title: Macrocyclic lactones differ in interaction with recombinant P-glycoprotein 9 of the parasitic nematode Cylicocylus elongatus and ketoconazole in a yeast growth assay.
abstract: Macrocyclic lactones (MLs) are widely used parasiticides against nematodes and arthropods, but resistance is frequently observed in parasitic nematodes of horses and livestock. Reports claiming resistance or decreased susceptibility in human nematodes are increasing. Since no target site directed ML resistance mechanisms have been identified, non-specific mechanisms were frequently implicated in ML resistance, including P-glycoproteins (Pgps, designated ABCB1 in vertebrates). Nematode genomes encode many different Pgps (e.g. 10 in the sheep parasite Haemonchus contortus). ML transport was shown for mammalian Pgps, Pgps on nematode egg shells, and very recently for Pgp-2 of H. contortus. Here, Pgp-9 from the equine parasite Cylicocyclus elongatus (Cyathostominae) was expressed in a Saccharomyces cerevisiae strain lacking seven endogenous efflux transporters. Pgp was detected on these yeasts by flow cytometry and chemiluminescence using the monoclonal antibody UIC2, which is specific for the active Pgp conformation. In a growth assay, Pgp-9 increased resistance to the fungicides ketoconazole, actinomycin D, valinomycin and daunorubicin, but not to the anthelmintic fungicide thiabendazole. Since no fungicidal activity has been described for MLs, their interaction with Pgp-9 was investigated in an assay involving two drugs: Yeasts were incubated with the highest ketoconazole concentration not affecting growth plus increasing concentrations of MLs to determine competition between or modulation of transport of both drugs. Already equimolar concentrations of ivermectin and eprinomectin inhibited growth, and at fourfold higher ML concentrations growth was virtually abolished. Selamectin and doramectin did not increase susceptibility to ketoconazole at all, although doramectin has been shown previously to strongly interact with human and canine Pgp. An intermediate interaction was observed for moxidectin. This was substantiated by increased binding of UIC2 antibodies in the presence of ivermectin, moxidectin, daunorubicin and ketoconazole but not selamectin. These results demonstrate direct effects of MLs on a recombinant nematode Pgp in an ML-specific manner.
authors: Institute for Parasitology and Tropical Veterinary Medicine, Freie Universität Berlin, Berlin, Germany.
authors: Maximiliane
authors: M
authors: Kaschny
authors: Institute for Parasitology and Tropical Veterinary Medicine, Freie Universität Berlin, Berlin, Germany.
authors: Janina
authors: J
authors: Demeler
authors: Institute for Parasitology and Tropical Veterinary Medicine, Freie Universität Berlin, Berlin, Germany.
authors: I Jana I
authors: IJ
authors: Janssen
authors: Schmalhausen Institute of Zoology, National Academy of Sciences of Ukraine, Kyiv, Ukraine.
authors: Tetiana A
authors: TA
authors: Kuzmina
authors: Zoetis, Paris, France.
authors: Bruno
authors: B
authors: Besognet
authors: Zoetis International Services, Paris, France.
authors: Theo
authors: T
authors: Kanellos
authors: INRA, ISP, Animal Infection and Public Health, Nouzilly, France.
authors: Dominique
authors: D
authors: Kerboeuf
authors: Institute for Parasitology and Tropical Veterinary Medicine, Freie Universität Berlin, Berlin, Germany.
authors: Georg
authors: G
authors: von Samson-Himmelstjerna
authors: Institute for Parasitology and Tropical Veterinary Medicine, Freie Universität Berlin, Berlin, Germany.
authors: Jürgen
authors: J
authors: Krücken
mesh_terms: D018435:ATP Binding Cassette Transporter, Subfamily B / Q000378:metabolism*; D000818:Animals; D000977:Antiparasitic Agents / Q000494:pharmacology*; D015153:Blotting, Western; D002469:Cell Separation; D004351:Drug Resistance / Q000187:drug effects / Q000502:physiology*; D007654:Ketoconazole / Q000494:pharmacology; D007783:Lactones; D047028:Macrocyclic Compounds / Q000494:pharmacology*; D008969:Molecular Sequence Data; D009348:Nematoda / Q000187:drug effects*; D010802:Phylogeny; D016133:Polymerase Chain Reaction; D015003:Yeasts / Q000187:drug effects / Q000254:growth & development
chemical_list: D018435:ATP Binding Cassette Transporter, Subfamily B; D000977:Antiparasitic Agents; D007783:Lactones; D047028:Macrocyclic Compounds; D007654:Ketoconazole