Lipid formulations of amphotericin B as first-line treatment of zygomycosis

Zygomycosis is a difficult to treat and frequently fatal infection affecting immunocompromised and, rarely, immunocompetent patients. The early diagnosis and immediate initiation of treatment with an antifungal agent in combination with surgical intervention has proved critical for the favourable outcome of the disease. Few antifungal agents are available for treatment. Amphotericin B (AmB) deoxycholate has been the drug of choice for many years and is usually given at high daily doses which can result in renal toxicity. Currently, lipid formulations of AmB (liposomal AmB (L-AmB), AmB lipid complex (ABLC), AmB colloidal dispersion (ABCD)), mainly L-AmB, rather than conventional AmB have become the standard therapy. The rationale behind the use of lipid formulations is that they decrease the nephrotoxicity associated with longterm AmB use. Although there is a developing consensus that high doses of lipid formulations of AmB should be the antifungal therapy of choice for all patients with zygomycosis, until now there have been no data available with which to define the appropriate dose. The duration of therapy remains an unresolved issue, regarding both lipid formulations of AmB as well as sequential or combination treatments consisting of lipid formulations of AmB with posaconazole, a drug which has now emerged as a new therapeutic option.     

Indomethacin treatment in amphotericin B induced nephrogenic diabetes insipidus

Nephrogenic diabetes insipidus (NDI) is a serious side effect of various drugs. Elevated renal prostaglandin E₂ levels have been found in patients with lithium-induced NDI and have been implicated in the pathogenesis. We report the case of a patient who developed NDI following treatment with amphotericin B. Prostaglandin levels were elevated. Indomethacin had an antidiuretic effect and normalized prostaglandin levels.Abbreviations NDI nephrogenic diabetes insipidus - DNDI drug-induced nephrogenic diabetes insipidius - ICU intensive care unit - PGE₂ Prostaglandin E₂ Correspondence to: K.-H. Meyer zum Buschenfelde     

Effect of amphotericin B on renal tubular acidification in the rat

Amphotericin B, a polyene antibiotic known to induce cation-selective pore formation in biological cell membranes, was given to rats by peritoneal injection (10 mg/kg for 21–26 days) or added to luminal perfusates (2×10^–5 M). Kinetics of tubular acidification and alkalinization after perfusion with alkaline or acid phosphate Ringer's solution was studied by means of double barrelled antimony/reference microelectrodes in cortical distal tubules. Stationary pH increased both in early and late distal segments. Acidification and alkalinization half-times decreased markedly from 15–18 s to 6–8 s, a value similar to that found in proximal tubule. Net H-ion secretion rates as well as H-ion back-flux approximately doubled after Amphotericin B. Apparent H-ion permeability of distal tubule epithelium measured during perfusion of lumen and peritubular capillaries with phosphate Ringer's solutions doubled both in early and late segments. These data show that amphotericin B produces a distal acidification defect which impairs formation of normal transepithelial pH gradients by increasing H-ion back-flux without reducing rates of net H-ion secretion.This work was supported by Fundação de Amparo à Pesquisa do Estado de São Paulo (Proc. 74/1306), FINEP, CNPq.     

Determination of amphotericin B, liposomal amphotericin B, and amphotericin B colloidal dispersion in plasma by high-performance liquid chromatography.

Amphotericin B is a potent polyene antifungal drug for intravenous treatment of severe infections. It is used as amphotericin B-deoxycholate and in order to reduce amphotericin B toxicity as lipid-formulated complex (liposomal or colloidal dispersion). A sensitive and specific analytical method is presented for the separation of lipid-complexed and plasma protein-bound amphotericin B in human heparinized plasma. This separation, which is required for pharmacokinetic studies, is achieved by solid-phase extraction (SPE) via Bond Elut C18. The protein-bound amphotericin B has a higher affinity to the SPE material and is therefore retained, whereas the lipid-complexed amphotericin B is eluted in the first step. The recovery of the SPE was >75% for high concentrations and >95% for low concentrations. Quantification was performed by reversed-phase HPLC using a LiChrosorb-RP-8 column, UV detection (lambda=405 nm) and a mixture of acetonitrile-methanol-0.010 M NaH2PO4 buffer (41:10:49, v/v) as mobile phase. The retention time for amphotericin B under the given conditions was 6.7 min. The calibration curves were found to be linear (r > or = 0.999) in two different ranges (5.0-0.50 microg/ml and 0.50-0.005 microg/ml). Intra- and inter-day precision and accuracy fulfilled the international requirements. No interference from other drugs (typical broad medication for intensive-care patients) or common plasma components was detected in >400 samples analyzed.     

