DREW J. WINSTON 1 AND RONALD W. BUSUTTIL

Transcription

1 688 TRANSPLANTATION Vol. 74, No. 5 stored platelets. Clin Chem 1990; 36: Fava RA, Casey TT, Wilcox J, Pelton RW, Moses HL, Nanney LB. Synthesis of transforming growth factor-beta 1 by megakaryocytes and its localization to megakaryocyte and platelet alpha-granules. Blood 1990; 76: Oberhammer FA, Pavelka M, Sharma S, et al. Induction of apoptosis in cultured hepatocytes and in regressing liver by transforming growth factor beta 1. Proc Natl Acad Sci USA 1992; 89: Sindram D, Porte RJ, Hoffman MR, Bentley RC, Clavien PA. Synergism between platelets and leukocytes in inducing endothelial cell apoptosis in the cold ischemic rat liver: a Kupffer cell-mediated injury. FASEB J 2001; 15: Lang-Rollin I, Schurek JO, Lang RE. Platelet-induced inhibition of adrenomedullin secretion. Eur J Pharmacol 2001; 427: Gawaz M, Neumann FJ, Dickfeld T, et al. Activated platelets induce monocyte chemotactic protein-1 secretion and surface expression of intercellular adhesion molecule-1 on endothelial cells. Circulation 1998; 98: Martinez-Mier G, Toledo-Pereyra LH, Ward PA. Adhesion molecules in liver ischemia and reperfusion. J Surg Res 2000; 94: 185. RANDOMIZED CONTROLLED TRIAL OF ORAL ITRACONAZOLE SOLUTION VERSUS INTRAVENOUS/ORAL FLUCONAZOLE FOR PREVENTION OF FUNGAL INFECTIONS IN LIVER TRANSPLANT RECIPIENTS 1 DREW J. WINSTON 1 AND RONALD W. BUSUTTIL Background. Liver transplant recipients at high risk for serious fungal infections frequently receive fluconazole or an amphotericin B preparation for antifungal prophylaxis. Because of concerns about fungal resistance with fluconazole, safety with amphotericin B, and the cost of lipid formulations of amphotericin, alternative prophylactic regimens are needed. In this randomized, controlled trial, we compared the efficacy and safety of oral itraconazole solution with intravenous/oral fluconazole for prevention of fungal infections. Methods. Adult liver transplant recipients were randomized to receive either oral itraconazole solution (200 mg every 12 hr) or intravenous/oral fluconazole (400 mg every 24 hr). Each study drug was started immediately before transplant surgery and continued for 10 weeks after transplantation. Patients were evaluated for fungal colonization, proven invasive or superficial fungal infection, drug-related side effects, and death. Results. Fungal colonization decreased from baseline to week 8 after transplantation in both the itraconazole patients (67% to 25%, P<0.001) and the fluconazole patients (77% to 30%, P<0.001). Proven fungal infection developed in 9 (9%) of 97 itraconazole patients and in 4 (4%) of 91 fluconazole patients (P 0.25). The number of proven invasive fungal infections (seven with itraconazole [7%], three with fluconazole [3%]) and proven superficial fungal infections (two with itraconazole [2%], one with fluconazole [1%]) were also similar in both groups of patients. Organisms causing infection were Candida glabrata (four patients), Candida albicans (three patients), and Aspergillus species (two patients) in the itraconazole group and C glabrata (two patients), Candida krusei (one patient), This work was supported by a research grant from Janssen Research Foundation. Address correspondence to: Drew J. Winston, MD, Room CHS, Department of Medicine, UCLA Medical Center, Los Angeles, California Received 17 October Revision Requested 3 December Accepted 20 March DOI: /01.TP F5 and Aspergillus species (one patient) in the fluconazole group. Mortality from fungal infection was very low and occurred in only 1 (0.5%) of 188 patients. Except for more frequent gastrointestinal side effects (nausea, vomiting, diarrhea) with itraconazole, both itraconazole and fluconazole were well tolerated and not associated with any hepatotoxicity. Mean trough plasma concentrations of itraconazole were greater than 250 ng/ml throughout the study and were not affected by H 2 -receptor antagonists or antacids. Conclusion. Oral itraconazole solution has adequate bioavailability in liver transplant recipients for effective antifungal prophylaxis. Similar to fluconazole, prophylactic oral itraconazole decreases fungal colonization and is associated with a low incidence of serious or fatal fungal infections. Except for gastrointestinal side effects, oral itraconazole solution is well tolerated and has no significant hepatotoxicity. Among solid organ transplant recipients, liver transplant recipients have one of the highest incidences of fungal infection. Depending on underlying risk factors, the reported incidence of serious invasive fungal infection in liver transplant patients has ranged from 7% to 42% (1, 2). The mortality rate from these infections is also quite high (25% to 60%). Candida and Aspergillus species are the predominant fungal pathogens in liver transplant recipients, but occasionally Cryptococcus and other opportunistic fungi may appear. Because of the relatively high incidence and severity of fungal infection in liver transplant recipients, antifungal agents are frequently used for prophylaxis in these patients (3). Fluconazole and intravenous amphotericin B have been the agents most widely used. Although data from randomized, controlled trials on the efficacy of prophylactic amphotericin B in liver transplant recipients are limited (4 6), we demonstrated in a large randomized, double-blind, placebocontrolled trial that prophylactic fluconazole administered at a dosage of 400 mg per day for 10 weeks after liver transplant decreases fungal colonization, prevents superficial and invasive fungal infections, and reduces mortality from fungal infection (7). Similar favorable results in

