Serum itraconazole concentrations and clinical responses in Candida-associated denture stomatitis patients treated with itraconazole solution and itraconazole capsules

L. J. Crossa,*, J. Bagga, D. Oliverb and D. Warnockb

a University of Glasgow Dental School, Level 2, 378 Sauchiehall Street, Glasgow G2 3JZ; b Public Health Laboratory Service Mycology Reference Laboratory, Bristol, UK


    Abstract
 Top
 Abstract
 Introduction
 Patients and methods
 Results
 Discussion
 References
 
The aim of this study was to compare the concentrations of itraconazole in serum and saliva after treatment with itraconazole cyclodextrin solution or itraconazole capsules in Candida-associated denture stomatitis patients without evidence of immunodeficiency. Forty patients were randomly assigned to receive either itraconazole cyclodextrin solution or itraconazole capsules, both at a dosage of 100 mg bd for 15 days. On completion of treatment palatal erythema was assessed and an oral rinse and imprint cultures were collected. Serum and saliva samples were collected at the same time and itraconazole concentrations measured using reverse-phase high-performance liquid chromatography. Itraconazole susceptibilities of Candida albicans and Candida glabrata strains isolated at baseline were measured by a broth microdilution method. Serum itraconazole concentrations achieved did not differ significantly between the two preparations (P = 0.39) although a significantly higher number of patients in the itraconazole cyclodextrin group (P < 0.001) had detectable levels of itraconazole in their saliva compared with the capsule group. Mycologically cured patients had slightly, though not significantly (P = 0.28), higher serum itraconazole concentrations than those from whom yeasts were not eradicated. It was concluded that both formulations of itraconazole were equally effective in treatment of denture stomatitis. Among immunocompetent patients, the absorption of the liquid preparation is no greater than that of the capsules. Therapeutic success in this group was achieved with lower serum itraconazole concentrations than have been reported for immunocompromised groups.


    Introduction
 Top
 Abstract
 Introduction
 Patients and methods
 Results
 Discussion
 References
 
Candida-associated denture stomatitis is characterized by chronic erythema and oedema of the palatal mucosa beneath an upper denture. It affects 50% of denture wearers.1 Treatment regimens reflect the multifactorial nature of the condition2 and include antifungal therapy, provision of new dentures, maintenance of good denture hygiene and removal of dentures for sleeping. Topical nystatin, amphotericin B and miconazole and systemically administered fluconazole have been used in the management of denture stomatitis but all have been associated with a high rate of relapse.3

Itraconazole has been effective in the management of oral candidosis among immunocompromised patients. The new cyclodextrin solution of itraconazole has been used successfully in AIDS patients unresponsive to itraconazole capsules4 and has been shown to result in higher serum concentrations than capsules in such patients,5 but the kinetics have been less fully studied among the immunocompetent. Until now, no clinical trials of either formulation of itraconazole have been undertaken in the management of oral candidosis in immunocompetent hosts. The present study compared the liquid and capsule preparations of itraconazole in treatment of denture stomatitis and correlated clinical outcome with serum and salivary concentrations of the drug.


    Patients and methods
 Top
 Abstract
 Introduction
 Patients and methods
 Results
 Discussion
 References
 
The study was approved by the local Ethics Committee. Thirty-one female and nine male patients (age range 29–81 years, mean 62 years) were recruited from the Prosthodontic and Oral Medicine Departments of Glasgow Dental Hospital and School NHS Trust. All were edentulous denture wearers with clinical evidence of denture stoma-titis and positive yeast cultures from both the palatal mucosa and dorsum of the tongue. Patients were excluded from the study if they were pregnant, lactating or of child-bearing age and not using reliable contraception; had impaired renal or hepatic function; had used an antifungal agent within the preceding 10 days; were taking any drug listed in the British National Formulary6 as interacting with the azole group of antifungal drugs; had a known sensitivity to the azole group of antifungals; or intended to donate blood during or up to 3 weeks after the study period.

The 40 patients were randomized to receive either cyclodextrin solution of itraconazole (Sporanox Liquid, Janssen-Cilag Ltd, High Wycombe, UK) at a dose of 100 mg bd for 15 days or itraconazole capsules (Sporanox, Janssen-Cilag Ltd) at the same dosage, also for 15 days. There were no significant differences in the age or sex distribution between those receiving the two preparations. The clinician examining the patients was blinded to the treatment code. Patients were instructed to take itraconazole capsules after food and itraconazole solution 1 h before food. Patients receiving itraconazole solution were asked to remove their dentures and briefly swill the solution around their mouth before swallowing.

