JAC Advance Access originally published online on November 14, 2006
Journal of Antimicrobial Chemotherapy 2007 59(2):317-320; doi:10.1093/jac/dkl472
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Oral administration of itraconazole solution has superior efficacy in experimental oral and oesophageal candidiasis in mice than its intragastric administration
Teikyo University Institute of Medical Mycology, 359 Otsuka Hachioji, Tokyo 192-0395, Japan
*Corresponding author. Tel: +81-426-78-3256; Fax: +81-426-74-9190; E-mail: hiroko-tei{at}umin.ac.jp
Received 24 March 2006; returned 5 May 2006; revised 12 September 2006; accepted 23 October 2006
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Abstract |
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Objective: The therapeutic activities of cyclodextrin-associated itraconazole oral solution (itraconazole OS) by two administration routes in experimental oral and oesophageal candidiasis in mice were examined and compared.
Methods: Using experimental oral and oesophageal candidiasis models in ICR mice, we investigated the efficacy of oral and intragastric administration of itraconazole OS and checked the concentration of itraconazole and its metabolite hydroxyitraconazole (OH-itraconazole) in tongues or oesophagus tissue after administration of itraconazole OS.
Results: Oral administration of itraconazole OS at doses of 0.8, 4.0 or 20 mg/kg/day clearly decreased the number of viable Candida albicans cells in the oral cavity of mice with oral candidiasis in a dose-dependent manner at 3 days after infection. Intragastric administration of itraconazole OS at doses of 4 and 20 mg/kg/day once a day were also effective but to a lesser degree than that of oral administration. In the oesophageal candidiasis model, oral administration of itraconazole OS displayed superior therapeutic efficacy to the intragastric route. In coincidence with the greater efficacy, itraconazole was detected in lesional tissues after oral administration of itraconazole OS.
Conclusions: Oral administration of itraconazole OS displayed therapeutic efficacy against murine oral and oesophageal candidiasis superior to that achieved by intragastric administration. This can be explained by there being higher concentrations of itraconazole in tongues or oesophagus tissues after administration of the suspension by the oral route.
Keywords: mucosal infections , intestinal tracts , antifungal chemotherapy , drug delivery
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Introduction |
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As clinical treatment for oropharyngeal and oesophageal candidiasis, oral administration of azole antifungals including itraconazole oral solution (itraconazole OS) is recognized as first choice. But pharmaceutical analysis of mechanisms of therapeutic actions of orally administered itraconazole OS in vivo has been very limited.1–3
Using murine oral and oesophageal candidiasis models, we evaluated the efficacy of itraconazole OS administered by oral or intragastrical routes on this condition and assessed the concentration of itraconazole and its metabolites in the tongue and oesophagus tissues after that administration.
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Materials and methods |
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Oral and oesophageal candidiasis in mice
Experimental procedures of the oral and oesophageal candidiasis models were described previously.4,5 All animal experiments were performed according to the guidelines for the care and use of animals approved by Teikyo University. Six-week-old female ICR mice (Charles River Japan, Inc., Kanagawa, Japan) were used for all animal experiments. Immunosuppressed mice were induced by subcutaneous treatment with a dose of 100 mg/kg of prednisolone (Mitaka Pharmaceutical Co., Tokyo) 1 day prior to oral or oesophageal infection. Tetracycline hydrochloride (Takeda Shering Purau Animal Health Co., Tokyo, Japan) in drinking water at a dose of 0.08% was given to the animals, beginning 1 day before infection. Ten minutes before Candida infection, the mice were anaesthetized by intramuscular injection in the foot with 100 µL of 0.25% chlorpromazine chloride (Wako Pure Chemical Industries Ltd, Osaka, Japan) and their sedated condition continued for about 3 h.
In the oral candidiasis model they were orally infected with 
2 x 108 cells/mL viable cells of Candida albicans TIMM2640 in 2.5% FCS-RPMI 1640 medium. Oral infection was performed using a cotton swab (baby cotton buds; Johnson & Johnson Co., Tokyo) rolled over all areas of the mouth.
