Amphotericin B

Amphotericin B use in children: conventional and lipid-based formulations

Necdet Kuyucu
Mersin University, Faculty of Medicine, Department of Pediatric Infectious Diseases, 33079, Mersin, Turkey [email protected]
Invasive fungal infections (IFIs) are important causes of morbidity and mortality in immunocompromised children. The increased incidence and high mortality rates associated with IFIs has led to development of novel antifungal agents to expand the breadth and effectiveness of treatment options available to clinicians. Since its initial approval in 1958, conventional amphotericin B deoxycholate had been considered the standard in treatment for IFIs. However, because of the dose-limiting toxicity of conventional amphotericin B deoxycholate, lipid formulations of amphotericin have been developed to potentially improve outcomes and mitigate the adverse effects associated with antifungal therapy. While less frequently employed today as prophylaxis (given the expanded availability of safer alternatives), amphotericin B is still considered a treatment option in select cases of severe or life-threatening IFIs. This article reviews the clinical use of amphotericin B for the prevention and treatment of IFIs.

Amphotericin B
Mechanism of action
The oldest antifungal class is the polyene macro- lides, amphotericin B (AMB) and nystatin. Initially approved for use in 1958, AMB deoxy- cholate (AMBd) is still an important, although not gold standard, therapeutic agent for many invasive fungal infections (IFIs) and the compar- ative agent for all newer antifungal agents [1–6]. AMB binds to ergosterol, the major sterol found in fungal cytoplasmic membranes, creating trans- membrane channels resulting in an increased permeability to monovalent cations. The result is the production of pores that allow concentra- tion-dependent leakage of intracellular cations (e.g., potassium and magnesium) and other cell components [7]. This leads to loss of membrane potential and subsequent fungal cell collapse [7]. Other mechanisms proposed include cell dam- age resulting from oxidative reactions linked to lipoperoxidation of the cell membrane [8]. While AMBd demonstrates a greater affinity for ergo- sterol, the nonselective binding to cholesterol (the primary sterol in human cell membranes) may contribute to AMBd-related adverse events.
Over the past few decades, new formula-
tions of AMB have been developed in an effort
to decrease nephrotoxicity while maintaining or possibly improving efficacy. Liposomes are composed of a phospholipid bilayer surrounding an aqueous core. Phospholipids have a bipolar character, containing a hydrophilic head and lipophilic tails. When placed in an aqueous medium, they arrange themselves so that the tails are shielded from the outside aqueous medium as well as the inner aqueous core [9]. Liposomal prep- arations have a number of potential advantages [10]: drugs with poor solubility can be prepared easily; the lipid bilayer may protect drugs from being destroyed by enzymatic degradation; lipo- somes may change pharmacokinetic profiles by slowly releasing drugs to provide for less frequent dosing; and drug delivery could be targeted to desired areas to improve efficacy and tolerability. Lipid-based AMB (LBF-AMB) has several advan- tages over the conventional AMB, namely up to a tenfold increase of daily dose, high tissue con- centration with better delivery to reticuloendo- thelial organs (lungs, liver and spleen), decrease in infusion-related side effects and decrease in renal toxicity. The first LBF-AMB is AMB col- loidal dispersion (ABCD; Amphotec), which contains roughly equimolar amounts of AMB and cholesteryl sulfate. The second formulation
ISSN 1478-7210

is liposomal AMB (L-AMB; Ambisome), which is composed of a small unilamellar liposome of approximately 55–75 µm in diameter, made up of a bilayer membrane of hydrogenated soy phosphatidylcholine and distearoyl phospatidylglycerol stabilized by cholesterol and combined with AMB in a 2:98:1, 0:4 ratio. The third formulation is AMB lipid complex (ABLC; Abelcet), which is a ribbon-like structure of a bilayered membrane formed by combining a 7:3 mixture of dimyristoyl phosphatidyl glycerol with AMB (drug:lipid ratio of 1:1). Although in vitro activity can vary between test conditions and isolates tested, studies have generally documented that the activity of AMB is rapidly fungicidal against susceptible organisms [11,12]. Fungicidal concentrations in vitro are generally one to three dilutions higher than that required for inhi- bition [13]. The antifungal activities of new LBF-AMB in vitro are comparable to those of standard AMB [10].
The immunomodulatory effect of AMB has been examined in numerous investigations [14,15] and summarized in detail else- where [16]. These effects include stimulation of proinflammatory cytokines: TNF-, IL-1 and -6, the chemokines IL-8, MCP-1, MIP1, nitric oxide, prostaglandins and ICAM-1 from murine and human immune cells in vitro and in vivo [14–16]. The cytokine release resulting from AMB administration may also be respon- sible (in part) for infusion-related reactions and differences may exist between preparations [16]. Other proposed mechanisms for such reactions include the release of prostaglandins [17]. Finally,
the antifungal activity of pulmonary alveolar macrophages and polymorphonuclear leukocytes against Aspergillus fumigatus may be augmented by the administration of AMB [15]. In addition to its rapidly fungicidal activity, AMB has demonstrated a pro- longed postantifungal effect (PAFE) against both Candida and Cryptococcus spp. [13,18]. The PAFE for Aspergillus, however, may be species-dependent, with significantly shorter PAFEs or lack of PAFE observed for Aspergillus terreus, Aspergillus ustus and Aspergillus nidulans [19]. In contrast to their activity against plank- tonic fungal cells, the in vitro activity of antifungals (including polyenes) can vary significantly in biofilms [20]. While AMBd’s activity in vitro against Candida spp. in biofilms is markedly reduced, both L-AMB and ABLC exhibited similar inhibitory activity in vitro against Candida biofilms in one report [21]. Similar findings were seen for ABLC in a rabbit model [22]. Numerous in vitro and animal model studies have been performed to assess the interaction of AMB with other antifungals (e.g., flucytosine, azoles and echinocandins). The lack of standards for synergy and antago- nism testing may limit the utility of such information and data may differ with the agents tested, model, test conditions and end point(s). The potential for AMB in combination with nonantifun- gals has also been explored. Examples include combinations with azithromycin (against Fusarium [23] and Aspergillus spp. [1]) and rifabutin (for both Fusarium and Aspergillus spp. [1]). However, the clinical significance of such interactions has not been established.

