Structure–Activity Relationship

Examples of polyene (i.e., multiple double-bond; see Figure 9-1) antifungal drugs include amphotericin B, nystatin, and pimaricin. Each antibiotic is produced by a different species of Streptomyces (family Actinomyces). Amphotericin B was developed in the 1960s and was so successful in fulfilling the need for a broad-spectrum antifungal that further advancement of antifungal therapy was largely ignored. These drugs are very large molecules, consisting of a macrolide containing a large lactone ring. The polyene contains three to eight double bonds, which represent the lipophilic portion of the molecule. The number of double bonds categorizes the polyenes into trienes, tetraenes (natamycin [paramycin]), and pentanes; amphotericin B and candicidin are heptanes, whereas nystatin is classified as a pseudo–heptane/tetraene.³ A hydroxylated hydrocarbon backbone represents the hydrophilic portion of the molecule. These compounds are insoluble in water and are unstable, and they will rapidly decompose if exposed to sunlight.

Mechanism of Action

Polyene macrolides bind with the sterol portion of the phospholipids that make up the fungal cell membrane. Amphotericin has a much higher affinity for ergosterol, the major sterol component of fungal cell membranes, than for cholesterol, the major sterol in mammalian cell membranes (Figure 9-3).¹ The interaction of the drug and the sterol results in the formation of channels or pores in the cell membrane (Figure 9-4). The result is an increase in cell permeability and disruption in proton gradient flow; loss of membrane fluidity may alter H+-ATPase activity.³ Altered K⁺/H⁺ exchange results in the loss of K⁺ and Mg²⁺ from the cell. Cellular metabolism is disrupted; internal acidification of the fungal cell and the loss of important organic molecules from the cell result in irreversible cell damage. The efficacy of some of the drugs can be related to their ability to bind to ergosterol. However, the polyenes also are associated with the formation of lethal reactive oxygen molecules. Polyenes also are defined according to the concentration of drug necessary to cause erythrocyte hemolysis, presumably because of cholesterol binding. Pimaricin causes lysis at low concentrations, limiting use to topical application, whereas amphotericin B and nystatin cause lysis at much higher concentrations.³

Figure 9-3 Cholesterol, the major sterol of mammalian cells, is structurally similar to ergosterol, the major sterol of fungal cell walls. Binding of amphotericin B to cholesterol results in complicated pharmacokinetics as well as host toxicity.

Figure 9-4 The mechanism of antifungal action and nephrotoxicity of amphotericin B reflect binding of the drug to ergosterol, the major sterol in fungal cell walls. Binding results in cell membrane permeability and the loss of critical micronutrients and electrolytes.

Amphotericin is fungistatic but can be fungicidal at high concentrations. At high concentrations the drug directly disrupts the fungal cell membrane. As with some other select antifungal drugs, amphotericin appears to have some immunomodulating characteristics. Both humoral and cell-mediated immunity may be enhanced, thus increasing the host’s ability to overcome infection. Amphotericin B activates macrophages and stimulates TNF-α and interleukin-1 (IL-1), both of which facilitate macrophage killing by way of nitric oxide–dependent pathways.^5,14

Spectrum of Activity and Pharmacodynamics

Despite the advent of the azole antifungal drugs, amphotericin B remains the most effective agent against most of the major fungal pathogens. The indications for amphotericin therapy include most systemic fungal diseases, including those caused by the dimorphic fungi (histoplasmosis, blastomycosis, cryptococcosis, and coccidioidomycosis) and disseminated sporotrichosis, phycomycosis, aspergillosis, and candidiasis. Amphotericin B has greater activity against some organisms (e.g., Candida and Aspergillus species and coccidioidal meningitis) than the newer azoles and particularly fluconazole. Amphotericin B is not effective against dermatophytes. Amphotericin MICs appear to correlate well with efficacy in animal models and human patients.¹⁰ The MICs of amphotericin B toward selected fungal microbes are listed in Table 9-4.^15,16 Although these are based on humans, they represent a reasonable target for isolates infecting animals as well. In addition to the MIC, the minimum fungicidal concentration (MFC) has been reported for selected organisms infecting humans (or animals).¹⁵ The MFC₅₀ and MFC₉₀; μg/mL) are, respectively, Blastomyces dermatitidis (0.125, 0.5); H. capsulatum (0.5, 2), and C. immitis (4, 16). In killing-curve studies, amphotericin B consistently exhibits concentration-dependent killing. However, killing is variable among organisms, being fungicidal toward Candida sp. but not toward many yeasts and other organisms. As a concentration-dependent drug, amphotericin B may exhibit a long post-antifungal effect (PAFE), which, like the postantibiotic effect of concentration antimicrobials, varies in duration with the organism and the relationship of the MIC to the plasma drug concentration (PDC).¹⁰ However, it can prolong the dosing interval of the drug. A PAFE of only 0.5 hours occurs at approximately 2 hours exposure at drug concentrations that reflect 0.5 times the MIC, but this duration is prolonged to approximately 10 hours at 32 times the MIC for Candida and Cryptococcus neoformans; the PAFE for the latter may exceed 12 hours.¹⁰

Table 9-4 Pharmacokinetic Data for Liposomal Amphotericin B Preparation in the Dog²²

Sufficient data exist to integrate Ambisome pharmocokinetics and pharmacodynamics toward selected microbes that infect the dog (Table 9-4; see also Table 9-2). Two targets are considered: the MFC and a 12-hour PAFE (targeting PDCs that equal the MIC₉₀ of the infecting microbe by 32). Based on the C_max of dogs for Ambisome measured at day 1 and day 30 of dosing, the MFC of B. dermatitidis and H. capsulatum will be achieved on the first day of dosing at 1 mg/kg. Assuming the PAFE as described in animal models would be exhibited by dimorphic fungal organisms infecting dogs, based on MIC₉₀ for B. dermatitidis of 0.05 μg/mL (see Table 9-2), a target of 1.6 μg/mL (32 times the MIC) is indicated to achieve a 10 to 12-hour PAFE. Again, the target will be reached in dogs on day 1 with a 1 mg/kg dose of Ambisome; the 12-hour PAFE could be added to the time the PDCs are above the MIC (which in turn, is based on half-life). For H. capsulatum, the MIC₉₀ is 1 μg/mL; accordingly, a target of 32 μg/mL (for at least 2 hours) might be indicated to achieve a 10 to 12-hour PAFE. Assuming linear kinetics, reaching the target on day 1 will require a dose of approximately 4 mg/kg, which is likely not to be tolerated (an exception may occur for lipid-based products). However, the target is likely to be approached with 30 days of dosing at 2.5 mg/kg. C. immitis is less susceptible to the killing effects of amphotericin B; the MFC₉₀ (11 μg/mL) is likely to be reached on day 1 of dosing at 2.5 mg/kg, but 30 days of dosing will be required to reach the MFC₉₀ or the 12-hour PAFE. These calculations were based on the MIC₉₀ of the respective organisms; if the MIC of the infecting microbe is known, then a lower dose might be reasonable.

Resistance

The incidence of resistance to amphotericin B is low and has been documented primarily for Candida. The development of resistance may be related to changes in the number of sterol components in the fungal cell wall, with low ergosterol content associated with resistance.³ Pythium insidiosum, classified as a pseudofungus, (the cause of what was previously and inappropriate referred to as fungal phycomycosis), as with other oomycetes, is largely devoid of cell membrane sterol. As such, they are inherently resistant to polyenes.¹ However, other mechanisms of resistance also exist, probably reflecting multiple mechanisms. Some organisms are able to resist concentrations that exceed 2 μg/mL. The azole derivatives may contribute to amphotericin B resistance by preventing the formation of ergosterol, the target of amphotericin B.¹

Amphotericin appears to act synergistically with the following: 5-flucytosine against cryptococcosis, tetracyclines against coccidioidomycosis, and imidazoles (discussed later) for a variety of fungal disorders. The use of synergistic combinations may enhance efficacy while reducing the potential of toxicity.

Preparations

Because amphotericin is unstable, it is prepared with desoxycholate as a lyophilized cake form (amphotericin B; previously Fungizone®; AmBD). Supplies were limited at the time of publication,⁸ perhaps reflecting declining use in human medicine in favor of newer lipid-based formulations.

Desoxycholate—also referred to as deoxycholate, a bile salt—is added to aid solubilization of the lyophilized cake form of the drug, which, when reconstituted, is in a colloidal suspension. Reconstitution should be with sterile water only; the solution will remain stable for 1 week if refrigerated. Storage should include protection from light, although protection during therapy is probably not necessary; potency is unaffected after light exposure of up to 24 hours. Labeled directions indicate that dilution should occur only with 5% dextrose to prevent drug precipitation and inactivation; Trissel’s Handbook on Injectable Drugs (http://online.statref.com/titles/titleinfopage.aspx?titleid=141) indicates that normal saline is incompatible with amphotericin B, leading to a 43% drop in drug concentrations within 2 hours. Although the drug has been mixed by veterinarians with 0.45% NaCl and 2.5% dextrose with no apparent change, the basis for this observation is unclear, and the prudent clinician would avoid the combination and provide sodium loading (see the section on therapeutic use) through a different catheter. In general, amphotericin B should not be mixed with other drugs; although known to be compatible with some, it is incompatible with many others. The drug should be administered only intravenously, with exceptions for localized treatment in selected body tissues or fluids (e.g., aqueous humor, cerebrospinal fluid [CSF]). For small animals, which do not require a complete vial, the reconstituted drug can be divided into smaller aliquots and frozen.

