Searching for effective and safe antifungals against cryptococcosis among halogenated carboxylic acids and their derivatives


Mariusz Dyląg1, Ewa Leniak2, Weronika Jasińska3, Maria Kozioł-Gałczyńska4

1Department of Genetics, Institute of Genetics and Microbiology, University of Wroclaw, Wroclaw, Poland
2Max Planck Institute of Molecular Plant Physiology, Am Mühlenberg 1, 14476 Potsdam-Golm, Germany
3Department of Life Sciences, Ben-Gurion University of the Negev, Beersheba, Israel
4Department of Dermatology, Venereology and Allergology, Wroclaw Medical University,
Chalubinskiego Street 1, 50-368 Wroclaw, Poland


Objectives:
The aim of these studies was the evaluation of anticryptococcal activity of chosen halogenated carboxylic acids and their derivatives. Equally important was the cytotoxicity assessment of the tested molecules toward mammalian cells. For the most active compounds were determined types of interaction with antifungal drugs commonly used in the therapy of cryptococcosis.

Methods:
The antifungal susceptibility testing was performed based on CLSI M27-A3 (2008) protocol. Evaluation of cytotoxicity of the tested compounds was performed on human lung (CCL-210TM and A549) and mouse fibroblast (L929) cell lines by sulphorodamine B (SRB) assay. To assess the interactions between amphotericin B (AMB) or fluconazole (FLC) and one of two the most active compounds 2D checkerboard microdilution test was performed in RPMI-1640 medium. In case of 3BP and 3-BrOP (data not shown) type of interaction with para-aminobenzoic acid (PABA) was investigated by spot tests method on caffeic acid agar (CAA) medium.

Results:
Our studies demonstrate very different anticryptococcal activity of the tested compounds. The highest antifungal effect was achieved for 3-bromopyruvate (3BP) and its propyl ester derivative (3-BrOP). In case of Cryptococcus neoformans tested strains the MIC values were in the range 0.12-0.15 mM and 0.06-0.08 mM, respectively. The highest susceptibility toward these compounds showed all the C. neoformans strains followed by Cryptococcus gattii and other environmental, non-pathogenic Cryptococcus spp. strains. Despite these results the cytotoxicity tests allow to continue more advanced studies only for 3BP and 3-BrOP. These compounds are non-toxic even at concentrations 200- and 400-times higher than determined MIC values, respectively. Moreover, for these pyruvate derivatives hipersynergistic and neutral effect with AMB and FLC was showed, respectively. Additionally, melanin synthesis inhibitor – PABA also act in synergistic manner with 3BP and 3-BrOP.

Conclusion:
Considering that cryptococcosis is responsible for 15-17% AIDS-related deaths globally and treatment options are limited mainly to AMB and FLC our results have strongly applicable character. We just showed that 3-bromopyruvate and its propyl ester derivative, especially in combination with amphotericin B could be developed as novel anticryptococcal drugs.