The majority of studies investigating the pharmacodynamics of polyenes have involved AMBd [24]. In time-kill studies employing in vivo and in vitro models for both Candida spp. and Aspergillus
spp., AMB displays concentration-dependent fungicidal activ- ity against susceptible Candida albicans, Cryptococcus neoformans and A. fumigatus [25]. In neutropenic pharmacokinetic/pharmaco- dynamic mouse models of disseminated candidiasis and pulmo- nary aspergillosis, peak plasma concentration (Cmax)/MIC was the parameter that provided the best correlation with outcome as measured by the residual organismal burden in tissue [26]. These laboratory findings indicate that large doses will be most effective and that achievement of optimal peak concentrations is impor- tant. Therefore, the dosage of AMB should not be uncritically reduced and infusion for durations longer than that recommended by the manufacturer should be avoided. Similar findings have been observed with L-AMB [27, 28]. However, the limited drug solubility, reversible binding in tissues and dose-dependent drug clearance of AMB may limit the benefit of dose-escalation in attempts at increasing the antifungal activity [29]. Data in humans to exam- ine the relationship between pharmacodynamics and outcomes are limited [30]. Subset analysis of data obtained in ten pediatric oncology patients treated with L-AMB for whom pharmacokinetic and susceptibility data were available suggested that a Cmax/MIC ratio exceeding 40 was more likely to achieve a complete ver- sus partial clinical response [31]. However, while such information might be important in empiric selection and dosing of antifungal therapy, neither AMB serum concentrations or in vitro susceptibility information is routinely available in clinical situations.

Clinical use of AMB is based on the limited pharmacokinetic information available for this agent in children. Dosage regi- mens are often dictated by toxicity rather than by the findings of controlled clinical studies [32]. AMB demonstrates a limited and erratic absorption following oral exposure. Low (yet detect- able) serum concentrations of AMB have been reported following oral administration [1]. While such absorption may be altered in the setting of mucositis, it is generally considered insufficient to treat systemic infections. The dose, frequency and infusion rate of AMB can influence plasma concentrations. Cmax following intravenous infusion of 1 mg/kg of AMBd have been reported to be approximately 2 µg/ml [5]. The high protein binding of AMB (in excess of 90%) to serum albumin and -1-acid glyco- protein is directly related to concentration [29]. Distribution has been described using a three-compartmental model [1,29] with the resulting volume of distribution of approximately 4 l/kg [1]. The disposition of the compound follows a three-compartment model, with rapid initial clearance from plasma followed by a biphasic pattern of elimination with a  half-life of 24–48 h and a pro- longed terminal () half-life of 15 days or more [26]. Tissue levels of AMB in laboratory animals are highest in the liver, spleen, bone marrow, kidney and lung; concentrations in body fluids other than plasma are generally low [26]. There was considerable vari- ation in the peak concentrations in serum among 12 patients in childhood (six with leukemia) [32]. Peak concentrations in serum occurred at the end of the infusion and ranged from 0.78 to
10.02 µg/ml. No correlation was found between dosage and peak concentration in serum, even when the dosages were corrected for

bodyweight [32]. Penetration of AMB into the CNS is thought to be minimal, with cerebrospinal fluid (CSF) concentrations being 0–4% of simultaneous serum concentrations [1]. However, such CSF concentrations are not likely to reflect higher concentrations in the meninges. Despite mostly undetectable concentrations in the CSF and comparatively low concentrations in brain tissue across all species, AMB is effective in the treatment of fungal infections of the CNS.
Metabolism plays a minor role in elimination of AMB [29]. Elimination of AMB is also poorly understood. For example, only 3% of the total dose is excreted as unchanged drug [29]. Clearance from plasma is slow and dose-dependent, with a ter- minal half-life of more than 15 days [29]. Concentrations in blood have been detected up to 4 weeks after an AMB treatment course and in urine for 4–8 weeks following completion of therapy. The pharmacokinetic differences between preparations of AMB have been reviewed in detail elsewhere [28,29,32,33]. In general, LBF- AMB exhibits a range of serum concentrations, with ABLC and
ABCD demonstrating similar Cmax values to AMBd when given at recommended dosages. By contrast, L-AMB exhibits both a higher Cmax and AUC relative to AMBd, ABLC and ABCD at
comparable doses, likely due to significant reduction in L-AMB’s
volume of distribution and total body clearance [32,33]. Reductions in unbound AMB have been reported with LBF-AMB relative to AMBd [29]. L-AMB may produce lower tissue concentrations in the liver, spleen, lung and kidney [3]. In contrast, L-AMB achieves higher concentrations in brain tissue [33]. ABLC achieves higher lung concentrations [28,34]. Studies have examined the pharmaco- kinetics of AMB (including LBF-AMB) in a variety of special populations, including patients with renal dysfunction [35]. AMB is poorly removed by dialysis. Studies have also been conducted to examine the pharmacokinetic profile of AMBd [36], L-AMB [1,31] and ABLC [37] in pediatrics. In general, significant differ- ences between adult and pediatric patients justifying alterations in weight-based dosing ranges have not been observed.

