Echinocandins in antifungal pharmacotherapy

Abstract

Objectives

Echinocandins are the newest addition of the last decade to the antifungal armamentarium, which, owing to their unique mechanism of action, selectively target the fungal cells without affecting mammalian cells. Since the time of their introduction, they have come to occupy an important niche in the antifungal pharmacotherapy, due to their efficacy, safety, tolerability and favourable pharmacokinetic profiles. This review deals with the varying facets of echinocandins such as their chemistry, in-vitro and in-vivo evaluations, clinical utility and indications, pharmacokinetic and pharmacodynamic profiles, and pharmacoeconomic considerations.

Key findings

Clinical studies have demonstrated that the echinocandins – caspofungin, micafungin and anidulafungin – are equivalent, if not superior, to the mainstay antifungal therapies involving amphotericin B and fluconazole. Moreover, echinocandin regimen has been shown to be more cost-effective and economical. Hence, the echinocandins have found favour in the management of invasive systemic fungal infections.

Conclusions

The subtle differences in echinocandins with respect to their pharmacology, clinical therapy and the mechanisms of resistance are emerging at a rapid pace from the current pool of research which could potentially aid in extending their utility in the fungal infections of the eye, heart and nervous system.

antifungal therapy, aspergillosis, candidiasis, echinocandins

Introduction

The three echinocandins, caspofungin, micafungin and anidulafungin, were the first-in-class antifungals that were developed to selectively target the fungal cell wall. As the mammalian cells lack cell wall, the echinocandins do not elicit their activity on the mammalian cells.^[1] This target specificity led to reduced instances of side effects and adverse events for the echinocandin therapy in comparison with the therapy with other mainstay antifungal agents such as amphotericin B and the azole class of antifungal drugs. This propelled the quick adoption of the echinocandin antifungals in the management of various invasive fungal infections and they came to be one the front-line agents in the antifungal therapy.^[2] The review discusses the various attributes of echinocandins such as their pharmacology, pharmacodynamics, preclinical and clinical evaluations with a particular emphasis on their clinical indications and manifestations.

The echinocandins were discovered as fermentation products, from pneumocandins, that had antifungal activity. However, these pneumocandin analogues had poor physicochemical properties. Hence, robust processes were undertaken for the modification of the analogues, with properties superior than their parent compounds. These efforts lead to the emergence of newer echinocandin analogues with improved physicochemical properties, broad spectrum of activity, pharmacokinetic (PK) profiles and a better balance between their potency and safety profiles.^[1]  Table 1 provides a summary on the improvements in the physicochemical properties and the enhancements in the antifungal potency of the newer echinocandin analogues in comparison with the earlier analogues.^[1,3–9] The development of the newer echinocandin analogues was a significant advancement in the antifungal therapy, due to the emergence of resistant strains of the fungal organisms.^[2]

Table 1

Summary on the improvements in the physicochemical properties and antifungal potency of the newer echinocandin analogues in comparison with the earlier analogues

	Echinocandin analogues	Molecular formula	Molecular weight (g/mol)	Aqueous solubility (mg/ml)	Log P	Potency (μg/ml)a
Early analogues	Echinocandin B	C52H81N7O16	1060.40	<0.1 (0.008)	5.4	0.01b >128c
	Cilofungin	C49H71N7O17	1030.14	<0.1	4.6	>3.2b NAc
	Pneumocandin Ao	C51H82N8O17	1079.26	<0.1	−0.5	0.06–8.0b >128c
	Pneumocandin Bo	C50H80N8O17	1065.23	<0.1	−1.1	0.25–4.00b >128c
Newer FDA-approved analogues	Caspofungin	C56H96N10O19	1213.44	28	−3.9	0.007–2.00b 0.25–>16c
	Micafungin	C56H70N9NaO23S	1292.27	>200	−1.5	0.004–2.00b 0.03–0.50 c
	Anidulafungin	C58H73N7O17	1140.25	0.05	2.9	0.06–2.00b 0.03–0.50c

NA, no activity. ^aPotency expressed as minimum inhibitory concentration (MIC). ^bMIC for Candida spp. ^cMIC for Aspergillus spp.

Amphotericin B and the azole antifungals – fluconazole, itraconazole, voriconazole and posaconazole – have been the mainstay chemotherapeutic agents for the fungal infections.^[10,11] However, the chemotherapy with these antifungal agents is associated with specific limitations such as nephrotoxicity, red blood cell (RBC) toxicity and arrhythmias for amphotericin B; hepatotoxicity associated with fluconazole; cardiotoxicity and gastrointestinal (GIT) disturbances attributed to itraconazole; voriconazole exhibiting neurological and hepatic toxicities; and posaconazole shown to elevate serum aminotransferase levels and cause mild-to-moderate hepatic toxicity.^[12–19] Development of cross-resistance has also been attributed to the use of these antifungal agents, due to their common site of action. Amphotericin B acts by binding to ergosterol, a component of fungal cell membrane, and permeabilizing it, causing cell leakage and subsequently eliciting fungicidal action.^[20] The azole antifungals act by inhibiting 14α-demethylase enzyme, which is essential for the synthesis of ergosterol.^[21] As azoles and amphotericin B share a common target – ergosterol – cases of emergence of cross-resistance among the azoles and between the azoles and amphotericin B have been reported.^[22,23] Moreover, amphotericin B is associated with non-selective mode of action and the azoles interact with cytochrome P-450 leading to significant drug–drug interactions (DDI).

Advent of the echinocandins in clinical practice has circumvented some of the aforementioned challenges in fungal therapy. All the echinocandins have a high antifungal activity, fewer DDI and lesser susceptibility to resistance in comparison with the other antifungals.^[24,25] Caspofungin was the first echinocandin to be approved by the US FDA and is commercially available as caspofungin acetate (Cancidas®; Merck, New Jersey, USA). Following the approval of caspofungin, micafungin (Mycamine®; Fujisawa Healthcare, Osaka, Japan) and anidulafungin (Eraxis®; Pfizer, New York, USA), were approved in 2005 and 2006, respectively.^[26,27] Their use has been indicated in the clinical treatment of invasive candidiasis and invasive aspergillosis.^[28–31]

Developmental history of echinocandins

Echinocandin B (Figure 1a) was one of the first lead compounds to be discovered at Sandoz (Novartis, New Jersey, USA) belonging to the class of echinocandins.^[32] Despite echinocandin B possessing antifungal activity, its potential application as a chemotherapeutic agent was dropped owing to its haemolytic action on the RBC.^[1] Consequently, cilofungin, a semisynthetic analogue of echinocandin B, was synthesized with significantly less haemolytic activity while retaining its antifungal activity. However, the clinical trials were halted due to the toxicity associated with its intravenous (IV) co-solvent system containing 26% polyethylene glycol.^[33,34] Subsequently, echinocandin congeners belonging to the pneumocandin class, such as pneumocandin A_o (Figure 1b) and pneumocandin B_o (Figure 1c), were synthesized and investigated. Although pneumocandin A_o retained the antifungal activity, it exhibited poor physicochemical properties and a narrow antifungal spectrum.^[1] Pneumocandin B_o was thus chosen as the starting compound for the synthesis of the first echinocandin antifungal agent – caspofungin acetate. Pneumocandin B_o was preferred over the other natural echinocandin analogues due to its superior potency and spectrum of antifungal activity.^[1,35] Pneumocandin B_o was repeatedly optimized for improving its potency, antifungal spectrum and commercial yield for the production of clinical quantities of caspofungin acetate. Caspofungin acetate was first synthesized in the year 1992 (Merck) and was then approved for clinical trials. Cancidas® was approved for invasive aspergillosis by the US FDA and later approved for oesophageal and invasive candidiasis, and finally as empirical therapy in fungal infections.^[36,37]

Figure 1

(a) Echinocandin B, (b) pneumocandin A_o and (c) pneumocandin B_o.

The parent compound for micafungin, FR901379, was developed at Fujisawa Pharmaceuticals in Japan. It was structurally similar to pneumocandin A_o, with the presence of a modified echinocandin B nucleus to incorporate a hydroxy-glutamine residue to improve its aqueous solubility. FR901379 was structurally optimized to incorporate a sulfate moiety, to overcome the aqueous insolubility of echinocandins. However, FR901379 was associated with reticulocyte lysis, and hence, further development of the lead compound was undertaken. Two important intermediates – FR179642 and FR131535 – were synthesized during the optimization process by bringing about modifications at the acyl side chain. These intermediates showed reduced lytic activity and enhanced anti-Candida and Aspergillus activity.^[9] The intermediates were further optimized to yield the final compound FK463 with isoxazole ring in the side chain, with no lytic activity and potent Candida and Aspergillus activity in vitro, and named micafungin.^[9]

Anidulafungin was modelled along cilofungin, by optimizing its chemical structure.^[38] The echinocandin B nucleus was structurally modified by deacylating it using Actinoplanes utahensis, to introduce a reactive amino group at the acyl side chain.^[39] This amino group was then treated with terphenyl acid to introduce the alkoxytriphenyl group on the echinocandin B nucleus at the reactive amino group.^[40] This introduction led to an enhanced antifungal potency with reduced RBC lytic activity of the newly synthesized echinocandin B nucleus with alkoxytriphenyl side chain. This echinocandin antifungal compound was then named as anidulafungin and approved for antifungal therapy in 2006.