Amphotericin also has been complexed to lipid mixtures in an attempt to reduce nephrotoxicity. Reticuloendothelial cells phagocytize the lipid component, facilitating directed delivery to the site of fungal infection. Drug uptake by the hepatic and splenic macrophages, bone marrow, and inflammatory tissues is thus enhanced. For liposomal products, some studies have shown that amphotericin B is selectively transferred from liposomes to fungal but not host cell membranes. Prolonged antifungal activity (compared with nonliposomal preparations) has been documented for these preparations. Each product is injectable and differs in the lipid makeup as well as the ratio of lipid to amphotericin B. Lipid-based products include a lipid complex (ABLC; Abelcet) suspension that forms a tightly packed ribbonlike structure and is the largest of the lipid-based products (250 nm); a colloidal dispersion of cholesterol sulfate, which forms 122 nm disks (Amphotec or Amphocil [ABCD]); and a single-layered liposomal product, Ambisome (LAmB; see Table 9-4), which is the simplest and smallest (60 to 70 nm) of the molecules.^17,18 The differences among these products have recently been reviewed.¹⁹ Amphotericin B also has been administered with a lipid vehicle (intralipid; AMB-IL) rather than 5% dextrose. Although nephrotoxocity appears to be reduced in some patients, it does not appear to be decreased in the patients at most risk for toxicity, and therefore liposomal lipid-based products should be considered.^18a Administration of AMB-IL also is not recommended because of mixed product is unstable. The differences among the lipid-based or lipid-containing products and their clinical use in humans have recently been reviewed.¹⁹

Amphotericin is also available as a topical preparation (Fungizone), including cream, lotion, and ointment forms.

Pharmacokinetics

Amphotericin is not water soluble and thus is not bioavailable after oral administration. It is more than 90% bound to circulating serum lipoproteins (including cholesterol). Penetration into the pleura, peritoneum, inflamed tissues, CSF, and aqueous humor may result in a drug concentration two thirds of that in the plasma. The metabolism and excretion of amphotericin is not well characterized and is complex, being complicated by binding to cholesterol, which is structurally similar to ergosterol (see Figure 9-3). Biliary elimination may be the primary method of excretion (in humans only 3% of the drug is eliminated in the urine); however, concentrations are detectable in both bile and urine for up to 7 weeks in human patients.

The pharmacokinetics of the various amphotericin B preparations, including those complexed to lipids, vary markedly in humans; interpretation of kinetics is complicated by the lack of discrimination between free amphotericin B and that complexed to the various carriers. Because of its small size, Ambisome is characterized by the slowest uptake, and the highest PDCs. In humans, peak PDCs (μg/mL) vary with the dose (indicated in mg/kg), reflecting differences in volume of distribution that vary on account of tissue uptake: AmBD, 2.9 μg/mL at 1 mg/kg); Albecet (ABLC), 1.7 μg/mL at 5 mg/kg); Amphotec (ABCD), 3.1 μg/mL at 5 mg/kg; and Ambisome (LAmB), 83 μg/mL at 5 mg/kg. For LAmB, concentrations appear to be both dose and time dependent in dogs (Table 9-4). An advantage of the liposomal product is that it accumulates at sites of infection and targets the fungal cell wall, with release of amphotericin into the cell.¹⁹ For ABLC (and LAmB), tissue or fungal phospholipases appear to release amphotericin B at the site of infection. However, the colloidal form of amphotericin B (Amphocil), which is taken up by reticuloendothelial cells, was demonstrated in humans to accumulate in lung tissues to a greater concentration than Ambisome, suggesting that it might be preferable for respiratory infections.²⁰ This finding requires further support by animal model or clinical studies. That LAmB may be preferred for central nervous system (CNS) infections is supported by the study and review of Ibrahim and coworkers,²¹ who address the potential inability of ABLC to penetrate the brain in mice or rabbit models. Comparing dosing ranges, LAmB was effective without nephrotoxicity at 5 to 20 mg/kg in animal models of disease, with highest tolerated doses being 30 to 50 mg/kg (as reviewed by Adler).¹⁹ Although ABLC was efficacious at doses ranging from 5 to 15 mg/kg, nephrotoxicity and decreased efficacy occurred at 15 mg/kg compared to 5 mg/kg. (as reviewed by Adler).¹⁹

The pharmacokinetics of Ambisone (LAmB) have been described in Beagles (n = 10 per group) as part of a toxicity study in humans and compared historically to AmBD (see Table 9-4).²² Dogs received 0.25 to 16 mg/kg/day as an intravenous infusion over 5 minutes, with PDCs determined on days 1, 14, and 30. Dogs receiving 8 and 16 mg/kg daily did not finish the treatment period because of severe weight loss (n = 17/20); this same pattern of weight loss was also described in dogs receiving AmBD at 0.75 mg/kg every other day (as reviewed by Bekersky and coworkers²²). However, dogs tolerated 4 mg/kg Ambisone daily with peak concentrations at this dose were markedly variable, ranging from 18 µg/mL at day 1 to 94 μg/mL at day 30. Concentrations of Ambisone were a hundredfold higher than other formulations: A dose of 0.6 to 0.75 of AmBD resulted in peak amphotericin concentrations of 1.25 to 4.4 μg/mL after multiple dosing. This compares to a peak concentration of 54 ± 16 μg/mL at 4 mg/kg daily. Ambisome kinetics were nonlinear, with clearance and distribution decreasing as dose increased, potentially reflecting saturation of distribution and clearance processes. For example, although dose only increased 64-fold in the study, area under the curve (AUC) on day 1 at 0.25 mg/kg was 2.6 μg^∗hr/mL (compared) to 2592 μg hr/mL at 16 mg/kg, representing a 2500-fold increase. Multiple dosing also resulted in accumulation at each dose, with 30-day to 1 day AUC ratios being 2.5 and 10 at low (0.25 mg/kg) and intermediate (4 and 8 mg/kg) doses, respectively. (see Table 9-4). Kinetics are likely to be equally complex in other dogs and cats, and lack of predictability may limit use.

Toxicities and Side Effects

Nephrotoxicity is the major side effect associated with the use of amphotericin B. Renal function becomes impaired in more than 80% of patients receiving amphotericin,^4,23,24 with serum creatinine at least doubling in 40% to 60% in human.²³ Although largely reversible, renal toxicity depends on total cumulative dose and duration of therapy. In human patients chronic renal disease occurs in close to 50% of patients receiving more than approximately 60 mg/kg total dose compared with only 17% who receive less than 57 mg/kg. The proportion decreases to 8% in those receiving less than 14 mg/kg total dose.²³ Although renal function usually returns to normal before completion of therapy, some residual damage often persists after discontinuation of the drug. Two mechanisms are important in renal toxicity. Intense arterial vasoconstriction occurs within 15 minutes of administration and lasts 4 to 6 hours. The mechanism is unknown but can lead to acute tubular nephrosis secondary to ischemia. Nerve degeneration, angiotensin II receptor blockage, adrenergic blockade, and potent vasodilators do not prevent renal vasoconstrictive effects. Amphotericin B may activate formation of vasoconstrictive (thromboxane) arachidonic acid metabolites, which suggests that direct vasoconstriction may be responsible. Amphotericin B can increase calcium fluxes. Histologic lesions are most profound in regions vulnerable to hypoxia or areas rich in oxygen.²³

Distal renal tubular toxic effects result from binding of membrane cholesterol in the tubular cell membrane (see Figure 9-4). Altered electrolyte fluxes result in acidification abnormalities (metabolic acidosis), hypokalemia, and concentrating defects (polyuria, polydipsia). Renal tubular acidosis is common, dose related, and generally reversible (although resolution may take several months) and generally precedes significant decreases in glomerular filtration rate (GFR). Potassium and magnesium wasting in humans can lead to substantial deficits.²³ Magnesium deficits appear at 2 weeks and are maximal at 4 weeks. Both potassium and magnesium should be monitored; electrolyte abnormalities may persist for several weeks after therapy is discontinued. Rarely, hyperkalemia has been reported in association with rapid infusion as potassium shifts from the intracellular compartments. The risk is greater in patients with renal failure. Concentration defects occur in essentially all human patients, are not related to azotemia, and may persist for months. Urine specific gravity may not be an effective measure by which to assess renal damage.

Ampohtericin B toxicity has been documented in dogs. Ceylan and colleagues²⁴ described the nephrotoxicity of AmBD in dogs (n = 18; cross-bred) receiving 0.5 mg/kg in 25 mL 5% dextrose in water as a 4- to 5-minute intravenous bolus; 1 mg/kg in 50 mL as a 4- to 5-minute bolus, or 2 mg/kg in 1000 mL over 4 to 5 hours. Dogs were treated for 12 days. Side effects (vomiting, diarrhea, anorexia, fever, tachycardia, and phlebitis) were evident in all dogs by day 3 but were worse in dogs receiving 1 mg/kg and least in dogs receiving 2 mg/kg. Indicators of renal damage (blood urea nitrogen, creatinine) were increased in all groups by day 5 but were highest in the 1 mg/kg group. Serum calcium was significantly decreased compared with baseline in the 1 mg/kg and 2 mg/kg groups. Hematologic indicators of anemia were evident in the 1 mg/kg group. Among the indicators of nephrotoxicity studied was urine γ-GGT, which was significantly higher than baseline in all groups by day 5 of therapy but did not differ among groups or times during treatment. On the basis of this study, the authors suggested that infusion over a 4- to 5-hour period (or fluid support) decreases the risk of aminoglycoside toxicity. However, no study has determined the long-term effects of any method of administration of amphotericin B on renal failure in dogs or cats.

In their pharmacokinetic study of LAmB (Ambisome) in Beagles at doses ranging from 0.25 to 16 mg/kg daily for 30 days, Bekersky and coworkers²² reported that no dogs died as a result of the drug; however, dose-dependent tubular nephrosis was present at 1 mg/kg, and 70% and 100% of dogs in the 8 and 16 mg/kg treatment group, respectively, were euthanized before the end of the study because of 25% or more body weight loss. Vomiting and diarrhea were evident in all groups, including a liposomal (i.e., without amphotericin B) control, but were worse in the 4, 8, and 16 mg/kg treatment groups. Azotemia was not present in the 1 mg/kg group. Azotemia was described as moderate and clinically significant toward the end of the 30-day treatment period in the 4 mg/kg group, with creatinine increasing from a baseline of 0.7.1 ± to 2.3 ± 0.4 mg/dl by study end. Serum potassium was not clinically affected. The authors concluded that dogs tolerated up to 4 mg/kg well, despite peak amphotericin B concentrations ranging from 18 to 94 μg/mL.