In the oesophageal candidiasis model the anaesthetized mice were intraoesophageally infected with C. albicans TIMM2640.5 The infection was performed by intraoesophageal inoculation of 4 x 107 viable C. albicans cells in 0.2 mL of 2.5% FCS-RPMI 1640 medium by a round-head needle (39 mm length, Fuchigami, Ltd, Kyoto) with a syringe. When the C. albicans cells were injected the needle head was placed at a site just under the pharynx.
Antifungal treatment
Itraconazole OS and beta-cyclodextrin were provided by Janssen Research Foundation, Beerse, Belgium. Itraconazole OS was administered to mice by two different routes throughout the experiment (once a day from 3 h after inoculation to day 2 post-infection). Oral administration was performed by spreading the drug in oral cavity by using a round-head needle with a syringe as described above, and intragastric dosing was done by the conventional method using the same type of needle. Doses of itraconazole OS for oral and intragastric administration were 0.8, 4 or 20 and 4 or 20 mg/kg/day, respectively, that were adopted in an effective range suggested by results of preliminary therapeutic experiments. Control groups of mice received the same volume of 40% beta-cyclodextrin on a time schedule the same as that of itraconazole OS. Therapeutic efficacy was evaluated on day 3 post-infection. The cfu of C. albicans recovered from oral cavity were determined and the white patch of each tongue and squamous disorder was scored as described previously.4
At the end of oesophageal candidiasis experiments, the oesophageal tubes were obtained and homogenized in 2 mL saline, and the homogenates were cultured to determine the cfu.
Pharmacokinetics
Murine tongues or oesophageal tubes were obtained from the infected mice on the last day in the itraconazole OS-treated mice with oral or oesophageal candidiasis model.
In other experiments, itraconazole OS was administered once to normal mice and their blood was collected until 8 h after administration. Concentrations of itraconazole and hydroxy itraconazole (OH-itraconazole), in plasma and tissues were measured by a method described previously.6
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Results |
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Efficacy of itraconazole solution administered through oral cavity or intragastrically for oral and oesophageal candidiasis
We confirmed that the MIC of itraconazole is 0.016 mg/L for C. albicans TIMM2640 by the microbioassay method described by NCCLS.7
The therapeutic effects of itraconazole OS given by two routes against murine oral candidiasis were examined. As shown in Figure 1(a), oral administration of itraconazole OS at doses of 0.8, 4.0 or 20 mg/kg/day clearly decreased the number of the viable C. albicans cells in the oral cavity in a dose-dependent manner. On the other hand, when itraconazole OS was administered directly to the stomach, a dose of 20 mg/kg/day significantly lowered the number of viable C. albicans cells but 4 mg/kg/day did not. Figure 1(b) shows that the symptom scores of the tongues of the mice were improved in all groups of itraconazole OS-treated mice. It was notable that tongue lesions of the mice given 4 mg/kg/day of itraconazole OS intragastrically showed only partially improved symptoms.
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We also examined the therapeutic effect of itraconazole OS given by the two routes for oesophageal candidiasis. Figure 1(c) shows that oral administration at doses of 0.8, 4.0 or 20 mg/kg/day clearly lowered C. albicans cell number in oesophagus tubes with dose-dependence. But when was administered to the stomach, only a dose of 20 mg/kg/day decreased the cell numbers but 4 mg/kg/day did not.
These results indicate that administration of itraconazole through the oral cavity, more effectively protected the mice from oral and oesophageal candidiasis than that given intragastrically.
Concentration of itraconazole and OH-itraconazole in tongues, oesophageal tissues or plasma after itraconazole OS administration by two different routes. In oral candidiasis experiments performed by the protocol described above, tongue samples were collected to estimate concentration of itraconazole and its metabolite, OH-itraconazole 3 days after infection, that is, 24 h after the last administration of itraconazole OS. The concentration (average ± SD) of itraconazole of the tongue tissues of the groups of mice given 4 or 20 mg/kg/day of itraconazole OS orally was 7.31 ± 2.47 or 53.5 ± 47.8 ng/g, but when given intragastrically, levels were below the limit of detection (<2.5 ng/g). OH-itraconazole in all samples tested was not detectable (<10 ng/g).