Chemistry

Echinocandin B comprising a cyclic lipopeptide core (nucleus) and a N-linked acyl fatty acid chain is the most important structural feature of the echinocandin class of antifungals. The hexacyclic lipopeptide core is composed of different amino acid residues such as 3, 4-dihydroxy-ornithine, 3-methyl-4-hydroxy-proline, 3,4-dihydroxy-homotyrosine and 3-hydroxy-proline, and two threonine residues (Figure 1a).^[1,41] These amino acid residues are vital to the antifungal activity and determine the physicochemical properties of the echinocandin B nucleus and its congeners. The homotyrosine amino acid residue is essential for the antifungal activity and for the inhibition of glucan synthase enzyme. The proline residues are vital for enhancement of the antifungal potency of the echinocandin drugs. The hydroxyl groups at the three amino acids residues that make up the core nucleus, do not contribute to the antifungal activity, but improve the stability and increase the water solubility.^[41] Hence, in developing the newer echinocandin B antifungal congeners, the core nucleus was retained without any modification or was slightly modified (caspofungin).

The linoleoyl fatty acid chain linked to the echinocandin B nucleus is crucial to the antifungal activity as it acts as an anchor for the drug at the fungal cell wall.^[42] It, however, was also responsible for its haemolytic activity. Thus, echinocandin B was structurally modified, primarily at the N-linked acyl fatty side chain, to improve the safety of the subsequent echinocandin drugs.

Pneumocandin A_o (Figure 1b) and pneumocandin B_o (Figure 1c) were the outcome of the structural modification of echinocandin B. Both these structures were characterized by modifying the nucleus to incorporate a 3-hydroxy-glutamine instead of a threonine residue and by replacing the linoleoyl side chain by a dimethylmyristoyl side chain, to reduce their susceptibility for RBC haemolysis. Additionally, pneumocandin B_o nucleus was devoid of the methyl group at the 4-hydroxy-proline residue that improved its aqueous solubility in comparison with pneumocandin A_o. All the commercially available echinocandin drugs contain either pneumocandin A_o or pneumocandin B_o, instead of echinocandin B, as their core structure that is responsible for their antifungal potency.

Caspofungin (Figure 2a) is a semisynthetic water-soluble analogue belonging to the echinocandin family derived from pneumocandin B_o. The presence of pneumocandin B_o nucleus in caspofungin overcomes the haemolytic activity that is associated with its natural parent compound – echinocandin B.^[43] The ethylenediamine substitution has little significance to the potency of caspofungin, but is required for improving its aqueous solubility.^[1,44]

Figure 2

Structures of the three echinocandins. (a) Caspofungin (with hydroxy-proline and ethylenediamine highlighted), (b) micafungin (with sulfate and diphenyl-substituted isoxazole ring side chain highlighted) and (c) anidulafungin (with pentoxytriphenyl side chain highlighted).

Micafungin (micafungin sodium; Figure 2b), like caspofungin, is a cyclic lipopeptide. Micafungin consists of pneumocandin A_o as its lipopeptide core, albeit with marked difference in the fatty acid side chain. The side chain comprises a 3,5-diphenyl-substituted isoxazole ring. This side chain leads to an appreciable reduction in the haemolytic activity associated with micafungin while maintaining its antifungal potency. This antifungal potency is observed against both Candida and Aspergillus species, in vitro and in vivo. The presence of sulfated tyrosine residue is essential to the high aqueous solubility of micafungin.^[9]

Anidulafungin (Figure 2c), like the other two echinocandins, consists of a lipopeptide nucleus composed of the amino acid residues present in echinocandin B nucleus. This core is essential for the antifungal activity, and the presence of hydroxyl groups and amino linkers is essential for the aqueous solubility of the drug. Anidulafungin consists of the alkoxytriphenyl side chain, which is instrumental for the intercalation of the drug in fungal cell wall.^[45] This side chain, however, is markedly lipophilic in comparison with the side chains of the other two echinocandins; as a result, the aqueous solubility of anidulafungin is lower than that of them.^[38]  Table 2 summarizes the above-mentioned structure activity relationships for caspofungin, micafungin and anidulafungin.

Table 2

Summary of the structure activity relationship for the three echinocandins

	Structure	Activity
Caspofungin, micafungin and anidulafungin (common to all)	Homotyrosine amino acid residue	Essential for the antifungal activity and for the inhibition of glucan synthase enzyme
	Proline residues	Enhancement of the antifungal potency of the echinocandin drugs
	The hydroxyl groups in the echinocandin B nucleus	Improve the stability and increase the water solubility
Caspofungin (specific)	Dimethylmyristoyl side chain	Reduces the susceptibility for RBC haemolysis and required for the intercalation the drug in fungal cell wall
	Presence of 4-hydroxy-proline residue	Improves aqueous solubility
	Ethylenediamine substitution	Improves aqueous solubility
Micafungin (specific)	3,5-Diphenyl-substituted isoxazole ring side chain	Reduces the haemolytic activity and retains the antifungal potency
	Presence of sulfated tyrosine residue	Improves aqueous solubility
Anidulafungin (specific)	Octyloxytriphenyl side chain	Required for the intercalation the drug in fungal cell wall

Mechanism of action

β-(1,3)-d-glucan synthase is a heteromeric glycosyltransferase enzyme complex present in the fungal cell membrane. It is composed of a catalytic Fks p subunit and a regulatory subunit belonging to the Rho GTPase family.^[46] Intracellularly the catalytic and regulatory subunits are bound to UDP-glucose and GTP, respectively. The complex in entirety polymerizes UDP-glucose to β-(1,3)-d-glucan, which is released extracellularly to be incorporated into the fungal cell wall. β-(1,3)-d-glucan constitutes about 30–60% of the fungal cell wall and is a vital component in maintaining the integrity and strength of the fungal cell wall.^[47–49]

Echinocandins elicit their antifungal activity by non-competitively binding to the Fks p subunit of the enzyme and blocking the β-(1,3)-d-glucan synthesis (Figure 3). This disruption in the β-(1,3)-d-glucan synthesis results in a leaky and a highly permeable cell wall that leads to an imbalance in the intracellular osmotic pressure of the fungal species. This results in the fungal cell lysis demonstrating their fungicidal action.^[50] This fungicidal effect is observed for all the three echinocandins on the fungal species belonging to Candida and Saccharomyces species.^[24,51–53] The three echinocandins, however, demonstrate a fungistatic effect on the Aspergillus species.^[30,54,55] This difference in the action is attributed to the variable glucan content in the different fungal species. The fungistatic action is elicited by morphological changes in the fungus, usually at the fungal hyphae and cell wall, which was previously evidenced in Aspergillus fumigatus.^[4]

Figure 3

Cartoon illustrating the binding of echinocandins on the glucan synthase enzyme complex. Echinocandins non-competitively inhibit the activity of glucan synthase and disrupts the β-(1,3)-d-glucan synthesis leading to fungal cell lysis.

Experimental studies

Several studies have been performed, both in vitro and in vivo, for evaluating the antifungal spectrum and potency for echinocandins.

Studies in vitro

In-vitro studies have been performed for assessing the antifungal spectrum of echinocandins. The susceptibility of the fungal species to echinocandins increases along with an increase in their concentration. The echinocandin antifungal activity is evaluated in terms of minimum inhibitory concentration (MIC) required to bring about a specific percentage reduction (usually 50, 80 or 90%) in the fungal load. The MIC values are determined by the E-test (Epsilometer test) and Clinical and Laboratory Standards Institute (CLSI) reference methods; however, these methods are not standardized methods in evaluating echinocandin antifungal activity and hence should be carefully construed.^[25,56]

As a class, the echinocandins show excellent antifungal activity against Candida species, including those that are less susceptible or resistant to other classes of antifungals such as amphotericin B and azoles.^[51,57–63] The activity of caspofungin was evaluated against 8197 Candida isolates by Pfaller et al.^[64] over a span of 4 years, and the MIC was found to be in the range of 0.007–2.00 μg/ml. This MIC range was in close agreement with a previous clinical study by Kartsonis et al.,^[65] in which the MIC range was determined to be 0.008–4 μg/ml, with approximately 95% of the MIC values in the range of 0.25–2.00 μg/ml. Caspofungin demonstrates excellent antifungal activity against most of the clinical Candida isolates such as C. albicans, C. glabrata, C. tropicalis, C. kefyr, C. pelliculosa, C. parapsilosis, C. krusei, C. guilliermondii and C. lusitaniae.^[64,66,67] However, certain Candida strains such as C. albicans, C. glabrata, C. tropicalis and C. krusei have shown reduced susceptibility accompanied with an increase in the MIC values.^[68–72] These incidences are attributed to the prolonged use of caspofungin. Micafungin, just like caspofungin, has shown potent antifungal activity against the aforementioned Candida species, with MIC values ranging from 0.004 to 2.00 μg/ml.^[30,73–75] In a clinical study by Ikeda et al.,^[76] micafungin has shown lower MIC value ranges for C. albicans, C. tropicalis and C. glabrata in comparison with caspofungin. Anidulafungin exhibits a potent action against C. albicans, C. glabrata, C. tropicalis, C. krusei and C. kefyr with low MIC range from 0.06 to 0.12 μg/ml, while C. parapsilosis, C. lusitaniae and C. guilliermondii isolates show a comparatively higher MIC range from 0.5 to 2.00 μg/ml.^[77,78] Anidulafungin also demonstrates biological activity against Candida biofilms, in vitro.^[79,80]  C. glabrata and C. parapsilosis isolates have shown clinical micafungin resistance with reduced susceptibility, in vitro.^[30,81–84] On the other hand, lowered susceptibility or resistance to anidulafungin is rare and has been reported only for C. glabrata and C. albicans, with the lowered susceptibility for the latter being evidenced in South Africa in <1% of the Candida fungal cases.^[83–85]

