title: Global analysis of the evolution and mechanism of echinocandin resistance in Candida glabrata.
abstract: The evolution of drug resistance has a profound impact on human health. Candida glabrata is a leading human fungal pathogen that can rapidly evolve resistance to echinocandins, which target cell wall biosynthesis and are front-line therapeutics for Candida infections. Here, we provide the first global analysis of mutations accompanying the evolution of fungal drug resistance in a human host utilizing a series of C. glabrata isolates that evolved echinocandin resistance in a patient treated with the echinocandin caspofungin for recurring bloodstream candidemia. Whole genome sequencing identified a mutation in the drug target, FKS2, accompanying a major resistance increase, and 8 additional non-synonymous mutations. The FKS2-T1987C mutation was sufficient for echinocandin resistance, and associated with a fitness cost that was mitigated with further evolution, observed in vitro and in a murine model of systemic candidemia. A CDC6-A511G(K171E) mutation acquired before FKS2-T1987C(S663P), conferred a small resistance increase. Elevated dosage of CDC55, which acquired a C463T(P155S) mutation after FKS2-T1987C(S663P), ameliorated fitness. To discover strategies to abrogate echinocandin resistance, we focused on the molecular chaperone Hsp90 and downstream effector calcineurin. Genetic or pharmacological compromise of Hsp90 or calcineurin function reduced basal tolerance and resistance. Hsp90 and calcineurin were required for caspofungin-dependent FKS2 induction, providing a mechanism governing echinocandin resistance. A mitochondrial respiration-defective petite mutant in the series revealed that the petite phenotype does not confer echinocandin resistance, but renders strains refractory to synergy between echinocandins and Hsp90 or calcineurin inhibitors. The kidneys of mice infected with the petite mutant were sterile, while those infected with the HSP90-repressible strain had reduced fungal burden. We provide the first global view of mutations accompanying the evolution of fungal drug resistance in a human host, implicate the premier compensatory mutation mitigating the cost of echinocandin resistance, and suggest a new mechanism of echinocandin resistance with broad therapeutic potential.
authors.affiliation: Department of Molecular Genetics, University of Toronto, Toronto, Ontario, Canada.
authors.forename: Sheena D
authors.initials: SD
authors.lastname: Singh-Babak
authors.forename: Tomas
authors.initials: T
authors.lastname: Babak
authors.forename: Stephanie
authors.initials: S
authors.lastname: Diezmann
authors.forename: Jessica A
authors.initials: JA
authors.lastname: Hill
authors.forename: Jinglin Lucy
authors.initials: JL
authors.lastname: Xie
authors.forename: Ying-Lien
authors.initials: YL
authors.lastname: Chen
authors.forename: Susan M
authors.initials: SM
authors.lastname: Poutanen
authors.forename: Robert P
authors.initials: RP
authors.lastname: Rennie
authors.forename: Joseph
authors.initials: J
authors.lastname: Heitman
authors.forename: Leah E
authors.initials: LE
authors.lastname: Cowen
mesh_terms: D000935:Antifungal Agents / Q000494:pharmacology* / Q000627:therapeutic use; D001483:Base Sequence; D019703:Calcineurin / Q000235:genetics / Q000378:metabolism; D041221:Candida glabrata / Q000187:drug effects* / Q000235:genetics* / Q000378:metabolism; D058387:Candidemia / Q000188:drug therapy / Q000382:microbiology; D000077336:Caspofungin; D025141:Drug Resistance, Fungal / Q000235:genetics*; D054714:Echinocandins / Q000494:pharmacology*; D019143:Evolution, Molecular; D005260:Female; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism; D016681:Genome, Fungal; D005964:Glucosyltransferases / Q000235:genetics; D018841:HSP90 Heat-Shock Proteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D055666:Lipopeptides; D008826:Microbial Sensitivity Tests; D008875:Middle Aged; D011110:Polymorphism, Genetic; D017422:Sequence Analysis, DNA
chemical_list: D000935:Antifungal Agents; D054714:Echinocandins; D005656:Fungal Proteins; D018841:HSP90 Heat-Shock Proteins; D055666:Lipopeptides; D005964:Glucosyltransferases; D019703:Calcineurin; D000077336:Caspofungin