We also examined the concentration of itraconazole in the oesophageal tubes of the mice in the oesophageal candidiasis similar to the case of oral candidiasis. Detectable levels of itraconazole (8.66 ± 5.17 ng/g), but no OH-itraconazole, were measured in the oesophageal tissues of the groups given 20 mg/kg/day of itraconazole OS orally, but not intragastrically (data not shown).
To investigate the pharmacokinetics of itraconazole after itraconazole OS administration, early change of blood concentration of itraconazole after a single itraconazole OS administration to normal mice was evaluated. As shown in Figure 2, the maximal concentrations of itraconazole in the plasma of the mice given itraconazole OS by oral and intragastric routes, detected 1 h after itraconazole OS administration, were 2200 and 500 ng/mL, respectively. Comparison of pharmacokinetics between the two groups of mice revealed a higher concentration of itraconazole and OH-itraconazole in the group receiving itraconazole OS intragastrically.
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Discussion |
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Here, we showed that oral administration of itraconazole OS elicited therapeutic responses superior to intragastric administration against murine oral and oesophageal candidiasis. Corresponding to these therapeutic efficiencies, concentration of itraconazole in tongues and oesophagus tissues remained at a detectable level even 24 h after the last oral administration of itraconazole OS, but not after intragastric administration. Therefore, it can be concluded that oral administration of itraconazole OS allowed a high distribution of itraconazole in oral and oesophageal lesions and achieved greater therapeutic efficiency.
The concentration profiles for itraconazole and OH-itraconazole in plasma of normal mice after itraconazole OS administration indicated that intragastric delivery increased plasma concentration of both compounds to higher levels than oral administration. These suggest that itraconazole in lesional tissues, which was detectable after oral administration of itraconazole OS, was not supplied from circulating blood.
Superior efficacy of oral intake of itraconazole OS can also be explained from another aspect: oral administration must increase the chance for interaction between C. albicans cells in situ and a high concentration of itraconazole for relatively short periods. This possible interaction of itraconazole OS to C. albicans cells or lesional tissue may allow rapid absorbance of itraconazole by the tissues, because highly lipophilic itraconazole is reported to be accumulated in cell membrane.8 Garcia et al.9 and our group10 reported that the growth of Candida cells, when pretreated with a relatively high concentration of itraconazole, was suppressed even in conditions without an antifungal agent.
We also showed that administration of itraconazole OS at a dose of 20 mg/kg/day into stomach by gastric gavage results in therapeutic activity against oral and oesophageal candidiasis. This suggests that a sufficient dose of itraconazole OS can elicit its anti-C. albicans activity without direct contact with lesional C. albicans cells, perhaps through transportation through the blood flow, although the possibility that a part of the itraconazole in gastric cavity may regurgitate to the C. albicans-infected lesions cannot be completely excluded.
Finally, we wish to discuss about the optimal manner of intake of itraconazole OS to assure obtaining the maximal therapeutic effect against oral or oesophageal candidiasis. Assuming our results obtained here can be applied to human case, itraconazole OS should be retained as long as possible in the mouth and swallowed slowly, a little at a time.
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None to declare.
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Acknowledgements |
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All authors thank financial support in part for this work by Grant-in-Aid (No. C15590401) for Scientific Research from the Ministry of Education, Culture, Science and Technology, Japan and Janssen Pharmaceutical K.K., Tokyo, Japan.
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References |
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1 Cartledge JD, Midgely J, Gazzard BG. (1997) Itraconazole solution: higher serum drug concentrations and better clinical response rates than the capsule formulation in acquired immunodeficiency syndrome patients with candidosis. J Clin Pathol 50:477–80.
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Groll AH, Wood L, Roden M, et al. (2002) Safety, pharmacokinetics, and pharmacodynamics of cyclodextrin itraconazole in pediatric patients with oropharyngeal candidiasis. Antimicrob Agents Chemother 46:2554–63.
7 National Committee for Clinical Laboratory Standards. (1997) Reference Method for Broth Dilution Antifungal Susceptibility Testing of Yeasts: Approved Standard M27-A. NCCLS, Wayne, PA, USA.
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Perfect JR, Savani DV, Durack DT. (1993) Uptake of itraconazole by alveolar macrophages. Antimicrob Agents Chemother 37:903–4.
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