Acute anaphylactic-type reactions such as vomiting, fever, and chills can occur with the use of amphotericin B and have been reported with lipid complex preparations as well.²⁵ Up to 30% of dogs receiving AmBD develop fever.²⁶ The frequent incidence of these reactions often leads to pretreatment for anaphylaxis (one-time use of a short-acting glucocorticoid does not enhance the toxicity of amphotericin B; see the discussion of “cocktail” in the following section).

Other side effects associated with amphotericin B include nausea and anorexia, thrombophlebitis, cardiac arrhythmias and related toxicities, hepatic dysfunction, and CNS signs (if given intrathecally) (see discussion on therapeutic use). Several side effects can be prevented by proper treatment (see following discussion of therapeutic use).

Drug Interactions

Amphotericin B interacts in an additive to synergistic fashion with a number of antifungal agents (see the discussion of combination therapy), although this effect seems to be most consistent with flucytosine. Although synergistic effects should be expected with azoles, this combination may be antagonistic if azole therapy is begun first.

In general, drugs that are renally active are not recommended for patients also receiving amphotericin B. In humans cyclosporine and tacrolimus (calcineurin inhibitors) will enhance nephrotoxicity. However, these immunomodulators tend to be safer, as far as nephrotoxicity is concerned, in both dogs and cats, and the combination may not be as dangerous. The nephrotoxicity of amphotericin also may be enhanced with catabolic drugs such as glucocorticosteroids, antineoplastic drugs, antiprostaglandins (glucocorticoids and nonsteroidal antiinflammatory drugs), and other nephrotoxic antibiotics. One study demonstrated that diuretic therapy during, but not just before, amphotericin B therapy increased the risk of nephrotoxicity more than twelvefold (as reviewed by Bagnis and Deray²³). Although diuretics have been used for a “protective” effect, the potential benefits are limited to mannitol, and even it may be associated with electrolyte disturbances. Combination with digoxin may enhance nephrotoxicity and hypokalemia. Declining renal function in association with amphotericin B therapy may affect clearance of co-administered drugs. Because it is renally excreted, 5-flucytosine toxicity may be enhanced if renal function declines with amphotericin B.

Therapeutic Use

Most of the advice and studies addressing the recommended method of administration of amphotericin B are based on the non-lipid preparation (AmBD).

Multiple strategies may help reduce toxicities or side effects to amphotericin B. The first focuses on prevention of an anaphylactoid reaction by pretreatment (to prevent vomiting, fever, chills, and anaphylaxis) with antihistamines (diphenhydramine 0.5 mg/kg, administered intravenously) and short-acting glucocorticosteroids (e.g., hydrocortisone sodium succinate, 0.5 mg/kg, administered intravenously). Because the reaction appears to be associated with direct mast cell degranulation (because of the cationic nature of amphotericin B), pretreatment with a small test dose (0.1 mg/kg for cats or 0.25 mg/kg for dogs diluted in 10 mL infused over 15 minutes) may help identify animals that are likely to have an adverse reaction during infusion. Whereas lipid products may be characterized by less nephrotoxicity, they are more likely than nonlipid products to cause vomiting, nausea, and phlebitis.³⁸ As such, pretesting is indicated with their use as well.

In addition to dilution protocols (see discussion under Toxicity), strategies are intended to minimize amphotericin-B induced nephrotoxicity. Although what constitutes amphotericin B nephrotoxicity may vary, an increase in creatinine of as little as 25% may be considered significant in some patients.¹⁸ Azotemia precedes tubular dysfunction in humans, allowing for early detection (i.e., before irreversible damage). Urine sediment initially should be monitored for evidence of nephrotoxicity. Serum chemistries tend to be less sensitive indicators of nephrotoxicity. Urine γ-gt has been used to assess lipid-based amphotericin B toxicity. The drug should be temporarily discontinued (24 to 48 hours) if the blood urea nitrogen level becomes abnormal.²⁷

First and foremost, pretreatment with sodium-containing fluids is particularly important for preventing renal toxicity, including that associated with renal arterial vasoconstriction; posttherapy treatment might also be prudent. Secondly, amphotericin B can be administered with a “cocktail” intended to protect the kidney. Administration of the dose diluted in 5% dextrose is accompanied by mannitol (0.5 mg/kg) to maintain glomerular filtration rate and sodium bicarbonate (1 to 2 mg/kg) to prevent cellular acidification defects. The mannitol and sodium bicarbonate should not be added directly to the amphotericin B solution but given through another catheter in order to avoid precipitation of amphotericin B. The ability of cocktails to prevent nephrotoxicity is controversial. The treatment is based on one small study in dogs³¹ for which mannitol was demonstrated to protect the kidney. However, a subsequent study (using isolated perfused rat kidneys) found mannitol to impart a protective effect, presumably by reducing renal tubular edema by virtues of its osmotic effect.³² Binding of cellular cholesterol prevented toxicity, supporting the presumed mechanism of direct cytotoxicty. This study also demonstrated a synergistic toxic effect of amphotericin B in the presence of hypoxia. Only one clinical trial, in humans, has addressed renoprotection by mannitol, and it failed to identify an effect. However, this clinical trial, although randomized, involved only 11 patients and is probably not conclusive (as reviewed by Bagnis and Deray^18,23) Although such cocktails generally are not harmful as long as the solutions are not mixed with amphotericin B, mannitol has been associated with electrolyte abnormalities in the human-medicine literature.²³ Deray¹⁸ indicates that mannitol decreases renal medullary blood flow and PO_2. As such, the use of mannitol remains controversial. A relatively recent study describes the nephroprotective effects of N-acetylcysteine (10 mg/kg daily) in a rat model of amphotericin-induced nephrotoxicity;³³ administration began 1 day before amphotericin was started. As previously described, amphotericin should not be mixed with solutions containing electrolytes, acidic solutions, or preservatives because these materials may cause drug precipitation. The rate at which amphotericin B is administered may also reduce the incidence of nephrotoxicity.

The third strategy by which amphotericin B toxicity might be reduced is administration with lipids or as a specialized delivery system (in lipids). Conventional amphotericin B is not recommended in humans in the presence of renal insufficiency, hypokalemia, hypomagnesemia, tubular acidoses, or polyuria.¹⁸ Lipid preparations are designed to deliver more drug selectively to the site of infection,³⁴ thus allowing higher doses (see the section on preparations). The liposomal products tend to be very expensive. In contrast to liposomes, fat emulsions are easy to prepare and administer and tend to be more cost effective. These products appear to be equal in efficacy to nonliposomal products but safer with regard to nephrotoxicity. However, the use of intralipid as the vehicle (AmB-IL) is associated with other difficulties that limit its use (see previous discussion).

The safety of liposomal or lipid-based products may allow delivery of higher doses without an increased risk of nephrotoxicity, with the incidence of nephrotoxicity reduced by 8% to 28%, depending on the preparation.^18,23 Although several studies are available that address the safety of these products individually, and LAmB (Ambisome) in particular,³⁵ none appears to have compared all three products. However, in general, the liposomal product LAmB (Ambisome) is recognized to be the safest with regard to nephrotoxicity. Ambisome achieves the highest concentrations in humans compared with other amphotericin preparations but is well tolerated renally.^17,19 In general, efficacy of LAmB is equal to or exceeds that of AmBD and is consistently better tolerated. Survival rates in humans for the product ranges from 99% to 100% (visceral leishmaniasis; three studies, n ≅ 400), 37% to 78% (aspergillosis; five studies, n ≅ 400), 84% to 93% for cryptococcosis (two studies, n ≅ 280), 98% for histoplasmosis (one study, n = 55), and 20% to 69% for zygomycosis (three studies, n ≅ 130).³⁵ That LAmB may be preferred for CNS infections is supported by the study and review of Ibrahim and coworkers,²¹ who found LAmB generally superior to ABLC in treating CNS infections, as indicated by response and decrease in fungal load. Abelcet (ABLC) and LamB were generally characterized as having the lowest incidence of all side effects (fever, chills, nausea, dyspnea, hypertension, hypotension, tachycardia, and nephrotoxicity) with the exception of hypokalemia (42% with LamB). Ambisome was associated with abnormal hepatic function tests in approximately 20% of human patients.¹⁷ Abelcet causes fewer infusion-related side effects in humans, and therefore a test dose is not necessarily indicated, whereas Amphotec is more likely to cause infusion side effects and should be given as a short infusion only.

One study reports the efficacy and safety of a liposomal product when used at higher than recommended cumulative doses for treatment of canine blastomycosis.³⁶ In another study, although the products were equally efficacious, the degree of nephrotoxicity between a fat emulsion and standard amphotericin B was no different.³⁷ Amphotericin B also has been administered with a lipid vehicle (intralipid) rather than 5% dextrose. Although nephrotoxocity appears to be reduced in some patients, it does not appear to be decreased in the patients at most risk for toxicity, and therefore liposomal lipid-based products should be considered.¹⁸ Further studies documenting the efficacy and safety of liposomal or fat emulsion products containing amphotericin B are needed.

