Integrating Phenotypic and Chemoproteomic Approaches to Identify Covalent Targets of Dietary Electrophiles in Platelets

Ivy A Guan; Joanna S T Liu; Renata C Sawyer; Xiang Li; Wanting Jiao; Yannasittha Jiramongkol; Mark D White; Lejla Hagimola; Freda H Passam; Denise P Tran; Xiaoming Liu; Simone M Schoenwaelder; Shaun P Jackson; Richard J Payne; Xuyu Liu

doi:10.1021/acscentsci.3c00822

Integrating Phenotypic and Chemoproteomic Approaches to Identify Covalent Targets of Dietary Electrophiles in Platelets

ACS Cent Sci. 2024 Jan 29;10(2):344-357. doi: 10.1021/acscentsci.3c00822. eCollection 2024 Feb 28.

Authors

Ivy A Guan^{1

2}, Joanna S T Liu^{2

3}, Renata C Sawyer^{1

2}, Xiang Li^{4

5}, Wanting Jiao^{6

7}, Yannasittha Jiramongkol^{1

8}, Mark D White¹, Lejla Hagimola³, Freda H Passam³, Denise P Tran⁹, Xiaoming Liu³, Simone M Schoenwaelder^{2

3}, Shaun P Jackson^{2

8}, Richard J Payne^{1

10}, Xuyu Liu^{1

2}

Affiliations

¹ School of Chemistry, Faculty of Science, The University of Sydney, Sydney, New South Wales 2006, Australia.
² The Heart Research Institute, The University of Sydney, Newtown, New South Wales 2042, Australia.
³ School of Medical Sciences, Faculty of Medicine and Health, The University of Sydney, Sydney, New South Wales 2006, Australia.
⁴ Department of Medicine, Washington University in St. Louis, St. Louis, Missouri 63110, United States.
⁵ McDonnell Genome Institute, Washington University in St. Louis, St. Louis, Missouri 63108, United States.
⁶ Ferrier Research Institute, Victoria University of Wellington, Wellington 6140, New Zealand.
⁷ Maurice Wilkins Centre for Molecular Biodiscovery, Auckland 1142, New Zealand.
⁸ Charles Perkins Centre, The University of Sydney, Sydney, New South Wales 2006, Australia.
⁹ Sydney Mass Spectrometry, The University of Sydney, Camperdown, New South Wales 2006, Australia.
¹⁰ Australian Research Council Centre of Excellence for Innovations in Peptide and Protein Science, The University of Sydney, Sydney, New South Wales 2006, Australia.

Abstract

A large variety of dietary phytochemicals has been shown to improve thrombosis and stroke outcomes in preclinical studies. Many of these compounds feature electrophilic functionalities that potentially undergo covalent addition to the sulfhydryl side chain of cysteine residues within proteins. However, the impact of such covalent modifications on the platelet activity and function remains unclear. This study explores the irreversible engagement of 23 electrophilic phytochemicals with platelets, unveiling the unique antiplatelet selectivity of sulforaphane (SFN). SFN impairs platelet responses to adenosine diphosphate (ADP) and a thromboxane A2 receptor agonist while not affecting thrombin and collagen-related peptide activation. It also substantially reduces platelet thrombus formation under arterial flow conditions. Using an alkyne-integrated probe, protein disulfide isomerase A6 (PDIA6) was identified as a rapid kinetic responder to SFN. Mechanistic profiling studies revealed SFN's nuanced modulation of PDIA6 activity and substrate specificity. In an electrolytic injury model of thrombosis, SFN enhanced the thrombolytic activity of recombinant tissue plasminogen activator (rtPA) without increasing blood loss. Our results serve as a catalyst for further investigations into the preventive and therapeutic mechanisms of dietary antiplatelets, aiming to enhance the clot-busting power of rtPA, currently the only approved therapeutic for stroke recanalization that has significant limitations.