Reduction in the nephrotoxicity of amphotericin B when administered in 20 % intralipid

Fungal infections have begun to rival those caused by bacteria as significant nosocomial problems (1). Improvement in the prognosis of patients with cancer, immunodeficiencies and connective tissue disorders has created a patient population that more susceptible to infection. The increase in the incidence of nosocomial fungal infection since 1980 has mirrored the expansion of the immunocompromised patient population (2). These infections are often severe, progress rapidly and are refractory to therapy. Despite the availability and relative success of newer sysBRIEF REPORT

temic antifungal agents such as fluconazole (3) and itraconazole (4), amphotericin B (AmB) has remained the treatment of choice for systemic fungal infections.However, the clinical use of AmB is complicated by the development of a variety of adverse events including fever, chills, nausea, vomiting, hypokalemia, hypomagnesemia and anemia (5).The most formidable of these effects, nephrotoxicity, may actually restrict AmB's effectiveness because of premature discontinuation of antifungal therapy or suboptimal dosing (6).
One approach to circumventing nephrotoxicity has been the formulation of preparations of AmB that are less toxic (7).As a result, AmB has been entrapped in liposomes, ambisome or amphotericin B lipid complex (ABLC), and complexed with cholesteryl sulfate, amphotericin B colloidal dispersion (ABCD).Ambisome is a liposomal preparation of small unilaminar vesicles with a particle size 80 nm, containing a 10% concentration of AmB in the membrane (8).ABLC is an AmB lipid complex with a particle size of 1.6 to 11 µm that includes an approximately 33% concentration of AmB in microparticulate polymorphic sheets and ribbons (8).Finally, ABCD is a stable complex of AmB and cholesteryl sulfate intercalated in a 1:1 ratio in disc-like structures with a diameter of 122 mn (8).All three formulations have been administered at higher dosages, 5 to 7 mg/kg/day, than AmB (9).These preparations have proven to be efficacious with reduced renal toxicity for the treatment of fungal infections in immunocompromised hosts (10)(11)(12).Nevertheless, the formulations are very expensive.An alternative, less expensive delivery system has been studied by Anaissie et al (11), Chavanet et al (13) and Caillot et al (14).This lipid emulsion mixture with AmB was demonstrated to reduce toxicity while maintaining efficacy (11,13,14).We, therefore, decided to conduct our own pilot study to assess the effect of AmB mixed with 20% intralipid on the renal function of immunocompromised patients.

PATIENTS AND METHODS
Ten immunocompromised patients (eight with an underlying hematological malignancy, one with diabetes mellitus and rheumatoid arthritis, and one with Crohn's disease) with a systemic fungal infection due to a Candida species were initially given AmB in aqueous solution.The presence of a systemic fungal infection was determined by the patient's attending physician based on microbiological documentation of Candida species from a body site in the clinical setting of fever, presumed to be infectious in etiology.
Aqueous AmB was given at a dose of 0.6 mg/kg/day intravenously after an initial test dose of 1 mg in 100 mL of dextrose and water solution infused intravenously over 1 h.AmB was reconstituted in 10 mL of dextrose and water solution and subsequently diluted in 500 mL of the same intravenous solution.The aqueous AmB solution was then infused intravenously daily over 4 h.Premedication with acetaminophen 650 mg orally, intravenous diphenhydramine 25 or 50 mg and intravenous meperidine 25 or 50 mg was given to each patient.
Each patient was crossed over from aqueous AmB to AmB in 20% intralipid (AmB-IL).This occurred during the treatment course in six cases and at the onset of nephrotoxicity in four patients.The same premedication was employed.Nephrotoxicity was defined as a 50% decrease from the baseline value in calculated creatinine clearance (15).
AmB-IL was prepared daily in the following manner.Based on a dosage of 1 mg/kg/day, the daily requirement of AmB to be administered was calculated.AmB (50 mg/vial) was then reconstituted in 10 mL of sterile water for injection to yield a final concentration of 5 mg/mL.The required daily dosage of AmB was then transferred from the reconstituted preparation of 5 mg/mL to a 100 mL bottle of 20% intralipid (Kabi Pharmacia Canada Inc) fat emulsion for parenteral use.This was mixed and shaken thoroughly for 2 mins.The mixture was protected from light and refrigerated.The AmB-IL formulation was infused intravenously over 2 h within 24 h of its preparation.No premedication was administered.Patient characteristics were recorded including age; sex; underlying diagnosis; site of infection; organism type; baseline and end of therapy serum creatinines, potassium, magnesium, asparate aminotransferase, alanine aminotransferase, gamma-glutamyl transpeptidase, alkaline phosphatase and bilirubin; duration of therapy with aqueous AmB and AmB-IL.The total dose administered of each formulation was also recorded.Mean total dose of each formulation of AmB, mean duration of treatment and mean change in calculated creatinine clearance based on initial and end of therapy values for each formulation of AmB, respectively, were compared by paired Student's t tests.