Inhibition of in vitro HBsAg production by amphotericin B and ketoconazole

The effects of amphotericin B, ketoconazole, and adenine arabinoside on production of hepatitis B surface antigen (HBsAg) particles by the human hepatoma cell line PLC/PRF/5 were examined. In addition, the effects of these drugs on cellular protein synthesis were determined. These drugs caused a dose-dependent decrease in HBsAg production that was paralleled by a decrease in cellular protein synthesis. Ketoconazole was the most active of these drugs and the most specific, causing a 72% reduction in HBsAg production with only a 38% reduction in protein synthesis. These data suggest that further studies evaluating ketoconazole for the treatment of chronic hepatitis B virus infection in animals are warranted.     

Fatal fat embolism following amphotericin B lipid complex injection

A case of amphotericin B lipid complex induced fatal fat embolism is described. A 41-year-old Caucasian man with AIDS was undergoing treatment for cryptococcal meningitis with amphotericin B. His course was complicated by renal failure necessitating a change in therapy to amphotericin B lipid complex (Abelcet). At approximately 48 h, the patient developed tachycardia, tachypnea, respiratory failure, decline in hematocrit, thrombocytopenia, and alteration in mental status. Autopsy findings included fat emboli involving heart, lungs, kidney, and brain. To our knowledge, this is the first case report of a fatal fat embolism caused by intravenous liposome drug delivery.     

Liposomal amphotericin B]

Liposomal amphotericin B (AmBisome) is a DDS (drug delivery system) formulation of amphotericin B (AMPH-B), and has been developed in an attempt to reduce the toxicity of AMPH-B while retaining its therapeutic efficacy. AMPH-B has been the "gold standard" of antifungal therapy over the past four decades. It has a broad spectrum of fungicidal activity against a number of clinically important pathogens including Aspergillus and Candida. The mechanism of action of AMPH-B involves binding to ergosterol, the principal sterol in fungal cell membranes. Binding to ergosterol causes an increase in fungal membrane permeability, electrolyte leakage, and cell death. AMPH-B has affinity for cholesterol in mammalian membranes, which leads to severe side-effects including kidney damage. AmBisome is a unilamellar vesicle composed of AMPH-B and phospholipid. Upon administration, AmBisome remains intact in the blood and distributes to the tissues where fungal infection may occur, and is disrupted after attachment to the outside of fungal cells, resulting in fungal cell death. AmBisome and AMPH-B show similar in vitro and in vivo antifungal activity and clinical efficacy. However, AmBisome has less infusion-related toxicity and nephrotoxicity than AMPH-B.     

Differential effects of amphotericin B and liposomal amphotericin B on inflammatory cells in vitro

OBJECTIVE AND DESIGN: To understand whether the pseudo-allergic reactions to amphotericin B (AmB) administration are due to AmB or to the solubilizing vehicles, a study was designed to evaluate the effects of AmB, liposomal AmB, (L-AmB), AmB-deoxycholate micellar complex (AmB-DC) and deoxycholate (DC) on the responses of rat serosal mast cells (RSMC) and of human basophils (HB), in vitro. MATERIALS AND METHODS: Serosal mast cells were obtained by density gradient centrifugations from male Wistar albino rats. Partially purified HB were obtained from healthy donors. The cells were exposed to AmB, L-AmB, AmB-DC and DC. Histamine release was measured fluorometrically, and the release of lactic dehydrogenase (LDH) was measured spectrophotometrically. HB activation was evaluated cytofluorimetrically by CD63 expression. RESULTS: AmB and L-AmB did not evoke histamine or LDH release from either RSMC or HB. CD63 expression was not induced in HB challenged with AmB and L-AMB. On the other hand, AmB-DC and DC produced a cytotoxic histamine release from both RSMC and HB, and a sustained increase of CD63 expression on HB. CONCLUSIONS: Only AmB-DC was able to induce the release of inflammatory mediators from RSMC and HB. Conceivably, the cytotoxic effect is accounted for by the micellar complexes with DC, which has been confirmed as a powerful histamine releaser, and held responsible for the pseudo-allergic reactions after AmB-DC administration. The data lend support to a better safety profile of L-AmB.     