Caspofungin primarily exhibits fungistatic action on the filamentous fungi such as the Aspergillus species. Caspofungin has a modest fungistatic action on the Aspergillus isolates such as A. fumigatus, A. flavus, A. niger, A. terreus and A. nidulans with MIC range from 0.25 to >16 μg/ml.^[86–89] However, it has been proven in vitro that co-administration of albumin and fluvastatin increases the caspofungin potency; albumin acts as a carrier molecule for caspofungin at the germinating hyphae of the filamentous fungi, whereas fluvastatin acts to inhibit the sterol synthesis, thereby providing a synergy to caspofungin.^[90,91] Micafungin and anidulafungin also demonstrate a fungistatic action on the aforementioned Aspergillus species with MIC ranging from 0.03 to 0.50 μg/ml for both the drugs, for all the Aspergillus species except A. flavus which shows MIC >16 μg/ml.^{[76,89,92–98]} As the Aspergillus species have lower glucan content than the Candida species, the potency of echinocandins on Aspergillus is not as pronounced as is seen in Candida species. Only one case of clinical Aspergillus isolate (A. fumigatus) resistant against caspofungin has been reported.^[99] However, a clinically resistant A. fumigatus with reduced susceptibility was engineered by Gardiner et al.^[100] Ironically, in comparison with the documentation of cases of echinocandin resistance in Candida, the reports for resistance in Aspergillus species remains scanty.

The echinocandins also exhibit biological activity against a host of other fungal species (Table 3). Caspofungin has exhibited antifungal activity against Exophiala jeanselmei, Fonsecaea pedrosoi, Paecilomyces variotii and Scedosporium apiospermum.^[101,102] However, it shows moderate activity against Penicillium, Rhizopus, Fusarium and Mucor species.^[103] Micafungin is active against dimorphic fungi; with greater antifungal activity against the mycelial form (MF) than the yeast form (YF).^[104] Micafungin is inactive against Cryptococcus, Fusarium and Zygomycetes species.^[104] Anidulafungin demonstrates antifungal activity against a plethora of fungal species such as B. spicifera, E. jeanselmei, F. pedrosoi, M. mycetomatis, P. marneffei, P. verrucosa, P. boydii, S. schenckii and W. dermatitidis.^[101] However, anidulafungin shows weak activity against Acremonium, Rhizopus and Scedosporium species.^[101] The echinocandins do not exhibit antifungal action against C. neoformans, which has been attributed to the presence of a capsule around the organism, which prevents the penetration of the drugs for their biological activity.^[105]

Table 3

Antifungal activity of the echinocandins against fungal species other than Candida and Aspergillus species with their MIC values

Echinocandin	Fungal species	MIC (μg/ml)
Caspofungin[102–104]	Exophiala jeanselmei	0.39–3.12
	Fonsecaea pedrosoi	0.09–0.19
	Paecilomyces variotii	≤0.09
	Scedosporium apiospermum	0.19–0.78
	Penicillium spp.	0.015–8
	Rhizopus spp., Fusarium spp., Mucor spp.a	>8
Micafungin[105]	Histoplasma capsulatum	0.0078–0.0625b; 32–>64c
	Paracoccidioides brasiliensis	4–16b; >64c
	Penicillium marneffei	0.0313–2b; 4–16c
	Sporothrix schenckii	0.5–1b; 16–>64c
	Coccidioides immitis	0.0156b; NAc
Anidulafungin[102]	Bipolaris spicifera	0.5–8
	Exophiala jeanselmei	0.125–2
	Fonsecaea pedrosoi	0.5–2
	Madurella mycetomatis	1
	Penicillium marneffei	0.5–2
	Pocillopora verrucosa	0.03–0.25
	Pseudallescheria boydii	2–8
	Sporothrix schenckii	2–8
	Wangiella dermatitidis	0.5–8
	Acremonium spp., Rhizopus spp., Scedosporium spp.d	>16

^aModerate antifungal activity. ^bMIC value against the mycelial form (MF) of fungi. ^cMIC value against the yeast form (YF) of fungi. ^dWeak antifungal activity.

Studies in vivo

The in-vitro antifungal activity of the echinocandins is also manifested in the in-vivo studies. In a murine model, both caspofungin and anidulafungin were found to reduce the fungal load of C. glabrata in the kidneys.^[106] Caspofungin has also been documented to reduce the C. glabrata fungal load in severe neutropenic murine models.^[107] It has exhibited potent in-vivo activity in conjunction with diclofenac sodium against C. albicans biofilms in rat model.^[108] In a study by Spreghini et al., all the three echinocandins were tested against Candida infection induced in murine model due to C. parapsilosis, C. orthopsilosis and C. metapsilosis. Both caspofungin and anidulafungin exhibited fungicidal activity against all the three strains, whereas micafungin was active only against C. metapsilosis, in vivo.^[109] The antifungal efficacy of the three echinocandins has been proven in neutropenic murine models infected with C. albicans, C. glabrata and C. parapsilosis.^[110] Micafungin has exhibited excellent in-vivo biological activity against C. tropicalis-infected murine model, where micafungin treatment was found to be more effective than amphotericin B therapy, and micafungin also elicited activity against fluconazole-resistant murine model.^[111] Micafungin efficacy has also been demonstrated in vivo against C. tropicalis, C. glabrata and C. albicans, and azole-resistant C. albicans murine models.^[112–115] Anidulafungin exhibited antifungal activity in murine models infected with C. tropicalis and in a rat model against C. albicans biofilms.^[116,117]

Echinocandins also exhibit antifungal activity against various Aspergillus species, in vivo. Caspofungin has exhibited dose-dependent efficacy against invasive aspergillosis in mice which was induced by A. fumigatus and A. niger composite strains, and also against A. fumigatus-induced infection in neutropenic mouse model.^[118–120] Caspofungin has demonstrated in-vivo antifungal activity against A. flavus and A. terreus.^[87,121] Studies have shown that posaconazole and albumin potentiate the fungistatic activity of caspofungin against Aspergillus species.^[90,121] Micafungin has exhibited antifungal potency against pulmonary aspergillosis in mouse model and in-vivo activity against amphotericin B-resistant A. terreus and itraconazole-resistant A. fumigatus in neutropenic mouse model.^[122–127] Anidulafungin has exhibited efficacy in A. niger-, A. fumigatus- and A. flavus-induced aspergillosis in murine models.^{[94,119,128,129]}

Micafungin has been found to be active against P. carinii and H. capsulatum in vivo, and anidulafungin has shown in-vivo potency against P. carinii-infected rat and murine models.^[130–132] There is a lack of in-vivo data on the susceptibility of other fungal species. In general, the echinocandins show a good degree of agreement between the in-vitro and in-vivo antifungal activity for Candida and Aspergillus fungal infections.

Paradoxical effect of echinocandins

The paradoxical effect, also referred as the eagle-like effect, is a characteristic effect shown by the echinocandins, especially by caspofungin on the Candida species, in vitro.^[133–136] This effect is associated with reduced growth at MIC, but at higher concentration (known as supra-MIC) the fungal growth continues, and at concentrations higher than supra-MIC, a reduction in fungal growth is again observed. This paradoxical effect has been well illustrated in the study by Chamilos et al., in which five different Candida isolates were utilized (C. albicans, C. krusei, C. parapsilosis, C. tropicalis and C. glabrata) along with three echinocandins. It was found that the paradoxical effect for caspofungin was the highest in comparison with the other two echinocandins and was observed in C. parapsilosis, C. albicans, C. tropicalis and C. krusei isolates. For micafungin, the paradoxical effect was observed only in C. tropicalis and C. krusei, while anidulafungin showed paradoxical growth only in C. albicans and C. tropicalis isolates. However, the paradoxical effect was absent in C. glabrata isolates for all the three echinocandins.^[133] A similar paradoxical effect was observed on C. dubliniensis isolates for all the three echinocandins; these differences in the paradoxical effects observed among different Candida species could be attributed to the differences in the genetic makeup of the varying Candida species.^[137] This paradoxical effect has also been demonstrated in A. fumigatus, A. flavus and A. terreus above their caspofungin MIC and was evidenced by an increase in their metabolic activity.^[138] The emergence and development of this paradoxical effect could be attributed to the activation of stress response pathways such as the calcineurin or chitin pathways to maintain the cell wall integrity.^{[136,139–142]} However, there are scanty and inconsistent reports on the clinical and in-vivo observation of this paradoxical effect, necessitating a further investigation of this phenomenon.^[143]

Resistance to echinocandins

Multidrug-resistant (MDR) transporters are an important reason behind the development of antifungal resistance. The clinically used azoles exhibit resistance mediated by MDR transporters in Candida species, as they are biological substrates for the MDR transporters.^[144–146] Echinocandins, however, are not the substrates for MDR transporters, and hence, the emergence of MDR transporter facilitated resistance is clinically rare for echinocandins in Candida species.^[51,114,147]

The manifestation of resistance to echinocandins has been studied extensively in the past decade and has been attributed to the mutation of the Fks subunits in the glucan synthase enzyme.^[71,148,149] In most of the Candida species, apart from C. glabrata, the development of resistance is due to the mutation in the Fks 1p subunit, whereas in the case of C. glabrata it is ascribed to both the Fks 1p and Fks 2p subunit genetic mutations.^[150–154] The mutations largely occur in the Fks 1p locus with substitutions of serine 645 for other amino acids such as proline, phenylalanine and tyrosine, and in Fks 2p, the mutations occur at serine and phenylalanine residues.^{[71,148,155,156]} These amino acid substitutions lead to the emergence of resistance and cross-resistance among the echinocandins.^[157] The Fks 3 subunit does not contribute to the development of echinocandin resistance.^[71,158,159] These mutations in the Fks 1p and Fks 2p loci lead to the reduced susceptibility of the Candida species to the echinocandins, with an increase in the MIC, and reduced glucan synthase enzymatic activity.^[148] In Aspergillus species, the mechanism of resistance was studied for caspofungin in A. fumigatus. The occurrence of amino acid mutation in the AfFKS1 gene (responsible for glucan synthesis in A. fumigatus) coupled with an escalation in chitin production for maintaining their cell wall, has been attributed to the development of resistance in Aspergillus species, particularly A. fumigatus.^[160,161] The development of resistance to echinocandins in other fungal species is still unknown.

