The novel isoxazoline ectoparasiticide fluralaner: Selective inhibition of arthropod γ-aminobutyric acid- and l-glutamate-gated chloride channels and insecticidal/acaricidal activity

Abstract

Isoxazolines are a novel class of parasiticides that are potent inhibitors of γ-aminobutyric acid (GABA)-gated chloride channels (GABACls) and L-glutamate-gated chloride channels (GluCls). In this study, the effects of the isoxazoline drug fluralaner on insect and acarid GABACl (RDL) and GluCl and its parasiticidal potency were investigated. We report the identification and cDNA cloning of Rhipicephalus (R.) microplus RDL and GluCl genes, and their functional expression in Xenopus laevis oocytes. The generation of six clonal HEK293 cell lines expressing Rhipicephalus microplus RDL and GluCl, Ctenocephalides felis RDL-A285 and RDL-S285, as well as Drosophila melanogaster RDLCl-A302 and RDL-S302, combined with the development of a membrane potential fluorescence dye assay allowed the comparison of ion channel inhibition by fluralaner with that of established insecticides addressing RDL and GluCl as targets. In these assays fluralaner was several orders of magnitude more potent than picrotoxinin and dieldrin, and performed 5-236 fold better than fipronil on the arthropod RDLs, while a rat GABACl remained unaffected. Comparative studies showed that R. microplus RDL is 52-fold more sensitive than R. microplus GluCl to fluralaner inhibition, confirming that the GABA-gated chloride channel is the primary target of this new parasiticide. In agreement with the superior RDL on-target activity, fluralaner outperformed dieldrin and fipronil in insecticidal screens on cat fleas (Ctenocephalides felis), yellow fever mosquito larvae (Aedes aegypti) and sheep blowfly larvae (Lucilia cuprina), as well as in acaricidal screens on cattle tick (R. microplus) adult females, brown dog tick (Rhipicephalus sanguineus) adult females and Ornithodoros moubata nymphs. These findings highlight the potential of fluralaner as a novel ectoparasiticide.

Novel GABA receptor pesticide targets

“…Several types of evidence establish that isoxazolines [57][58][59][60][61][62][63][64][65][66] and meta-diamides (including mDA-7 [24,67] and BPB-1 [68]) are NCAs but at a different high affinity site(s) than those for compounds acting at the NCA-IA, NCA-IB and AVE targets ( …”

“…A 2012 compilation considering the chronology and numbers for introduction of the current insecticides gave the GABA-R targeting compounds as only 1.7% of the total with half of them introduced by 1955 [81]. Newly introduced GABAergic insecticides are flu and afoxolaner used for flea and tick control on cats and dogs [58,[62][63][64][65]. There are also important crop pests highly sensitive to NCA-IIs [24,60,82] indicating possible expanded use on optimization.…”

Contemporary Asymmetric Phase Transfer Catalysis: Large-Scale Industrial Applications

“…52 For instance, Bravecto (106, fluralaner) is an insecticide developed by Nissan Chemical Industries, Ltd. and Merck, which was approved by US Food and Drug Administration (FDA) in 2014 as a flea treatment in dogs. 53 To address the synthetic challenge of this structural motif, an elegant cascade conjugate addition/cyclization approach 54 was reported by Matoba at Nissan Chemical Industries, Ltd. 55a By using the cinchona-derived PTC 108a, the desired trifluoromethyl-substituted 2-isoxazoline 109 was obtained in 94% yield and 54% ee from an E/Z isomeric mixture of 107 (Scheme 19). Later, the asymmetric synthesis of structurally similar compound 112 was reported by Toyama and co-workers also at Nissan Chemical Industries, Ltd. By using the different PTC 111, the desired cyclization product 112 was obtained in 97% yield and 94% ee (Scheme 19).…”

The speed of kill of fluralaner (Bravecto¿) against Ixodes ricinus ticks on dogs

“…Fluralaner's mode of action is antagonism of ligandgated chloride channels (both gamma-aminobutyric acid-(GABA) receptor and glutamate-receptor) that potently inhibit the arthropod nervous system [10], resulting in paralysis and death of fleas and ticks [11]. Fluralaner has a significantly high selectivity for arthropod versus mammalian neurons [10,12] and is well tolerated by dogs that are at least 8 weeks old, including MDR 1 (−/−) Collies [13,14].…”