Liposomal amphotericin B twice weekly as antifungal prophylaxis in paediatric haematological malignancy patients

Data on antifungal prophylaxis in paediatric cancer patients at high risk for invasive fungal disease (IFD) are scant. Intermittent administration of liposomal amphotericin B (LAMB) has been shown to be safe and effective in adult patients with haematological malignancies. We prospectively evaluated the safety and efficacy of prophylactic LAMB at a dosage of 2.5 mg/kg twice weekly in children at high risk for IFD. Efficacy was compared with that in a historical control group of patients with similar demographic characteristics not receiving LAMB prophylaxis. A total of 46 high-risk patients (24 boys; mean age, 7.7 years) with 187 episodes of antifungal prophylaxis were analysed. The median duration of neutropenia (<500/μL) was 10 days. LAMB was discontinued in four patients because of acute allergic reactions. Median values for creatinine and liver enzymes at end of treatment did not differ significantly from those at baseline. Hypokalaemia (<3.0 mmol/L) occurred with 13.5% of the prophylactic episodes, but was usually mild and always reversible. No proven/probable IFD occurred in patients receiving LAMB prophylaxis. In comparison, five proven and two probable IFDs were observed in 45 historical controls not receiving LAMB prophylaxis (p 0.01). LAMB prophylaxis had no impact on the use of empirical antifungal therapy. Systemic antifungal prophylaxis with LAMB 2.5 mg/kg twice weekly is feasible and safe, and seems to be an effective approach for antifungal prophylaxis in high-risk paediatric cancer patients.     

Sequential treatment of deep fungal infections with amphotericin B deoxycholate and amphotericin B colloidal dispersion

Amphotericin B colloidal dispersion (ABCD) is a novel lipid formulation of amphotericin B designed to diminish toxic effects of the drug. In the following report, nine cases of suspected (n=4) and proven (n=5) deepCandida infection, treated sequentially with amphotericin B deoxycholate and ABCD, are presented. The treatment was successful in seven cases. During treatment with amphotericin B deoxycholate, a rise in serum creatinine was observed in seven patients, hypokalemia in five, and metabolic acidosis in four. After replacing amphotericin B deoxycholate with ABCD, laboratory parameters improved in four of the seven patients with increased creatinine, in four of the five patients with hypokalemia, and in two of the four patients with metabolic acidosis. Infusion-related rigors were observed in four patients receiving amphotericin B deoxycholate and in one patient treated with ABCD. Reversible elevation of liver enzymes was found in one patient receiving ABCD. In this study ABCD proved less toxic than amphotericin B deoxycholate. The efficacy of ABCD alone cannot be assessed because of previous treatment with amphotericin B deoxycholate, but sequential treatment of deepCandida infections with amphotericin B deoxycholate and ABCD seems to be an effective therapeutic modality, especially in patients requiring prolonged administration of amphotericin B.     

Assessment of nephrotoxicity in patients receiving amphotericin B lipid complex: a pharmacosurveillance study in Spain