RESULTS
Ten immunocompromised patients were initially treated with aqueous AmB and were then crossed over to AmB-IL.Table 1 details the characteristics of the patients.Six patients with underlying acute leukemia and two patients with lymphoma had recently completed antineoplastic chemotherapy.One patient, with a history of diabetes mellitus, rheumatoid arthritis and renal insufficiency, developed candiduria and candidemia.Another patient developed a central venous access device-related candidemia while receiving total parenteral nutrition for Crohn's disease.There were six female and four male patients, mean age 52 years.Causative pathogens were Candida albicans in six patients, Candida tropicalis in one, Candida lusitaniae in one, C albicans with Torulopsis glabrata in one, and C albicans with Candida lambica in one.Sources of infection were urine, bloodstream and gastrointestinal tract (neutropenic enteropathy).
The mean total dose of AmB-IL exceeded that of aqueous AmB (649±165 mg versus 344±105 mg, P=0.06), as did the mean duration of treatment (13.2±2.5 versus 9±2.1 days; P=0.31) (Table 2).However, during the aqueous AmB treatment, the mean creatinine clearance for the group dropped by 23.5±9.2mL/min (Figure 1).Thereafter, during AmB-IL therapy, the mean creatinine clearance rose by 10.7±7.7 mL/min  (P=0.03)(Figure 2).Nevertheless, in contrast with the improvement in creatinine clearance observed with AmB-IL, it had no significant beneficial effect on other biochemical laboratory parameters commonly affected by AmB (Table 3).

DISCUSSION
Our crossover study clearly demonstrated that AmB-IL was less nephrotoxic than aqueous AmB.Even though our immunocompromised patient sample received a higher concentration of AmB-IL than aqueous AmB (1 mg/kg/day versus 0.6 mg/kg/day), a larger mean total dose and longer length of therapy, mean creatinine clearance rose significantly during the AmB-IL phase of treatment.Although it is unclear whether AmB nephrotoxicity is dose-dependent (6), our findings clearly delineate the improvement in creatinine clearance associated with AmB-IL administration, even with administration of more of the latter preparation.Some investigators have demonstrated a renal sparing effect with AmB-IL compared with aqueous AmB (10); nevertheless, others have still noted significant reductions in creatinine clearance with a fat emulsion preparation (11).This latter finding may have been related to the addition of flucytosine (11).
However, there are a number of limitations to our observations.First, because aqueous AmB was initiated when sepsis was recognized, other factors such as concomitant hypoten-sion or other nephrotoxins may have contributed to the nephrotoxicity observed.Second, we could not assess the efficacy of the AmB-IL preparation.This is relevant because it has been speculated that AmB-IL produces less toxicity by trapping and binding AmB molecules in the lipid portion, thus reducing the availability of free and active AmB compound (7).Similarly, serum concentrations of free AmB in the AmB-IL mixture are lower than aqueous AmB (10).It may be necessary to infuse higher doses of AmB-IL to attain the efficacy rates achieved by aqueous AmB (7,10).Third, we did not demonstrate the permanent stability and absolute safety of our mixture.Concerns have been raised that AmB-IL may contain particles at least 10 µm in diameter (16) and that these particles may lodge in blood vessels.Aqueous AmB consistently had particles of less than 10 µm.Finally, it would have been advantageous for all our patients to cross over from aqueous AmB to AmB-IL at a specified AmB dose.
In this crossover pilot study where patients acted as their own controls, AmB-induced nephrotoxicity was significantly reduced with AmB-IL.This mixture warrants further evaluation of its long term stability, safety, clinical efficacy and maximum tolerated dose.Because the costs of AmB-IL are substantially less than ABLC, ABCD or liposomal AmB, the mixture may prove to be a welcome addition to our antifungal armamentarium.

Figure 1 )Figure 2 )
Figure 1) Baseline and ending creatinine clearance values for patients treated with aqueous amphotercin B. Creatinine clearance was calculated using the Cockcroft and Gault formula