A fourth strategy that can be used to minimize adverse events associated with amphotericin B is administration using alternative routes. Localized mycotic infections have been treated with localized administration of amphotericin, which may reduce the incidence of nephrotoxicity. Subconjunctival, intravitreal, intrathecal, intranasal (human aspergillosis: 5 mg/mL in water administered as aerosol), and intraperitoneal routes have been reported. Oral administration has been used for treatment of gastrointestinal candidiasis and presumably might be used for other gastrointestinal fungal disorders.²⁹ Both LamB and ABLC have been demonstrated to have enhanced efficacy when administered as an aerosol for pulmonary infections (as reviewed by Adler).¹⁹ Amphotericin B (AmBD) can be mixed in sterile water to 200 mg/kg and infused into the bladder for fungal cystitis. For fungal infections of the CNS, the drug can be given (0.2 to 0.5 mg in either 5 mL of CSF or 10% dextrose) intrathecally (under general anesthesia) two to three times per week.²⁷

Combination antifungal therapy is a fifth stategy that is strongly encouraged to enhance efficacy and thus decrease the duration of antifungal exposure to the host. For example, as reviewed by Adler,¹⁹ efficacy of LamB is enhanced when combined with micafungin (zygomycosis, aspergillosis) and voriconazole (aspergillosis).

Although a sixth strategy whereby amphotericin B toxicity can reduced includes variation in doses, frequency of administration, concomitant therapy, and duration of therapy, no protocol has proven to be superior to others. Some literature supports rapid intravenous administration (bolus) in less debilitated dogs; slow intravenous administration might be more prudent in cats. A dose (0.25 to 0.5 mg/kg) can be diluted in 300 to 1000 mL of 5% dextrose and administered in an indwelling catheter over 2 to 6 hours, or it can be diluted in as little as 10 to 60 mL (30 recommended) and given over 2 to 10 minutes through a butterfly catheter²⁷ (flush catheter after infusion). The slow infusion method has the added advantage of additional fluids, which may reduce the incidence of nephrotoxicity,²⁸ especially in debilitated animals. The advantages of dilution and slow infusion is preferred and was previously discussed (see Toxicity). It is recommended that the small bolus be preceded or followed up with supplemental fluid, preferably normal saline. One report of an uncontrolled clinical trial describes the successful and apparently safer administration of amphotericin B after twice- or thrice-weekly subcutaneous administration (0.5 to 0.8 mg/kg diluted in 400 to 500 mL of fluid such that amphotericin B is less than 20 mg/L) for several months for treatment of cryptococcosis in dogs and cats.³⁰ However, only five successful cases were described, and further confirmation should be expected before this route is routinely embraced.

The sequence of repetitive treatments is also controversial. Some authors recommend alternate-day therapy, whereas others recommend daily therapy at a smaller dose. Doses also vary. Daily doses range from 0.15 to 0.5 mg/kg every other day (e.g., on Monday, Wednesday, Friday) until a cumulative dose of 4 to 12 mg/kg (depending on the organism or if therapy is combined with another antifungal) AmBD has been reached. Starting at a low dose (0.15 to 0.25 mg/kg) and gradually increasing the dose until the desired daily dose has been reached may reduce the severity of side effects. For particularly resistant infections, a dose of 1 mg/kg has been used on an alternate-day basis.

Other Polyenes

The spectrums of piramicin (natamycin) and nystatin are similar to that of amphotericin B. Pirimacin is used primarily to treat fungal keratitis, although efficacy toward Aspergillus may be questionable.³ Aerosolization of piramycin for treatment of susceptible fungal disorders also should be considered. The toxicity of nystatin precludes parenteral administration, although a liposomal product is being formulated. Currently, use of nystatin is limited to superficial and mucosal mycoses.

Structure–Activity Relationship

The azole derivatives (imidazoles and triazoles) include a large number of predominantly synthetic drugs. These drugs consist of a five-member ring with other aromatic rings attached by a carbon nitrogen bond. Imidazoles contain two nitrogen atoms and include clotrimazole, econazole, enilconazole, miconazole, and ketoconazole. Triazoles contain three nitrogen atoms and include fluconazole, itraconazole, and voriconazole (see Figure 9-1).^4,39-41 Among the newer azoles are posaconazole and voriconazole. Voriconazole, a synthetic deriviative of fluconazole, is the first of the second-generation triazole compounds to be approved by the Food and Drug Administration. Voriconazole contains a fluorine molecule and a methyl group, which greatly enhances its spectrum compared with that of fluconazole.⁴² Other triazoles have been patented. Most compounds generally are not available as solutions because they tend to be insoluble in water (an exception is fluconazole). They are, however, soluble in organic solvents such as propylene glycol.

Mechanism of Action

The azoles target fungal sterol ergosterol. However, in contrast to amphotericin B, the imidazole derivatives do not bind to ergosterol but rather block its synthesis. The azoles inhibit fungal cytochrome P450 enzymes with sterol14α-demethylase as the primary fungal target; however, other synthesizing enzymes also are targeted.³ Decreased cell membrane ergosterol alters cell membrane function but also is accompanied by an increase in 14-methylsterols, compounds that are potentially toxic. Cell membrane function fluidity decreases as cell permeability increases, resulting in a fungistatic effect (fluconazole and ketoconazole). At higher concentrations, selected drugs (miconazole, econazole, clotrimazole) also interfere with cell membrane fluidity and physiochemical intracellular processes (e.g., secretory vesicles, mitochondrial respiration), resulting in fungicidal effects. Efficacy of selected agents against gram-positive organisms may reflect this latter effect.³ Chitin synthesis increases in concert with decreased ergosterol synthesis, but its irregular distribution contributes to altered cell wall function. Because azoles also generate and detoxify intracellular hydrogen peroxide, selected drugs also express antibacterial, antiprotozoal, and anthelmintic activities. The imidazoles are also characterized by immunomodulatory effects, which may facilitate effective therapy.⁵ Because their mechanism of action depends on cell wall synthesis, the onset of action of the imidazoles may result in a lag time to therapeutic efficacy. In addition, a long elimination half-life of some members of this class (e.g., itraconazole) results in a lag time as steady-state concentrations are achieved.

Spectrum of Activity and Pharmacodynamics

Although the imidazole derivatives are more selective in their cellular activity than amphotericin B (i.e., impairing the synthesis of rather than binding to ergosterol), their spectrum of activity is broad and includes the dermatophytes (“ringworm”: Microsporum and Trichophyton species), yeasts, dimorphic fungi (blastomycosis, histoplasmosis, cryptococcosis, coccidioidomycosis), Eumycetes, Actinomyces, and some Phycomycetes.^{1,4,11,27,40,43} The efficacy against these organisms varies. Studies comparing the efficacies of the azoles in animals are limited at the time of this publication, although several are pending in the human-medicine literature.

In vitro MIC data regarding the relative susceptibility of selected organisms to itraconazole emphasize the variable susceptibility of fungal organisms to these drugs. Table 9-2 provides MIC₉₀ data for selected organisms and itraconazole. For dermatophytes, the MIC₉₀ for Microsporum species is 250 μg/mL versus for Trichophyton species 4 μg/mL. For other Eumycetes, MIC₉₀ concentrations range from 0.130 μg/mL to more than 128 μg/mL.^11,44 Fluconazole generally is considered to be fungistatic in action. This has been demonstrated for Candida sp. and C. neoformans.¹⁰ However, more recent data suggest that fungicidal activity may occur toward Candida. Like fluconazole, itraconazole more consistently exerts a time-dependent, fungistatic effect toward most organisms. The MFC₅₀ and MFC₉₀ (μg/mL) have been reported for B. dermatitidis (0.125, 3, respectively), H. capsulatum (2, 16), and C. immitis (>16 for both).¹⁵ However, it may exert fungicidal effects towards Aspergillus. The long half-lives of the imidazoles will facilitate convenience of dosing despite their time-dependent killing effects. Efficacy of itraconazole will be enhanced by the formation of its active metabolite, hydroxyl-itraconazole, which may surpass the parent.⁴⁵ As with other azoles, the spectrum of the third-generation imidazoles, posaconazole (similar in structure to itraconazole) and voriconazole (similar in structure to fluconazole), includes a variety of infecting fungal organisms. Their spectrums are very similar.¹⁰ Posaconazole has been demonstrated to be more potent than fluconazole toward Candida and should be effective against the dimorphic fungal organisms,⁴⁶ although little information is available regarding its use. As with other azoles, voriconazole is fungistatic toward yeasts, exhibiting time-dependent killing, but for some filamentous organisms may be -cidal. Its spectrum includes Candida sp., with MIC generally being 1 to 2 log lower than fluconazole; however, MICs are higher for fluconazole-resistant strains than for nonresistant strains. Voriconazole also is effective against C. neoformans, Trichosporon beigelii, and Saccharomyces cerevisiae. Additionally, it is very effective against Aspergillus sp., including some strains resistant to amphotericin B; indeed, it is approved for the treatment of Aspergillosis in humans. Time-killing studies with Aspergillus demonstrate that, in general, amphotericin B is more efficacious but itraconazole less efficacious than voriconazole. Activity against B. dermatitidis, C. immitis, and H. capsulatum appears to be “reasonable” but “less” toward S. schenckii. Li¹⁵ compared the MICs of amphotericin B, itraconazole, and voriconazole and found voriconazole to be more active in vitro than amphotericin B for the mold forms of H. capsulatum, B. dermatitidis, and C. immitis, with activity being fungicidal (MFC₅₀, MFC₉₀, respectively) toward B. dermatitidis (0.125, 4) and some H. capsulatum (8, ≥ 32) (see Table 9-2). Fungicidal activity toward C. immitis is difficult to achieve (>32 for the MFC₅₀). Activity Aspergillus also is fungicidal (MFC not provided) toward several dematiaceous molds.¹⁵ Many dematiaceous and hyaline molds resistant to amphotericin B (e.g., Scedosporium, Fusarium, Paecilomyces, Alternaria) are susceptible to voriconazole. However, zygomycetes are not susceptible.⁴² Cutaneous infections by Leishmania species are clinically susceptible to ketoconazole.⁴⁷ Clotrimazole and miconazole are common drugs used topically for treatment of dermatophytosis (e.g., Conofite) or yeast (e.g., otic preparations such as Otomax). Both drugs, as well as other imidazoles (e.g,) appear to exhibit efficacy against gram-positive organisms including staphylococci and anaerobes.

