Towards novel acaricide development against cattle fever tick: GPCR target validation by RNAi and chemical leads

Authors: PIETRANTONIO, PATRICIA - Texas A&M University; XIONG, CAIXING - Texas A&M University; Temeyer, Kevin

Submitted to: Meeting Abstract
Publication Type: Abstract Only
Publication Acceptance Date: 9/28/2021
Publication Date: N/A
Citation: N/A

Interpretive Summary: The southern cattle tick, Rhipicephalus (Boophilus) microplus, is considered the most economically damaging tick to worldwide livestock because it transmits potentially fatal diseases to cattle. The ticks have developed resistance to most available pesticides, so research to develop new tick control methods is critically needed. Ongoing cooperative research between Texas A&M University and the U.S. Department of Agriculture is investigating tick hormone receptors as potential targets for development of novel acaricides. Recombinant DNA and gene silencing was utilized to identify and silence tick genes encoding tick hormone receptors which indicated that interfering with the function of these receptors would reduce tick survival or reproduction. In addition, chemical analogs of the peptide hormones were synthesized and tested by Texas A&M researchers to identify hormone receptor antagonists potentially useful as novel acaricides. A patent application was filed for development of the pharmacore hormone analog as a novel acaricide.

Technical Abstract: The project advances the discovery of novel chemistries to control the tick vector R. microplus. Innovation is in the validation of G protein-coupled receptors (GPCRs) as acaricide targets, and in advancing tick neurobiology and endocrinology using a multidisciplinary approach. Aims: 1. Define pharmacological profiles of tick GPCRs expressed in CHO-K1 cells using peptide ligands and small-molecule chemical libraries. 2. Validate GPCRs as targets for tick control by RNAi silencing. 3. Perform chemical validation with tick bioassays of discovered compounds. We focused on the tick kinin signaling system that is hypothesized to regulate water balance, metamorphosis, and feeding. We silenced the pyrokinin (PK) and periviscerokinin receptors and verified PK activity on the cognate receptor. Methods: 1. To determine the identity and function of the tick kinin and pyrokinin neuropeptides, their cDNA was cloned from R. microplus synganglia and predicted the endogenous peptides; we also sequenced the kinin gene. Predicted kinins and pyrokinins and designed pyrokinin analogs were tested on the receptors expressed in CHO-K1 cells. 2. RNAi in females was performed using pyrokinin or periviscerokinin receptor dsRNA constructs validated in a luciferase cell assay system. 3. High-throughput screening (HTS) identified antagonists of the kinin receptor (KR). Results: The kinin gene encodes 17 kinins. Fourteen kinins tested on the kinin receptor were highly active (nM level). Thirty-six kinin receptor antagonists were identified by HTS; the most potent had an IC50 = 600 nM. Structure-activity relationships of the potent antagonists identified a pharmacophore needed for antagonism; a potent kinin antagonist has paralytic action on tick reproductive tissue and an international patent application was filed in 2021. PK and analog can elicit muscle contractions in ticks. Both KR- and PKR-RNAi caused delays in oviposition and hatching (P <0.05). Conclusions: Tick neuropeptide GPCRs can be interfered with dsRNA and small molecules. Peptidomimetics and small molecules aid to elucidate functions of tick neuropeptides. Financial Support: U.S. Department of Agriculture, National Institute for Food and Agriculture