All patients were reviewed after 15 days, on completion of their course of antifungal medication and those who had failed to complete the course were excluded. Erythema of the palatal mucosa was assessed objectively by means of an electro-optical instrument called an erythema meter.7 The instrument is based on the principle that when white light is reflected off a cutaneous or mucosal surface, haemoglobin in the vasculature selectively absorbs green light but has little effect on red. An erythema meter index, defined as the ratio of red to green reflected light, can be determined. The difference between the ‘erythema index’ measured before and after treatment is related to reduced vasodilation following a reduction in the degree of inflammation. An oral rinse and imprint cultures were collected and inoculated on to plates of Sabouraud's dextrose agar (Life Technologies, Paisley, UK) and CHROMagar Candida agar (Life Technologies). All plates were incubated at 37°C for 48 h. One colony of each morphological type on CHROMagar Candida was plated for purity, identified and stored at –70°C. The proportion of each morphological type was also recorded for each patient. Four further colonies from the Sabouraud plates were selected at random from the specimens available. Each isolate was formally identified and stored at –70°C for further investigation. Candida albicans was identified on the basis of germ-tube production. A sucrose assimilation test was performed on germ-tube-positive yeasts to distinguish C. albicans from Candida stellatoidea. Germ-tube-negative yeasts were identified by the API 32C test (bioMerieux SA, Marcyl’Etoile, France). A blood sample (10 mL) was collected into a plain bottle and at least 1 mL of whole saliva was collected into a sterile universal container to enable measurement of itraconazole concentrations in serum and saliva. The time intervals between last dose and specimen collection are given in the TableGo. Itraconazole concentrations were measured by reverse-phase high-performance liquid chromatography with a lower limit of detection of 0.01 mg/L and coefficients of variation of 2.2–7.8% over the concentration range 0.02–1.6 mg/L.8 The standard curve was linear from 0.01 to 10 mg/L.


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Table. Serum and salivary itraconazole concentrations for each patient in each treatment group and the time between the last dose of itraconazole and specimen collection
 
In vitro susceptibility testing of the strains of C. albicans and Candida glabrata isolated at baseline was undertaken in RPMI broth by the broth dilution method proposed as the NCCLS standard.9 C. albicans and C. glabrata isolates with MIC values in the region of <=0.125 mg/L were classed as susceptible, those with MICs of 0.25–0.5 mg/L were classed as susceptible dependent upon dose and those with MICs of >=1.0 mg/L were classed as resistant.9

Statistical analysis

Data were analysed with the Minitab statistical program, Release 11 for Windows. Two-sample t-tests were used for both within-group and between-group comparisons. A {chi}2 test was used to compare the frequency of zero and non-zero salivary itraconazole concentrations in each treatment group. P < 0.05 was considered significant.


    Results
 Top
 Abstract
 Introduction
 Patients and methods
 Results
 Discussion
 References
 
The TableGo shows the serum and salivary concentrations of itraconazole measured for patients in each of the two study groups on completion of the 15 day course of treatment. The median serum itraconazole concentration in the cyclodextrin group was 0.74 mg/L (interquartile range 0.46–1.18) compared with 0.61 mg/L (interquartile range 0.37–0.93) in the capsule group. With a two-sample t-test, no significant difference was found between the serum itraconazole concentrations with the two formulations (P = 0.39). The median salivary itraconazole concentration in the cyclodextrin group was 0.12 mg/L (interquartile range 0–0.53) compared with 0 (no interquartile range) in the capsule group. The high incidence of zero values in the capsule group is obvious (TableGo) and the difference in the frequency of zero and non-zero values in the two groups was highly significant (P < 0.001). There was little evidence of any correlation between the serum and salivary concentrations of itraconazole among patients in the cyclodextrin group (P = 0.704).

The relationship between the serum itraconazole concentration and the mean reduction in erythema meter index for patients in both treatment groups on review following completion of treatment is shown in Figure 1Go. There was a significant (P = 0.033), moderately positive relationship (95% confidence interval 0.03–0.61) between the serum itraconazole concentration and the mean reduction in erythema meter index recorded following treatment for patients in both the treatment groups. The sample correlation coefficients were 0.370 for the cyclodextrin group and 0.302 for the itraconazole capsule group, revealing that, on completion of treatment, the reductions in palatal inflammation related to the serum itraconazole concentrations were very similar for both forms of itraconazole. However, given the low correlation coefficients between the serum itraconazole concentrations and mean reduction in erythema meter index, this result should be considered with some caution.



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Figure 1. Relationship between serum itraconazole concentration and mean reduction in erythema meter indices for patients on completion of treatment (P = 0.033). Cyclodextrin group {diamondsuit}; capsule group {blacksquare}.

