Oral Presentation 28th Annual Lorne Proteomics Symposium 2023

Chemoproteomic Target Deconvolution Approaches in Giardia duodenalis Using Kinase Inhibitors (#16)

Alex Lam 1 , Samantha Emery-Corbin 1 , Louise Baker 1 , Guillaume Lessene 1 , Subash Adhikari 1 , Jumana Yousef 1 , Aaron Jex 1 2
  1. Walter and Eliza Hall Institute of Medical Research, Melbourne, VICTORIA, Australia
  2. Faculty of Veterinary and Agricultural Sciences, The University of Melbourne, Melbourne, VIC, Australia

Giardia duodenalis is a gastrointestinal parasite causing ~200 million symptomatic infections annually, disproportionately affecting children in lower socioeconomic tiers where it causes malnutrition, stunting in physical and cognitive growth. Chemotherapeutic treatments for such infections are limited to nitroheterocyclic antibiotics such as metronidazole. However, high doses of these drugs are toxic and treatment failure related to drug-resistance occurs in up to 20% of cases. This highlights a need for a novel and safer chemotherapeutic option for treating Giardia infections.

 

The Giardia kinome is an attractive druggable space, as the NEK (Never in Mitosis A related kinases) family of kinases have been expanded in this genus’ otherwise reduced kinome (198/278 of annotated kinases). We performed a medium-throughput in vitro screen of a published kinase inhibitor library and identified a putative compound class that is effective against Giardia at sub-micromolar concentrations. As a next step, we have undertaken drug-target identification using the Proteome Integral Solubility Alteration (PISA) assay and affinity-chromatography “pulldown” with MS-based identification. In the former, we shortlisted 63 proteins demonstrating dose-dependent thermal-(de)stabilisation resulting from ligand binding. Four of these 63 proteins were classified as pseudokinases (3 NEKs), presumed catalytically inactive kinases that are the likely primary targets of our compound. These will be cross-referenced with hits generated through our pulldown dataset. Further, in silico binding predictions of our lead compound with the four kinases identified in PISA produced comparable ligand-protein intermolecular interactions to those described in the literature. Together, these orthogonal approaches have allowed us to identify credible candidate targets for the compound in Giardia, which we will now explore through target-based assays and additional compound chemistries.