Resistance

Resistance to the azoles has been sporadic, generally occurring in immunocompromised patients receiving long-term therapy. Failed drug accumulation in the cell is a major mechanism of resistance (e.g., Candida sp., C. neoformans, Aspergillus flavus, and A. fumigatus), caused by either decreased influx or formation of efflux proteins. Two major efflux transport proteins have been identified, particularly for Candida sp. toward fluconazole: the multidrug-resistant protein, major facilitator superfamily (MFS; a proton-motive-force–based mechanism; e.g., CaMDR1) and the ATP-binding cassette (e.g., CDR1 and 2). Resistance to azoles also may reflect altered interaction between the drug and targeted fungal CYP 450 enzyme; fluconazole appears more susceptible than itraconazole to mutations that alter drug-receptor fit. Gene amplification leading to enhanced synthesis of the target protein also has been identified as a mechanism of resistance to azole derivatives. Finally, fungal organisms may circumvent the synthetic pathway inhibited by azoles or compensate for altered enzyme activity; for example, inhibition of ergosterol may result in the accumulation of 14-methylated sterols that are less toxic than other compounds.³ Resistance to fluconazole appears to be increasing. Widespread use of fluconazole for treatment of Candida has been associated with increased resistance of the drug for selected species. Whereas many molds (e.g., A. fumigatus) are inherently resistant to fluconazole,³ resistance of Aspergillus isolates to itraconazole is rare. Combinations with nonantagonistic antifungal agents might be considered for treatment of infections by organisms associated with antifungal resistance.

Pharmacokinetics

Only selected pharmacokinetic information is available for the imidazoles in dogs or cats (Table 9-5).Oral absorption of the imidazole derivatives varies with the drug and among animals. For many drugs oral preparations are not available. For example, enilconazole (imazalil) is not orally bioavailable. For other imidazoles the rate of absorption varies from 1 to 4 hours. Oral absorption often depends on gastric pH, product preparation, and the presence of other drugs.⁴⁸ Absorption of itraconazole and ketoconazole is enhanced by gastric acidity; these drugs should be administered with food.⁴⁹ Alkalinizing drugs administered orally will decrease their absorption. Cyclodextrins are used to form complexes with the drugs, thus rendering the liphophilic drug soluble in solution, making oral solutions available for some drugs (e.g., Sporonix solution). In general, oral bioavailability should be anticipated to be better for these solutions than for capsules. Peak PDCs of itraconazole occur between 1 to 5 hours in cats and dogs. Bioavailability of capsules is approximately 20% in dogs and may be as little as 10% in cats, compared with close to 50% for the solution in cats and dogs.^50,51 Decreased bioavailability may be responsible for therapeutic failure associated with low PDCs in some animals (cats and dogs). Unpredictable oral administration of itraconazole prepared as a capsule limits its usefulness in humans; the solution, on the other hand, generates more predictable concentrations.¹⁷ Care should be taken with compounded preparations; oral absorption generally is not verified for these products, and adequate oral bioavailability should not be assumed. Fluconazole is characterized by the best oral bioavailability among the imidazoles, being completely absorbed in cats;⁵² however, its efficacy, compared with that of itraconazole, is limited.

Table 9-5 Pharmacokinetic Data for Selected Imidazoles in the Dog

Distribution to tissues also varies among the imidazoles. Ketoconazole is up to 99% protein bound; the highest tissue levels occur in the liver, lung, and kidney (and cerumen). Itraconazole is also very highly protein bound in humans.⁴ There is minimal penetration of the CSF by ketoconazole, although fluconazole penetrates the CSF well, with serum to CSF PDCs ranging from 0.58 to 0.89 μg/mL. The volume of distribution of ketoconazole is only 0.87 L/kg in dogs compared with 17 L/kg 5 L/kg for itraconazole in dogs⁵¹ and cats, respectively.⁵⁰ The volume of distribution of fluconazole is 1.14 L/kg in cats, with high concentrations occurring in the CSF and aqueous humor.⁵² The difference in distribution volume reflects, in part, distribution and accumulation to fat.⁵¹ Drug concentrations of itraconazole in the skin may exceed that in plasma by threefold to tenfold, with drug detectable 2 to 4 weeks after therapy is discontinued.⁵¹ Although distribution of itraconazole to the CSF appears to be limited, therapeutic concentrations appear to be achieved in patients suffering from cryptococcal meningitis. Among the azole derivatives, fluconazole has the best tissue distribution pattern and can achieve effective concentrations in CSF.

With the exception of fluconazole, the azole derivatives are eliminated by extensive oxidative (cytochrome P450) metabolism with excretion as inactive metabolites into the bile and urine. However, metabolism of itraconazole generates an active hydroxylated metabolite, whose AUC may exceed that of the parent compound.⁵³ Metabolism may be dose dependent; elimination rate constants are lower and half-lives are longer at higher doses and with longer therapy. In contrast to the other imidazoles, fluconazole is eliminated principally (70%) in the urine. The half-life of the imidazoles varies, with that of ketoconazole being relatively short (1.4 hours in dogs). Fluconazole and itraconazole have longer half-lives, ranging from 22 to 32 hours in humans. The half-life of fluconazole in cats is 25 hours.⁵² The half-life of itraconazole in dogs is 51 hours (itraconazole)⁵² versus 40 to 70 hours in cats (itraconazole);⁵⁰ The longer drug elimination half-life must be taken into account because it results in a longer time to steady-state concentration and maximum therapeutic effect. However, it also allows the flexibility of once-daily (10 mg/kg) rather than twice-daily (5 mg/kg) dosing.

The disposition of voriconazole is complicated. Its disposition has been reported in dogs as part of a preclinical study (see Table 9-5).⁵⁵ Beagles (n = 4) received multiple doses (8) of 6 mg/kg orally or 3 mg/kg intravenously using a crossover design; disposition was studied on day 1 and day 30 for each route. The AUC after single and multiple dosing (μg∗h/mL) after the intravenous dose was 32 and 18, respectively, and after the oral dose 89 and 52, respectively, suggesting autoinduction. Other relevant parameters after single oral dosing were as follows: C_max, 6.5 μg/mL at T_max of 3 hours and apparent bioavailability of 138%. The drug was 51% protein bound, indicating a concentration of approximately 3.25 μg/mL of active drug. For a safety study, Beagles (n = 6) also received oral doses of 3, 6, or 12 mg/kg for 1 month. The apparent volume of distribution was 1.3 L/kg and clearance was 24 mL/min/kg, resulting in a calculated half-life of 6.3 hours. The C_max increased in a slightly disproportionate dose-dependent manner, being 1.69 ± 0.83 μg/mL at 3 mg/kg and 10.3 ± 5.1 μg/mL at 12 mg/kg 30 days after oral dosing. However, cytochrome P450 increased in a dose-related manner, resulting in a 1.7-fold increase in relative liver weight at the highest compared with the lowest dose. Autoinduction in dogs, which does not occur in humans, resulted in an increased clearance after multiple dosing, with these effects dissipating approximately 1 month after the drug was discontinued. For example, the mean C_max after a single oral dose of 3 mg/kg in dogs was 6.5 μg/mL but only 1.7 after 30 days dosing at the same dose. Although 55% to 87% of the drug ultimately was eliminated in the urine in dogs, only 5% was as the parent drug, indicating extensive hepatic metabolism. A major pathway of metabolism was generation of the N-oxide and hydroxylation metabolites as well as glucuronidation. The dose of voriconazole recommended for dogs can be based on pharmacokinetic–pharmacodynamic integration. The MIC₉₀ of most infecting microbes for which data are available will be achieved after 30 days at 3 mg/kg, even accounting for 50% protein binding in dogs. An exception occurs, however, for H. capsulatum, for which a higher dose is indicated. With a half-life of 6 hours, based on time-dependent killing, at least twice the MIC of the infecting microbe should be targeted to allow for a 12-hour dosing interval. In humans dosing regimens are designed to reach 3 to 6 μg/mL in the plasma; accordingly, a dose of 3 to 6 mg/kg twice daily is recommended. However, the half-life in humans is long (6 to 24 hours, depending on the dose), and steady-state concentration requires 5 to 6 days of dosing. As such, a loading dose consisting of a double daily dose is recommended for the first day of therapy. This is not necessary in dogs: the shorter half-life precludes accumulation to a steady state. However, the shorter half-life will necessitate twice-daily dosing. Because of autoinduction in dogs, dosing regimens should not be extrapolated for cats from dogs without the support of pharmacokinetic studies.

Voriconazole is cleared primarily by hepatic metabolism to inactive metabolites by CYP 2C19 (the primary isoenzyme), 2C9, and 3A4 being involved. Selected humans are considered “poor metabolizers” of the drug because of variation in CYP 2C19. In humans the dose is halved in the presence of mild to moderate liver disease. The relevance of this to dogs, which autoinduce, is not known.

Very limited pharmacokinetic information is available for dogs receiving posaconazole (see Table 9-5).⁵⁶ It was studied in two different vehicles (cyclodextrin or methylcellulose) during preclinical investigations. Absorption from the cyclodextrin vehicle was better, resulting in a higher C_max, greater AUC, and better oral bioavailability. Although the half-life following intravenous administration was approximately 8 hours, the effective half-life is 15 hours after oral administration, probably reflecting slow absorption, as is indicated by a T_max of 8 to 9 hours. Food enhanced absorption, increasing C_max and AUC fourfold. Although C_max increased more than two fold with multiple dosing, AUC was the same, indicating accumulation is not likely to be a clinical concern.