In the past decade, cases of clinical emergence of echinocandin resistance have been on the rise steadily for numerous Candida species, especially C. albicans, C. krusei, C. glabrata and C. tropicalis.^{[160,162–166]} The cases for resistance in Aspergillus and other fungal species, however, remain scanty as susceptibility testing is less frequently performed due to the lack of standardized techniques for evaluation.^[167]

Clinical trials

Caspofungin

There have been many clinical trials that have been undertaken to evaluate the clinical efficacy of caspofungin. A multicentre, double-blind, randomized trial was conducted by Villanueva et al. to assess the efficacy, safety and tolerability of caspofungin in comparison with amphotericin B in clinical cases of Candida oesophagitis. It was found that 50 mg/day and 70 mg/day doses of caspofungin showed clinical success rates of 74 and 89%, respectively, in comparison with amphotericin B (0.5 mg/kg per day) dose which showed 63% clinical success. Also, the amphotericin B chemotherapy was discontinued in 24% of the patients due to drug-related adverse effects, whereas in the caspofungin group (50 and 70 mg/day) it was 4 and 7%, respectively. This report provided the first demonstration of clinical utility for caspofungin in 2001.^[168] Villanueva et al. also conducted another randomized double-blinded study that compared caspofungin (50 mg) and fluconazole (200 mg) therapeutic regimen for oesophageal candidiasis, in HIV-infected patients. In this study, it was found that the success rate for caspofungin and fluconazole was 81 and 85%, respectively, with no adverse drug–drug interactions reported in the caspofungin group and one reported in the fluconazole group. The extent of resolution of the clinical symptoms was similar for both the groups; however, higher relapse was reported in the caspofungin group compared to the fluconazole group, with no statistically significant difference between the groups. The study concluded that caspofungin appeared to be as efficacious and generally as well tolerated as fluconazole in HIV-infected patients with Candida oesophagitis.^[169] The clinical efficacy of caspofungin in Candida oesophagitis was further corroborated by Arathoon et al.^[170] in his randomized, double-blind, multicentre study of caspofungin vs amphotericin B in the treatment of oesophagitis in HIV-infected patients. The efficacy of caspofungin was found to be similar to the efficacy of amphotericin B in the treatment of invasive candidiasis and invasive aspergillosis, with infrequent reports of drug-related nephrotoxicity and hepatotoxicity in the caspofungin group.^[171,172] A clinical trial was initiated for evaluating the safety of caspofungin, in which 1205 patients were given daily doses of caspofungin (35–100 mg), and it was found that caspofungin had lower cases of drug-related adverse effects and lower discontinuations in the therapy, with no dose-related toxicity in comparison with amphotericin B deoxycholate, liposomal AmB and fluconazole.^[173] In a clinical report by Aoun et al.,^[174] caspofungin showed a good response rate with overall better safety profile than liposomal amphotericin B formulation in the therapy of invasive aspergillosis in neutropenic patients.

Robust clinical trials were also undertaken to evaluate the safety and efficacy of caspofungin therapy. A multicentre and double-blind trial for a high-dose caspofungin (150 mg/day) and a standard-dose caspofungin (50 and 70 mg/day) against invasive candidiasis was conducted by Betts et al. in 2009. A total of 204 patients were included in the study with 104 patients receiving the standard dose and 100 receiving the high dose. Significant drug-related adverse events occurred in 1.9% of patients receiving the standard dose and 3.0% of patients receiving the high dose; favourable response of 71.6 and 77.9% in patients with the standard dose and high dose, respectively, was observed, and it was concluded that caspofungin was safe for the chemotherapy of invasive candidiasis, even at higher doses (150 mg/day).^[175] The safety and PK profile of caspofungin, liposomal amphotericin B and a combination of both were investigated in a risk-stratified, randomized, multicentre phase II clinical trial in patients with allogeneic haematopoietic stem cell with granulocytopenia and refractory fever. The patients received caspofungin (50 and 70 mg/day), liposomal amphotericin B (3 mg/kg per day) or the combination of both. Safety and relapse potential were assessed 14 days after the end of therapy and PK analyses were conducted on the 1st and 4th day. All three regimens were well tolerated. Discontinuation of therapy was observed in individual caspofungin and individual amphotericin B group only. A similarity was observed for the adverse drug effects in all the three groups which did not lead to the discontinuation of the therapy. The combination therapy in immunocompromised patients was concluded to be similar in efficacy and PK profile as the monotherapy with either of the drugs.^[176] Caspofungin has also shown clinical efficacy in invasive candidiasis caused due to non-C. albicans species, such as C. tropicalis, C. parapsilosis, C. glabrata, C. krusei, C. guilliermondii and C. lusitaniae, with favourable safety profile and without any discontinuation in therapy due to drug-related adverse effects.^[177] In the clinical study by Maertens et al., caspofungin was found to be efficacious and well tolerated in patients with invasive aspergillosis (caused by A. fumigatus) accompanied by active malignancies, neutropenia, allogeneic haematopoietic stem cell transplantation, HIV infection and organ transplantation.^[178] The efficacy of caspofungin has also been reported in Germany from a multinational multicentre study to prospectively assess the outcome of caspofungin therapy in proven or probable cases of invasive aspergillosis.^[179]

The safety and efficacy profile of caspofungin (50 mg) and micafungin (150 mg) against Candida and Aspergillus infections were directly compared in the prospective, randomized, double-blind study conducted by Kohno et al. The proportion of patients who developed significant drug-related adverse events was 5% in the caspofungin group and 10% in the micafungin group. The response rate in the caspofungin group was 100, 100 and 46.7% in patients with oesophageal candidiasis, invasive candidiasis and chronic pulmonary aspergillosis including aspergilloma, whereas it was 83.3, 100 and 42.4% in the micafungin group, respectively. In this clinical study, it was concluded that there was no statistical difference in the safety and clinical outcomes between caspofungin and micafungin and both were equally efficacious.^[180]

The antifungal potency and efficacy of caspofungin was also evaluated in the paediatric population. To evaluate the efficacy and safety of caspofungin in immunocompromised patients, a multicentre retrospective study was conducted on 64 immunocompromised paediatric patients. No discontinuation in therapy was observed owing to the drug-related adverse effects and the clinical side effects were mild to moderate and observed in 53.1% of the patients. The survival was 75% at the end of caspofungin chemotherapy and 70% at 3 months post-therapy, respectively. This clinical trial evidenced that caspofungin displayed favourable safety and efficacy in immunocompromised paediatric patients.^[181] Caspofungin has also shown antifungal activity against azole-resistant candidaemia in a neonatal case, in which a dose of 3 mg/kg was found to be the most effective with no possible side effects.^[182] Caspofungin has also demonstrated clinical effectiveness in treating refractory candidaemia caused due to C. albicans, C. parapsilosis and C. tropicalis in 12 neonatal cases, which was resistant to other antifungal therapy.^[183] In another cohort study, caspofungin and liposomal amphotericin B have shown similar efficacy as antifungal prophylactic agents in paediatric patients undergoing allogeneic haematopoietic stem cell transplantation.^[184] Two independent studies on paediatric and adolescent patients proved that the caspofungin therapy was effective in treating fungal infections due to Candida and Aspergillus, with no discontinuation in therapy due to adverse side effects.^[185,186] In another study, caspofungin has shown to be effective against C. parapsilosis biofilms in paediatric patients.^[187]

In a clinical study by Mattiuzzi et al., the prophylactic potential of caspofungin was compared against IV itraconazole in patients suffering from acute myeloid leukaemia. Two hundred patients were evaluated for the prophylactic activity in the said open-label, randomized study.^[188] About 51 and 52% patients completed the therapy in the itraconazole and caspofungin group, respectively. The contraction of fungal infections was higher in the caspofungin group (58%) vs the itraconazole (42%) group. The incidence of deaths in the caspofungin group was twice as high as seen in the itraconazole group. Additionally, itraconazole did not exhibit any cardiotoxicity. Thus, the prophylactic activity of caspofungin was found to not be superlative over the itraconazole therapy. Additionally, itraconazole can also be taken orally and has activity against other species such as Trichosporon, giving it a slight edge in clinical therapy over the caspofungin prophylactic therapy.

All the above-mentioned clinical trials are summarized in Table 4. These trials have primarily elucidated the efficacy and safety of caspofungin for the treatment of fungal infections, both in adults and in paediatric populations, mainly for Candida and Aspergillus species. However, its evaluation as a prophylactic agent is warranted.

