Identification of proteins and cellular pathways targeted by 2-nitroimidazole hypoxic cytotoxins
Faisal Bin Rashed,
Alexandru Cezar Stoica,
Dawn MacDonald,
Hassan El-Saidi,
Carolynne Ricardo,
Bhumi Bhatt,
Jack Moore,
Diana Diaz-Dussan,
Nirilanto Ramamonjisoa,
Yvonne Mowery,
Sambasivarao Damaraju,
Richard Fahlman,
Piyush Kumar,
Michael Weinfeld
Affiliations
Faisal Bin Rashed
Department of Oncology, University of Alberta, Edmonton, AB, T6G2R3, Canada
Alexandru Cezar Stoica
Department of Oncology, University of Alberta, Edmonton, AB, T6G2R3, Canada
Dawn MacDonald
Department of Oncology, University of Alberta, Edmonton, AB, T6G2R3, Canada
Hassan El-Saidi
Department of Oncology, University of Alberta, Edmonton, AB, T6G2R3, Canada; Department of Pharmaceutical Chemistry, Faculty of Pharmacy, University of Alexandria, El Sultan Hussein St. Azarita, Alexandria, Egypt
Carolynne Ricardo
Department of Oncology, University of Alberta, Edmonton, AB, T6G2R3, Canada
Bhumi Bhatt
Department of Laboratory Medicine & Pathology, University of Alberta, Edmonton, AB, T6G2R3, Canada
Jack Moore
Alberta Proteomics and Mass Spectrometry Facility, University of Alberta, Edmonton, AB, T6G2R3, Canada
Diana Diaz-Dussan
Department of Chemical & Materials Engineering, University of Alberta, Edmonton, AB, T6G2R3, Canada
Nirilanto Ramamonjisoa
Department of Oncology, University of Alberta, Edmonton, AB, T6G2R3, Canada
Yvonne Mowery
Radiation Oncology, School of Medicine, Duke University, Durham, NC, 27708, United States
Sambasivarao Damaraju
Department of Laboratory Medicine & Pathology, University of Alberta, Edmonton, AB, T6G2R3, Canada
Richard Fahlman
Department of Biochemistry, University of Alberta, Edmonton, AB, T6G2R3, Canada
Piyush Kumar
Department of Oncology, University of Alberta, Edmonton, AB, T6G2R3, Canada; Corresponding authors. Department of Oncology, University of Alberta, 116 St & 85 Ave, Edmonton, AB, T6G2R3, Canada.
Michael Weinfeld
Department of Oncology, University of Alberta, Edmonton, AB, T6G2R3, Canada; Corresponding authors. Department of Oncology, University of Alberta, 116 St & 85 Ave, Edmonton, AB, T6G2R3, Canada.
Tumour hypoxia negatively impacts therapy outcomes and continues to be a major unsolved clinical problem. Nitroimidazoles are hypoxia selective compounds that become entrapped in hypoxic cells by forming drug-protein adducts. They are widely used as hypoxia diagnostics and have also shown promise as hypoxia-directed therapeutics. However, little is known about the protein targets of nitroimidazoles and the resulting effects of their modification on cancer cells. Here, we report the synthesis and applications of azidoazomycin arabinofuranoside (N3-AZA), a novel click-chemistry compatible 2-nitroimidazole, designed to facilitate (a) the LC-MS/MS-based proteomic analysis of 2-nitroimidazole targeted proteins in FaDu head and neck cancer cells, and (b) rapid and efficient labelling of hypoxic cells and tissues. Bioinformatic analysis revealed that many of the 62 target proteins we identified participate in key canonical pathways including glycolysis and HIF1A signaling that play critical roles in the cellular response to hypoxia. Critical cellular proteins such as the glycolytic enzyme glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and the detoxification enzyme glutathione S-transferase P (GSTP1) appeared as top hits, and N3-AZA adduct formation significantly reduced their enzymatic activities only under hypoxia. Therefore, GAPDH, GSTP1 and other proteins reported here may represent candidate targets to further enhance the potential for nitroimidazole-based cancer therapeutics.