2 September 15, 2002 WINSTON AND BUSUTTIL 689 transplant patients have been reported by other investigators (8 10). Itraconazole is another azole antifungal agent with excellent in vitro activity against many opportunistic fungi, including Aspergillus and some Candida species that are resistant to fluconazole (11, 12). Itraconazole is now available in a hydroxypropyl- -cyclodextrin solution, which greatly enhances its absorption after oral administration (12). Itraconazole has been used successfully for antifungal prophylaxis in neutropenic patients and possesses a safety profile superior to either amphotericin B or the lipid formulations of amphotericin B (13 15). However, there are very limited data on the relative efficacy and safety of prophylactic itraconazole in liver and other solid organ transplant recipients. Thus, we performed this randomized trial comparing oral itraconazole solution with intravenous/oral fluconazole for prevention of fungal infections in liver transplant recipients. PATIENTS AND METHODS Patients Patients of either gender were eligible for the study if they met the following criteria: (1) age between 18 and 75 years; (2) undergoing orthotopic liver transplantation; (3) no clinical or microbiological evidence of fungal infection at time of study entry; and (4) no allergy to imidazole or azole antifungal agents. Patients receiving any investigational drug at time of transplantation or within 1 month before study entry were excluded. Similarly, patients receiving rifampin, rifabutin, phenobarbital, carbamazepine, terfenadine, astemizole, cisapride, vincristine, triazolam, or oral midazolam were not eligible for the study. Women were required to have a negative pregnancy test. Informed consent approved by the UCLA Human Subject Protection Committee was obtained from each patient or appropriate relative. Transplantation Procedures The surgical procedures and posttransplantation medical care used in liver transplant recipients at the UCLA Medical Center have previously been published (1, 16). Baseline immunosuppressive therapy usually consisted of tacrolimus or cyclosporine in combination with corticosteroids, azathioprine, or mycophenolate. Acute rejection was documented by liver biopsy and treated with boluses of intravenous corticosteroids. Immediately before transplant surgery, patients received oral erythromycin and neomycin for decontamination of the gastrointestinal tract. Perioperative intravenous antibacterial prophylaxis was given for 24 hr and consisted of ampicillin-sulbactam alone, vancomycin with ceftizoxime, or vancomycin plus an aminoglycoside (for patients allergic to penicillin). During surgery, the abdominal cavity was irrigated with a solution of 25 mg of amphotericin B in 100 ml of sterile water. Antiviral prophylaxis consisted of either 100 days of intravenous ganciclovir given at a dose of 6 mg/kg per day (for a cytomegalovirus [CMV]-seronegative patient with a CMV-seropositive donor) or 2 weeks of intravenous ganciclovir at a dose of 6 mg/kg per day followed by 800 mg of oral acyclovir four times a day for 12 weeks (for a CMV-seropositive patient) (17, 18). Trimethoprim-sulfamethoxazole twice a week or pentamidine once every 2 weeks (for patients allergic to sulfa) was used for Pneumocystis carinii prophylaxis. Study Drugs and Design Eligible patients were randomly assigned to receive prophylaxis with either itraconazole or fluconazole. Each study medication was given as a single dose preoperatively and then continued for 10 weeks after transplantation. If a patient required another transplantation during the study period, prophylaxis was given for an additional 10 weeks from the date of the repeat transplantation. Patients randomized to itraconazole received an oral itraconazole solution at a loading dose of 200 mg every 8 hr for 24 hr followed by 200 mg every 12 hr. Patients unable to take medications by mouth immediately after the transplant surgery received the itraconazole solution through a nasogastric tube. Patients randomized to fluconazole initially received intravenous fluconazole at a dose of 400 mg once daily and were then switched to oral fluconazole at a dose of 400 mg once daily when they were able to swallow oral tablets. If patients became unable to take oral tablets, they were changed back to intravenous fluconazole. Itraconazole oral solution was supplied in 150-mL bottles containing 10 mg of itraconazole per milliliter. The fluconazole was provided at a concentration of 2 mg per milliliter for intravenous administration and in 200-mg tablets for oral administration. The daily dose of fluconazole was decreased by 50% for a creatinine clearance of 20 to 50 ml/min and by 75% for a creatinine clearance less than 20 ml/min. There was no adjustment of the itraconazole dose in patients with renal failure or hepatic dysfunction. After transplantation, prophylaxis with the study drug was discontinued if a documented invasive fungal infection developed, a serious adverse side effect definitely related to the study drug occurred, or the patient died. Patients who developed a documented superficial fungal infection could be treated with topical clotrimazole while continuing to receive the study drug. Treatment with the study drug was also continued when empirical therapy with intravenous amphotericin B was administered for suspected but undocumented invasive fungal infection. Laboratory Procedures Complete blood counts with differential white blood cell counts and platelet counts, blood urea nitrogen levels, serum creatinine and electrolyte determinations, urinalyses, and liver function studies (aspartate aminotransferase, alanine aminotransferase, alkaline phosphatase, and total bilirubin levels) were obtained at study entry and at least once weekly during weeks 1, 2, 4, 8, and 10 of the study period. Serum tacrolimus levels measured by an enzyme immunoassay or serum cyclosporine levels determined by high-pressure liquid chromatography were also followed during the duration of the study (19, 20). In patients receiving itraconazole, trough plasma levels of itraconazole and its active metabolite, hydroxy-itraconazole, were measured by high-performance liquid chromatography during weeks 1, 2, 4, 8, and 10 of the study (21). All patients were evaluated throughout the study for clinical symptoms and signs of adverse events related to the study drug. Surveillance cultures of the oropharynx, axillae, inguinal skin folds, urine, and stool or perirectal area were done at study entry and then once weekly during weeks 1, 2, 4, 8, and 10 of the study to determine the presence of fungal colonization. Cultures of blood and other suspected sites of fungal infection were also obtained whenever a patient s clinical condition suggested the possibility of infection. Minimum inhibitory concentrations (MICs) of itraconazole and fluconazole were determined for yeasts and filamentous fungi isolated from patients with documented fungal infection to evaluate the possible emergence of resistant organisms. Antifungal susceptibility testing was performed by a central reference laboratory according to guidelines of the National Committee for Clinical Laboratory Standards (22, 23). Definitions of Fungal Colonization and Infection Fungal colonization was defined as the presence of a fungus in one or more surveillance cultures in the absence of any clinical symptoms or signs of infection. Superficial fungal infections were diagnosed by the isolation of a fungus from the skin, oropharynx, gastrointestinal tract, or vagina in association with signs of inflammation, ulcerations, plaques, exudates, or other manifestations of infection not explainable by other pathogens. Invasive fungal infections were di-

3 690 TRANSPLANTATION Vol. 74, No. 5 agnosed by the presence of fungus in the blood, pulmonary tissues or secretions, sinuses, soft tissues, peritoneal cavity, or other organ structure in association with symptoms and signs of infection not explainable by other pathogens. Statistical Analysis All statistical tests were performed as two-tailed tests. Differences in proportions were compared by the Fisher s exact test (24). Ninetyfive percent confidence intervals were determined for differences by using the normal approximation. The Cochran-Mantel Haenszel test was used to compare the proportion of patients in each treatment group with positive surveillance cultures. Comparisons of times to specific events were performed by using the Kaplan-Meier method and the log-rank test (25). Differences in the median trough serum cyclosporine or tacrolimus levels in patients on or off study drug were compared by the Wilcoxon signed rank test. All patients who received a liver transplant and then at least one dose of study drug after transplantation were included in the efficacy analysis (modified intention-to-treat analysis). RESULTS Patient Characteristics Two hundred forty-six consecutive adults hospitalized for orthotopic liver transplantation at the UCLA Medical Center were evaluated for the study. Two hundred four patients were considered eligible for the study and randomized to receive antifungal prophylaxis with either itraconazole or fluconazole. Sixteen patients (6 itraconazole patients, 10 fluconazole patients) were excluded from analysis after randomization. Reasons for exclusion were cancellation of transplantation because of inadequate donor organ (six cases), cardiovascular instability (three cases), pulmonary hypertension (one case), or metastatic tumor (one case); death in the operating room or immediately after transplant surgery (three cases); concurrent use of another systemic antifungal drug (one case); and a simultaneous pancreatic islet cell transplant (one case). The characteristics of the other 188 patients are summarized in Table 1. Ninety-seven patients received itraconazole, and 91 patients received fluconazole. The two groups of patients were similar in terms of age, gender, underlying liver disease, and possible risk factors for fungal infection (United Network Organ Sharing [UNOS] classification, pretransplantation corticosteroid therapy or renal failure, fungal colonization at baseline, repeat transplantation, other abdominal surgery, CMV infection after transplantation, baseline immunosuppressive agents, and graft rejection). The median number of on-study days was 70 for itraconazole patients (range, 4 to 83 days) and 69 for fluconazole patients (range, 3 to 79 days). The median duration of intravenous dosing of fluconazole was 6 (range, 2 to 71) days. The median duration of oral dosing was 63 (range, 1 to 73) days in the fluconazole group and 70 (range, 4 to 83) days in the itraconazole group. Fungal Infections The types of proven fungal infections and their causative organisms are summarized in Table 2. Proven fungal infection occurred in 9 (9%) of 97 itraconazole patients and in 4 (4%) of 91 fluconazole patients (P 0.23, 95% confidence interval 12 to 2.3). The number of proven invasive fungal infections (seven with itraconazole, three with fluconazole) TABLE 1. Patient characteristics Characteristic Itraconazole Fluconazole Number of patients Median age (range), yr 53 (24 72) 50 (19 72) Gender, n (%) Male 51 (53) 52 (57) Female 46 (47) 39 (43) Underlying liver disease, n (%) Chronic hepatitis C 38 (39) 30 (33) Chronic hepatitis B 11 (11) 9 (10) Chronic hepatitis B and C 3 (3) 1 (1) Chronic non-a, non-b hepatitis 4 (4) 1 (1) Alcoholic liver disease 9 (9) 14 (15) Cryptogenic cirrhosis 10 (10) 14 (15) Primary biliary cirrhosis 6 (6) 7 (8) Sclerosing cholangitis 7 (7) 5 (5) Autoimmune liver disease 3 (3) 6 (7) Fulminant hepatic failure 4 (4) 4 (4) Hepatocellular carcinoma 1 (1) 0 (0) Isoniazid toxicity 1 (1) 0 (0) UNOS Classification, n (%) 1 (Life support in intensive care) 28 (29) 29 (32) 2 (Continuous hospitalization) 24 (25) 19 (21) 3 (Continuous medical care) 