The identification of new pathways and compounds that can enhance caspofungin activity against Aspergillus fumigatus

Abstract

Invasive Pulmonary Aspergillosis (IPA), mainly caused by Aspergillus fumigatus, is a major infectious complication with mortality rates as high as 90% depending on the immunological status of the host. Azoles are fungicidal drugs for A. fumigatus and the main antifungal agents recommended for IPA. Echinocandins, such as caspofungin (CSP), represent a second line therapy and target the fungal cell wall by inhibiting 1,3-beta-D-glucan synthase, which is responsible for the assembly of beta-D-glucan, a major component of the fungal cell wall. CSP is fungistatic against A. fumigatus and treatment with CSP leads to hyphae with defects in growth and morphology. One of the objectives of our project is to investigate how the signal transduction for the fungal response to CSP is mediated via oxylipins. The fungal oxylipin 5,8-dihydroxyoctadecadienoic acid (5S,8R-diHODE) is important for cellular differentiation and lateral branching in A. fumigatus. CSP induces A. fumigatus hyperbranching and phenocopies the increased lateral branching observed when this fungus is exposed to 5S,8R-diHODE. Preliminary results from our laboratory indicate that the null mutant for ppoA gene that encodes the fatty acid oxygenase 5,8-linoleate diol synthase (5,8-LDS) responsible for the formation of 5S,8R-diHODE is resistant to CSP while the strain overexpressing ppoA is more sensitive to CSP than the corresponding wild-type strain. We also observed that the null mutant for ppoC (ppoC encodes a fatty acid dehydrogenase) has lost the Caspofungin Paradoxical Effect (CPE). We propose to identify through a combination of RNAseq, metabolomics and molecular genetics which genes are modulated by 5S,8R-LDs, 8R-HODE, and CSP. We will identify these genes by comparing the corresponding wild-type strain with the null mutants of ppoA and ppoC, and also by using representative strains from a collection of about 70 worldwide A. fumigatus clinical isolates with different degrees of CSP sensitivity and tolerance. As a second objective, we will also evaluate A. fumigatus wild-type and null ppoA mutant growth alone or synergistically with CSP and libraries of compounds and chemical probes that could synergize and potentiate CSP. Finally, as a third objective, we will investigate a collection of 11 transcription factors previously implicated in caspofungin resistance and CPE and will determine if they are modulated by oxylipins. This can open new avenues for the identification of new metabolic pathways involved in caspofungin resistance and tolerance and can provide the development of novel CSP-based antifungal therapy. (AU)