title: Addiction to MTH1 protein results in intense expression in human breast cancer tissue as measured by liquid chromatography-isotope-dilution tandem mass spectrometry.
abstract: MTH1 protein sanitizes the nucleotide pool so that oxidized 2'-deoxynucleoside triphosphates (dNTPs) cannot be used in DNA replication. Cancer cells require MTH1 to avoid incorporation of oxidized dNTPs into DNA that results in mutations and cell death. Inhibition of MTH1 eradicates cancer, validating MTH1 as an anticancer target. By overexpressing MTH1, cancer cells may mediate cancer growth and resist therapy. To date, there is unreliable evidence suggesting that MTH1 is increased in cancer cells, and available methods to measure MTH1 levels are indirect and semi-quantitative. Accurate measurement of MTH1 in disease-free tissues and malignant tumors of patients may be essential for determining if the protein is truly upregulated in cancers, and for the development and use of MTH1 inhibitors in cancer therapy. Here, we present a novel approach involving liquid chromatography-isotope-dilution tandem mass spectrometry to positively identify and accurately quantify MTH1 in human tissues. We produced full length (15)N-labeled MTH1 and used it as an internal standard for the measurements. Following trypsin digestion, seven tryptic peptides of both MTH1 and (15)N-MTH1 were identified by their full scan and product ion spectra. These peptides provided a statistically significant protein score that would unequivocally identify MTH1. Next, we identified and quantified MTH1 in human disease-free breast tissues and malignant breast tumors, and in four human cultured cell lines, three of which were cancer cells. Extreme expression of MTH1 in malignant breast tumors was observed, suggesting that cancer cells are addicted to MTH1 for their survival. The approach described is expected to be applicable to the measurement of MTH1 levels in malignant tumors vs. surrounding disease-free tissues in cancer patients. This attribute may help develop novel treatment strategies and MTH1 inhibitors as potential drugs, and guide therapies.
authors.affiliation: Biomolecular Measurement Division, National Institute of Standards and Technology, Gaithersburg, MD 20899, USA; Department of Toxicology, Faculty of Pharmacy, Gazi University, Ankara, Turkey.
authors.forename: Erdem
authors.initials: E
authors.lastname: Coskun
authors.affiliation: Biomolecular Measurement Division, National Institute of Standards and Technology, Gaithersburg, MD 20899, USA.
authors.forename: Pawel
authors.initials: P
authors.lastname: Jaruga
authors.affiliation: Science for Life Laboratory, Division of Translational Medicine and Chemical Biology, Department of Medical Biochemistry and Biophysics, Karolinska Institutet, Stockholm, Sweden.
authors.forename: Ann-Sofie
authors.initials: AS
authors.lastname: Jemth
authors.affiliation: Science for Life Laboratory, Division of Translational Medicine and Chemical Biology, Department of Medical Biochemistry and Biophysics, Karolinska Institutet, Stockholm, Sweden.
authors.forename: Olga
authors.initials: O
authors.lastname: Loseva
authors.affiliation: Biomolecular Measurement Division, National Institute of Standards and Technology, Gaithersburg, MD 20899, USA.
authors.forename: Leona D
authors.initials: LD
authors.lastname: Scanlan
authors.affiliation: Biosystems and Biomaterials Division, National Institute of Standards and Technology, Gaithersburg, MD 20899, USA.
authors.forename: Alessandro
authors.initials: A
authors.lastname: Tona
authors.affiliation: Biomolecular Measurement Division, National Institute of Standards and Technology, Gaithersburg, MD 20899, USA.
authors.forename: Mark S
authors.initials: MS
authors.lastname: Lowenthal
authors.affiliation: Science for Life Laboratory, Division of Translational Medicine and Chemical Biology, Department of Medical Biochemistry and Biophysics, Karolinska Institutet, Stockholm, Sweden.
authors.forename: Thomas
authors.initials: T
authors.lastname: Helleday
authors.affiliation: Biomolecular Measurement Division, National Institute of Standards and Technology, Gaithersburg, MD 20899, USA. Electronic address: miral@nist.gov.
authors.forename: Miral
authors.initials: M
authors.lastname: Dizdaroglu
mesh_terms: D000595:Amino Acid Sequence; D001943:Breast Neoplasms / Q000378:metabolism*; D045744:Cell Line, Tumor; D002853:Chromatography, Liquid / Q000379:methods*; D045643:DNA Repair Enzymes / Q000737:chemistry / Q000378:metabolism*; D005260:Female; D006801:Humans; D006868:Hydrolysis; D008969:Molecular Sequence Data; D009587:Nitrogen Isotopes; D010455:Peptides / Q000378:metabolism; D010744:Phosphoric Monoester Hydrolases / Q000737:chemistry / Q000378:metabolism*; D053719:Tandem Mass Spectrometry / Q000379:methods*; D014357:Trypsin / Q000378:metabolism
chemical_list: D009587:Nitrogen Isotopes; D010455:Peptides; D010744:Phosphoric Monoester Hydrolases; D014357:Trypsin; C078207:8-oxodGTPase; D045643:DNA Repair Enzymes
keywords: Extreme expression; Isotope-dilution mass spectrometry; MTH1 protein; Nucleotide pool; Stable isotope-labeled MTH1