45 (46) 43 (47) 4 (Stable at home) 0 (0) 0 (0) Corticosteroids within 4 weeks of 2 (2) 2 (2) transplantation, n (%) Pretransplantation renal failure, 18 (19) 16 (18) (serum creatinine 2.0 mg/ dl), n (%) Fungal colonization at baseline, 64/96 (67) 70/91 (77) a (number colonized/number cultured) Repeat transplant, n (%) 12 (12) 12 (13) Other abdominal surgery during 22 (23) 13 (14) b study, n (%) CMV infection after 1 (1) 5 (6) transplantation, n (%) Initial immunosuppressive agents, n (%) Tacrolimus plus corticosteroids 55 (57) 47 (52) Tacrolimus, mycophenolate, and 13 (13) 11 (12) corticosteroids Tacrolimus, azathioprine, and 11 (11) 8 (9) corticosteroids Cyclosporine plus corticosteroids 3 (3) 8 (9) Cyclosporine, azathioprine, and 15 (16) 17 (18) corticosteroids Patients with graft rejection, n (%) 23 (24) 28 (31) a P b P and proven superficial fungal infections (two with itraconazole, one with fluconazole) were also similar in both treatment groups. Kaplan-Meier estimates of the percentage of patients in each study group who had a proven fungal infection after transplantation revealed no significant difference (Fig. 1, P 0.19). Except for one fungemia and two superficial infections caused by Candida albicans in the itraconazole group, all proven infections were caused by Candida glabrata, Candida krusei, or Aspergillus species. Of the 13 fungal isolates causing infection, 6 were susceptible in vitro to itraconazole (MIC 1.0 g/ml, range to 0.5 g/ml) and 5 were susceptible to fluconazole (MIC 64 g/ml, range to 16

4 September 15, 2002 WINSTON AND BUSUTTIL 691 TABLE 2. Incidence of proven fungal infections Variable Itraconazole Fluconazole Number of patients Patients with proven fungal 9 (9) 4 (4) a infection, n (%) Patients with proven invasive 7 (7) 3 (3) infection, n (%) C glabrata fungemia 3 1 C krusei fungemia 0 1 C albicans fungemia 1 0 C glabrata abdominal 1 0 abscess Aspergillus pneumonia 1 1 Aspergillus wound infection 1 0 Patients with proven 2 (2) 1 (1) superficial infection, n (%) C glabrata of oral cavity 0 1 C albicans of oral cavity 1 0 C albicans of skin 1 0 a P 0.25 (95% confidence interval, 12 to 2.3). FIGURE 1. Time to development of proven fungal infections. Survival distributions of time to development of proven invasive or superficial fungal infection in itraconazole and fluconazole patients were defined by using Kaplan-Meier product-limit estimates. P 0.19 for the difference in time of onset of proven fungal infection between itraconazole and fluconazole patients. g/ml). All four organisms causing infection in the fluconazole group (two C glabrata, one C krusei, one Aspergillus species) were resistant in vitro to fluconazole (MIC 64 g/ ml). On the other hand, only four of the nine organisms causing infection in the itraconazole group (three C glabrata, one C albicans) were resistant to itraconazole (MIC 1.0 g/ml). Four patients in the fluconazole group were given empiric antifungal therapy for treatment of a suspected fungal infection. Three of these patients received empiric intravenous amphotericin B for a suspected systemic fungal infection because they had persistent fever and overall clinical deterioration despite antibiotic therapy. One fluconazole patient received topical nystatin for a localized skin rash associated with negative cultures. All four patients improved on antifungal therapy, but a fungal infection was never documented. None of the patients in the itraconazole group had suspected fungal infection. Thus, the total number of patients with proven or suspected fungal infection was the same in the itraconazole group (nine patients) and the fluconazole group (nine patients). The incidence of fungal infection in each treatment group was also evaluated in patients with the highest risk for fungal infection. At the UCLA Medical Center, factors previously found to be associated with a high risk for fungal infection in liver transplant recipients are UNOS classification of 1, fungal colonization at time of transplantation, and repeat transplantation (7). Seventy itraconazole patients and 73 fluconazole patients had one or more of these risk factors for fungal infection. Proven fungal infection occurred in 7 (10%) of 70 high-risk itraconazole patients (5 invasive infections, 2 superficial infections) and in 2 (3%) of 73 high-risk fluconazole patients (2 invasive infections) (P 0.09). Among low-risk patients without any significant risk factors for fungal infection, proven fungal infection developed in 2 (7%) of 27 itraconazole patients (2 invasive infections) and in 2 (11%) of 18 fluconazole patients (1 invasive infection, 1 superficial infection) (P 1.0). During long-term follow-up of patients for 1 year after transplantation, two additional itraconazole patients developed fungal infection (C glabrata fungemia, oral C albicans infection) several months after completion of the prophylactic itraconazole. No additional fungal infections occurred among the fluconazole patients. There were no late-onset Aspergillus infections. Fungal Colonization The effects of prophylaxis with itraconazole or fluconazole on fungal colonization are shown in Table 3. Fungal colonization decreased in both the itraconazole and fluconazole patients during the study period. By week 8, the incidence of fungal colonization had decreased in the itraconazole group from 67% (64 of 96 patients) at baseline to 25% (21 of 83 patients) (P 0.001) and in the fluconazole group from 77% (70 of 91 patients) at baseline to 30% (24 of 79 patients) (P 0.001). Both itraconazole and fluconazole were especially effective in eliminating colonization with C albicans. Both study drugs also decreased colonization with other Candida species except C glabrata. The incidence of colonization with C krusei decreased in the itraconazole group (5% at baseline versus 0% at week 8, P 0.03) but