Table 4

Summary of the outcomes from clinical trials of caspofungin

Research group	Outcome
Villanueva et al.	Efficacy, safety and tolerability of caspofungin were statistically comparable to amphotericin B in cases of Candida oesophagitis
Villanueva et al.	Efficacy and tolerability of caspofungin was statistically comparable to fluconazole in cases of oesophageal candidiasis in HIV patients
Arathoon et al.	Efficacy of caspofungin was similar to amphotericin B in invasive candidiasis and aspergillosis in HIV patients
Betts et al.	Caspofungin at higher dose (150 mg/day) was safe as clinical doses (50 and 70 mg/day) in the treatment of invasive candidiasis
Groll et al.	Clinical outcomes from caspofungin, liposomal amphotericin B and their combination therapy were found to be similar in patients with allogeneic haematopoietic stem cell granulocytopenia
Kartsonis et al.	Caspofungin was safe and efficacious in the treatment of non-C. albicans species
Maertens et al.	Caspofungin was efficacious and well tolerated in cases of invasive aspergillosis
Kohno et al.	No statistical difference in the safety and clinical outcomes between caspofungin and micafungin in aspergilloma
Groll et al., Benjamin et al., Bedard et al., Doring et al.	Caspofungin therapy is effective, safe and well tolerated in paediatric and adolescent patients suffering from invasive candidiasis and aspergillosis
Mattiuzzi et al.	Prophylactic activity of caspofungin was found to not be superlative to IV itraconazole

Micafungin

The clinical efficacy and safety of micafungin has been validated by various clinical trials in adult as well as paediatric population. The efficacy and safety of micafungin was investigated in patients with deep-seated mycosis in the study by Kohno et al. The overall clinical response rates were 60, 67, 55, 100 and 71% in invasive pulmonary aspergillosis, chronic necrotizing pulmonary aspergillosis, pulmonary aspergilloma, candidaemia and oesophageal candidiasis, respectively. Antifungal activity was observed against A. fumigatus, A. flavus, A. terreus, A. niger, C. albicans, C. glabrata and C. krusei. Drug-related adverse effects were reported in 30% of the patients and they were not dose-dependent. Thus, it is concluded micafungin therapy is effective and safe in patients with deep-seated mycosis.^[189] To evaluate micafungin in the treatment of newly diagnosed and refractory candidaemia, an international, open-label, non-comparative, clinical trial was undertaken in paediatric, neonatal and adult patients totalling to 126 patients. Overall success rate was 83.3% with individual rates being 85.1, 93.8, 86.4 and 83.3% for C. albicans, C. glabrata, C. parapsilosis and C. tropicalis, respectively, without any drug-related adverse effects.^[190] The safety of micafungin in paediatric patients was evaluated, and it was observed that adverse effects were recorded in 93.2% of patients, with only 4.7% of patients experiencing serious adverse effects due to micafungin. The study showed that micafungin was well tolerated by paediatric population.^[191]

A multicentre, randomized, open-label phase III study by Huang et al. compared micafungin (50 mg/day) efficacy and safety with itraconazole (5 mg/kg per day) against the prophylaxis of invasive fungal infections in 287 neutropenic patients undergoing haematopoietic stem cell transplants in China. The micafungin group exhibited a success rate of 92.6% and itraconazole group demonstrated 94.6% success rate, indicating the non-inferiority of micafungin chemotherapy against itraconazole regimen. The discontinuation of therapy was higher in the itraconazole (67.3%) than in the micafungin (82.9%) group, with discontinuation, because an adverse effect (4.4% vs 21.1%) and drug-related adverse effects (8% vs 26.5%) were shown between micafungin and itraconazole. Thus, it was concluded that micafungin was as effective as itraconazole and comparatively safer in preventing invasive fungal infections in patients with neutropenia.^[192] So also, it has been documented in a clinical study that micafungin and itraconazole combination therapy is clinically effective and safe in treating pulmonary aspergilloma.^[193] Andes et al. compared the outcomes of two independent, double-blind multicentre, randomized clinical trials of micafungin, which were carried out using different dosing regimens (centre 1: 150 mg daily dose, centre 2: 300 mg every alternate day). No significant differences were found in (a) the clinical success rate between the centres (78.8% in centre 1 vs 87.1% in centre 2) and (b) the relapse rates between the centres (12.2% in centre 1 vs 5.6% in centre 2), indicating that micafungin at higher doses every alternate day could be a promising alternative to the conventional daily therapy.^[194] This study was further corroborated by a clinical study by Mehta et al., which, on PK analyses of a higher micafungin dose (3 mg/kg, with normal micafungin regimen being 1 mg/kg daily) every alternate day, showed safety and efficacy of this dosing regimen in paediatric patients undergoing haematopoietic stem cell transplantation.^[195] A prospective, multicentre, open-label study was conducted to establish the efficacy and safety of micafungin prophylactic therapy of invasive fungal infections during neutropenia in children and adolescents undergoing allogeneic haematopoietic stem cell transplantation. 82.3% of the patients completed the regimen, with a success rate in 90.2% patients, and 23.8% patients showed drug-related adverse effects. Only 1.4% discontinued the micafungin therapy due to adverse effects. The study concluded that micafungin therapy could be a promising clinical approach in preventing fungal infections during allogeneic haematopoietic stem cell transplantation.^[196]

The safety and efficacy of micafungin were evaluated in HIV-compromised patients suffering from oesophageal candidiasis. In the study by Pettengell et al.,^[197] dose-dependent activity of micafungin was evaluated for testing the efficacy and safety of micafungin. It was found that the minimum effective concentration for micafungin was 12.5 mg, and it showed a significant resolution of oesophageal candidiasis symptoms at 75 and 100 mg doses, with about 94.7% fungal resolution at the latter dose. In another randomized, double-blind, parallel-group clinical study, the efficacy of micafungin (100 and 150 mg/day) was concluded to be similar to the standard fluconazole therapy (200 mg/day), with equivalent success rates and outcomes.^[198] In yet another study, the IV micafungin (150 mg) therapy was found to be similar in efficacy with the standard IV fluconazole (200 mg) therapy in the management of oesophageal candidiasis.^[199]

To evaluate the prophylactic potential of micafungin against invasive fungal infections, a comparative study between micafungin and fluconazole was undertaken in patients with neutropenia and haematopoietic stem cell transplantation. A randomized, double-blind, multicentre, phase III trial was conducted which comprised 882 adult and paediatric patients.^[200] In this clinical trial, the success rate with micafungin was found to be higher than the fluconazole treatment in a 4-week efficacy study (80.0% vs 73.5%).

A systematic review of the clinical outcomes of all 17 micafungin trials was carried out by Cornely et al. and Ullman et al., and it was found that micafungin had a good safety profile, with lower drug-related adverse events, and an efficacy similar to other potent antifungal agents such as caspofungin or fluconazole in treating invasive fungal infections.^[201,202]  Table 5 summarizes the key aspects of the micafungin clinical trials in establishing its efficacy and safety as an antifungal agent.

Table 5

Summary of the outcomes from clinical trials of micafungin

Research group	Outcome
Kohno et al.	Efficacy and safety of micafungin in systemic candidiasis and aspergillosis
Ostrosky-Zeichner et al.	Efficacy and safety of micafungin in refractory candidaemia in neonatal, paediatric and adult patients
Groll et al.	Micafungin is safe and well tolerated in paediatric population
Huang et al.	Non-inferiority of micafungin to itraconazole as a prophylactic agent
Andes et al.	Higher dose of micafungin (300 mg/alternate day) is equally efficacious as daily conventional dose of (150 mg/day)
Mehta et al.	Efficacy of micafungin in paediatric patients when a higher dose (3 mg/kg) was given every alternate day in comparison with the conventional daily dose (1 mg/kg)
De Wet et al., Walsh et al.	Efficacy of micafungin was found to be similar to fluconazole in oesophageal candidiasis and also as a prophylactic agent

Anidulafungin

Two clinical studies were conducted by Krause et al. in 2004 to evaluate the efficacy and therapeutic potency of anidulafungin in Candida infections. The first trial was a randomized, double-blind study comparing IV anidulafungin and oral fluconazole in the treatment of oesophageal candidiasis. The overall success rate for anidulafungin therapy (97.2%) was found to be statistically non-inferior to the therapy success rate for fluconazole therapy (98.8%). Both the drugs exhibited similarity in the extent of adverse effects (anidulafungin: 9.3% and fluconazole: 12.0%). From the study, it was established that anidulafungin was similar in efficacy and safety to fluconazole in the treatment of oesophageal candidiasis.^[203] The second clinical study evaluated the safety and efficacy of anidulafungin by varying its dosing regimen in invasive Candida infections. Anidulafungin was given daily at varying doses (50, 75 and 100 mg). The efficacy of the drug for the varying doses was evaluated at the follow-up visits by the patients after the end of therapy. It was found that the overall success rates at the end of therapy were 84, 90 and 89% in the 50, 75 and 100 mg groups, respectively, while the success rates at the follow-up visits were 72, 85 and 83%. No dose-dependent side effects were observed, and 9.0% of the deaths were reported in complicated and comorbid patients which were thought to be either probably or possibly related to the drug-related adverse events.^[204] Both these studies validated the clinical efficacy and safety of anidulafungin in invasive fungal infections.