 
Figure 2Go shows a comparison of the serum itraconazole concentrations among mycologically cured patients and non-cured patients on completion of the course of antimycotic therapy. There was a large overlap between the serum itraconazole concentrations measured both for those patients who were free of yeasts on review following completion of treatment and for those who were not mycologically cured. However, the serum itraconazole concentrations of the mycologically cured patients were greater than 0.6 mg/L in all but three of the 17 cases, whereas the majority (12 of 19) of the mycologically non-cured patients had serum itraconazole concentrations of less than 0.5 mg/L (Figure 2Go). Nevertheless, the serum itraconazole concentrations seen in the patients who were mycologically cured on review following treatment were not significantly different, on average, from those not cured (P = 0.28).



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Figure 2. Comparison of serum concentrations of itraconazole in mycologically cured and non-cured patients on completion of treatment (treatment groups combined).

 
A wide range of yeast species was isolated from the mouths of the denture stomatitis patients before treatment. Of the 40 study participants, 18 were infected with more than one species of yeast. C. albicans was present in samples from all 40 patients. The second most commonly isolated species, C. glabrata was found in 12 patients at baseline and persisted post-treatment in all but three of these cases, two from the cyclodextrin itraconazole group and one from the itraconazole capsule group. Other species including Candida tropicalis, Candida krusei, Candida parapsilosis, Candida guillermondii and Saccharomyces cerevisiae were present in samples from only a few of the patients at baseline.

Baseline C. albicans isolates tested had an MIC50 of 0.125 mg/L and an MIC90 of 0.25 mg/L. However, C. glabrata isolates had an MIC50 of 2 mg/L and an MIC90 of 4 mg/L.


    Discussion
 Top
 Abstract
 Introduction
 Patients and methods
 Results
 Discussion
 References
 
The solubility and subsequent absorption of itraconazole following oral administration of the capsule formulation is dependent on gastric acidity, the bioavailability of the itraconazole being reduced in the fasting state or under conditions of hypochlorhydria. In contrast to the capsules, the cyclodextrin solution of itraconazole is more bioavailable when taken optimally with or before food.10 In order to ensure optimal bioavailability of each formulation of itraconazole in this trial, patients were requested to take the cyclodextrin solution of itraconazole 1 h before food and the itraconazole capsules with food. However, it is claimed that the cyclodextrin solution of itraconazole offers improved solubility over itraconazole capsules, resulting in improved absorption and enhanced bioavailability.10 To investigate this claim, patients from each treatment group in this clinical trial provided serum and whole saliva samples on completion of the 15 day course of itraconazole. No significant difference was found in the serum itraconazole concentrations achieved in patients from the two treatment groups. The reduction in palatal inflammation resulting from the itraconazole therapy was found to be very similar for both formulations of the medication.

The serum itraconazole concentrations in those patients who were mycologically cured on completion of treatment were compared with the concentrations found in those patients from whom yeasts could still be isolated. The patients were grouped according to their mycological response, irrespective of treatment group, as it was now apparent that both formulations of itraconazole had similar efficacy in the treatment of denture stomatitis. Considerable overlap was found between the serum itraconazole concentrations of the mycologically cured and non-cured patients. However, patients who were free of yeasts on completion of treatment did have slightly higher serum concentrations of itraconazole, although this proved not to be statistically significant (P = 0.28). Nevertheless, it is interesting to note that the majority of mycologically cured patients in this clinical trial had serum itraconazole concentrations of greater than 0.6 mg/L whereas most of the mycologically non-cured patients had serum itraconazole concentrations of 0.5 mg/L or less. In contrast to this, Cartledge et al.5 found higher serum drug concentrations and better clinical response rates with the cyclodextrin solution of itraconazole than with itraconazole capsules in AIDS patients. The bioavailability of itraconazole capsules in immunocompromised patient groups is variable and known to be lower than in non-immunocompromised individuals, largely resulting from low gastric acidity or chemotherapy-induced gastro-intestinal toxicity associated with patients in the former group.10 Cartledge et al.5 also suggested that a serum itraconazole concentration of less than 1.0 mg/L on day 7 was predictive of therapeutic failure. Therapeutic success was achieved with much lower serum itraconazole concentrations in the denture stomatitis patients. This may reflect the immunocompetent status of the patients enrolled in this study or the multifactorial aetiology of denture stomatitis.

C. albicans species isolated from trial participants at baseline all proved to be susceptible to itraconazole. The MICs (0.06–0.5 mg/L) related well to the serum itraconazole concentrations discussed above. However, MICs of itraconazole for C. glabrata isolates tended to be suscep-tible dependent on dose or resistant. There was no correlation between itraconazole sensitivity and clinical outcome.