Preparations

Ketoconazole, itraconazole, and fluconazole are available for oral administration. Solutions are available for some products. Although their safety has not been documented for animals, fluconaozle (Diflucan) is available as an intravenous preparation that appears to be safe in cats when administered as a slow intravenous drip at 5 mg/kg. The solubility of imidazoles is poor and potentially toxic; solubilizing agents may cause adverse reactions. Cats also appear to tolerate a slow intravenous drip of itraconazole (5 mg/kg) with no adverse effects, although neither product is commercially available. Ketoconazole also is available in a topical preparation and a shampoo. Clotrimazole and miconazole are recommended only for localized dermatophyte or yeast infections susceptible to topical treatment. Clotrimazole has been used topically to treat nasal aspergillosis. Enilconazole is a topically effective azole that has been used to treat nasal aspergillosis but is available in the United States only as a 13.8% poultry dip. It is available in Canada as a 10% solution approved for use in dogs and horses. The poultry dip has been used topically in the United States at a dilution of 1:50 in water in dogs and cats with no apparent adverse effects. Terconazole is a new, topically active triazole that apparently has not yet been used for animals. However, an otic preparation containing posaconazole (with orbifloxacin and an anti-inflammatory) has recently been approved for dogs.

Drug Interactions

The azoles may interact synergistically with a number of antifungal agents. Synergism with polymyxin B is benefited in otic preprations.³ Ketoconazole and, presumably, other azole antifungals have synergistic antifungal activities with 5–flucytosine against Candida and Cryptococcus and with amphotericin against a variety of organisms. However, timing of amphotericin B and azole therapy is important. Azoles impair ergosterol synthesis, and therefore their use before amphotericin B may decrease its efficacy which is dependent on active cell wall synthesis. Amphotericin B should begin either simultaneously with (azoles are characterized by a lag time to effect) or before azole administration. Enhanced efficacy has also been demonstrated for terbenifine and topical therapy (see Therapeutic Use).

Because the efficacy of the azoles depends on interaction with P450 (an oxidative enzyme responsible for drug metabolism), drug interactions at the level of drug or steroid metabolism should be anticipated in the patient. The azoles are both inhibitors and inducers of CYP isoenzymes; the extent to which an azole inhibits metabolism often depends on its relative affinity for host compared to fungal CYP 450.^3,57 All of the clinically relevant imidazoles inhibit some CYP. These include: CYP3A4 (ketoconazole, itraconazole, fluconazole, voriconazole; (ketoconzole has a higher affinity than itraconazole); 2C19 and 2C9 (ketoconazole, fluconazole, voriconazole); 2D6 (ketoconazole); 2C8 (ketoconazole and voriconazole); and 1A2 and 2E1 (ketoconazole). However, as has been demonstrated in dogs, voriconazole is an autoinducer and species differences in CYP interactions may be profound. When present, inhibition can be clinically relevant. It can be beneficial (e.g., the combination of ketoconazole with cyclosporine in an attempt to prolong cyclosporine clearance in order to decrease the daily cost of this drug (see Chapter 31).⁵⁸ In the author’s laboratory, the effect of ketoconaozle on cyclosporine concentrations appears to be quite variable, ranging from no effect to a dramatic increase. Kukanich and Borum⁵⁹ found that ketoconazole (approximately 13 mg/kg for 5 days) did not appear to affect the intravenous disposition of morphine in Greyhounds.

However, more commonly, inhibition is associated with adverse reactions. The author is aware of two cases of marked increases in phenobarbital concentrations (to over 85 µg/mL) in epileptic dogs receiving fluconazole or itraconazole, respectively, for treatment of Malassezia; one of the dogs was euthanized resulting from (assumed) liver disease. More recently, a dog receiving deracoxib and also developed a perforated duodenal ulcer receiving ketaconazole for a skin yeast infection. Itraconazole appears to cause autoinhibition; changes in the elimination half-life of intractonaozle have been documented in cats receiving long-term therapy (>6 weeks).⁵⁰ In humans, the risk of drug interactions involving voriconazole is high, perhaps more so than with the other azoles. Their magnitude and impact in dogs and cats needs to be clarified. Clinically relevant drug interactions have been documented for a number of drugs administered in concert with voriconazole; avoidance of other drugs administered by the liver would be prudent.⁴²

Not all drug interactions involve inhibition of CYP. Clotrimazole is a potent inducer of CYP3A and miconazole of CYP 1A and 2E; even ketoconazole is an inducer of CYP 2, although it is less potent than clotrimazole (see Chapter 2 for drugs metabolized by CYP isozymes). As previously discussed, voriconazole is an inducer of CYP in dogs. Ketoconazole interferes with sex hormones and corticosteroids by displacing them from globulins and perhaps by interfering with their synthesis. Ketoconazole inhibits lanosterol14-demethylase (cholesterol synthesis; CYP51) and two hydroxylase enzymes responsible for steroid metabolism as well as a key enzyme involved in testosterone synthesis. As a result of its effects on steroid synthesis, ketoconazole has been used to treat hyperadrenocorticism and to impair testosterone synthesis in patients with prostatic hypertrophy or prostatic cancer. Ketoconazole has caused lightening of the hair coat of some dogs.⁶⁰ Willard and coworkers⁶⁰ reported depressed basal cortisol and testosterone concentrations and ACTH response by cortisol at 30 mg/kg/day. A rebound response was seen after ketoconazole was discontinued. Serum progesterone concentrations were also decreased. Aldosterone was not decreased. Cats receiving 30 mg/kg/day for 30 days developed dry hair coat and weight loss but no changes in testosterone or progesterone concentrations.⁶¹

Finally, not all clinically relevant drug interactions involving imidazoles reflect effects on cytochrome P450. The imidazoles in general are substrates for P-glycoprotein, and ketoconazole is a known inhibitor.⁶² Accordingly, other drugs that serve as substrates for P–glycoprotein are likely to be absorbed to a greater extent. Thus ketoconazole can affect (increase) cyclosporine A concentrations following oral administration without affecting cyclosporine elimination half-life as has been documented in the author’s laboratory.

Toxicities and Side Effects

In general, the imidazoles are not characterized by the complex toxicities that are associated with amphotericin B. Because the azoles interfere with synthesis of ergosterol rather than binding the sterol, the host toxicities typical of those induced by amphotericin do not occur.⁶³ Gastrointestinal toxicities are the most common and are not severe.^{4,40,27,54,64} Nausea and vomiting can usually be prevented by administration of the drug with food. Hepatotoxicity with ketoconazole has been reported in humans. Mayer and colleagues⁶⁵ retrospectively reported adverse effects of ketoconazole in dogs. Medical records in Australia (n = 296), Germany (n = 35), and the United States (n = 301) were reviewed, and adverse events were reported in 92 (14.6%). Doses ranged from 2.6 to 33.4 mg/kg, with an average daily dose of 11.2 mg/kg. The frequency of adverse events was as follows: vomiting (7.1%), anorexia (4.9%), diarrhea (1.1%), and lethargy (1.9%). Uncommon side effects included pruritis (0.6%) and ataxia, polyuria, and polydipsia; causal relationships were difficult to establish for the uncommon reactions.

Side effects to itraconazole are limited to gastrointestinal symptoms (nausea and vomiting), which may be related in part to the vehicle if associated with the oral solution (a cyclodextrin carrier).³ One case of cutaneous drug eruption typical of erythema multiforme caused by itraconazole has been reported in a dog;⁶⁷ idiopathic vasculitis has also been reported.⁵⁴ Fluconazole is associated with very few side effects; hematologic disorders may occur particularly in profoundly ill patients; otherwise, side effects appear to be limited to gastroinestinal and cutaneous reactions.

Isolated cases of hepatotoxicity have been reported with itraconazole and fluconazole. A retrospective study of dogs with blastomycosis found 5% to 10% of dogs treated with itraconaxole (5 to 10 mg/kg bid) developed hepatotoxicity with the effect dose dependent.¹¹⁹ A dose dependency has been documented in one case with fluconazole. Patients with impaired liver function may be predisposed to worsening hepatic function induced by the azole antifungal drugs. The occurrence of liver disease in animals treated with itraconazole is controversial. Cats receiving 5 and 10 mg/kg twice daily showed no adverse effects (including weight loss) after receiving itraconazole for 6 weeks.⁵⁰ However, one study of enilconazole applied as a 0.2% solution once every 3 days to Persian cats in a cattery indicated potential gastrointestinal signs in the cats, including salivation, anorexia, increased liver enzymes, and emesis and muscle weakness.⁶⁶ Nausea, vomiting, skin rash, thrombocytopenia, and hypokalemia have also been reported with fluconazole therapy. The author dealt with acute hepatopathy in a cat treated topically with a commercial over the counter preparation of clotrimazole; the owner treated a large cutaneous wound topically several times a day. The hepatopathy resolved within several days of discontinuing therapy.