Spectrum of activity
Amphotericin B possesses a broad spectrum of antifungal activ- ity against a variety of yeasts and molds [38]. Potency differences in vitro are consistently observed between AMBd and the three lipid-based formulations (LBFs): ABLC, ABCD and L-AMB. LBF-AMB generally exhibits a fivefold reduction in potency in vitro (when expressed as mg/kg) relative to AMBd [27]. The relevance of these in vitro differences remains uncertain, since higher MICs for LBF-AMB may not account for release from the lipid carrier. By contrast, recent data from the European Committee on Antimicrobial Susceptibility Testing (EUCAST) reported increased potency for L-AMB (relative to AMBd) for several strains of Candida spp. [13]. The reasons for such findings, however, were not clear. While Clinical Laboratory Standards Institute (CLSI) standards exist for susceptibility testing of AMB against yeasts by both macrodilution and microdilution meth- ods, no breakpoints have been approved [1]. CLSI-recommended disk diffusion methods for susceptibility testing of yeasts do not include AMB [1]. In addition, the utility of in vitro testing
may be limited by the general lack of correlation between in vitro susceptibility and treatment outcome in patients with IFIs [30]. AMB is highly active in vitro against most Candida spp. (includ- ing C. albicans, Candida glabrata, Candida tropicalis and Candida parapsilosis), with MICs generally ranging between 0.5 and 1 µg/ml [39]. More recently, MICs from a variety of Candida spp. tested according to methodology described by the EUCAST ranged from 0.06 to 2 µg/ml [40]. While Candida lusitaniae is thought to be intrinsically resistant to AMB, in vitro data are often conflict- ing and the clinical significance of this resistance is questionable [1,40,]. The clinical data about AMB resistance in C. lusitaniae infections are very contradictory; AMB is generally found to be nonefficacious in treatment of this organism. However, this may result from lack of standardization in susceptibility tests [41–43]. AMB displays favorable activity in vitro against Cryptococcus spp., Malassezia spp. and Saccharomyces spp. However, Trichosporon beigelii displays variable in vitro susceptibility to AMB [1]. AMB is highly active in vitro against fungi responsible for endemic mycoses, such as Coccidioides spp., Blastomyces spp., H. capsulatum and Paracoccidioides brasiliensis [1]. It also exhibits activity in vitro against most Sporothrix schenkii, although strain-dependent resist- ance has been reported [1]. In addition to its activity against yeasts, AMB has also demonstrated significant activity against a variety of molds, including most Aspergillus spp. (with the exception of A. ter- reus) [38,40]. AMB is active in vitro against the Zygomycetes [44]. While Scedosporium apiospermum may be susceptible to AMB, Scedosporium prolificans is generally resistant [7]. Although AMB may be active against Fusarium spp., MICs of 0.25–8 µg/ml have been reported for Fusarium solani and therefore some strains of this species are considered resistant [7]. Cladophialophora isolates have demonstrated variable susceptibility to AMB. Finally, intrin- sic resistance has been reported in both Malassezia furfur and Paecilomyces lilacinus [7].

Adverse effects
It is perhaps the adverse event profile that most limits the cur- rent use of AMB [45–49]. A variety of electrolyte abnormalities have been associated with AMB administration, most commonly hypokalemia and hypomagnesemia [1,26]. Recent clinical studies report hypokalemia in approximately 10–20% of patients receiv- ing various AMB formulations. Hyperkalemia is less frequently reported and has been more frequently associated with rapid infusions [26].
Infusion-related reactions related to intravenous adminis- tration of AMB occur frequently, ranging from 20 to 90% (depending largely upon population, preparation, administra- tion and the use of premedications) [1,26]. Such effects usually occur during the infusion or within 1–3 h following therapy. These include headache, fever, chills and rigors. Gastrointestinal complaints (e.g., nausea, vomiting and abdominal discomfort) may also occur during or directly following administration. Less common reactions during or immediately following infu- sion include bronchospasm, hypotension, thrombophlebitis and cardiac arrhythmias. Hypertension has also been reported [48]. Anaphylaxis associated with AMB administration has rarely