increased slightly in the fluconazole group (1% at baseline versus 3% at week 8, P 0.32). Colonization with Aspergillus was low (4% or less) throughout the study in both groups of patients. Plasma Itraconazole Concentrations Mean trough plasma concentrations of itraconazole and its active metabolite, hydroxy-itraconazole, are summarized in Table 4. The mean trough plasma concentration of itraconazole was 282 ng/ml after 1 week of treatment and increased further to 428, 774, 1313, and 1610 ng/ml after 2, 4, 8, and 10 weeks of treatment, respectively. These mean trough plasma concentrations of itraconazole are all greater than 250 ng/ml, which has been considered the minimum level required for efficacy (26). H 2 -receptor antagonists, proton pump inhibitors, or antacids were used in 16 patients receiving itraconazole but had no appreciable effect on the plasma concentrations of itraconazole. On the other hand, the mean trough plasma concentrations of itraconazole after 2, 4, 8,

5 692 TRANSPLANTATION Vol. 74, No. 5 Patients cultured TABLE 3. Fungal colonization at baseline and after transplantation a Other Candida species b Aspergillus species Any fungus C albicans C krusei C glabrata Variable n n (%) n (%) n (%) n (%) n (%) n (%) Baseline Fluconazole (77) 56 (62) 1 (1) 19 (21) 12 (13) 4 (4) Itraconazole (67) 52 (54) 5 (5) 17 (18) 9 (9) 1 (1) Week 1 Fluconazole (66) 39 (44) 4 (4) 27 (30) 5 (6) 3 (3) Itraconazole (66) 37 (38) 5 (5) 30 (31) 10 (10) 4 (4) Week 2 Fluconazole (44) c 16 (18) 6 (7) 23 (26) 3 (3) 1 (1) Itraconazole (60) 27 (29) 4 (4) 34 (36) 9 (10) 1 (1) Week 4 Fluconazole (35) 6 (7) 5 (6) 19 (22) 4 (5) 2 (2) Itraconazole (47) 10 (11) 1 (1) 23 (26) 7 (8) 4 (4) Week 8 Fluconazole (30) 3 (4) 2 (3) 17 (22) 4 (5) 1 (1) Itraconazole (25) 0 (0) 0 (0) 14 (17) 2 (2) 0 (0) Week 10 Fluconazole (30) 2 (3) 2 (3) 15 (19) 2 (3) 2 (3) Itraconazole (33) 3 (3) 0 (0) 21 (24) 4 (5) 2 (2) a Colonization was defined as isolation of fungus from any surveillance culture in absence of symptoms or signs of infection. b Candida guilliermondi, Candida parapsilosis, Candida tropicalis, Candida lusitaniae, Candida lipolytica, Candida kefyr, Candida lambica. c P 0.04 for difference in colonization at week 2. All other differences in colonization for each week are not significant (P 0.1). TABLE 4. Mean trough plasma concentrations of itraconazole and its metabolite, hydroxyitraconazole Time after transplantation Number of patients Plasma concentration (ng/ml SD) Itraconazole Hydroxyitraconazole Week Week Week Week Week and 10 weeks of treatment were measured in 8 of the 9 patients who failed itraconazole prophylaxis and were generally lower (246, 543, 170, and 193 ng/ml, respectively) than the mean trough levels (446, 797, 1351, and 1690 ng/ ml, respectively) in 71 patients who did not develop a fungal infection. None of these differences are statistically significant (P 0.05). Four itraconazole patients who developed nosocomial fungal infections caused by itraconazole-sensitive organisms had low mean trough plasma itraconazole concentrations of 258, 285, and 696 ng/ml after 1, 2, and 4 weeks of treatment. Side effects Adverse clinical events or laboratory abnormalities possibly or probably related to the study drug occurred in 49 (51%) of 97 itraconazole patients and in 19 (21%) of 91 fluconazole patients (P 0.001, Table 5). Gastrointestinal side effects (nausea, diarrhea, vomiting, and abdominal pain) developed more often in patients who received itraconazole. The other types of side effects in the itraconazole and fluconazole patients were similar. Abnormal liver function tests possibly related to study drug developed in three itraconazole patients and one fluconazole patient. However, no patients were removed from the study because of concerns about TABLE 5. Side effects possibly or probably related to study drug Variable Itraconazole Fluconazole Number of patients Patients with side effects, n (%) 49 (51) 19 (21) a Side effects, n (%) Gastrointestinal Nausea 26 (27) 5 (6) a Diarrhea 18 (19) 3 (3) b Vomiting 9 (9) 1 (1) c Abdominal pain 5 (5) 0 (0) d Neurological (headaches, dizziness) 4 (4) 3 (3) Cutaneous (rash) 1 (1) 1 (1) Elevated liver function tests 3 (3) 1 (1) Elevated serum creatinine 0 (0) 3 (3) Others (anorexia, insomnia, nervousness, fever) 10 (10) 3 (3) a P b P c P d P hepatotoxicity from itraconazole or fluconazole. Two itraconazole patients withdrew from the study because of gastrointestinal side effects related to itraconazole. No fluconazole patients withdrew from the study because of an adverse event related to fluconazole. There were no significant differences in serum cyclosporine or tacrolimus levels between the itraconazole and fluconazole patients. Serum levels of both cyclosporine and tacrolimus did become lower after itraconazole or fluconazole was discontinued. The median trough serum levels of cyclosporine in the itraconazole and fluconazole patients were 262 ng/ml and 260 ng/ml, respectively, while on study drug, and 157 ng/ml and 154 ng/ml, respectively, after the study drug was stopped (P 0.001). Similarly, the median trough serum levels of tacrolimus in the itracon-