Reboli et al. conducted a clinical trial to compare the efficacy of IV anidulafungin and fluconazole in invasive candidiasis. The success rate was 75.6 and 60.2% in the anidulafungin and fluconazole group, respectively. A similarity was observed in the adverse events and death rates (anidulafungin: 23% and fluconazole: 31%) of the two groups. This study corroborated the earlier clinical study by Krause et al.,^[205] indicating the similarity in effectiveness of anidulafungin and fluconazole (mainstay for invasive fungal infections) in the treatment of invasive candidiasis. However, in a prospective, randomized clinical study following the Infectious Diseases Society of America (IDSA) guidelines showed that the response to anidulafungin therapy was better than the fluconazole therapy in critically ill patients.^[206] Dowell et al. undertook a study to evaluate the safety and PK profile of voriconazole and anidulafungin combination, and also for the drugs individually. It was found that dose-dependent toxicities did not occur, and all the reported adverse events were mild and within the known safety profiles of both drugs. PK parameters were not affected by co-administration, and the combination was found to be clinically safe and efficacious.^[207] The prophylactic effect of anidulafungin was compared with fluconazole in a randomized, double-blind study in liver transplant patients who were at a high risk of invasive fungal infections. Similarities were seen in the incidence rates for fungal infections, graft rejection, fungal-free survival and mortality in the two groups. However, anidulafungin prophylaxis was associated with less invasive Aspergillus infections. Thus, anidulafungin was evaluated to be an effective candidate in the chemotherapy of diseases associated with increased risk for Aspergillus infection.^[208]

In the year 2012, Ruhnke et al. performed a prospective, multicentre study to evaluate the efficacy and safety of anidulafungin in critically ill patients admitted in ICU (due to postabdominal surgery, solid tumour, renal/hepatic insufficiency, solid organ transplant, neutropenia and age ≥ 65 years) and also suffering from candidaemia and invasive candidiasis. The evaluation for the safety and efficacy of the anidulafungin regimen was carried out at the end of therapy, at the end of IV therapy and at 2 and 6 weeks after the end of therapy. The success rate was 69.5, 70.7, 60.2 and 50.5% at the end of therapy, at the end of IV therapy and at 2 and 6 weeks after the end of therapy, respectively. Anidulafungin related adverse effects accounted to 15.3%, with serious adverse events accounting to 1.9%. The study concluded with the premise that anidulafungin was found to be an effective treatment in critically ill patients suffering from invasive candidiasis.^[209]

The safety and PK evaluation of anidulafungin was carried out in paediatric population (between 2 and 17 years), in a multicentre, cohort study. Two anidulafungin doses (0.75 and 1.5 mg/kg per day) were received by the patients. It was found that both the doses were well tolerated with no serious adverse events, except in two cases, which resolved on slowing the anidulafungin infusion rate. The PK profile in children at both the doses was similar to an adult receiving 50 or 100 mg/day dose. As the drug was found to be well tolerated and safe in children, this could be considered as a potential therapeutic regimen in neutropenic paediatric population.^[210] This has been the only clinical study in paediatric patients for evaluating the anidulafungin therapy, and there is a need of more clinical evaluations for this drug in paediatric population.

Table 6 summarizes the aforementioned clinical outcomes. From it, it can be concluded that anidulafungin has shown safety and potency in clinical therapies, like the other two echinocandins, indicating its utility in invasive fungal infections. However, further studies exploring the relevance of anidulafungin in fungal treatments are warranted because, in a recent report by Krcmery et al., it was found that there was a development of C. glabrata fungal infection in haematology patients treated with anidulafungin. Hence, a critical evaluation of development of resistance or nature of relapse is necessary.^[211]

Table 6

Summary of the outcomes from clinical trials of anidulafungin

Research group	Outcome
Krause et al., Reboli et al.	Anidulafungin therapy was found to be statistically non-inferior to fluconazole therapy in oesophageal candidiasis
Dowell et al.	Anidulafungin and voriconazole combination therapy is safe and efficacious in systemic fungal infections
Winston et al.	Anidulafungin was found to be safe and efficacious as a prophylactic in Aspergillus infections
Ruhnke et al.	Anidulafungin is safe and effective in the treatment of invasive candidiasis in critically ill patients
Walsh et al.	Efficacy of anidulafungin in the treatment of fungal infection in neutropenic paediatric population

Commercial echinocandin formulations – Cancidas™, Mycamine™ and Eraxis™

For the antifungal therapy, the echinocandins are available commercially as lyophilized powders that need to be reconstituted before use. They are prescribed to be administered as IV infusions, and not as IV bolus injections. This could be attributed to the histamine-mediated or histamine-like reactions observed after the administration of fast IV infusions of the echinocandin drugs.^[212] Hence, they are given as a slow infusion over 1 h to avoid any potential unwarranted side effects.

The drugs are reconstituted and administered to the patients using an infusion bag, containing a specific electrolyte solution (Table 7). All the formulations contain excipients for pH adjustment, such as glacial acetic acid, citric acid, tartaric acid and sodium hydroxide (Table 7). These excipients provide an acidic pH to the formulations (Cancidas™: pH: 6.6; Mycamine™: pH: 5–7; Eraxis™: pH: 4.5–4.6), to ensure their chemical stability (as they undergo ionization and ring-opening at neutral or alkaline pH).^[213,214] The bulking agents such as sucrose, mannitol and fructose are used to prevent hydrolysis of the drugs by sequestering water, to form an easily dispersible cake upon lyophilization and to prevent the dimerization of the drug leading to instability in solution.^[1] As the osmolality of the reconstituted drug solution is low, it is administered using an electrolyte solution, to maintain iso-osmolal conditions, during the IV administration.^[213]

Table 7

A summary on the marketed echinocandin formulations^[26,27,214]

	Caspofungin	Micafungin	Anidulafungin
Marketed as	Cancidas™; Merck and Co.	Mycamine™; Astellas Pharma	Eraxis™; Pfizer
Commercial availability	50-mg and 70-mg vials containing lyophilized powder for reconstitution	50-mg and 100-mg vials containing lyophilized powder for reconstitution	50-mg and 100-mg vials containing lyophilized powder for reconstitution
Active pharmaceutical ingredient	Caspofungin acetate	Micafungin sodium	Anidulafungin
Excipients	Sucrose (39 mga/54 mgb), mannitol (26 mga/36 mgb), glacial acetic acid or sodium hydroxide	Lactose (200 mgc), citric acid or sodium hydroxide	Fructose (50 mgd/100 mge), mannitol (250 mgd/500 mge), polysorbate 80 (125 mgd/250 mge), tartaric acid (5.6 mgd/11.2 mge), sodium hydroxide or hydrochloric acid
Administration	IV infusion	IV infusion	IV infusion
Reconstitution of lyophilized formulation using	0.9% Sodium Chloride Injection, Sterile Water for Injection, Bacteriostatic Water for Injection with methylparaben and propylparaben, or Bacteriostatic Water for Injection with 0.9% benzyl alcohol	0.9% Sodium Chloride Injection USP	20% (w/w) Dehydrated Alcohol in Water for Injection
Electrolyte solution in infusion bag	0.9, 0.45 or 0.225% Sodium Chloride Injection USP; Lactated Ringer's Injection	0.9% Sodium Chloride Injection USP or 100 ml of 5% Dextrose Injection USP	0.9% Sodium Chloride Injection USP or 100 ml of 5% Dextrose Injection USP
Stability	RS: 1 h at ≤25 °C IS: 24 h at ≤25 °C or 48 h at 2–8 °C	RS and IS: 24 h at ≤25 °C	RS and IS: 24 h at ≤25 °C
Specific precautions	Its contact should be avoided with dextrose solutions, as it is incompatible with dextrose solutions	Existing IV line should be flushed with 0.9% Sodium Chloride Injection USP, before its infusion	The rate of infusion should always be <1.1 mg/min, to avoid histamine-mediated reactions

RS, reconstituted solution; IS, infusion solution. ^a50 mg of Cancidas™ vial; ^b70 mg of Cancidas™ vial; ^c50 and 100 mg of Mycamine™ vial; ^d50 mg of Eraxis™ vial; ^e100 mg of Eraxis™ vial.

Clinical usage

All the three echinocandins have shown in-vitro and in-vivo activity against various fungal species, and their potency, efficacy and safety have also been corroborated in the aforementioned plethora of clinical studies. However, the clinical use of echinocandins has been limited to Candida and Aspergillus fungal infections, due to the lack of consistent activity and outcome in other fungal infections.

The IDSA has recommended the use of the echinocandins in the treatment of invasive aspergillosis and invasive candidiasis in both the neutropenic and non-neutropenic patients. Their use has been especially important in the azole-resistant candidaemia infections, and candidiasis due to C. glabrata and C. parapsilosis.^[28,29]

Caspofungin has been widely used for treating candidaemia, oesophageal candidiasis, invasive aspergillosis and febrile neutropenia in both adult and paediatric population. The dosage regimens for both the populations involve the IV administration of a loading dose followed by a maintenance dose. Micafungin, on the other hand, is given as a single-dose IV infusion in adult and paediatric populations, in the chemotherapy of candidaemia and oesophageal candidiasis, and for fungal prophylaxis. Micafungin has been the only clinically approved echinocandin used as a prophylactic agent for fungal infections in haematopoietic stem cell transplant patients due to its proven efficacy of micafungin in clinical studies for fungal prophylaxis.^[200] Anidulafungin is used as a therapeutic agent only in the adult populations infected with candidaemia and oesophageal candidiasis. The use of anidulafungin has not been approved in paediatric therapy due to the lack of clinical studies. Tables 8 and 9 summarize the clinical therapeutic regimens for caspofungin, micafungin and anidulafungin.