This study assessed the risk of haematological, renal and hepatic toxicity associated with amphotericin B lipid complex (ABLC; Abelcet) in a multicentre, open-label, non-comparative study of 93 patients from 17 different hospitals who received ABLC because of proven or suspected systemic fungal infection or leishmaniasis. Most (66%) patients had onco-haematological diseases. Optimum treatment with ABLC comprised a slow (2-h) infusion dose of 5 mg/kg/day for a minimum period of 14 days. Biochemical and haematological parameters were measured pre-, during and post-treatment. In the overall patient group, the mean serum creatinine concentration was similar pre- and post-study (1.00 +/- 1.14 mg/dL vs. 1.20 +/- 1.19 mg/dL; p > 0.05). There were no significant changes pre- and post-treatment in concentrations of haemoglobin, potassium, transaminases and bilirubin. There was no significant correlation between the dose administered and the concentrations of serum creatinine (Spearmann 0.22). There was no greater nephrotoxicity in the patients with previous renal failure, or in those who had received amphotericin B previously. There were serious adverse events in five patients, but other alternative causes that could explain these events were present in three of these patients. Fevers or chills were experienced by 23% of the patients during the ABLC infusion, but only in one case did this necessitate the suspension of treatment. It was concluded that ABLC is a drug with low nephrotoxicity, even when administered to patients with pre-existing renal insufficiency. Adverse events were generally slight or moderate, and were managed easily with appropriate pre-medication.     

EUCAST Technical Note on Aspergillus and amphotericin B,itraconazole, and posaconazole

The European Committee on Antimicrobial Susceptibility Testing Subcommittee on Antifungal Susceptibility Testing (EUCAST-AFST) has determined breakpoints for amphotericin B, itraconazole and posaconazole for Aspergillus species. This Technical Note is based on the EUCAST amphotericin B, itraconazole and posaconazole rationale documents (available on the EUCAST website: http://www.eucast.org/antifungal_susceptibility_testing_afst/rationale_documents_for_antifungals/). The amphotericin B and itraconazole breakpoints are based on epidemiological cut-off values and clinical experience. The posaconazole breakpoints are also based on pharmacokinetic and pharmacodynamic data. Breakpoints will be reviewed regularly or when new data emerge.     

Safety of amphotericin B colloidal dispersion

The safety of amphotericin B colloidal dispersion (ABCD) was tested in five open-label Phase I/II clinical trials in 572 selected patients who had a fungal infection secondary to a severe underlying disease. In 442 cases ABCD was administered after therapy with amphotericin B, which had been withdrawn in 192 of them because of toxicity. One hundred and forty patients had pre-existing nephrotoxicity. ABCD doses of up to 6 mg/kg/day resulted in no differences in serum creatinine levels, even in patients with pre-existing renal failure. ABCD therapy resulted in no difference in liver function as measured by SGOT, alkaline phosphatase and total bilirubin levels in serum. Apart from thrombocytopenia, there was no significant alteration in hematological or other biochemical parameters in the blood. Adverse events attributable to ABCD requiring discontinuation of therapy occurred in 70 patients (12.2%). The most frequent of these were infusion-related adverse events, which occurred in 5.4% of patients. As a consequence, the maximum tolerated dose was set at 7.5 mg/kg/day. These studies show clearly that ABCD can be administered safely to patients without the risk of renal toxicity, even when renal impairment has already developed following therapy with conventional amphotericin B deoxycholate.     

In vivo emergence of high-level resistance during treatment reveals the first identified mechanism of amphotericin B resistance in Candida auris

ObjectiveCandida auris has emerged as a health-care-associated and multidrug-resistant fungal pathogen of great clinical concern. As many as 50% of C. auris clinical isolates are reported to be resistant to amphotericin B, but no mechanisms contributing to this resistance have been identified. Here we describe a clinical case in which high-level amphotericin B resistance was acquired in vivo during therapy and undertake molecular and genetic studies to identify and characterize the genetic determinant of resistance.MethodsWhole-genome sequencing was performed on four C. auris isolates obtained from a single patient case. Cas9-mediated genetic manipulations were then used to generate mutant strains harbouring mutations of interest, and these strains were subsequently subjected to amphotericin B susceptibility testing and comprehensive sterol profiling.ResultsA novel mutation in the C. auris sterol-methyltransferase gene ERG6 was found to be associated with amphotericin B resistance, and this mutation alone conferred a >32-fold increase in amphotericin B resistance. Comprehensive sterol profiling revealed an abrogation of ergosterol biosynthesis and a corresponding accumulation of cholesta-type sterols in isolates and strains harbouring the clinically derived ERG6 mutation.ConclusionsTogether these findings definitively demonstrate mutations in C. auris ERG6 as the first identified mechanism of clinical amphotericin B resistance in C. auris and represent a significant step forward in the understanding of antifungal resistance in this emerging public health threat.     