been reported [1]. Rapid infusion of AMBd (i.e., less than 4–6 h) may increase the incidence of infusion-related reactions [1,26]. In addition, the formulation of AMB may influence the frequency of infusion-related reactions [1,26]. For example, infusion-related reactions on day 1 of empiric therapy without premedications were reported in 88.5% of febrile neutropenic patients receiv- ing ABLC 5 mg/kg/day compared with 52 and 48% of those receiving L-AMB 3 or 5 mg/kg/day, respectively (p < 0.001) [1]. Infusion-related reactions have also been reported more frequently among subjects receiving ABCD than those receiving AMBd [1]. Select reactions have also been reported to occur more frequently with certain formulations. For example, a triad of hypoxia, back pain and chest pain has been reported following administration of L-AMB [1]. A prospective analysis found a 20% mean overall frequency (range: 0–100%) of acute infusion-related reactions among 84 patients at 64 centers [1]. While these reactions rarely required discontinuation of therapy, slowing the infusion rate had no effect on the infusion-related reactions described. ABCD administration has been associated with hypoxia, dyspnea and res- piratory distress, which may necessitate cessation of therapy and the need for supportive care [1]. Reactions in individual patients may also be formulation specific and may not necessarily recur upon rechallenge with a different formulation. Numerous strate- gies have been proposed to minimize the frequency and severity of AMB-related infusion reactions, including the administration of premedications and the use of L-AMB. Renal dysfunction second- ary to AMB administration is often the treatment-limiting adverse effect of AMB. Proposed mechanisms include direct interaction with epithelial cell membranes (causing cellular disruption) and renal vasoconstriction (with resulting reductions in renal blood flow). Manifestations may include renal tubular acidosis, casts in the urine, azotemia, oliguria, and magnesium and potassium wasting [1]. The incidence of AMB-induced nephrotoxicity varies widely between studies because of differences in definition, study population, underlying risk factors, duration of therapy and use of premedications. However, such reactions (generally described as a doubling of the baseline creatinine value) have been reported in up to 50% of patients receiving AMBd [50]. Risk factors include underlying renal dysfunction, formulation, concomitant nephro- toxins and dosing (daily and cumulative). Significant economic and clinical consequences can result [1,50]. Numerous strategies have been employed in attempts to reduce the incidence and sever- ity of AMB-induced renal dysfunction. These include careful patient selection and (whenever possible) minimizing concomi- tant nephrotoxins. Use of saline loading [48,49], aggressive fluid resuscitation and continuous infusions [51] have also been investi- gated. More common today in patients at increased risk or expe- riencing AMBd-related nephrotoxicity is the use of LBF-AMB, which exhibits a reduced incidence of nephrotoxicity when com- pared with AMBd [52,53]. For example, ABLC-associated nephro- toxicity (defined as a doubling of baseline serum creatinine) was observed in 13% of 3514 patients with suspected or proven IFI in a multicenter, open-label retrospective study [52]. Studies compar- ing the incidence of nephrotoxicity between preparations have also been evaluated. For example, doubling of serum creatinine
was significantly less frequent with L-AMB 3 mg/kg/day than with AMBd 0.6 mg/kg/day when given as empiric therapy in persistently febrile neutropenic patients (18.7 vs 33.7%, respec- tively; p < 0.001) [54]. L-AMB (3 or 5 mg/kg/day) has also been compared with ABLC (5 mg/kg/day) in this patient population, with rates of doubling of serum creatinine from baseline observed in 29.4, 25.9 and 42%, respectively [55]. Other reports comparing L-AMB and ABLC for a variety of indications and ranges of doses for both agents (i.e., between 3 and 5 mg/kg/day) failed to detect significant differences between these preparations [53]. ABCD 6 mg/kg/day demonstrated a reduced incidence of nephrotoxic- ity relative to AMBd 1–1.5 mg/kg/day in patients with invasive aspergillosis (IA; 12.5 vs 38.4%, respectively) [56]. Elevations in liver function tests have less frequently been associated with AMB administration [57]. Anemia (usually normochromic or normo- cytic) has been reported as secondary to AMB administration and may be a consequence either of direct inhibition of erythropoietin
[1] or secondary to renal toxicity.

Overview of clinical applications
Amphotericin B’s status as the ‘gold standard’ for the preven- tion and treatment of IFIs has been put into question by the recent introduction of new options, namely extended-spectrum triazoles and echinocandins [1]. Despite these new alternatives, AMB is continually cited by numerous consensus guidelines as an option for serious and treatment-refractory IFIs [1,3–6]. With few exceptions, AMBd is frequently replaced by LBF-AMB, largely based on the potential for increased safety [3]. However, control- led studies examining LBF-AMB as primary therapy for IFIs are limited. In addition, while ABCD may be associated with increased infusion-related reactions [56], clinically significant dif- ferences between other LBF-AMB (i.e., L-AMB and ABLC) may be less clear [1].

