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Cutaneous and mucocutaneous Candida infections are considered to be important targets for antimicrobial photodynamic therapy (PDT). Clinical application of antimicrobial PDT will require strategies that enhance microbial killing while minimizing damage to host tissue. Increasing the sensitivity of infectious agents to PDT will help achieve this goal. Our previous studies demonstrated that raising the level of oxidative stress in Candida by interfering with fungal respiration increased the efficiency of PDT. Therefore, we sought to identify compounds in clinical use that would augment the oxidative stress caused by PDT by contributing to reactive oxygen species (ROS) formation themselves. Based on the ability of the antifungal miconazole to induce ROS in Candida, we tested several azole antifungals for their ability to augment PDT in vitro. Although miconazole and ketoconazole both stimulated ROS production in Candida albicans, only miconazole enhanced the killing of C. albicans and induced prolonged fungistasis in organisms that survived PDT using the porphyrin TMP-1363 and the phenothiazine methylene blue as photosensitizers. The data suggest that miconazole could be used to increase the efficacy of PDT against C. albicans, and its mechanism of action is likely to be multifactorial., (© 2011 Wiley Periodicals, Inc. Photochemistry and Photobiology © 2011 The American Society of Photobiology.)

Background and Objective: The fungus Candida albicans commonly causes mucosal and cutaneous infections in patients with impaired immunity. We investigated the effectiveness of the photosensitizer meso-tetra (N-methyl-4-pyridyl) porphine tetra tosylate (TMP-1363) in the photodynamic treatment (PDT) of C. albicans infection in vitro and its selectivity in an animal model., Materials and Methods: The efficacy of TMP-1363 in PDT of C. albicans in vitro was compared to that of methylene blue (MB) using a colony forming unit (CFU) assay. In vivo infection in the mouse was established by inoculation of C. albicans yeast in the intradermal space of the ear pinna. Two days post-infection, 0.3 mg ml(-1) TMP-1363 was administered topically. Thirty minutes after TMP-1363 application, the ears were irradiated at 514 nm using a fluence of 90 J cm(-2) delivered at an irradiance of 50 mW cm(-2) . The ears were excised 2 hours post-irradiation, homogenized, and the organism burden was determined by a CFU assay. In vivo wide field and confocal fluorescence imaging assessed the localization of the photosensitizer in relationship to C. albicans., Results: Photosensitization with TMP-1363 resulted in a greater than three-log increase in killing of C. albicans in vitro compared to MB. In vivo fluorescence imaging demonstrated a high degree of selective labeling of C. albicans by TMP-1363. PDT of infection using TMP-1363 resulted in a significant reduction in CFU/ear relative to untreated controls. Infected ears subjected to PDT displayed complete healing over time with no observable damage to the pinna., Conclusion: Our in vitro and in vivo findings support TMP-1363-mediated PDT as a viable therapeutic approach for the PDT of candidiasis., (Copyright © 2011 Wiley-Liss, Inc.)

Respiratory deficiency increases the sensitivity of the pathogenic fungi Candida albicans and Candida glabrata to oxidative stress induced by photodynamic therapy (PDT) sensitized by the cationic porphyrin meso-tetra (N-methyl-4-pyridyl) porphine tetra tosylate (TMP-1363). Since disruption of electron transport chain (ETC) function increases intracellular levels of reactive oxygen species in yeast, we determined whether interference with ETC assembly or function increased sensitivity to TMP-1363-PDT in C. albicans, C. glabrata and the non-pathogenic yeast Saccharomyces cerevisiae. Metabolic inhibitor antimycin A and defined genetic mutants were used to identify ETC components that contribute to the sensitivity to PDT. Inhibition of cytochrome bc(1) (Complex III) with antimycin A increases mitochondrial levels of reactive oxygen species. PDT performed following pre-treatment with antimycin A reduced colony forming units (CFU) of C. albicans and C. glabrata by approximately two orders of magnitude relative to PDT alone. A S. cerevisiae mitochondrial glutaredoxin grx5 mutant, defective in assembly of Fe-S clusters critical for Complex III function, displayed increased sensitivity to PDT. Furthermore, C. glabrata and S.cerevisiae mutants in cytochrome c oxidase (Complex IV) synthesis and assembly were also significantly more sensitive to PDT. These included suv3, encoding an ATP-dependent RNA helicase critical for maturation of cytochrome c oxidase subunit transcripts, and pet117, encoding an essential cytochrome c oxidase assembly factor. Following PDT, the reduction in CFU of these mutants was one to two orders of magnitude greater than in their respective parental strains. The data demonstrate that selective inhibition of ETC Complexes III and IV significantly increases the sensitivity of C. albicans, C. glabrata and S. cerevisiae to PDT sensitized with TMP-1363., (2010 Elsevier B.V. All rights reserved.)