Efficacy of voriconazole plus amphotericin B or micafungin in a guinea-pig model of invasive pulmonary aspergillosis

The efficacy of voriconazole in combination with amphotericin B or micafungin was studied in a transiently neutropenic guinea-pig model of invasive pulmonary aspergillosis. Guinea-pigs treated with the antifungal drugs, alone or in two-drug combinations, had an improved survival rate and reduced fungal burden in the lungs compared to untreated control animals. The efficacy of monotherapy and combination therapy was similar; activity was neither enhanced nor reduced with the two-drug combinations. Further studies of efficacy, dosing and optimal regimens for antifungal combinations are warranted.     

Compassionate use of amphotericin B lipid complex (Abelcet) in life-threatening fungal infections: report of 30 courses

Objective: To report a single-center experience of compassionate use of amphotericin B lipid complex (ABLC) in patients with proven or suspected fungal infection who were or would have been unable to tolerate conventional amphotericin B.
Methods: Twenty-eight patients receiving 30 courses of ABLC for 22 proven invasive mycosis episodes (11 aspergillosis, seven candidosis, four miscellaneous) and eight suspected episodes are described. Seven patients were given ABLC first-line therapy because of conditions precluding the use of amphotericin B deoxycholate (Am B). Twenty-one patients, initially given Am B, were shifted to ABLC because of failure in four, nephrotoxicity of AM B alone or in combination with another drug in 15, and acute side effects in two. The initial dose of ABLC was 5 mg/kg per day; this could be lowered to 3 mg/kg per day or transiently interrupted in cases of impairment of renal function.
Results: A mean cumulative dose of 6107 mg (660–16 050) was given over a mean duration of 22 days (4–49). Clinical response rate was 63% (14/22), with mycologic eradication in 37% (9/17) in proven infections. For proven aspergillosis, corresponding rates were 54% (6/11) and 20% (2/10), and in proven candidosis 71% (5/7) and 60% (3/5), respectively. Twenty-one courses were complicated by one or more side effects: fever and chills (11), impairment of renal function requiring a transient reduction of drug dosage (14), hypotension (1). However, for the whole group, creatinine clearance before and after 2, 4 and 6 weeks of treatment remained quite stable.
Conclusions: ABLC, with its low toxicity, enabled us to treat patients who were or would have been unable to tolerate an efficacious dose of Am B. No conclusions about efficacy can be drawn from this small-size, compassionate study. Well-designed studies to compare efficacy and safety of conventional amphotericin B and the various lipidic formulations should be implemented.     

A comparison of AmBisome to amphotericin B for treatment of systemic candidiasis in very low birth weight infants

PURPOSE: Amphotericin B is considered the treatment of choice for systemic candidiasis, but adverse effects may limit its use. An alternative option for the treatment of candidiasis includes lipid preparations of amphotericin B. This study investigated the safety and efficacy of AmBisome, a lipid formulation of amphotericin B containing liposomal structures, for the treatment of systemic candidiasis in very low birth weight infants (VLBWI). MATERIALS AND METHODS: Data from 26 VLBWI treated with AmBisome in the study group (AmBisome group) from October 2003 to July 2006 were compared with data from 20 VLBWI treated with amphotericin B as a historical control (Amphotericin group). This study was a prospective, historical control, multi-center trial. RESULTS: Candida spp. was isolated in 73% (19/26) of the cases for the AmBisome group and 90% (18/20) of the cases for the Amphotericin group. The fungal eradication rate and the time to eradication was 84% (16/19) and 9+/-8 days in the AmBisome group, and 89% (16/18) and 10+/-9 days in the Amphotericin group, respectively (p=0.680 vs p=0.712). The major adverse effects were lower in the AmBisome group (renal toxicity, 21% vs 55%, p=0.029; hepatotoxity, 25% vs 65%, p=0.014, AmBisome group vs Amphotericin group, respectively). There was no significant difference in mortality attributed to systemic candidiasis (12% in the AmBisome group, 10% in the Amphotericin group, p=0.868). CONCLUSION: AmBisome is effective and safe for treating systemic fungal infections in VLBWI.