Patients in the cyclodextrin group had significantly higher (P < 0.001) concentrations of itraconazole in their saliva compared with patients in the itraconazole capsule group, the majority of whom had no itraconazole detectable in their saliva at all. Only two patients in the capsule group had unusually high salivary itraconazole concentrations. These results were so aberrant that they may be attributed to laboratory or sampling error. The detection of itraconazole in the saliva of patients treated with the cyclodextrin formulation suggests an additional topical effect of cyclodextrin which would provide an obvious advantage of this preparation in the treatment of oropharyngeal candidosis. However, in this study, despite the significantly higher salivary itraconazole concentrations achieved with the cyclodextrin solution of itraconazole, the advantage of a topical effect was not apparent, as both formulations were equally effective in the management of denture stomatitis in terms of mycological response and reduction in erythema.

There was no relationship between the serum and saliva itraconazole concentrations of the group on the cyclodextrin preparation (P = 0.704). This implies that the presence of itraconazole in the saliva following administration of the cyclodextrin solution is due to persistence of the drug following direct contact with the buccal mucosa, to which itraconazole adheres and is subsequently released,10 rather than systemic absorption followed by secretion of itraconazole into saliva. This is confirmed by the fact that no active metabolite has been found in saliva.10

In conclusion, the absorption of itraconazole with the liquid preparation was no greater than with the capsules, but itraconazole was more likely to be found in the saliva on treatment with the liquid preparation. Itraconazole cyclodextrin solution and itraconazole capsules were equally effective in the treatment of denture stomatitis.


    Acknowledgments
 
Grant Carnegie and Nigel Blatchford are thanked for their helpful advice. Financial support for the itraconazole assays was provided by Janssen-Cilag Ltd.


    Notes
 
* Corresponding author. Tel: +44-141-211-9831; Fax: +44-141-353-1593; E-mail: L.Cross{at}dental.gla.ac.uk Back


    References
 Top
 Abstract
 Introduction
 Patients and methods
 Results
 Discussion
 References
 
1 . Budtz-Jörgensen, E., Stenderup, A. & Grabowski, M. (1975). An epidemiologic study of yeasts in elderly denture wearers. Community Dentistry and Oral Epidemiology 3, 115–9.[ISI][Medline]

2 . Budtz-Jörgensen, E. (1974). The significance of Candida albicans in denture stomatitis. Scandinavian Journal of Dental Research 82, 151–90.[ISI][Medline]

3 . Budtz-Jörgensen, E. (1990). Candida-associated denture stomatitis and angular cheilitis. In Oral Candidosis, (Samaranayake, L. P. & MacFarlane, T. W., Ed.), pp. 156–83. Wright, London.

4 . Cartledge, J. D., Midgeley, J., Youle, M. & Gazzard, B. G. (1994). Itraconazole cyclodextrin solution: effective treatment for HIV-related candidosis unresponsive to other azole therapy. Journal of Antimicrobial Chemotherapy 33, 1071–3.[ISI][Medline]

5 . Cartledge, J. D., Midgeley, J. & Gazzard, B. G. (1997). Itraconazole solution: higher serum drug concentrations and better clinical response rates than the capsule formulation in acquired immunodeficiency syndrome patients with candidosis. Journal of Clinical Pathology 50, 477–80.[Abstract]

6 . British Medical Association and Royal Pharmaceutical Society of Great Britain. (1996). Section 5.2—Antifungal Drugs. British National Formulary 32, 262–6.

7 . Cross, L. J., Bagg, J. & Moseley, H. (1998). Evaluation of an optical instrument for objective assessment of oral mucosal erythema. Journal of Oral Rehabilitation 25, 496–501.[ISI][Medline]

8 . Warnock, D. W., Turner, A. & Burke, J. (1988). Comparison of high performance liquid chromatographic and microbiological methods for determination of itraconazole. Journal of Antimicrobial Chemotherapy 21, 93–100.[Abstract]

9 . Rex, J. H., Pfaller, M. A., Galgiani, J. N., Bartlett, M. S., Espinel-Ingroff, A., Ghannoum M. A. et al. (1997). Development of interpretive breakpoints for antifungal susceptibility testing: conceptual framework and analysis of in vitro–in vivo correlation data for fluconazole, itraconazole and candida infections. Clinical Infectious Diseases 24, 235–47.[ISI][Medline]

10 . Janssen-Cilag Ltd. (1996). Sporanox liquid. In Sporanox (Itraconazole) in Systemic Fungal Infection. Product Monograph, pp. 75–86. Janssen-Cilag Ltd, Buckinghamshire, UK

Received 25 August 1998; returned 3 April 1999; revised 10 June 1999; accepted 19 September 1999