6 September 15, 2002 WINSTON AND BUSUTTIL 693 azole and fluconazole patients were 9.9 ng/ml and 9.0 ng/ ml, respectively, while on study drug, and 7.3 ng/ml and 6.5 ng/ml, respectively, after the study drug was stopped (P 0.001). Survival Twelve itraconazole patients (12%) and 7 fluconazole patients (8%) died during the study. Causes of death in the itraconazole group were liver graft failure (five cases), cardiovascular disease (one case), hemorrhage (one case), aplastic anemia (one case), graft-versus-host disease (one case), bacterial infection (two cases), and Aspergillus pneumonia (one case). Causes of death in the fluconazole group were liver graft failure (one case), cardiovascular disease (two cases), hemorrhage (one case), central pontine myelinolysis (one case), aplastic anemia (one case), and bacterial infection (one case). Thus, only 1 (0.5%) of the 188 patients in this study died from fungal infection. The five itraconazole patients and one fluconazole patient who died from liver graft failure had either primary nonfunction of the liver graft or hepatic artery thrombosis. This difference in deaths from liver graft failure between the two groups was not significant (P 0.21). DISCUSSION Based on the results of randomized controlled trials and uncontrolled observations, fluconazole or amphotericin B is commonly used for antifungal prophylaxis in liver and other solid organ transplant recipients (3 8, 27). Nevertheless, both fluconazole and amphotericin B have certain limitations as prophylactic agents. Although prophylactic fluconazole reduces the incidence of colonization, superficial infection, and invasive fungal infection caused by many Candida species, it has no effect on colonization or infection caused by C glabrata, C krusei, oraspergillus species, which are usually resistant to fluconazole (7 10). Standard amphotericin B or the lipid formulations of amphotericin B are very active against Aspergillus and most Candida species, including fluconazole-resistant strains of Candida (4 6). However, because these drugs are toxic, must be administered intravenously, and can be expensive (lipid preparations of amphotericin), they are not optimal agents for prophylaxis. Thus, there is a need for effective regimens that combine the safety of fluconazole with the antifungal activity of amphotericin B for Aspergillus and other common fungal pathogens. In this trial, liver transplant recipients were randomized to receive antifungal prophylaxis with either oral itraconazole solution or intravenous/oral fluconazole. Similar to fluconazole, oral itraconazole significantly reduced fungal colonization and was associated with a relatively low incidence of serious invasive fungal infections (7%) among a group of liver transplant patients at very high risk for infection. There was only one death from fungal infection among the 97 patients receiving prophylactic itraconazole. Given the effectiveness of prophylactic fluconazole demonstrated in a previous randomized, double-blind, placebo-controlled trial involving UCLA liver transplant patients (7), the inclusion of a control group not receiving antifungal prophylaxis in this study was considered inappropriate and unethical. Furthermore, preliminary results from another randomized trial comparing oral itraconazole solution with placebo for prophylaxis in liver transplant recipients has shown a reduction in fungal infection requiring systemic therapy from 24% in the placebo group to 4% in the itraconazole group (28). A major advantage of itraconazole over fluconazole is itraconazole s activity against Aspergillus. The reported incidence of invasive aspergillosis in liver transplant recipients is between 2% and 8% (2, 3). The mortality rate from these infections exceeds 90%. There were only 3 Aspergillus infections (2 with itraconazole, 1 with fluconazole) in the 188 patients (1.6%) enrolled in the study. Thus, evaluation of the possible impact of prophylactic itraconazole on the incidence of invasive aspergillosis was hampered by the low observed incidence of this infection in a sample size of 188 patients. However, even in other controlled trials of prophylactic itraconazole involving more than 900 patients with hematological malignancies, it has been difficult to demonstrate a reduction in Aspergillus infections in favor of oral itraconazole (13 15). Nonetheless, it would seem reasonable to consider antifungal prophylaxis with itraconazole instead of fluconazole in those liver transplant recipients with risk factors for invasive aspergillosis (pretransplantation fulminant hepatic failure, Aspergillus colonization, poor liver graft function requiring excessive immunosuppression, renal failure necessitating dialysis) (27, 29 31). Prophylaxis with a lipid formulation of amphotericin B has been used to prevent invasive fungal infections caused by Aspergillus and other organisms in liver transplant recipients requiring dialysis (6). A previous concern about the use of oral itraconazole for prophylaxis in transplant patients was the erratic bioavailability of the capsule formulation of itraconazole. Absorption of itraconazole capsules is enhanced by normal gastric acidity and the presence of food (12). In gravely ill patients who are unable to eat, have altered gastric acidity, or are receiving concomitant H 2 -receptor antagonists and antacids, the poor bioavailability of itraconazole capsules can be problematic. The oral itraconazole solution used in this study contains hydroxypropyl- -cyclodextrin, which greatly increases the bioavailability of itraconazole and eliminates the need for food or gastric acidity for optimal absorption (12). Mean trough plasma concentrations of itraconazole were 250 ng/ml throughout the 10-week duration of this study and were not affected by the concomitant use of H 2 -receptor antagonists, proton pump inhibitors, or antacids. Although it is difficult to define the most ideal plasma itraconazole concentration needed for efficacy, a plasma concentration of 250 ng/ml has been correlated with effective prophylaxis in neutropenic patients (26). The liver transplant patients who failed prophylaxis in this study and developed a documented fungal infection had mean plasma concentrations of itraconazole that were generally 250 ng/ml and lower than the levels in patients without infection. Thus, certain more critically ill patients or other patients unable to adequately take oral medications may have impaired absorption of even the oral solution of itraconazole and require close monitoring of plasma itraconazole concentrations. Alternatively, these patients may benefit from a new intravenous formulation of itraconazole (32), which was not available for this study but has been used effectively for empiric antifungal therapy in febrile neutropenic patients and for prophylaxis in allogeneic bone marrow transplant patients (33, 34). Both itraconazole and fluconazole were generally well tolerated. There was no appreciable hepatotoxicity with either

