Chapter 1

44
Antifungal Agents and Resistance

C
ompared 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
Many antifungals are too toxic for
characteristics, such as poor diffusion into tissues. Of the agents in current clinical use, the
use
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 Treatment is most needed for
to the host. The remaining small group of deep mycoses that are uncontrolled by the host’s dissemination in immunocompro-
immune system require the prolonged use of antifungals. This, combined with the fact that mised persons
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.

ANTIFUNGAL AGENTS

CYTOPLASMIC MEMBRANE
MM 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 chan-
nels, which penetrate the membrane and lead to leakage of essential small molecules from Ergosterol binding forms
the cytoplasm and cell death. Their binding affinity for the ergosterol of fungal membranes is membrane channels
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 Active against most fungi
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 intra-
venously 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 mam-
malian as well as fungal membranes. Infusion is commonly followed by chills, fever, head-
Insoluble compound must be
ache, and dyspnea. The most serious toxic effect is renal dysfunction and is observed in
infused in suspension
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
Therapy must be titrated against
amphotericin B is limited to progressive, life-threatening fungal infections. In such cases,
toxicity
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.
713
714 PART IV Pathogenic Fungi

TABLE 44–1 Features of Antifungal Agents

AGENT ACTION RESISTANCE ROUTE CLINICAL USE
Polyenes
Nystatin Cytoplasmic membrane Sterol modification Topical Most fungi
pores
Amphotericin B Cytoplasmic membrane Sterol modification Intravenous Aspergillus, Candida,
pores ­Cryptococcus, Histoplasma,
­Sporothrix, Coccidioides
Azoles
Ketoconazole Ergosterol synthesis Efflux, demethylase Oral Candida, dermatophytes,
(demethylase) ­alteration, bypass, ­dimorphic fungib
­overproductiona
Fluconazole Ergosterol synthesis Efflux, demethylase Oral, intravenous Candida, Cryptococcus,
(demethylase) ­alteration, bypass, ­Histoplasmae, Coccidioidese
overproductiona
Itraconazole Ergosterol synthesis Efflux, demethylase Oral, intravenous Aspergillus, Sporothrix, ­Candida,
(demethylase) ­alteration, bypass, Blastomyces, Histoplasma,
overproductiona ­Coccidioides
Voriconazole Ergosterol synthesis Oral, intravenous Candida, Aspergillus, some
(demethylase) ­saprophytic molds
Posaconazole Ergosterol synthesis Oral Candida, Aspergillus
(demethylase) ­(prophylaxis), Zygomycetes
Clotrimazole Ergosterol synthesis Unknownc Topical Candida, dermatophytes
(demethylase)
Miconazole Ergosterol synthesis Unknownc Topical Candida, dermatophytes
(demethylase)
Allylamines
Terbinafine Ergosterol synthesis ?Efflux Oral, topical Dermatophytes
(squalene epoxidase)
Flucytosine
DNA synthesis, RNA Permease or modifying Oral Candida and Cryptococcusf
transcription enzymesd mutation
Echinocandins
Caspofungin Glucan synthesis (glucan Altered synthetase Intravenous Candida, Aspergillus
synthetase)
Micafungin Glucan synthesis (glucan Altered synthetase Intravenous Candida, Aspergillus
synthetase)
Anidulafungin Glucan synthesis (glucan Altered synthetase Intravenous Candida, Aspergillus
synthetase)
Nikkomycins Chitin synthesis (chitin Developmental
synthetase)
Griseofulvin Microtubule disruption Unknown Oral Dermatophytes
Potassium iodide Unknown Unknown Oral Sporothrix schenckii

5FC, 5-Flucytosine.
a
Most work is with fluconazole and Candida; other azoles are to be assumed to be similar.
b
Generally less absorbed and less active than fluconazole or itraconazole.
c
Probably similar to other azoles, but resistance to the concentrations in topical preparations may differ.
d
Cytosine deaminase and uracil phosphoribosyltransferase (the enzymes that modify 5FC to active forms).
e
Itraconazole generally preferred.
f
Only in combination with amphotericin B owing to resistance mutation.
 ANTIFUNGAL AGENTS AND RESISTANCE CHAPTER 44 715

Echinocandins

Glucan
Chitin
Nikkomycin

Azoles Ergosterol
Amphotericin B

Flucytosine FIGURE 44–1.  Action of antifun-

gal agents. The sites where the major
antifungal agents act in the cell wall
(echinocandins, nikkomycin), cytoplasmic
membrane (azoles, amphotericin B) and
genome (flucytosine) are shown.