Voriconazole is associated with a number of side effects. Interestingly, voriconazole (but no other azole) causes vision disturbances in humans. Disturbances are characterized by loss of color discrimination, blurred vision, bright spots, wavy lines, and photophobia; up to 30% of human patients are afflicted, although the drug rarely is discontinued because of this effect. Visual hallucinations are reported in 5% of human patients. Experimentally, voriconazole produced dose-dependent changes in the electroretinogram of dogs at plasma levels comparable to those in humans.⁶⁸ Skin rashes also commonly occur; although most are mild, occasional severe reactions (e.g., Stevens–Johnson syndrome) occur. Clinical signs resolve when the drug is discontinued. As with other azoles, voriconazole is associated with increased hepatic leakage enzymes as well as serum alanine phosphatase; increased liver weight occurred experimentally in dogs after 1 month of dosing at 3 mg/kg.⁶⁸ Hepatotoxicity should be assumed to be dose and duration dependent. Although most changes are asymptomatic, severe hepatopathy has been reported in humans receiving voriconazole, with effects apparently dose dependent. Hepatic function might be measured before and 2 weeks into therapy and then every 2 to 4 weeks. The use of N-acetylcysteine, S–adenosylmethionine, or other hepatoprotectants might be considered. Again, because dogs appear to autoinduce, the impact of voriconazole on the liver is not clear. Other gastrointestinal side effects include nausea, vomiting, diarrhea, and abdominal pain.⁴² Although voriconazole is not nephrotoxic, nor is it eliminated in the urine, the carrier of the intravenous preparation may accumulate in patients with impaired renal function, and therefore the intravenous preparation should not be used in these patients.⁴²

Therapeutic Use

In general, itraconazole and fluconazole are more efficacious against many organisms than ketoconazole; however, the spectrum of fluconazole is limited compared with that of itraconazole. Ketoconazole has been used effectively for dermatophyte infections; mucocutaneous candidiasis; and many systemic mycoses in both dogs and cats. Ketoconazole has been reported to be effective in the treatment of dermatophytosis,^70,71 blastomycosis,⁷² histoplasmosis,⁷³ coccidioidomycosis,⁷⁴ and cryptococcosis.⁷⁵ Ketoconazole probably should not be used alone for treatment of canine blastomycosis; recommendations are to use amphotericin in addition to ketoconazole. Higher doses also are indicated for systemic cryptococcosis and coccidioidomycosis. Ketoconazole also has proved effective for treatment of Malassezia dermatitis, and candidiasis. It is available as a topical shampoo that can be useful for treatment of dermatophytosis or Malassezia. Ketoconazole has little efficacy (43%) against Aspergillosis species, fluconazole more efficacy (although some isolates, including A. fumigatus, are inherently resistant), and itraconazole most efficacy (60% to 70%). Fluconazole has been used successfully to treat ketoconazole-resistant strains of Candida. Equal efficacies of itraconazole and fluconazole have been shown for cryptococcal meningitis, despite relatively poor penetration of the CSF by itraconazole. Both are equally effective in Candida-induced pyelonephritis. Comparison of ketoconazole and fluconazole reveals fluconazole to be more active against coccidioidal meningitis.

Among the imidazoles, itraconazole and fluconazole are being used more consistently than the others for systemic treatment of susceptible fungal infections. For itraconazole⁷⁶ conditions successfully treated include blastomycosis⁵⁴ (including ocular⁷⁷), histoplasmosis,⁷⁸ cryptococcosis (including meningitis),^79–81 sporotrichosis,⁸² aspergillosis,⁸³ dermatophytosis,⁶⁴ dermatophytic pseudomycetomas,^84,85 phaeohyphomycosis,⁸⁶ and cutaneous Alternaria.⁸⁷ Efficacy against Aspergillosis is better than that clinically recognized for any other agent (not including newer drugs), and although resistance is rare, treatment failure rates of up to 50% have been reported for itraconazole. In animals administration of itraconazole at a rate of 5 mg/kg twice daily is efficacious in the treatment of blastomycosis and histoplasmosis. After administration of 10 mg/kg, the C_max for itraconazole in dogs was 13.5 ± 8.5 μg/mL versus 3.55 ± 2.81 μg/mL at 5 mg/kg. Although the MIC₉₀ of most infecting fungal organisms will be achieved at 5 mg/kg, at the higher 10 mg/kg dose, the MFC for itraconazole will be achieved for B. dermatitidis, almost reached for H. capsulatum, but not reached for C. immitus. However, the incidence of adverse effects may be greater at this higher dose.⁵⁴ Concentrations will be higher at steady state. The efficacy of itraconazole against coccidioidomycosis is equivocal, requiring long-term therapy. Relapse of disease appears to be common. Despite the larger MIC for dermatophytes compared to other susceptible fungal organisms, itraconazole at 1.5 to 3 mg/kg every 24 hours was effective in 8 of 15 cats in one uncontrolled clinical trial for treatment of dermatophytosis.⁶⁴ Fluconazole is only modestly effective toward sporotrichosis; itraconazole should be considered first-line therapy.⁶⁹

Itraconazole has been used to treat canine blastomycosis (5 mg/kg/day); sporotrichosis (7.5 mg/kg/day); and, in conjunction with surgery, nasal aspergillosis (10 mg/kg/day). In cats it has proved effective for treatment of cryptococcosis and histoplasmosis. The dermatologic pharmacokinetics of itraconazole support pulse therapy, which has been used in human medicine for treatment of selective dermatologic fungal disorders. Treatment occurs for 2 consecutive weeks of daily administration each month for 3 consecutive months.⁸⁸ One report of itraconazole used to treat dermatophytosis noted similar success with this technique.⁶⁴

There are few reports regarding the efficacy of fluconazole for treatment of fungal infections in animals; infections that have been treated include blastomycosis,⁸⁹ cryptococcosis,⁷⁴ and nasal aspergillosis.⁹⁰ Efficacy has, however, been demonstrated toward a variety of fungal disorders in humans. Pulse dosing (once weekly) of fluconazole also has been described for treatment of skin infections in people.

Enilconazole has excellent in vitro activity against a number of organisms, but its topical use is limited to dermatophytes and nasal aspergillosis. Miconazole also is limited to topical use; combination with polymyxin B yields synergistic activity.

Newer azoles are likely to prove even more efficacious than fluconazole and itraconazole for the treatment of aspergillosis and coccidioidomycosis. Voriconazole is approved for treatment of selected infections, but in particular aspergillosis. The maintenance dose in humans is approximately 2 to 5 mg/kg twice daily. In human patients who have not responded adequately to traditional therapy for treatment of aspergillosis, 38% had a partial to complete response to voriconazole used as salvage therapy. In a study of invasive aspergillosis in humans, 53% of patients receiving voriconaozle responded (partial to complete) within 12 weeks compared with only 32% of patients receiving amphotericin B; survival rates were 71% and 58%, for each drug, respectively.⁴² Because of superior clinical response, safety, and survival, treatment of immuncompromised human patients with voriconazole was demonstrated to be economically superior to treatment with amphotericin B.⁹¹ For Pseudallescheria/Scedosporium, major pathogens in human immunocompromised hosts normally resistant to amphotericin B and Fusarium sp., response rate was 30% to 63% and 50%, respectively, for the two species. Despite excellent in vitro efficacy against C. neoformans and good CNS penetration, voriconazole is not recommended for treatment of Cryptococcosis, in part because of its failure in human patients. The same may be true for blastomycosis, histoplasmosis, and coccidiodomycosis. Although effective in animal models, successful therapy with voriconazole for these organisms has not been demonstrated in humans. A salvage approach might be considered for these organisms.⁴²

A single case report describes the successful treatment with posoconazole of a fungal disorder caused by Mucor (Zygomycetes class; Mucorales order) species on the nose of a 15-year-old cat.^91a After poor response to fluconazole, the cat was treated at 5 mg/kg daily for 3 months, with initial response evident in 2 weeks and continued response for the remainder of the 3-month treatment period.

Terbinafine was shown to enhance efficacy when combined with itraconaozle or fluconazole when treating candidiasis characterized by low susceptibility to the azoles.⁹² Indeed, combination with benzoyl peroxide topically enhanced treatment of candidiasis associated with Pseudomonas and Staphylococcus aureus infections in humans.⁹³

Structure–Activity Relationship

5-Flucytosine (FLU; 5-fluorocytosine) was originally developed as an anticancer drug much the same as its sister anticancer drug, 5-fluorouracil. It is a water-soluble powder.

Mechanism of Action

As an antimetabolite, FLU interferes with DNA synthesis after its conversion to 5–fluorouracil, a substitution compound that prevents synthesis in the fungal cell.^4,40 The compound enters the cell by way of cytosine permease, which also takes up adenine, guanine, hypoxanthine, and cytosine.³ The enzyme responsible for conversion of FLU to 5–fluorouracil is a cytosine deaminase, an enzyme whose absence in mammalian cells renders FLU relatively specific for fungal cells. The effect of FLU depends on subsequent metabolism: if converted by pyrimidine processing enzymes to a uridine monophosphate derivative, inhibition of thymidylate synthase and DNA synthesis yields -cidal effects. Alternatively, it can be converted by way of a pyrimidine salvage pathway resulting in metabolism to a uridine-5’-triphosphate derivative. Subsequent incorporation into RNA results in impaired protein synthesis and fungistatic effects (Candida, Cryptococcosis). Many organisms are intrinsically resistant to FLU because they either lack the permease enzyme or have a defective deaminase enzyme. Resistance also develops relatively rapidly, particularly for aspergillosis followed by cryptococcosis and candidiasis (especially Candida krusei).³ Secondary resistance usually reflects a decrease in an enzyme responsible for formation of uridine monophosphate. Use in combination with another antifungal agent reduces the development of resistance.

Spectrum of Activity

The spectrum of activity of FLU is limited and includes cryptococcosis, candidiasis and some cladosporiosis, aspergillosis, chromomycosis, and sporotrichosis. It has been the treatment of choice for cryptococcosis in humans.^4,40 Combination therapy is usually indicated (e.g., amphotericin, ketoconazole). When FLU is used alone, resistance develops rapidly. Synergism occurs with amphotericin B and probably with ketoconazole (or other imidazoles).

Pharmacokinetics

Oral absorption of FLU is rapid and close to complete. Peak plasma concentrations occur in 1 to 2 hours. Distribution is large, to total body water. Protein binding is minimal, and CSF concentrations reach up to 90% of plasma concentrations. Penetration of aqueous humor and joints is good. The half-life of FLU is 3 to 6 hours. Most of the drug is excreted into the urine unchanged. Renal clearance is similar to that of creatinine and thus may be significantly decreased if renal dysfunction is present. Doses will probably need to be modified for patients with renal disease.

Preparations

Flucytosine is available as an oral preparation.

Side Effects

Because FLU interferes with DNA synthesis, body systems composed of rapidly dividing cells are adversely affected. Bone marrow depression is manifested as anemia, leukopenia, and thrombocytopenia (pancytopenia). This toxicity may be serious and is more common in patients with renal disease. Gastrointestinal toxicity is manifested as nausea, vomiting, and diarrhea, but it is not usually serious. Reversible, erythemic, alopecic dermatitis has been reported in dogs.