Nystatin usage for the treatment of infections due to Candida spp. is restricted to the treatment of mucocutaneous forms. This may include oropharyngeal, cutaneous, mucocutaneous and vulvo- vaginal infections. The efficacy of AMBd in infections caused by many Candida spp. has been established in invasive candidiasis (IC), including candidemia, osteomyelitis, disseminated candidia- sis, endophthalmitis and endocarditis [1]. In their recent therapeu- tic guidelines, the expert panel of the Infectious Diseases Society of America (IDSA) does not, in general, explicitly differentiate between adults and pediatric patients [58]. However, special guid- ance is given for neonatal candidiasis. Neonates with Candida- positive cultures from physiologically sterile body fluids and/or urine should undergo CSF and retinal exams. The genitourinary tract, liver and spleen should be imaged if physiologically ster- ile body fluid cultures show persistently positive results. AMBd (1 mg/kg/day) is recommended for neonates with disseminated candidiasis. If candiduria is excluded, L-AMB (3–5 mg/kg/day) can be an option. Alternatively, fluconazole (12 mg/kg) may be used. The duration of therapy should be at least 3 weeks. Owing to a lack of data in neonates, echinocandins should generally

be restricted to patients with refractory infection or toxicity of fluconazole or AMB. This recommendation may also relate to the comparatively high prevalence of C. parapsilosis, a species with comparatively weak susceptibility to echinocandins in young children. For infections with this species, echinocandins may not be the optimal choice since the MICs of the strains often reach the susceptibility breakpoint of 2 µg/ml proposed by the CLSI [59]. There is no general evidence of increased failure rates with echi- nocandins in C. parapsilosis. However, C. parapsilosis fungemia was found to be associated with prior caspofungin therapy in retrospective data sets of hematologic patients [60], and the IDSA guidelines do not recommend first-line use of echinocandins in patients with documented C. parapsilosis infection [58]. Removal of intravascular catheters is strongly advised. Flucytosine is not routinely recommended in cases of Candida meningitis owing to insufficient comparative data, although it has been used exten- sively in combination with AMB in this indication. In neonates, AMBd is still a recommended option in disseminated candidiasis (IDSA evidence level A-II) [56] owing to a scarcity of meaning- ful data with newer antifungals and owing to its more favorable tolerability in this population as compared with older patients. In a recent retrospective study, transient nephrotoxicity was reported in 16% of neonates (<90 days of age) receiving at least three doses of AMBd, while 17% developed hypokalemia [61].
Fluconazole was established as a standard option for the treat-
ment of IC after a randomized trial in adult patients had shown it has therapeutic equivalence to AMB and superior tolerability [62]. In a small randomized study of fluconazole versus AMB in pedi- atric IC patients (n = 23), the case–fatality rate was substantially lower with fluconazole (33 vs 45%) and fluconazole was better tolerated [63]. It should be noted that fluconazole is inactive against Candida krusei and must be considered as inadequate therapy in patients with Candida glabrata infections, which however, occur at low incidence in pediatric patients.
A study comparing caspofungin and AMB therapy in 224 adults with IC has been conducted. Response to caspofungin (n = 104; 73.4%) was slightly better than response to AMB (n = 115; 61.7%) [64]. More recently, caspofungin (n = 556) was compared with L-AMB (n = 539) in febrile neutropenic patients and the
overall success (33%) was virtually identical in both groups [65].
In pediatric patients, caspofungin was prospectively investigated in a multicenter trial including 37 cases (age: 3 months–17 years) of IC infections, mostly candidemia [66]. In patients receiving primary therapy (n = 30), favorable outcomes were seen in 81% of cases (complete responses).
Micafungin was shown to be noninferior to AMB and caspo- fungin in two randomized trials in adult patients with invasive Candida infections [67, 68]. The efficacy of micafungin in pediatric patients with IC was prospectively investigated in a substudy of a double-blind, randomized trial comparing micafungin (2 mg/kg) with L-AMB (3 mg/kg) as a first-line therapy of IC (n = 106) [53]. A total of 57 patients who were 2 years of age or younger and of whom 19 were premature infants, were incuded. A successful out- come was reported for 73% of patients treated with micafungin versus 76% receiving L-AMB. Treatment success was independent
of neutropenic status or prematurity. Consistent with the good safety profile of the echinocandins, adverse events leading to dis- continuation of study therapy were significantly less frequent in the micafungin group.
In general, efficacy for AMBd is comparable to these new agents. However, newer agents are generally better tolerated than AMBd. In the case of voriconazole, an alternative exists for continued oral therapy once the patient is stable. LBF-AMB has also been used in the treatment of IC. The efficacy of ABLC has been reported in open-label studies [69]. For example, cure (30%) or improvement (30%) was noted in the treatment of IC in ABLC-treated patients in a retrospective observational trial [69]. In noncomparative studies, ABLC has been found to be an effec- tive antifungal agent in children. In an open-label pediatric trial, complete or partial therapeutic response was observed in 70% (38 of 54) and 81% (22 of 27) of patients with candidiasis, respec- tively [70]. A retrospective study of 46 children treated with ABLC reported an overall response rate of 83 (38 of 46) and 89% (17 of 19) against candidiasis [71]. Use of L-AMB for the treatment of IC has also been reported in pediatric patients. Few published data exist on the use of lipid formulations of AMB in neonates. One study that included 40 preterm neonates noted that L-AMB was associated with clinical resolution in greater than 70% of patients with candidiasis [72]; other uncontrolled studies have confirmed the high response rates. In three other studies, 21 of 21, 35 of 37 and 20 of 24 neonates with candidiasis cleared their infections
[72]. Published experience with ABCD is somewhat limited. It
has been studied in an open-label, Phase I [56] and retrospective analysis of open-labeled trials. Early trials comparing ABCD with azoles (fluconazole) demonstrated comparable efficacy but ABCD was less well tolerated [1]. One published experience in the neonatal setting includes 16 very-low-birth-weight infants with IC and a serum creatinine concentration of 1.2 mg/dl or greater [26]. Infants received 3 mg/kg ABCD on day 1, followed by 5 mg/kg/day. A total of 13 of 14 evaluable patients cleared the organism after therapy with ABCD alone (n = 8) or in combina- tion with another agent (n = 5) and overall survival was 75%. ABCD was well tolerated without infusion-related reactions, increases in serum creatinine or hepatotoxicity. More recently, L-AMB was demonstrated to be equally effective but less well tolerated than micafungin for candidemia and IC in adults [64]. ABLC and L-AMB are US FDA-approved as second-line therapy for use in proven candidiasis in patients intolerant or refractory to AMBd. The potential role of AMBd as part of combination therapy (with fluconazole) was examined in a randomized study in non-neutropenic patients with candidemia [73]. In this trial, 30-day success rates were not different between subjects receiving fluconazole plus placebo (57%) or fluconazole in combination with AMBd (69%; p = 0.08). The rate of clearance from the bloodstream trended toward improvement in patients receiving combination therapy. The availability of equally efficacious and better tolerated agents limit the role of AMBd for candidiasis. Current expert guidelines about the treatment of IC limit the role of AMB in severe, refractory infections, CNS infections and in pregnant patients [1].