MM 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 develop-
ment or evaluation. Their activity is based on inhibition of the enzyme (14 α-demethylase) Inhibit enzyme crucial for synthesis
responsible for conversion of lanosterol to ergosterol, the major component of the fungal of membrane ergosterol
cytoplasmic membrane. This leads to lanosterol accumulation and the formation of a defec-
tive 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 sys-
temic mycoses. Although nausea, vomiting, and elevation of hepatic enzymes complicate
Less toxic than amphotericin B
the treatment of some patients, the azoles are much less toxic than amphotericin B. Fluco-
nazole was the first azole with good central nervous system penetration, but itraconazole
Itraconazole and voriconazole
is now generally preferred for fungal meningitis. Azoles are also effective for superficial
prime systemic agents
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. Clotrima-
zole and miconazole are available in over-the-counter topical preparations.

MM 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 Inhibit ergosterol synthesis
includes an oral and topical agent, terbinafine used in the treatment of dermatophyte (ring-
worm) infections.

NUCLEIC ACID SYNTHESIS

MM Flucytosine
5-Flucytosine (5FC) is an analog of cytosine. It is a potent inhibitor of RNA and DNA Enzymatically modified form makes
synthesis. 5FC requires a permease to enter the fungal cell, where its action is not direct defective RNA
but through its enzymatic modification to other compounds (5-fluorouracil, 5-fluorode-
oxyuridyic acid, 5-fluoruridine). These metabolites then interfere with DNA synthesis and Inhibits DNA synthesis
cause aberrant RNA transcription.
716 PART IV
Active against yeasts but not molds
Resistance develops during therapy
if used alone
Inhibit synthease crucial for glucan
synthesis
Current use is Candida, Aspergillus
Inhibit chitin synthesis
Microtubule disruption interferes
with cell division
Active against dermatophytes
Iodide inhibits Sporothrix
Pathogenic Fungi
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 resis-
tance 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 clini-
cal use and the nikkomycins, which block chitin synthesis, are in development.
MM 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 morpho-
logic 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 neo-
formans whose cell wall glucans have a slightly different structure is resistant. Since there
are no similar human structures, toxicity is minimal. The newest echinocandins, micafun-
gin and andiulafungin, have the same mode of action and a similar spectrum.
MM 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 lay-
ers of the skin. Clinical effectiveness has been demonstrated for all causes of dermatophyte infec-
tion, 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 infec-
tion. 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.
RESISTANCE TO ANTIFUNGAL AGENTS
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
 ANTIFUNGAL AGENTS AND RESISTANCE CHAPTER 44 717

various fungi have added technical variables to testing, but standardized methods are now Concepts are similar to bacterial
available. Comparison of MICs with drug pharmacology allows classification of fungi as resistance
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 antifun- Laboratory methods are variable
gal 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 enzy-
matic 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.

MM 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 Membrane ergosterol decreased
been studied show a decrease in the ergosterol content of the membrane. This limits the
primary binding sites.

MM 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 inef-
fective. This happens readily under the selective pressure of 5FC use. It is one of the few Multiple enzymes can mutate
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.

MM 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 Azole pumped out
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 Enzyme target altered
azole for its enzyme target. Multiple mutations can have an additive effect.
Two metabolic mechanisms compensate for the drug’s presence without altering its tar-
get or directly inactivating it. Upregulation of the target demethylase allows its action to Demethylase upregulated or
continue despite binding of some of the enzyme by the azole. Some resistant strains have bypassed
been shown to accomplish ergosterol synthesis by an alternate pathway, thus bypassing the
azole affected mechanism.

MM 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 Mutant 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, sub-
cutaneous, and systemic mycoses involves balancing probable efficacy against toxicity. The fac-
tors 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 susceptibil-
ity 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.