Chapter 44: Antifungal Agents and Resistance

Compared with antibacterial agents, relatively few antimicrobials are available for treatment of fungal infections. Many substances with antifungal activity have proved to be unstable, to be toxic to humans, or to have undesirable pharmacologic characteristics, such as poor diffusion into tissues. Of the agents in current clinical use, the newer azole compounds have the broadest spectrum with significantly lower toxicity than earlier antifungal agents. An even newer class of cell wall active agents offers hope for the selective toxicity that β-lactams provide for antibacterial therapy.

Fortunately, most fungal infections are self-limiting and require no chemotherapy. Superficial mycoses are often treated, but topical therapy can be used, thus limiting toxicity to the host. The remaining small group of deep mycoses that are uncontrolled by the host's immune system require the prolonged use of antifungals. This, combined with the fact that most of the patients have underlying immunosuppression, makes them among the most difficult of all infectious diseases to treat successfully. The characteristics of currently used antifungal agents are discussed next and summarized in Table 44–1. They are discussed in the text that follows in relation to their target of action, as illustrated in Figure 44–1.

CYTOPLASMIC MEMBRANE

Polyenes

The polyenes nystatin and amphotericin B are lipophilic and bind to ergosterol, the dominant sterol in the cytoplasmic membrane of fungal cells. After binding, they form annular channels, which penetrate the membrane and lead to leakage of essential small molecules from the cytoplasm and cell death. Their binding affinity for the ergosterol of fungal membranes is not absolute and includes sterols such as cholesterol, which are present in human cells. This is the basis of the considerable toxicity that limits their use. Almost all fungi are susceptible to amphotericin B, and the development of resistance is too rare to be a consideration in its use.

At physiologic pH, amphotericin B is insoluble in water and must be administered intravenously as a colloidal suspension. It is not absorbed from the gastrointestinal tract. The major limitation to amphotericin B therapy is the toxicity created by its affinity for mammalian as well as fungal membranes. Infusion is commonly followed by chills, fever, headache, and dyspnea. The most serious toxic effect is renal dysfunction and is observed in virtually every patient receiving a therapeutic course. Experienced clinicians learn to titrate the dosage for each patient to minimize the nephrotoxic effects. For obvious reasons, use of amphotericin B is limited to progressive, life-threatening fungal infections. In such cases, despite its toxicity it retains a prime position in treatment often by administration of an initial course of amphotericin followed by a less toxic agent. Preparations that complex amphotericin B with lipids have been used as a means to limit toxicity. The even greater toxicity of nystatin limits its use to topical preparations.

Azoles

The azoles are a large family of synthetic organic compounds, which includes members with antibacterial, antifungal, and antiparasitic properties. The important antifungal azoles for systemic administration are ketoconazole, fluconazole, itraconazole, and voriconazole. Clotrimazole and miconazole are limited to topical use. Other azoles are under development or evaluation. Their activity is based on inhibition of the enzyme (14 α-demethylase) responsible for conversion of lanosterol to ergosterol, the major component of the fungal cytoplasmic membrane. This leads to lanosterol accumulation and the formation of a defective membrane. Effects on the precursors of some hormones may cause endocrine side effects and restricts use in pregnancy. All antifungal azoles have the same mechanism of action. The differences among them are in avidity of enzyme binding, pharmacology, and side effects.

Ketoconazole, the first azole, has now been supplanted by the later azoles for most systemic mycoses. Although nausea, vomiting, and elevation of hepatic enzymes complicate the treatment of some patients, the azoles are much less toxic than amphotericin B. Fluconazole was the first azole with good central nervous system penetration, but itraconazole is now generally preferred for fungal meningitis. Azoles are also effective for superficial and subcutaneous mycoses in which the initial therapy either fails or is not tolerated by the patient. In general, itraconazole and, more recently, voriconazole are the primary azoles used together with, or instead of, amphotericin B for serious fungal infections. Clotrimazole and miconazole are available in over-the-counter topical preparations.

Allylamines

The allylamines are a group of synthetic compounds that act by inhibition of an enzyme (squalene epoxidase) in the early stages of ergosterol synthesis. The allylamine group includes an oral and topical agent, terbinafine used in the treatment of dermatophyte (ringworm) infections.

NUCLEIC ACID SYNTHESIS

Flucytosine

5-Flucytosine (5FC) is an analog of cytosine. It is a potent inhibitor of RNA and DNA synthesis. 5FC requires a permease to enter the fungal cell, where its action is not direct but through its enzymatic modification to other compounds (5-fluorouracil, 5-fluorodeoxyuridyic acid, 5-fluoruridine). These metabolites then interfere with DNA synthesis and cause aberrant RNA transcription.

Flucytosine is well absorbed after oral administration. It is active against most clinically important yeasts, including Candida albicans and Cryptococcus neoformans, but has little activity against molds or dimorphic fungi. The frequent development of mutational resistance during therapy limits its application to mild yeast infections or its use in combination with amphotericin B for cryptococcal meningitis. The combination reduces the probability of expression of resistance and allows a lower dose of amphotericin B to be used. The primary toxic effect of flucytosine is a reversible bone marrow suppression that can lead to neutropenia and thrombocytopenia. This effect is dose related and can be controlled by drug monitoring.

CELL WALL SYNTHESIS

The unique chemical nature of the fungal cell wall, with its interwoven layers of mannan, glucan, and chitin (Figure 44–1), makes it an ideal target for chemotherapeutic attack. Although such antifungal agents have only recently (2002) entered the armamentarium, they are most welcome. The echinocandins, which block glucan synthesis, are now in clinical use and the nikkomycins, which block chitin synthesis, are in development.