Mucosal infections caused by the pathogenic fungus Candida are a significant infectious disease problem and are often difficult to eradicate because of the high frequency of resistance to conventional antifungal agents. Photodynamic treatment (PDT) offers an attractive therapeutic alternative. Previous studies demonstrated that filamentous forms and biofilms of Candida albicans were sensitive to PDT using Photofrin as a photosensitizer. However, early stationary phase yeast forms of C. albicans and Candida glabrata were not adversely affected by treatment. We report that the cationic porphyrin photosensitizer meso-tetra (N-methyl-4-pyridyl) porphine tetra tosylate (TMP-1363) is effective in PDT against yeast forms of C. albicans and C. glabrata. Respiratory-deficient (RD) strains of C. albicans and C. glabrata display a pleiotropic resistance pattern, including resistance to members of the azole family of antifungals, the salivary antimicrobial peptides histatins and other types of toxic stresses. In contrast to this pattern, RD mutants of both C. albicans and C. glabrata were significantly more sensitive to PDT compared to parental strains. These data suggest that intact mitochondrial function may provide a basal level of anti-oxidant defense against PDT-induced phototoxicity in Candida, and reveals pathways of resistance to oxidative stress that can potentially be targeted to increase the efficacy of PDT against this pathogenic fungus.

Treatment of mucocutaneous and cutaneous Candida albicans infections with photosensitizing agents and light, termed photodynamic therapy (PDT), offers an alternative to conventional treatments. Initial studies using the clinically approved photosensitizer Photofrin demonstrated the susceptibility of C. albicans to its photodynamic effects. In the present study, we have further refined parameters for Photofrin-mediated photodynamic action against C. albicans and examined whether mechanisms commonly used by microorganisms to subvert either antimicrobial oxidative defenses or antimicrobial therapy, including biofilm formation, were operative. In buffer and defined medium, germ tubes preloaded with Photofrin retained their photosensitivity for up to 2 hours, indicating the absence of degradation or export of Photofrin by the organism. The addition of serum resulted in a gradual loss of photosensitivity over 2 hours. In contrast to an adaptive response by germ tubes to oxidative stress by hydrogen peroxide, there was no adaptive response to singlet oxygen-mediated stress by photodynamic action. C. albicans biofilms were sensitive to Photofrin-mediated phototoxicity in a dose-dependent manner. Finally, the metabolic activity of C. albicans biofilms following photodynamic insult was significantly lower than that of biofilms treated with amphotericin B for the same time period. These results demonstrate that several of the mechanisms microorganisms use to subvert either antimicrobial oxidative defenses or antimicrobial therapy are apparently not operative during Photofrin-mediated photodynamic treatment of C. albicans. These observations provide support and rationale for the continued investigation of PDT as an adjunctive, or possibly alternative, mode of therapy against cutaneous and mucocutaneous candidiasis.

ABSTRACT Cutaneous and mucocutaneous Candida infections are considered to be important targets for antimicrobial photodynamic therapy (PDT). Clinical application of antimicrobial PDT will require strategies that enhance microbial killing while minimizing damage to host tissue. Increasing the sensitivity of infectious agents to PDT will help achieve this goal. Our previous studies demonstrated that raising the level of oxidative stress in Candida by interfering with fungal respiration increased the efficiency of PDT. Therefore, we sought to identify compounds in clinical use that would augment the oxidative stress caused by PDT by contributing to reactive oxygen species (ROS) formation themselves. Based on the ability of the antifungal miconazole to induce ROS in Candida, we tested several azole antifungals for their ability to augment PDT in vitro. Although miconazole and ketoconazole both stimulated ROS production in Candida albicans, only miconazole enhanced the killing of C. albicans and induced prolonged fungistasis in organisms that survived PDT using the porphyrin TMP-1363 and the phenothiazine methylene blue as photosensitizers. The data suggest that miconazole could be used to increase the efficacy of PDT against C. albicans, and its mechanism of action is likely to be multifactorial. [ABSTRACT FROM AUTHOR]

ABSTRACT Cutaneous and mucocutaneous Candida infections are considered to be important targets for antimicrobial photodynamic therapy (PDT). Clinical application of antimicrobial PDT will require strategies that enhance microbial killing while minimizing damage to host tissue. Increasing the sensitivity of infectious agents to PDT will help achieve this goal. Our previous studies demonstrated that raising the level of oxidative stress in Candida by interfering with fungal respiration increased the efficiency of PDT. Therefore, we sought to identify compounds in clinical use that would augment the oxidative stress caused by PDT by contributing to reactive oxygen species (ROS) formation themselves. Based on the ability of the antifungal miconazole to induce ROS in Candida, we tested several azole antifungals for their ability to augment PDT in vitro. Although miconazole and ketoconazole both stimulated ROS production in Candida albicans, only miconazole enhanced the killing of C. albicans and induced prolonged fungistasis in organisms that survived PDT using the porphyrin TMP-1363 and the phenothiazine methylene blue as photosensitizers. The data suggest that miconazole could be used to increase the efficacy of PDT against C. albicans, and its mechanism of action is likely to be multifactorial. [ABSTRACT FROM AUTHOR]