Randomized, phase 1/2, double-blind pioglitazone repositioning trial combined with antifungals for the treatment of cryptococcal meningitis - PIO study.

BACKGROUND: Cryptococcosis affects more than 220,000 patients/year, with high mortality even when the standard treatment [amphotericin B (AMB), 5-flucytosin (5-FC) and fluconazole] is used. AMB presents high toxicity and 5-FC is not currently available in Brazil. In a pre-clinical study, pioglitazone (PIO - an antidiabetic drug) decreased AMB toxicity and lead to an increased mice survival, reduced morbidity and fungal burden in brain and lungs. The aim of this trial is to evaluate the efficacy and safety of PIO combined with standard antifungal treatment for human cryptococcosis. METHODS: A phase 1/2, randomized, double blind, placebo-controlled trial will be performed with patients from Belo Horizonte, Brazil. They will be divided into three groups (placebo, PIO 15 mg/day or PIO 45 mg/day) and will receive an additional pill during the induction phase of cryptococcosis' treatment. Our hypothesis is that treated patients will have increased survival, so the primary outcome will be the mortality rate. Patients will be monitored for survival, side effects, fungal burden and inflammatory mediators in blood and cerebrospinal fluid. The follow up will occur for up 60 days. CONCLUSIONS: We expect that PIO will be an adequate adjuvant to the standard cryptococcosis' treatment. TRIAL REGISTRATION: ICTRP/WHO (and International Clinical Trial Registry Plataform (ICTRP/WHO) (http://apps.who.int/trialsearch/Trial2.aspx?TrialID=RBR-9fv3f4), RBR-9fv3f4 (http://www.ensaiosclinicos.gov.br/rg/RBR-9fv3f4). UTN Number: U1111-1226-1535. Ethical approvement number: CAAE 17377019.0.0000.5149.

N-acetylcysteine reduces amphotericin B deoxycholate nephrotoxicity and improves the outcome of murine cryptococcosis.

Cryptococcosis is a life-threatening fungal infection, and its current treatment is toxic and subject to resistance. Drug repurposing represents an interesting approach to find drugs to reduce the toxicity of antifungals. In this study, we evaluated the combination of N-acetylcysteine (NAC) with amphotericin B (AMB) for the treatment of cryptococcosis. We examined the effects of NAC on fungal morphophysiology and on the macrophage fungicidal activity 3 and 24 hours post inoculation. The therapeutic effects of NAC combination with AMB were investigated in a murine model with daily treatments regimens. NAC alone reduced the oxidative burst generated by AMB in yeast cells, but did not inhibit fungal growth. The combination NAC + AMB decreased capsule size, zeta potential, superoxide dismutase activity and lipid peroxidation. In macrophage assays, NAC + AMB did not influence the phagocytosis, but induced fungal killing with different levels of oxidative bursts when compared to AMB alone: there was an increased reactive oxygen species (ROS) after 3 hours and reduced levels after 24 hours. By contrast, ROS remained elevated when AMB was tested alone, demonstrating that NAC reduced AMB oxidative effects without influencing its antifungal activity. Uninfected mice treated with NAC + AMB had lower concentrations of serum creatinine and glutamate-pyruvate transaminase in comparison to AMB. The combination of NAC + AMB was far better than AMB alone in increasing survival and reducing morbidity in murine-induced cryptococcosis, leading to reduced fungal burden in lungs and brain and also lower concentrations of pro-inflammatory cytokines in the lungs. In conclusion, NAC + AMB may represent an alternative adjuvant for the treatment of cryptococcosis.

Pioglitazone as an adjuvant of amphotericin B for the treatment of cryptococcosis.

Approximately 180,000 people worldwide die from cryptococcosis each year, probably due to the ineffectiveness and toxicity of drugs currently available to treat the disease. Amphotericin B (AMB) is effective for killing the fungus, but has serious adverse effects linked to excessive production of reactive oxygen species which compromise renal function. Pioglitazone (PIO) is a peroxisome proliferator-activated receptor-γ agonist widely repositioned as an adjuvant of various drugs that have toxic effects due to its antioxidant and anti-inflammatory effects. This study evaluated PIO in combination with AMB for the treatment of cryptococcosis. PIO was found to reduce serum creatinine and glutamic-oxalacetic transaminase levels in mice treated with PIO+AMB. In vitro, PIO was able to control harmful oxidative bursts induced by AMB without compromising the antifungal effect. In vivo, PIO+AMB increased the survival rate compared with AMB alone, and improved the morbidity of the animals. PIO+AMB was more efficient than AMB alone for inhibiting fungal transmigration from the lungs to the brain, and killing yeasts that reached the central nervous system, avoiding the establishment of meningoencephalitis. In a phagocytosis assay, PIO did not influence the engulfment and fungicidal activity of macrophages induced by AMB, but reduced the oxidative bursts after the reduction of fungal burden, pointing to control of the pathogen without leading to excessive stress which can be damaging to the host. In conclusion, PIO+AMB was found to ameliorate cryptococcosis in a murine model, indicating that it is a promising therapeutic adjuvant for combating and controlling this fungal infection.