Table 8

Echinocandin chemotherapy regimen in adult patients^{[26–29,214]}

	Caspofungin	Micafungin	Anidulafungin
Candidaemia
D	LD: 70 mg; MD: 50 mg	100 mg	LD: 200 mg; MD: 100 mg
Du	Until at least 14 days after the last positive fungal culture	Therapy ranges from 10 to 47 days, with an average of 15 days	Until at least 14 days after the last positive fungal culture
Oesophageal candidiasis
D	50 mg	150 mg	LD: 100 mg; MD: 50 mg
Du	Until 7–14 days after symptom resolution	Therapy ranges from 10 to 30 days, with an average of 15 days	Therapy should be continued for at least 14 days, and then for a minimum of 7 days following symptom resolution
Invasive aspergillosis
D	LD: 70 mg; MD: 50 mg	NA	NA
Du	Therapy based on the severity of the patient's symptoms
Febrile neutropenia
D	LD: 70 mg; MD: 50 mg	NA	NA
Du	With fungal infections for a minimum of 14 days, and continued for at least 7 days after resolution of symptoms
Fungal prophylaxis
D	NA	50 mg	NA
Du		Therapy ranges from 6 to 51 days with an average of 19 days

LD, loading dose given IV; MD, maintenance dose given IV; D, dosage of the drug during the regimen; Du, duration of the therapeutic regimen.

Table 9

Echinocandin chemotherapy regimen in paediatric patients^[26,27,214]

	Caspofungina	Micafunginb	Anidulafungin
Candidaemia
D	LD: 70 mg/m2 MD: 50 mg/m2	2 mg/kg 100 mg/day (Max. D)	NA
Du	Until at least 14 days after the last positive fungal culture	Therapy should be continued for at least 14 days and continued for 7 days after two sequential negative blood cultures
Oesophageal candidiasis
D	LD: 70 mg/m2 MD: 50 mg/m2	3 mg/kg (weight < 30 kg) 2.5 mg/kg (weight > 30 kg) 150 mg/day (Max. D)	NA
Du	MD given until at least 7–14 days after symptom resolution	Therapy should be continued for at least 14 days and continued for 7 days after two sequential negative blood cultures
Invasive aspergillosis
D	LD: 70 mg/m2 MD: 50 mg/m2	NA	NA
Du	Therapy based on the severity of the patient's symptoms
Febrile neutropenia
D	LD: 70 mg/m2 MD: 50 mg/m2	NA	NA
Du	With fungal infections for a minimum of 14 days, and continued for at least 7 days after resolution of symptoms
Fungal prophylaxis
D	NA	1 mg/kg 100 mg/day (Max. D)	NA
Du		Until at least 7 days after neutrophil recovery

LD, loading dose given IV; MD, maintenance dose given IV; Max. D, maximum dose permitted; D, dosage of the drug during the regimen; Du, duration of the therapeutic regimen. ^aPaediatric patients 3 months or older; ^bPaediatric patients 4 months or older.

Pharmacokinetics

Adult population

Caspofungin, micafungin and anidulafungin due to their high molecular weight exhibit poor oral bioavailability, and hence, they are administered as IV formulations. However, enfumafungin – a novel echinocandin in clinical trials – is orally active and could potentially be the first echinocandin to be administered orally.^[215] All the three echinocandins undergo extensive protein binding, which is responsible for their long half-lives. They have variable tissue penetration with prolonged residence times; however, they do not show penetration into brain, cerebrospinal fluid, prostrate and eyes (except micafungin, which penetrates into the eye).^[209,216] Their poor penetration into the aforementioned sites is attributed to their high molecular weights and extensive tissue protein binding.^[25]

Upon their administration, the echinocandins (except anidulafungin) are taken up primarily by the liver where they undergo hepatic metabolism via hydrolysis and N-acetylation pathways, to form inactive metabolites.^[24] These metabolites are primarily eliminated by the bile in faeces over a long period of time. The echinocandins are non-dialysable and do not undergo renal metabolism; hence, the unchanged drug is eliminated in the urine.^[25] Caspofungin undergoes hepatic metabolism via peptide hydrolysis and N-acetylation, and its metabolites are eliminated in both the urine and faeces.^[217] Micafungin, however, undergoes metabolism via the arylsulfatase and catechol-O-methyltransferase (COMT) enzyme pathway in the liver, and its metabolites are eliminated in the faeces and urine.^[218,219] Anidulafungin, on the other hand, does not undergo hepatic metabolism, but undergoes a slow metabolic degradation over a period of time at physiological conditions to form a ring-opened chemical moiety that is then eliminated primarily in the faeces.^[27] As they are not the substrates for renal metabolism, the dose adjustments for echinocandins in renal insufficiency are not warranted. The echinocandins are poor substrates for both P-glycoprotein transporters and cytochrome P 450 enzyme family.^[220]  Table 10 compares the clinical PK parameters among the three echinocandins.

Table 10

Comparison between the PK parameters of caspofungin, micafungin and anidulafungin^{[27,30,209,217,219]}

Parameter	Caspofungin	Micafungin	Anidulafungin
Oral bioavailability	(−)	(−)	(−)
Linear PK	(+)	(+)	(+)
Plasma protein binding (%)	97	99	99
Mean terminal elimination half-life (h)	9–11	12–17	24
Plasma clearance (ml/min)	10–12.5	≈10.5	≈16.67
Volume of distribution (l)	≈9.67	18–19	30–50
Distribution half-life (h)	≈5.5	<1	0.5–1
Mean Cmax (μg/ml) (after 50 mg single dose)	12.1	10.9	7.7
Steady-state AUC0–24 (μg/h per ml) (after 70 mg single dose)	100.5	111.3	42.3

C_max, maximum concentration of the drug during the dosing regimen; AUC_0–24, area under the curve (AUC) from time 0 to 24 h for the drug.

Special population

Hepatic insufficiency

The echinocandins have been studied in patients with mild (Child–Pugh score: 5–6), moderate (Child–Pugh score: 7–9) and severe (Child–Pugh score: 10–12) hepatic insufficiency. After the administration of single 70 mg caspofungin dose in mild hepatic insufficiency patients, the increase in AUC was about 55%, which was not significantly different than that in the control group, and hence, no dose adjustment is needed. However, when 70 mg LD is given with 50 mg daily MD, the AUC in the moderate hepatic insufficiency group increased by 75% necessitating lowering of the MD to 35 mg daily, without changing the LD.^[214] In micafungin regimen, the clinical dose of 100 mg was given by IV infusion in patients with moderate and severe hepatic insufficiency and it was found that the AUC lowered by 22 and 30% in both the groups, respectively, without affecting the therapy and safety of the patients. Hence, the adjustment of dose for micafungin is not warranted.^[26] Anidulafungin is not a substrate for hepatic metabolism, and hence, its dose adjustment in mild, moderate and severe hepatic insufficiency is not required.^[27]

Renal insufficiency

As the echinocandins are not a substrate for renal metabolism, their dose adjustments in mild, moderate or severe renal insufficiency are not warranted. A similarity in the PK parameters for the renal insufficiency (mild, moderate and severe) group and control group was observed in a clinical study performed for evaluating the effect of renal insufficiency on echinocandin PK parameters. As all the echinocandins are non-dialysable, they may be administered without any regard to the haemodialysis regimen.^[26,27,214]

Paediatric population

Micafungin is currently approved for antifungal therapy in neonates and paediatric populations. A clinical study was undertaken to evaluate the micafungin PK parameters in paediatric population by Seibel et al. in 2005. The micafungin therapy was conducted between doses 0.5–4.0 mg/kg per day, and the PK parameters exhibited a dose linearity, with no adverse side effects. Clearance, volume of distribution and half-life did not significantly change during the dosage regimen, and the overall PK profile was similar to that observed in adults. The clearance, however, increased with a decrease in age of the paediatric patients.^[221]

Limited studies have been performed on paediatric patients for caspofungin. In a clinical study on paediatric patients, children between 2–17 years were included. Caspofungin was given either as a 1 mg/kg per day dose or as a 50 mg/m² per day or 70 mg/m² per day dose, and the resulting PK parameters were compared to adult patients receiving 50 or 70 mg/day caspofungin doses. AUC_0–24 was significantly smaller for paediatric patients receiving 1 mg/kg per day calculated caspofungin dose; whereas the paediatric group receiving 50 mg/m² per day dose had AUC_0–24 similar to the adults receiving 50 mg/day dose. Thus, a caspofungin dose of 50 mg/m² per day in paediatric population shows similar PK parameters with an equivalent response as seen in a 50 mg/day adult dose.^[186]

Anidulafungin is currently not approved for paediatric administration. However, an evaluation of anidulafungin PK parameters in immunocompromised paediatric patients (2–17 years), showed similar PK parameters and response at 0.75 and 1.5 mg/kg per day to adult populations receiving 50 and 100 mg/day anidulafungin doses.^[27,222]

Geriatric population

No dose adjustments are required in geriatric populations for any of the echinocandins.^[26,27,214] In the case of caspofungin, about 28% of increase in AUC was observed in patients above 65 years of age upon administration of a single 70 mg caspofungin dose. However, it was not significantly different from the younger control group (20–24 years of age), and hence, no adjustment for caspofungin dose is warranted. In the case of anidulafungin, a slight difference was observed in the rate of clearance between the elderly (>65 years) and non-elderly (<65 years) group, with no significant differences in the other PK parameters. Similarities in PK parameters for micafungin were also observed between the elderly (>65 and >75 years) and the young control group (20–24 years), not requiring the adjustment of micafungin doses.