In contrast to adult patients, large prospective comparative studies have not been performed in pediatric patients with IA. Moreover, pediatric subgroups have not been analyzed in pub- lished studies that include a broader age range. For example, the large randomized trial of voriconazole versus AMB in IA included adolescents from 12 years of age [74]. The results of the adolescent subpopulation were, however, not reported sepa- rately. Thus, clinicians treating pediatric IA are largely left with the results of uncontrolled trials, observatory surveys, salvage therapy data and extrapolations from adult studies to guide their treatment choices.
The last decade has seen a substantial expansion of the range of mold-active antifungals. A recent multicenter analysis of 139 pediatric IA cases recorded from the years 2000–2005 revealed that L-AMB (57.3%), voriconazole (52.7%) and caspofungin (42.0%) were the most commonly used therapeutic agents in mold infections [75].
Since the largest randomized therapeutic trial performed to date on IA established the significant superiority of voriconazole versus AMB in terms of response rates and survival [74], voriconazole has been the first-line treatment of choice for IA, which is reflected in several current therapeutic guidelines, including those of the IDSA, the European Conference on Infections in Leukaemia and the German Society of Hematology and Oncology [76,77]. Of note, voriconazole has been shown to provide unprecedented response rates in cerebral aspergillosis, most likely due to its unique ability to penetrate CNS compartments [78].
Amphotericin B deoxycholate, the traditional mainstay of anti- fungal therapy, is still in use in pediatric patients despite its dose- limiting nephrotoxicity. However, a recent multicenter survey in pediatric IA patients [75] indicates that it is being replaced more and more by newer antifungal preparations, mostly L-AMB and voriconazole, owing to their more favorable tolerability profile and therapeutic superiority demonstrated in the adult population, respectively.
Liposomal AMB is recommended by the IDSA for treat- ment of IA as an alternative to voriconazole in adults based on a randomized dose-comparison trial investigating L-AMB 10 versus 3 mg/kg/day [76]. While in clinical practice L-AMB is an established option in pediatric patients with IA, published prospective data on its use for IA in this age group are limited. For example, Groll et al. report on the – mostly empirical – use of L-AMB in pediatric recipients of allogeneous stem cell trans- plants, involving 84 patients including four with proven or prob- able IA of whom two were successfully treated [79]. According to pharmacokinetic data, equal per weight dosages produce similar exposure in pediatric patients beyond 1 month of age and in adults [80].
Amphotericin B lipid complex has not been studied in rand- omized comparative trials for IA. Data on its use in this indi- cation mostly stem from the ClearCase collection database [75]. For example, Ito et al. report on 85 patients from this database treated with ABLC with a success rate of 31%; first-line therapy was successful in 11 out of 27 patients (41%) [81].
Cryptococcal infections
While mild-to-moderate forms of cryptococcosis outside the CNS may be managed by an azole such as fluconazole or itraconazole, AMB has become the standard of care for the initial therapy of severe, disseminated disease and infections involving the CNS [3]. The efficacy of AMBd for the treatment of cryptococcal menin- gitis has been established by randomized controlled trials in both HIV and non-HIV patient populations [1,82,83]. Combination therapy of AMBd with flucytosine has improved clinical efficacy, time to sterilization of the CSF and reduction in relapse rates [82]. In HIV-infected patients, AMBd (0.7 mg/kg/day) combined with flucytosine 100 mg/kg/day for 2 weeks followed by azole main- tenance therapy was superior to AMBd plus placebo, but more prolonged courses of flucytosine were of no additional benefit
[83]. Current expert treatment guidelines recommend that AMBd
be combined with flucytosine as initial ‘induction therapy’ for cryptococcal meningitis in both HIV-infected and non-HIV- infected persons [3]. However, the optimal dose of AMBd for this indication is unknown. The potential role of AMBd dose escalation for induction therapy of cryptococcal meningitis was recently examined in HIV-positive patients [84]. Initial doses of 0.7–1.0 mg/kg/day (plus 5-fluorocytosine) for 2 weeks were fol- lowed by fluconazole. In this study, increasing doses demonstrated more rapid fungicidal activity. There is less published experience with LBF-AMB for cryptococcal infections. A comparison of L-AMB 4 mg/kg/day and AMBd 0.7 mg/kg/day as induction therapy in HIV-infected patients (n = 28) concluded that L-AMB was more effective than AMBd in sterilizing the CSF (p < 0.05)
[85]. However, overall clinical response was similar between the
two treatments. The efficacy of ABLC has also been reported for cryptococcosis [86] and cryptococcal meningitis [87]. In patients with HIV-associated cryptococcal meningitis (n = 21), success- ful treatment was reported in 86% of subjects receiving ABLC 5 mg/kg/day [87]. However, rates of CSF sterilization at 6 weeks were only 58%. The current IDSA treatment guidelines recom- mend a dose of L-AMB of 4 mg/kg/day be used as the LBF- AMB, although such treatment is not currently FDA approved
[3]. In addition, while initial treatment of serious cryptococcal disease in solid organ transplant recipients has not been evalu- ated by prospective controlled clinical trials, LBF-AMB (either L-AMB or ABLC) may also be considered over AMBd in this population owing to the frequent coadministration of nephrotoxic calcineurin inhibitors [88].