Echinocandins

Echinocandins act by inhibition of a glucan synthetase (1,3-β-D-glucan synthetase) required for synthesis of the principal cell wall glucan of fungi. Its action causes morphologic distortions and osmotic instability in yeast and molds that are similar to the effect of β-lactams on bacteria. The first such agent to be licensed is caspofungin, which has good activity against Candida and Aspergillus and a wide range of other fungi. Cryptococcus neoformans whose cell wall glucans have a slightly different structure is resistant. Since there are no similar human structures, toxicity is minimal. The newest echinocandins, micafungin and andiulafungin, have the same mode of action and a similar spectrum.

Nikkomycins

Nikkomycins have a mechanism of action analogous to echinocandins. They inhibit chitin synthetase, which polymerizes the N-acetylglucosamine subunits that make up chitin. The result is inhibition of chitin synthesis. The agent in development, nikkomycin Z, has activity against dimorphic fungi such as Coccidioides immitis and Bastomyces dermatitidis but not against yeast or Aspergillus.

Other Antifungal Agents

Griseofulvin is a product of a species of the mold Penicillium. It is active only against the agents of superficial mycoses. Griseofulvin is actively taken up by susceptible fungi and acts on the microtubules and associated proteins that make up the mitotic spindle. It interferes with cell division and possibly other cell functions associated with microtubules. Griseofulvin is absorbed from the gastrointestinal tract after oral administration and concentrates in the keratinized layers of the skin. Clinical effectiveness has been demonstrated for all causes of dermatophyte infection, but the response is slow. Difficult cases may require 6 months of therapy to affect a cure.

Potassium iodide is the oldest known oral chemotherapeutic agent for a fungal infection. It is effective only for cutaneous sporotrichosis. Its activity is somewhat paradoxical, because the mold form of the etiologic agent, Sporothrix schenckii, can grow on medium containing 10% potassium iodide. The pathogenic yeast form of this dimorphic fungus appears to be susceptible to molecular iodine.

DEFINITION OF RESISTANCE

The concepts, definitions, and laboratory methods described in Chapter 23 for bacterial resistance are generally applicable to fungi. Quantitative susceptibility is measured by the minimal inhibitory concentration (MIC) under conditions that favor the growth of fungi. The wide range of growth rates and diversity of growth forms (yeast, hyphae, conidia) in the various fungi have added technical variables to testing, but standardized methods are now available. Comparison of MICs with drug pharmacology allows classification of fungi as susceptible or resistant, but these results do not yet predict clinical outcome with the same certainty they do with bacteria. Because of its specialized nature, the availability of antifungal susceptibility testing is restricted to major centers and reference laboratories.

MECHANISMS OF RESISTANCE

The same resistance mechanisms observed in bacteria are also found in fungi. A major addition is the much greater use of metabolic means such as efflux pumps and changes in synthetic pathways by fungi. The most glaring difference is the complete absence of enzymatic inactivation of antifungals as resistance mechanism. Perhaps, related to this is the absence in fungi of powerful means for gene transfer such as conjugation and transposition.

Polyene Resistance

Because amphotericin B binds directly to the cytoplasmic membrane, the only means to resist this action is to change the membrane composition. The uncommon strains that have been studied show a decrease in the ergosterol content of the membrane. This limits the primary binding sites.

Flucytosine Resistance

Flucytosine requires a permease for entry into the cell and then multiple enzymes to modify it to the active metabolites. Mutation in any one of these enzymes renders the drug ineffective. This happens readily under the selective pressure of 5FC use. It is one of the few antimicrobials in which emergence of resistance during therapy of an acute infection is predictable. It is the reason its use is limited to combinations with other antifungals.

Azole Resistance

There are four major mechanisms of resistance that cross all the azole agents. The two most prominent are efflux pumps and altered target. The efflux pumps transport drug that has entered the cell back outside. Some pumps act for all azoles and others act on only one. Alteration of subunits of the demethylase enzyme by mutation decreases the affinity of the azole for its enzyme target. Multiple mutations can have an additive effect.

Two metabolic mechanisms compensate for the drug's presence without altering its target or directly inactivating it. Upregulation of the target demethylase allows its action to continue despite binding of some of the enzyme by the azole. Some resistant strains have been shown to accomplish ergosterol synthesis by an alternate pathway, thus bypassing the azole affected mechanism.

Echinocandin Resistance

Although the echinocandins are relatively new, resistance has already been observed with their use. The mechanism is altered target. Mutations in subunits of the glucan synthetase target have been correlated with increases in MIC of up to a thousandfold.

SELECTION OF ANTIFUNGALS

As with all chemotherapy, the selection of antifungal agents for treatment of superficial, subcutaneous, and systemic mycoses involves balancing probable efficacy against toxicity. The factors to be considered are the following: (1) the threat of morbidity or mortality posed by the specific infection, (2) the immune status of the patient, (3) the toxicity of the antifungal, and (4) the probable activity of the antifungal agent against the fungus. In the case of superficial mycoses, the risks of appropriate therapy are small, and various topical agents may be tried. At the other extreme, an immunocompromised patient will most likely be treated aggressively with systemic agents for proven or even suspected systemic fungal infection. Since susceptibility tests are usually not available, the decisions regarding which agents to use are usually made and sustained on an empiric basis. Even when guided by in vitro testing, treatment failures are common particularly in the immunocompromised. It is hoped that the addition of the new cell wall active agents to the regimen will have a favorable effect on these outcomes.