Pregnant and lactating mothers

The echinocandins have been categorized as pregnancy class C drugs, as caspofungin and anidulafungin cross the placental barrier. Thus, the echinocandins should be cautiously used in pregnant women. So also, the echinocandins are found in the milk of lactating rats that had been treated with echinocandins, and hence, their administration should be avoided in lactating women. However, it is also imperative that more robust clinical studies should be undertaken in evaluating the efficacy and safety of echinocandins in pregnant and lactating mothers.^[26,27,214]

Clinical considerations

Safety and adverse drug reactions

The marketed echinocandins are generally well tolerated and safe in clinical practices for the treatment of fungal infections. The use of echinocandin antifungals are contraindicated in the patients who are sensitive to its use. All the three echinocandins exhibit a similarity in their adverse side effects.

All the three marketed echinocandins have been reported to exhibit hypersensitivity associated anaphylactic shock, and in such clinical scenarios, withdrawal of echinocandin chemotherapy is essential. Histamine-mediated reactions such as rash, facial swelling, angio-oedema, sensation of warmth, bronchospasm, urticaria, flushing, pruritus, dyspnoea and hypotension have been associated with the infusion mediated echinocandin therapy; hence, careful monitoring of the infusion is necessary. All the three echinocandins have exhibited rare and isolated cases of hepatic side effects such as hepatic dysfunction, hepatitis and hepatic failure, and therefore, careful monitoring of patients with hepatic abnormalities should be ensured.^[26,27,214]

In addition to these common side effects, caspofungin shows diarrhoea, pyrexia, increased alanine aminotransferase (ALT)/aspartate aminotransferase (AST) levels and decreased blood alkaline phosphatase levels.^[214] Micafungin has additionally exhibited diarrhoea, nausea, vomiting, pyrexia, thrombocytopenia, headache and isolated cases of haematological side effects (acute intravascular haemolysis, haemolytic anaemia and haemoglobinuria).^[26] Anidulafungin exhibited the following adverse effects: cardiac (atrial fibrillation, bundle branch block right, sinus arrhythmia and ventricular extrasystoles), ocular (vision blurred, eye pain and visual disturbance), GIT (nausea, abdominal pain upper, faecal incontinence and loose stools), hyperkalaemia, hypercalcaemia, hypernatraemia, headache, convulsions, and an increase in AST, blood amylase, blood creatinine and lipase levels. However, these side effects were observed in less than 1% of the patient population.^[27]

As the occurrence of these adverse effects is rare and not frequent (for caspofungin and micafungin <10% patients, and anidulafungin ≤1.0% patients), all the three echinocandins are considered clinically safe.

Drug–drug interactions

As previously discussed, the echinocandins are poor substrates for CYP450 isoenzymes and do not interact with P-glycoprotein transporters. Hence, they do not affect or do not get affected by other drugs to show erratic changes in their PK profiles. Nonetheless, some DDI still persist.

Caspofungin PK profile has been reported to be affected by ciclosporin, tacrolimus and rifampin most commonly. Ciclosporin increased the caspofungin AUC by approximately 35%, but caspofungin was not found to increase the ciclosporin plasma concentration. Caspofungin reduced the tacrolimus 12-h blood concentration, 12-h maximum concentration and AUC_0–12 by 26, 16 and 20%, respectively. Rifampin, however, affected the caspofungin trough concentration by reducing it by approximately 30%. Co-administration of CYP enzyme inducers such as efavirenz, nevirapine, phenytoin, dexamethasone and carbamazepine are known to decrease caspofungin concentration. Caspofungin has been found to not affect the PK profiles of amphotericin B, nelfinavir, itraconazole and mycophenolate, and conversely, these drug also do not affect the PK parameters of caspofungin.^[214]

The effect of micafungin administration was evaluated on a number of drug classes such as antifungals (amphotericin B, fluconazole, itraconazole, voriconazole), immunosuppressants (tacrolimus, sirolimus, mycophenolate mofetil, ciclosporin), steroids (prednisolone), calcium channel blockers (nifedipine), antiprotease inhibitors (ritonavir) and antimycobacterials (rifampin). None of these drugs affected micafungin PK parameters; however, micafungin was found to affect the PK profiles of nifedipine (increase in AUC and maximum concentration by 18 and 42%, respectively), sirolimus (increase in AUC by 21% with no increase in maximum concentration) and itraconazole (increase in AUC and maximum concentration by 22 and 11%, respectively). Micafungin did not alter the PK profiles of mycophenolate mofetil, ciclosporin, tacrolimus, prednisolone, fluconazole and voriconazole.^[26]

The interactions of anidulafungin have been studied with ciclosporin, voriconazole and tacrolimus. It has been found that the co-administration of anidulafungin causes an approximate 22% increase in the AUC for ciclosporin with no significant change in its maximum concentration. However, anidulafungin causes no significant changes in the AUC and maximum concentration of voriconazole and tacrolimus with its simultaneous administration. The administration of amphotericin B and rifampin with anidulafungin causes no effect on its PK profile.

Table 11 gives a brief overview over the adverse effects and DDI associated with each of the echinocandins. Overall, the echinocandins have shown clinical success in the management of fungal infections, by exhibiting acceptable safety with minimum chances of adverse drug reactions and DDI.

Table 11

Summary of the clinical adverse side effects and drug–drug interaction for the three echinocandins^[26,27,214]

	Safety	Specific adverse side effects	Drug–drug interaction
			Interaction	No interaction
Caspofungin	Safe	Diarrhoea, pyrexia, increased (ALT)/(AST) levels and decreased blood alkaline phosphatase levelsa	Ciclosporin, tacrolimus and rifampin	Amphotericin B, nelfinavir, itraconazole and mycophenolate
Micafungin	Safe	Diarrhoea, nausea, vomiting, pyrexia, thrombocytopenia, headache and isolated cases of acute intravascular haemolysis, haemolytic anaemia and haemoglobinuriab	Nifedipine, sirolimus and itraconazole	Mycophenolate mofetil, ciclosporin, tacrolimus, prednisolone, fluconazole and voriconazole
Anidulafungin	Safe	Atrial fibrillation, bundle branch block right, sinus arrhythmia, ventricular extrasystoles, vision blurred, eye pain, and visual disturbance, nausea, abdominal pain upper, faecal incontinence, loose stools, hyperkalaemia, hypercalcaemia, hypernatraemia, headache, convulsions, and an increase in AST, blood amylase, blood creatinine and lipase levelsc	Ciclosporin	Amphotericin B, rifampin, voriconazole and tacrolimus

^aIncidence in adults (10% or more) and in paediatric population (≤10%). ^bIncidence in adults (10% or more) and in paediatric population (<15%). ^cIncidence in adults (≤1.0%).

Pharmacoeconomic considerations

Pharmacoeconomic analyses reveal that the echinocandin antifungal pharmacotherapy maybe more cost-effective than the other current therapies involving amphotericin B and the azoles.^[223,224] The pharmacoeconomic analyses involved cohesive evaluation of various parameters such as duration of therapy, modification in the regimen of the therapy, stage of the initiation of the therapy, cost of the drugs during the course of the therapy, clinical complications and therapeutic success of the therapy, in deciding the cost-effectiveness of the echinocandin therapy. Caspofungin therapy has shown cost-effectiveness in the management of clinical fungal infections in comparison with the standard amphotericin B therapy.^[225,226] Similarly, economic analyses of micafungin therapies in fungal infections (candidaemia, invasive candidiasis and suspected ICU fungal infections) vs amphotericin B and fluconazole therapy, yielded results in favour of micafungin for better economy in treating of the fungal infections.^[227,228] So also, the use of anidulafungin in fungal infections such as candidaemia and invasive candidiasis has been reported to be more cost-effective than the standard fluconazole therapy in Australia, the United Kingdom and Spain.^{[223,229,230]} In accordance to these various pharmacoeconomic evaluations, it can be concluded that all the three marketed echinocandins seem to be a cost-effective therapy in the successful management of antifungal infections.

Conclusion

The echinocandins are a novel and an important class of antifungals that can potentially overcome some of the drawbacks of the current therapeutic agents. The echinocandin antifungals have shown success in the treatment of invasive candidiasis and aspergillosis with better safety and tolerability profiles in both adult and paediatric patient populations. Hence, they are worthy of being considered as front-line agents in the management of invasive fungal infections. However, in spite of the availability of abundant data on the echinocandins, encompassing their various facets and attributes, there are still numerous unexplored areas for this class of drugs. These include delineation of the differences among the echinocandins with respect to their pharmacology and clinical outcomes, better understanding of the echinocandin resistance mechanisms, potential combination therapy of the echinocandins with other antifungal agents, clinical trials for the evaluation of anidulafungin chemotherapy in paediatric patients, a better in-vitro–in-vivo correlation for the activity of echinocandins against fungal species other than Candida and Aspergillus, potential concomitant administration of two different echinocandins for antifungal treatment against Candida and Aspergillus species, and an in-depth global pharmacoeconomic analyses for the marketed echinocandins. Moreover, the prospective use of echinocandins in treating fungal infections affecting the cardiac, ocular and nervous sites should be explored to extend the usage of echinocandins beyond their current application in the systemic invasive fungal infections. A better understanding of the echinocandins in these myriad niches would further aid in giving a clarity to the locus of echinocandins in the consortium of the antifungal agents.