Limited options exist for the treatment of invasive zygomycoses. While AMB maintains activity in vitro against Zygomycetes, treatment outcomes (especially in the immunocompromised host) remain poor [89,90]. AMBd [91] or lipid-based formulations of AMB [91,92] are frequently prescribed in this clinical setting, especially as initial therapy and frequently in combination with surgical intervention [90,91]. The introduction of newer agents with activity against zygomycoses (i.e., posaconazole) may impact the role of AMB in the future to use in the initial management of severe infections [90].

Endemic mycoses
Current expert treatment guidelines identify AMBd as the first-line therapy for management of severe histoplasmosis infections (e.g., pulmonary and CNS infections, mediastinitis and disseminated disease) [4]. AMBd is, however, rarely used today, except in severe cases [93]. In general, higher doses of AMBd (i.e., 0.7–1 mg/kg/day) have been employed as initial therapy, with reduced doses (i.e., 0.5–
0.6 mg/kg/day) for patients unable to tolerate higher doses. Patients who improved or those with stable disease following initial AMBd therapy (usually 2 weeks) can often be transitioned to azole therapy. LBF-AMB may also have a role, especially for patients intolerant to AMBd [4]. ABLC was evaluated in 25 patients, with efficacy (cure and improvement) observed in 84% [92]. In a randomized, double- blind multicenter study for disseminated histoplasmosis compar- ing AMBd (0.7 mg/kg/day) to L-AMB (3 mg/kg/day) as initial (i.e., 2 weeks) induction therapy for moderate-to-severe disease in AIDS patients, clinical success rates were 64 and 88%, respectively (p = 0.014) [94]. Fewer deaths and improved treatment tolerability were reported in patients receiving L-AMB. However, no differ- ence in time to defervescence, rate of blood culture conversion or change in H. capsulatum antigen levels was observed. Similar to its role in histoplasmosis and despite lack of published data from large controlled clinical trials, AMBd is considered as primary therapy for pulmonary, disseminated and CNS blastomycosis based on published guidelines [5]. AMBd is also the preferred therapy for immunocompro- mised or pregnant patients [5]. Cure rates for AMBd have been reported to range between 70 and 91% [5] with LBF-AMB 3–5 mg/kg/day or AMBd 0.7–1 mg/kg/day and can be administered for 1–2 weeks or until improvement is observed, followed by a transition to an oral agent [5]. Low relapse rates have been reported when cumulative doses of AMBd greater than 1 g have been employed. Similar to histoplasmosis, higher doses of AMBd (0.7–1.0 mg/kg/day) should be considered for disseminated disease with blastomycosis, in which cumulative doses of 1.5–2.5 g or higher (at least 2 g) for CNS disease are used. Published experiences with LBF-AMB are limited. Some open-label experience exists with ABLC, with cure or improvement in nine out of 14 (64%) cases [92]. Despite this relative lack of pub- lished data, LBF-AMB are generally recommended for CNS disease [5]. In general, the use of AMB for the treatment of sporotrichosis is restricted to pregnant patients or those with osteoarticular, pulmo- nary or disseminated infections [6]. LBF-AMB 3–5 mg/kg daily or AMBd 0.7–1.0 mg/kg daily can be used in such settings [6]. Similar to other guidelines, LBF-AMB has been preferred by some clinicians for CNS disease. In a similar circumstance, fluconazole and itraconazole have largely replaced the need for the use of AMB in the treatment of coccidioidomycosis [1]. AMBd 0.5–1.5 mg/kg intravenously daily or on alternating days may be considered in patients with rapidly progress- ing disease, hypoxia and/or respiratory failure, or in pregnant women. Experience with LBF-AMB for the treatment of coccidioidomycosis is largely limited to case reports [92].

