Immunotherapy in Paracoccidioidomycosis: evaluation of combined therapy with antifungals and MDSCs depletion by 5-fluorouracil.

Abstract

Paracoccidioidomycosis (PCM) is a systemic fungal infection caused by the fungus Paracoccidiodes brasiliensis. It is an endemic disease in the Americas, with a higher incidence in Brazil. The lungs are affected in almost all adult patients, in addition to the oral, pharyngeal and laryngeal mucosal tissues. Current treatment is based on the use of the antifungals itraconazole, amphotericin B and fluoconazole. However, long-term treatment causes treatment abandonment and/or sequelae, due to drug toxicity, chronic inflammatory processes and fibrosis, which can profoundly affect organ function. Previous studies have revealed that host immunity is strongly regulated by several suppressive mechanisms mediated by tolerogenic plasmacytoid dendritic cells, the enzyme 2,3 indoleamine dioxygenase (IDO1) and regulatory T cells. IDO1 activity has also been demonstrated to orchestrate local and systemic immunosuppressive effects through the recruitment and activation of myeloid lineage-derived suppressor cells (MDSCs), a heterogeneous population of myeloid cells with a strong ability to suppress T cell responses. Immune responses and tissue repair in healthy individuals and expand during a microbial infection. In this context, 5-Fluorouracil (5-FU) appears, a chemotherapy drug that has the ability to stimulate the immune responses. Cancer studies show that 5-FU selectively eliminates circulating MDSCs through apoptosis without harming either B and T lymphocytes or NK cells. In PCM, the administration of 5-FU in infected mice resulted in a controlled disease, with reduced fungal load in target organs and longer survival time of the animals. Importantly, disease improvement was associated with an increase in protective immunological responses, such as Th1 and Th17. Therefore, this project aims to evaluate the combined therapy of 5-FU with antifungals already used in the treatment of PMC, in order to evaluate its effect on modulating the host's immune system and effectiveness in eliminating the fungus. To this end, C57BL/6 mice will be inoculated with 1x106 P. brasiliensis yeast. After 6 weeks, the animals will be treated with 5-FU in combination or not with amphotericin B, fluconozole or itraconazole. After 2 weeks of treatment, the animals will be evaluated for the course of the disease by CFU, histopathology and morbidity. Additionally, ELISA and flow cytometry will be performed in order to verify the immunological response resulting from the use of this new therapeutic procedure. In an additional protocol, the animals will also be evaluated 3 weeks after the end of treatments. In this way, we intend to develop a protocol that aims to interrupt the course of the disease by controlling fungal growth associated with the restoration of the immune response which, together, could result in significant improvement or even cure of this chronic infection.