7 694 TRANSPLANTATION Vol. 74, No. 5 itraconazole or fluconazole. Itraconazole was associated with more gastrointestinal side effects (nausea, vomiting, and diarrhea). The hydroxypropyl- -cyclodextrin used in the oral itraconazole solution has an osmotic effect, which may cause diarrhea and other gastrointestinal side effects (12 15). Oral itraconazole solution was also associated with more gastrointestinal side effects in one study of antifungal prophylaxis in neutropenic patients (14) but not in other studies involving neutropenic patients or patients with human immunodeficiency virus infection (13, 15, 35). Like fluconazole and other azole compounds, itraconazole may increase serum levels of cyclosporine or tacrolimus by inhibiting the cytochrome P-450 enzyme system (36, 37). This drug interaction likely explains the drop in serum cyclosporine and tacrolimus levels noted after the patients in this study discontinued itraconazole or fluconazole. A potential problem with use of azole antifungals for prophylaxis is the emergence of resistant fungi. Long-term itraconazole prophylaxis given for 1 year to patients with human immunodeficiency virus has been associated with a reduction in susceptibility of mucosal C albicans isolates to itraconazole and cross-resistance to fluconazole (38). The use of routine prophylactic fluconazole in allogeneic bone marrow transplant recipients over a 4-year period at one large transplant center was associated with colonization by azole-resistant Candida species but a very low incidence of candidemia (39). In this study, all four fungal infections occurring in the fluconazole patients were caused by organisms (two C glabrata, one C krusei, one Aspergillus species) resistant to fluconazole. Four of the nine infections in the itraconazole group were caused by organisms resistant to itraconazole (C glabrata 3, C albicans 1). Thus, the prospect of azole-resistant organisms causing infection during prophylaxis with itraconazole or fluconazole must be considered and is an argument for limiting prophylaxis to those patients at highest risk for infection. Of note, despite the routine use of prophylactic azole antifungal agents in UCLA liver transplant recipients for the past 6 years, we have not observed an increase in the rate of invasive fungal infection caused by C glabrata or C krusei. In summary, oral itraconazole solution has adequate bioavailability in liver transplant recipients, which allows its use as an effective agent for antifungal prophylaxis. Similar to fluconazole, prophylactic oral itraconazole solution decreases fungal colonization and is associated with a low incidence of serious or fatal fungal infections. Except for gastrointestinal side effects, oral itraconazole solution is well tolerated and has no significant hepatotoxicity. Serum cyclosporine and tacrolimus levels decrease after prophylaxis with either fluconazole or itraconazole is discontinued and require close monitoring so appropriate adjustments of cyclosporine and tacrolimus dosages can be made. Acknowledgments. The authors thank the following study coordinators for their valuable assistance during this study: Maureen Burns, RN, Nancy Dolan, RN, and Linda Reyes, RN. We are also grateful to Dr. Ronghua Yang, Larry Broach, Katie Harding, and Dr. Gerhard Leitz for performing data and statistical analyses; Dr. David Bruckner and the staff of the UCLA clinical microbiology laboratories for technical assistance; Ed Arriola and the staff of the UCLA pharmacy for administrative assistance; Katharine Fry for preparation of the manuscript; and the UCLA liver transplant physicians and nurses for their help and care of the patients during the study. REFERENCES 1. Winston DJ, Emmanouilides C, Busuttil RW. Infections in liver transplant recipients. Clin Infect Dis 1995; 21: Paya CV. Fungal infections in solid-organ transplantation. Clin Infect Dis 1993; 16: Paya CV. Prevention of fungal and hepatitis virus infections in liver transplantation. Clin Infect Dis 2001; 33(suppl 1): S Mora NP, Klintmalm G, Solomon H, Goldstein RM, Gonwa TA, Husberg BS. Selective amphotericin B prophylaxis in the reduction of fungal infections after liver transplant. Transplant Proc 1992; 24: Tollemar J, Höckerstedt K, Ericzon BG, Jalanko H, Ringdén O. Liposomal amphotericin B prevents invasive fungal infections in liver transplant recipients: a randomized, placebo-controlled study. Transplantation 1995; 59: Singh N, Paterson DL, Gayowski T, Wagener MM, Marino IR. Preemptive prophylaxis with a lipid preparation of amphotericin B for invasive fungal infections in liver transplant recipients requiring renal replacement therapy. Transplantation 2001; 71: Winston DJ, Pakrasi A, Busuttil RW. Prophylactic fluconazole in liver transplant patients: a randomized, double-blind, placebo-controlled trial. Ann Intern Med 1999; 131: Lumbreras C, Cuervas-Mons V, Jara P, et al. Randomized trial of fluconazole versus nystatin for the prophylaxis of Candida infection following liver transplantation. J Infect Dis 1996; 174: Goodman JL, Winston DJ, Greenfield RA, et al. A controlled trial of fluconazole to prevent fungal infections in patients undergoing bone marrow transplantation. N Engl J Med 1992; 326: Slavin MA, Osborne B, Adams R, et al. Efficacy and safety of fluconazole prophylaxis for fungal infections after marrow transplantation: a prospective, randomized, double-blind