ABSTRACT
In recent decades, the incidence of candidemia in tertiary hospitals worldwide has substantially increased. These infections are a major cause of morbidity and mortality; in addition, they prolong hospital stays and raise the costs associated with treatment. Studies have reported a significant increase in infections by non-albicans Candida species, especially C. tropicalis. The number of antifungal drugs on the market is small in comparison to the number of antibacterial agents available. The limited number of treatment options, coupled with the increasing frequency of cross-resistance, makes it necessary to develop new therapeutic strategies. The objective of this study was to evaluate and compare the antifungal activities of three semisynthetic naphthofuranquinone molecules against fluconazole-resistant Candida spp. strains. These results allowed to us to evaluate the antifungal effects of three naphthofuranquinones on fluconazole-resistant C. tropicalis. The toxicity of these compounds was manifested as increased intracellular ROS, which resulted in membrane damage and changes in cell size/granularity, mitochondrial membrane depolarization, and DNA damage (including oxidation and strand breakage). In conclusion, the tested naphthofuranquinones (compounds 1-3) exhibited in vitro cytotoxicity against fluconazole-resistant Candida spp. strains.
Subject(s)
Antifungal Agents/pharmacology , Candida/drug effects , Drug Resistance, Fungal/drug effects , Fluconazole/pharmacology , Naphthoquinones/pharmacology , Animals , Antifungal Agents/chemical synthesis , Antifungal Agents/chemistry , Candida/classification , Candida/genetics , Candida tropicalis/drug effects , Candida tropicalis/genetics , Candida tropicalis/metabolism , Cell Line , Cell Survival/drug effects , DNA Damage , DNA, Fungal/chemistry , DNA, Fungal/genetics , DNA, Fungal/metabolism , DNA, Ribosomal Spacer/chemistry , DNA, Ribosomal Spacer/genetics , Fibroblasts/cytology , Fibroblasts/drug effects , Membrane Potential, Mitochondrial/drug effects , Microbial Sensitivity Tests , Models, Chemical , Molecular Sequence Data , Molecular Structure , Naphthoquinones/chemical synthesis , Naphthoquinones/chemistry , Phosphatidylserines , RNA, Ribosomal, 5.8S/genetics , Reactive Oxygen Species/metabolism , Sequence Analysis, DNAABSTRACT
Naked plasmid DNA and DNA/liposome complexes are currently being considered as gene therapy treatments for cystic fibrosis (CF) pulmonary disease. Current methods of gene delivery to the airways result only in transient correction of the CF ion transport defect, and disease treatment is likely to require repeated administrations of vector. However, it is unclear if repeat administration will be tolerated by CF individuals. Technologies including TaqMan (Applied Biosystems) real-time quantitative reverse transcription-polymerase chain reaction (RT-PCR) can be used to determine the efficacy of gene transfer formulations. TaqMan RT-PCR assays were designed and optimised to detect plasmid vector-derived and endogenous gene expression. Subsequently, these assays were used to quantify vector-derived mRNA after delivery of naked DNA and DNA/liposome formulations expressing human and murine cystic fibrosis transmembrane conductance regulator (CFTR) to the mouse airways. Vector-derived mRNA was detected in samples following the delivery of naked DNA or DNA/liposomes to the mouse airways, and no reduction in vector-derived mRNA was observed upon repeat administration, a finding that is consistent with the murine and human CFTR being tolerated by the mouse. Although it remains to be seen if CF patients can tolerate long-term expression of wild-type CFTR, these data demonstrate that TaqMan RT-PCR is an effective tool to accurately quantify transgene expression in the airways.