Neutropenic fever
Numerous studies have examined the efficacy and safety of AMB in the treatment of fever in neutropenic patients. Early published experience with AMBd helped establish a role of antifungal therapy
in empiric regimens for patients persistently (>7 days) febrile despite broad-spectrum antibacterials [1]. The addition of AMBd substan- tially reduced rates of infection and/or shock (two out of 18 or 11% of patients) compared with broad-spectrum antibacterials alone (six out of 16 patients) or discontinuation of broad-spectrum antibacterials (nine out of 16 patients). Later studies confirmed this by treatment success (defined by absence of fever and infection) in AMBd-treated patients when compared with no treatment (69 vs 53%, respectively)
[1]. More recently, AMBd has been compared with voriconazole [95],
fluconazole [96,97] and caspofungin [64] in this patient population. LBF-AMB has also been evaluated for persistent fever in neutropen- ics. L-AMB (3 mg/kg/day) was compared with AMBd (0.6 mg/ kg/day) among 660 patients in a large, double-blind, multicenter, ran- domized trial [55]. Similar rates of success (based on a composite end point of defervescence, survival, treatment of baseline infection and absence of breakthrough IFI or toxicity requiring treatment discon- tinuation) of 50% (172 out of 343) for L-AMB versus 49% (170 out of 344) for AMBd were reported. However, L-AMB-treated patients experienced fewer proven breakthrough fungal infections (3.2 [11 out of 343] vs 7.8% [27 out of 344]; p = 0.009). In addition, a reduction in infusion-related reactions (including fever [17 vs 44%] and chills or rigors [18 vs 54%]) was observed in L-AMB versus AMBd treatments. Other studies have compared L-AMB to AMBd [55,98,99] and vorico- nazole [1]. ABCD (4 mg/kg/day) therapy has also been compared with AMBd (0.8 mg/kg/day) in a prospective, randomized, double-blind study among neutropenic adults and children [1]. Success rates were 50 (49 out of 98) and 43.2% (41 out of 95), respectively (p = 0.31), while documented or suspected breakthrough fungal infections were reported as 14.3 and 14.7%, respectively. Although ABLC has been compared with both L-AMB [1,54] and AMB [1] in this patient population, such evaluations were designed primarily to evaluate the safety and tolerability rather than efficacy. Given the expanded options of alternative therapies and the underlying risks for toxici- ties associated with AMB, published guidelines for the empiric management of fever in neutropenic oncology patients restrict the role of AMB [1]. Examples include treatment of patients at high risk and/or with clinical evidence of nasal and/or sinus infection (in whom CT scan or MRI findings suggest the potential for IFIs such as aspergillosis or mucormycosis) or those receiving prior azole therapy at risk of invasive mold infections. In these cases, either L-AMB or ABLC should be considered.

Expert commentary & five-year view
Amphotericin B continues to be the broadest-spectrum antifun- gal with cidal activity against many pathogens and a very low potential for treatment-emergent resistance. Its efficacy has been established for numerous IFIs in a variety of patient populations. Despite the recent introduction of new options for prevention and treatment of IFIs, AMB is still prominent in numerous treatment guidelines for severe, life-threatening infections. Requirements for parenteral administration, along with treatment-related adverse effects, restrict the widespread use of AMB. Of par- ticular note is the significant clinical and economic impact of AMB-induced nephrotoxicity. The introduction of LBF-AMB has expanded the population that can safely receive the drug.

However, increased efficacy with LBF-AMB is less clear. Increases in LBF-AMB drug acquisition cost (relative to AMBd) may be offset by reductions in costs associated with AMB nephrotoxicity. Despite these advantages, AMBd may continue to play a role in select uses and patient populations, such as neonates and chil- dren, intrathecal use in patients with Coccidioides immitis men- ingitis or in patients otherwise at low risk of AMB nephrotoxicity or in azole/echinocandin-resistant fungal strains.
Financial & competing interests disclosure
The author has no relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript. This includes employment, consultancies, honoraria, stock ownership or options, expert testimony, grants or patents received or pending, or royalties.
No writing assistance was utilized in the production of this manuscript.

Key issues

Over the past few decades, new formulations of amphotericin B were developed in an effort to decrease nephrotoxicity while maintaining or possibly improving efficacy. These new formulations include amphotericin B colloidal dispersion, liposomal amphotericin B and amphotericin B lipid complex.
Although penetration of amphotericin B into the CNS is thought to be minimal and concentrations in the cerebrospinal fluid are mostly undetectable, amphotericin B, especially liposomal amphotericin B, is effective in the treatment of fungal infections of the CNS.
Amphotericin B is highly active in vitro against most Candida spp. (including Candida albicans, Candida glabrata, Candida tropicalis and Candida parapsilosis), Cryptococcus spp., Malassezia spp., Saccharomyces spp., Fusarium spp., a variety of molds (including most Aspergillus spp.) and endemic mycoses (Coccidioides spp., Blastomyces spp., Histoplasma capsulatum and Paracoccidioides brasiliensis).
Amphotericin B is still an important, although not gold standard, therapeutic agent for many invasive fungal infections and the comparative agent for all newer antifungal agents.
Current expert guidelines about the treatment of invasive candidiasis limit the role of amphotericin B to severe and/or refractory infections, CNS infections and pregnant patients.
Liposomal amphotericin B is recommended by the Infectious Diseases Society of America for treatment of invasive aspergillosis as an alternative to voriconazole.
Amphotericin B has become the standard of care for the initial therapy of severe or disseminated cryptococcal disease and/or cryptococcal infections involving the CNS.
Amphotericin B is considered as the primary therapy for pulmonary, disseminated and CNS histoplasmosis and blastomycosis.

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