study. J Infec Dis 1995; 171: Sheehan DJ, Hitchcock CA, Sibley CM. Current and emerging azole antifungal agents. Clin Microbiol Rev 1999; 12: De Beule KL. Itraconazole: pharmacology, clinical experience, and future development. Int J Antimicrob Agents Chemother 1996; 6: Menichetti F, Del Favero A, Martino P, et al. Itraconazole oral solution as prophylaxis for fungal infections in neutropenic patients with hematologic malignancies: a randomized, placebo-controlled, double-blind, multicenter trial. Clin Infect Dis 1999; 28: Morgenstern GR, Prentice AG, Prentice HG, et al. A randomized controlled trial of itraconazole versus fluconazole for the prevention of fungal infections in patients with haematological malignancies. Br J Haematol 1999; 105: Harousseau JL, Dekker AW, Stamatoullas-Bastard A, et al. Itraconazole oral solution for primary prophylaxis of fungal infections in patients with hematological malignancy and profound neutropenia: a randomized, double-blind, double-placebo, multicenter trial comparing itraconazole and amphotericin B. Antimicrob Agents Chemother 2000; 44: Busuttil RW, Shaked A, Millis JM, et al. One thousand liver transplants: the lessons learned. Ann Surg 1994; 219: Winston DJ, Wirin D, Shaked A, Busuttil RW. Randomised comparison of ganciclovir and high-dose acyclovir for long-term cytomegalovirus prophylaxis in liver-transplant recipients. Lancet 1995; 346: Seu P, Winston DJ, Holt CD, Kaldas F, Busuttil RW. Long-term ganciclovir prophylaxis for successful prevention of primary cytomegalovirus (CMV) disease in CMV-seronegative liver transplant recipients with CMV-seropositive donors. Transplantation 1997; 64: Jusko WJ, Thomson AW, Fung J, et al. Consensus document: therapeutic monitoring of tacrolimus (FK-506). Ther Drug Monit 1995; 17: Huang WY, Lipsey AI, Cheng MH. Comparison of cyclosporine determinations in whole blood by three different methods: HPLC, 125 I RIA, and 3 H RIA. Am J Clin Pathol 1987; 87: Warnock DW, Turner A, Burke J. Comparison of high performance liquid chromatographic and microbiological methods for determination of itraconazole. J Antimicrob Chemother 1988; 21: National Committee for Clinical Laboratory Standards. Reference method for broth dilution antifungal susceptibility testing of yeasts. Tentative standard. Wayne, PA: National Committee for Clinical Laboratory Standards, (NCCLS document M27-A). 23. Espinel-Ingroff A, Bartlett M, Bowden R, et al. A multicenter evaluation of the standardization of antifungal susceptibility testing of filamentous fungi. J Clin Microbiol 1997; 35: 139.

8 September 15, 2002 WINSTON AND BUSUTTIL SAS Institute Inc. SAS Version 6.12 Software. Cary, NC. 25. Kaplan EL, Meier P. Nonparametric estimation from incomplete observations. J Am Stat Assoc 1958; 53: Boogaerts MA, Verhoef GE, Zachee P, Demuynck H, Verbist L, De Beule K. Antifungal prophylaxis with itraconazole in prolonged neutropenia: correlation with plasma levels. Mycoses 1989; 32: Singh N. Antifungal prophylaxis for solid organ transplant recipients: seeking clarity amidst controversy. Clin Infect Dis 2000; 31: Colby WD, Sharpe MD, Ghent CN, et al. Efficacy of itraconazole prophylaxis against systemic fungal infection in liver transplant recipients. Program and abstracts of the 39th interscience conference on antimicrobial agents and chemotherapy. Washington, DC: American Society for Microbiology, 1999: Kusne S, Torre-Cisneros J, Manez R, et al. Factors associated with invasive lung aspergillosis and significance of positive Aspergillus culture after liver transplantation. J Infect Dis 1992; 166: Collins LA, Samore MH, Roberts MS, et al. Risk factors for invasive fungal infections complicating orthotopic liver transplantation. J Infect Dis 1994; 170: Patel R, Portela D, Badley AD, et al. Risk factors of invasive Candida and non-candida fungal infections after liver transplantation. Transplantation 1996; 62: Boogaerts MA, Maertens J, Van Der Geest R, et al. Pharmacokinetics and safety of a 7-day administration of intravenous itraconazole followed by a 14-day administration of itraconazole oral solution in patients with hematologic malignancy. Antimicrob Agents Chemother 2001; 45: Boogaerts M, Winston DJ, Bow EJ, et al. Intravenous and oral itraconazole versus intravenous amphotericin B deoxycholate for empirical antifungal therapy of persistent fever in neutropenic cancer patients receiving broad-spectrum antibacterial therapy: a randomized controlled trial. Ann Intern Med 2001; 135: Winston DJ, Maziarz RT, Chandrasekar P, et al. Long-term antifungal prophylaxis in allogeneic bone marrow transplant patients: a multicenter, randomized trial of intravenous/oral itraconazole versus intravenous/oral fluconazole (Abstract 2002]. Blood 2001: 98: 479a. 35. Graybill JR, Vasquez J, Darouiche RO, et al. Randomized trial of itraconazole oral solution for oropharyngeal candidiasis in HIV/AIDS patients. Am J Med 1998; 104: Back DJ, Tjia JF. Comparative effects of the antimycotic drugs ketoconazole, fluconazole, itraconazole and terbinafine on the metabolism of cyclosporin by human liver microsomes. Br J Clin Pharmacol 1991; 32: Leather HL, Boyette R, Wingard JR. Evaluation of the pharmacokinetic drug interaction between intravenous itraconazole and intravenous tacrolimus or intravenous cyclosporin in allogeneic bone marrow transplant patients. Blood 2000; 96: 390a. 38. Goldman M, Cloud GA, Smedema M, et al. Does long-term itraconazole prophylaxis result in in vitro azole resistance in mucosal Candida albicans isolates from persons with advanced human immunodeficiency virus infection? Antimicrob Agents Chemother 2000; 44: Marr KA, Seidel K, White TC, Bowden RA. Candidemia in allogeneic blood and marrow transplant recipients: evolution of risk factors after the adoption of prophylactic fluconazole. J Infect Dis 2000; 181: 309.