 6 IV 37 Poor liver function 3 IV and PO 69 Poor liver function 4 PO 182 Normal liver function 3 PO 289 All values reported are the means for each group. PO, per orally. 'Excluding the pretransplant studies. tBased on an assumed oral bioavailability of 20%. mL/h 3.75 6.0 14.7 24.1 Dose Excreted Absorbed Amount of in Bile Dose Excreted eyA Excreted as eyA in Bile as CyA (mgl 1%1 (%1 0.082 0.059 0.059 0.075 0.018 0.053 t 0.141 0.025 0, 124t 0,789 0.170 O.85O t 288 from 9,280 to 61,300 ng/ mL, as measured by RIA. The maximum bile CyT concentration measured by RIA was 18 to 36 times higher than the CyA concentration by HPLC mea-surements. The total amount of CyT excreted in the bile when measured by RIA ranged from 0.980 to 6.900 mg, with a mean of 3.018 mg. The ratio of the total amount of CyT to CyA excreted in bile ranged from 23:1 to 39:1, with a mean of 30: 1. DISCUSSION Cyclosporine and steroids are the primary immunosuppressants used in heart, liver, and kidney transplant patients. Very little infor-mation is available on CyA kinetics in trans-plant recipients. Urinary excretion of CyA and CyT is a minor elimination pathway in dogs, rats, and humans. In dogs, most of the orally administered radioactive CyA is elimi-nated in the feces. 7 More than 50% of the intravenously administered radioactive CyA is excreted in rat bile.7 In humans, indirect evidence for the involvement of bile in CyT elimination has been obtained from the analy-sis of ileostomy drainage.8 This drainage accounted for 61.5% (measured as CyT) of the orally administered dose of CyA in one patient. As the drug was administered orally, unabsorbed drug may account partially for this observation. Moreover, RIA was used to analyze the ileostomy drainage. At present, no data on the amount of intact drug excreted in the bile are available. In our study, we obtained a direct estima te of the biliary excretion of CyA and CyT in adult liver transplant patients with aT-tube. The concentration of CyA and CyT in bile was greater than the concentrations in blood. However, less than 1 % of the administered dose of CyA was excreted in the bile during one dosing interval. The amount of CyA excreted in bile was highly variable and was partially dependent on the funtional status of the liver as well as the volume of bile. Patients with poor liver function had a low bile output and a low concentration of drug in their bile. Patients with normal liver function had the VENKATARAMANAN ET AL highest bile output and higher drug concen-trations in their bile. One patient with liver disease who was studied pretransplant excreted the least amount of CyA in his bile. This may be due to a combination of poor absorption (bioavailability was 3%) and poor excretory capacity of the diseased liver in this patient. There was no significant difference in the percentage of the dose excreted in the bile following oral and rv administration of CyA in patients with poor liver function. Increases in the blood CyA and CyT concentrations following T-tube clamping could be rational-ized on the basis of a possible increase in the absorption of CyA due to increased amounts of bile being <;lvailable to facilitate the absorp-tion of fat-soluble cyclosporine. Analysis of bile by RIA revealed the pres-ence of large amounts of CyT. The CyT:CyA concentration ratio of 18-36:1 in the bile as compared to a ratio of 2-5: 1 in blood indicates that relatively greater amounts of CyA metabolites are excreted in the bile. The amount of CyT excreted in the bile as deter-mined by RIA is 30 times greater than that measured by HPLC. RIA measurements of bile and blood are an underestimate of the total amount of CyA and its metabolites, as various CyA metabolites do not interact with the antibodies used in the RIA to the same extent that the intact parent CyA does. In our patients with normal liver function, bile collection represented more than half of the estimated normal bile output. Using a normal bile output of 500 mL in a l2-hour dosing interval and a mean CyA bile concen-tration of 2.73 Jlg/mL (as observed in patients with normal liver function), one can estimate that approximately 1.37 mg of CyA is excreted in the bile. This represents 0.29% of the orally administered dose (mean oral dose in patients with normal liver function, 467 mg). CyA is incompletely absorbed following oral administration. Assuming a bioavailabil-ity of 20%, one can ca1cula te the percentage of the dose absorbed that is excreted in the bile as 1.46. Therefore, a patient receiving 500 mg CYCLOSPORINE BILIARY EXCRETION of oral CyA with a bioavailability of 20% would excrete approximately 1.46 mg of CyA in the bile. The amount of CyT excreted would be nearly 30 times greater and thus, a total of 43.8 mg of CyT will be excreted in the bile. The low amount of parent CyA excreted in bile indicates that intact CyA is not likely to undergo any significant enterohepatic recy-cling. These results differ from an earlier report that suggested the possibility of signifi-cant enterohepatic recycling of CyA in humans. This discrepancy may be explained partially because of the use of RIA assay to measure CyA in plasma and ileostomy drain-age. CyA metabolites may, however, undergo enterohepatic recycling and hence produce secondary plasma CyT peaks. Secondary blood peaks have also been observed in a few of our patients when CyA was measured by HPLC. From our present results, it appears 289 that mechanisms other than enterohepatic recirculation may be responsible for such observa tions. In conclusion, very little intact CyA is excreted in the bile of liver transplant recip-ients. In contrast, a large amount of CyT (CyA plus metabolites) is excreted in the bile. Intact CyA is not likely to undergo any signif-icant enterohepatic recycling in patients fol-lowing liver transplantation. SUMMARY The biliary excretion of Cy A was studied in 13 patients with externalized bile flow. Analy-sis of bile by HPLC revealed that less than 1 % of an administered dose of Cy A is excreted in the bile. RIA analysis of bile revealed that large amounts of CyT are excreted in the bile. Enterohepatic recycling of CyA is unlikely in liver transplant patients, but CyA metabolites may be subject to this process. REFERENCES I. Borel JF, Feurer C, Gubler HU, et al: Agents Actions 6:468, 1976 2. CaIne RY, RoUes K, White D1G, et al: Lancet 2:1033,1979 3. Starzl TE, Weil R, Iwatsuki S, et al: Surg Gynecol o bstet 15 1: 17, 1980 4. Starzl TE, Klintmalm GB, Porter KA, et al: N Engl 1 Med 305:266, 1981 5. Pennock 1L, Oyer PE, Reitz BA, et al: 1 Thorac Cardiovasc Surg 83: 168, 1982 6. Griffith BP, Hardesty RL, Deeb M, et al: Ann Surg 196:324, 1982 7. Beveridge T: In White D1G (ed): Cyclosporin A. Amsterdam, Elsevier-North Holland, 1982, p 35 8. Kahan BD, Ried M, Newburger J: Transplant Proc 15:446,1983 9. Sawchuk RT, Cartier LL: Clio Chern 27:1368, 1981 10. Donatsch P, Abisch E, Hornberger M, et al: J Immunoassay 2:19, 1981 

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Biliary excretion of cyclosporine in liver transplant patients

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