Structure–Activity Relationship

Griseofulvin (see Figure 9-1) is produced from a Penicillium species bacterium. The drug is insoluble in water.

Mechanism of Action

Griseofulvin enters fungi through an energy-dependent transport system. Griseofulvin inhibits fungal mitosis by binding to the microtubules that form the mitotic spindle. The formation of microtubules from tubulin is inhibited. Formation of cytoplasmic microtubules responsible for transport of endogenous compounds also is inhibited. Other drugs, such as colchicine and vincristine, which also bind to and inhibit the microtubule, do so at a site that is different from that of griseofulvin. Griseofulvin also probably inhibits nucleic acid and fungal wall synthesis. It is not certain if griseofulvin is fungistatic or fungicidal. Resistance probably reflects decreased drug uptake.

Spectrum of Activity

The spectrum of activity of griseofulvin reflects the presence of an energy-dependent transport system in the fungal organism. Those with prolonged energy-dependent transport systems are susceptible, whereas those with independent systems of short duration are not. Efficacy is limited to dermatophytes: Microsporum, Trichophyton, and Epidermophyton. Because of its distribution into keratin, griseofulvin remains the drug of choice for fungal infections of the nails.

Pharmacokinetics

Oral absorption varies because of water insolubility and depends on particle size and preparation. Absorption is increased in the presence of fat. The rates of dissolution and disaggregation alter the bioavailability of different products. Bioavailability of the ultramicrosize is at least 50% greater than that of the microsize. Although griseofulvin penetrates the stratum corneum, it does not achieve effective concentrations topically. Griseofulvin is widely distributed to most tissues, but it is deposited and concentrated in keratin precursor cells. Thus it is incorporated in new keratin of skin, nails, and hair and (in humans) is secreted in perspiration. Although new keratin formed during treatment with griseofulvin is resistant to fungus, griseofulvin does not destroy fungi that infect the outer layers of the skin. New hair, skin, or nail growth accompanied by shedding of older growth is necessary before the fungus is affected; new growth is the first to be free of disease. Thus skin infections require 4 to 6 weeks of therapy, whereas toenails may require up to a year of therapy. Long hair breeds probably should be treated for a longer period of time compared to short hair breeds.

Hepatic metabolism of griseofulvin by dealkylation is significant; metabolites are not active. The half-life reportedly is 24 hours in the dog. Half of the drug is excreted as metabolites in the urine. The rest is excreted through the bile unchanged in the feces.

Preparations

Griseofulvin is available for oral use as either a microsize (particle size 10 μm) or ultramicrosize (particle size 2.7 μm; e.g., Fulvicin, Gris-PEG) tablets. The drug should be administered with a fatty meal, particularly if the microsize preparation is used. Duration of therapy is at least 4 to 6 weeks (new hair growth must occur) and possibly longer. The drug should be administered at least once a day despite initial reports that recommend one weekly administration.

Side Effects and Drug Interactions

Side effects to griseofulvin are not uncommon. Nausea, vomiting, and diarrhea can be minimized by administration of the dose in divided increments with a meal. Hepatotoxicity may occur, and use in liver disease should be avoided. Idiosyncratic toxicity has been reported in the cat, manifested as gastrointestinal upset, neurologic disease, and bone marrow suppression.⁹⁸ The reaction appears to be both dose and duration independent. Signs may not be reversible, depending on the severity. Cats with feline immunodeficiency disorders may be more likely to develop neutropenia.⁹⁹ Certain feline breeds (e.g., Persian, Siamese, Abyssinian) may be more commonly affected.⁹⁸ At very high doses, the drug is teratogenic and carcinogenic in animals. The drug should not be given during the first two trimesters of pregnancy. Use with a shampoo (miconazole or chlorhexidine) enhances efficacy.

Griseofulvin is a potent inducer of microsomal enzymes. The clinical sequelae of this drug interaction are not well known, although increased metabolism of other drugs should be anticipated.

Dermatologic Fungal Infections: Dermatophytosis

A number of fungal organisms inhabit the hair coats of dogs and cats. Alternaria, Cladosporium, and yeasts may be associated with dermatitis. Dermatophytes can be isolated from normal animals or can be a cause of infection. Dermatophyte infections generally are self-limiting, with ability to mount an inflammatory response being an important determinant of infection control. Accordingly, drugs such as glucocorticoids, which mute the inflammatory response, predispose a patient to dermatophyte infection; dermatophytosis infection is 3 times more prevalent in cats infected with feline immunodeficiency virus than in noninfected cats.⁶⁶ The route of drug administration (topical versus systemic therapy) depends on the extent of infection, with the exception of Trichophyton infections, which should be treated systemically.

Topical therapy is indicated for all patients with dermatophytosis and may be the sole therapy for local, nondiffuse lesions. Hair coat preparation before medication should include clipping and bathing to remove hair and crusts. Several medicaments are available as shampoo, ointment, or cream formulations for topical therapy (see Table 9-5). Active ingredients include povidone–iodine, chlorhexidine, and imidazole. Other active ingredients that can be applied topically include captan, lime sulfur, and sodium hypochlorite. Short-term topical glucocorticoid therapy might be considered to control acute inflammation when present. Topical administration of enilconazole emulsion has been useful for treatment of feline dermatophytosis.

Moriello^66,107 retrospectively evaluated in vitro and in vivo studies of treatments for dermatophytosis in dogs and cats. Topical treatments consistently found to be effective when administered once or twice weekly were lime sulfur (1:16), 0.2% enilconazole rinse (twice weekly; response in as early as 5 weeks), and a combined 2% miconazole–chlorhexidine shampoo. Captan, chlorhexidine (as sole agent), and povidine–iodine were generally ineffective.

Response to systemic therapy is likely to be more rapid if combined with topical therapy.⁶⁶ Systemic therapy should probably be preceded with a total body clip. Griseofulvin is the treatment of choice for long-term systemic antifungal therapy of dermatophytosis,¹⁰⁹ although expense mandates that a diagnosis of dermatophytosis be confirmed. Care should be taken that the proper dose and duration of therapy are followed; treatment probably should be longer for longer-haired animals. Cure was reported in 63 to 70 days in one study with a mean of 41 days at 50 mg/kg in another study (as reviewed by Moriello).⁶⁶ For infections that do not respond to griseofulvin, an imidazole can be used. Ketoconazole has been used successfully, particularly when dosed at 10 mg/kg daily for 20 days. Itraconazole has, however, proved more efficacious for treatment of dermatophytes in human patients and has proved efficacious experimentally in cats infected with M. canis.¹¹⁰ Itraconazole was effective at 10 mg/kg once daily for 56 days or for 28 days followed by a week–on week–off pulse dosing for 56 to 70 days.¹¹¹ Interestingly, low-dose itraconazole (1.5 to 3 mg/kg) in 15-day cycles required a shorter time period at 1 to 3 cycles or 15 to 45 days. However, only 8 of 15 cats were successfully treated in this study, and two of these cats required a second cycle. Using an open clinical trial, a protocol using itraconazole (10 mg/kg once daily) and lime sulphur rinses (cat coat saturated with rinse at 8 ounces per gallon of water) every 7 days for 21 days was associated with successful treatment of dermatophytosis in shelter cats.^111a Cure was longer with higher fungal load (18.4+ 9.5 days) compared to smaller loads (14.5+5.7 days). Cats tolerated the rinses well. Efficacy of lufenuron was not substantiated in controlled studies. Terbinafine has demonstrated efficacy toward dermatophytosis in dogs and cats, although higher doses (>30 to 40 mg/kg) generally are required to achieve a mycologic cure. Time to cure in dogs varied from 21 to more than 126 days and from 28 to 84 days in dogs. A lower dose (10 to 30 mg/kg) may also be effective in both dogs and cats, although treatment should be expected to be approximately 60 to 90 days.¹⁰⁷ Use of terbinafine at 30 mg/kg for 2 weeks resulted in a cure of 11 of 12 cats with M. canis, and at 8.25 mg/kg daily, it eradicated spores from asymptomatic carrier cats (as reviewed by Bossche³). The use of systemic terbenafine at either a low (10 to 20 mg/kg) or high (30 to 40 mg/kg) dose once daily was compared with placebo treatment in cats (n = 9 per group) ranging between 1.5 and 4.5 months of age that were experimentally infected with M. canis. Cats were treated for 120 days. Response in the low-dose treatment group did not differ from that of the control group but did in the high-dose group; the number of cats cured per group was not provided. PDCs did not differ between the two dose groups, but concentrations in the hair were significantly higher with the higher dose.¹¹² Drug accumulated with multiple dosing, with concentrations at 3 months higher than at 2 months, suggesting peak effects may take up to 3 months (or more). Median concentrations in plasma at 9 and 120 days were 1.4 mg/L and 4.1 mg/L respectively, in the low-dose cats compared with 1.7 and 5.5 mg/L respectively, in the high-dose cats. Median concentrations in the hair at 9 and 120 days were 1 mg/L and 1.2 mg/L, respectively, in the low-dose cats compared with 1.9 and 3.6 mg/L respectively, in the high-dose cats. The highest concentration achieved in the hair in any one cat was 7.92 μg/mL; this compares with humans receiving only 6 mg/kg, for which concentrations reach 2.40 to 55 μg/g.

Additional topical therapies for treatment of dermatophytosis or other dermatologic fungal disorders are also addressed in Chapter 22. Environmental cleansing may be important to treatment of dermatophytosis. This might be accomplished with either 2% chlorhexidine or 0.5% sodium hypochlorite.

Yeast or Yeastlike Infections

Systemic Fungal Diseases

Therapeutic success with antifungal drugs can be enhanced by long-term therapy, generally one to several months beyond the resolution of clinical signs; avoidance of immunosuppressive drugs; and use of combination therapy, particularly for infections that are difficult to penetrate or are life or organ threatening.

Subcutaneous Mycoses