Development of an IPRP-LC-MS/MS method to determine the fate of intracellular thiamine in cancer cells

• Published in: Journal of chromatography. B, Analytical technologies in the biomedical and life sciences Vol. 1124; pp. 247 - 255
• Main Authors: Kim, Jaeah, Jonus, Hunter C., Zastre, Jason A., Bartlett, Michael G.
• Format: Journal Article
• Language: English
• Published: Netherlands Elsevier B.V 15-08-2019
• Subjects: Cell Line, Tumor; Chromatography, High Pressure Liquid - methods; Hexylamine; Humans; Liquid chromatography-mass spectrometry; Stable isotope-labeled thiamine; Tandem Mass Spectrometry - methods; Thiamine; Thiamine - analysis; Thiamine - metabolism; Thiamine kinetics; Thiamine pyrophosphate; Thiamine pyrophosphate; Hexylamine; Stable isotope-labeled thiamine; Thiamine kinetics; Liquid chromatography-mass spectrometry; Thiamine
• DOI: 10.1016/j.jchromb.2019.05.037
• EISSN: 1873-376X
• ISSN: 1570-0232

Understanding the mechanisms underlying cancer cell survival is critical toward advancing drug discovery efforts in this field. Supplemental vitamins have been proposed to play a role in cancer cell metabolism because the increased supply of nutrients is thought to provide cofactors supporting the higher metabolic rate of cancer cells. Particularly, the role of thiamine (vitamin B1) in many biochemical pathways that supports cancer cell metabolism has been investigated. Consequently, the analysis of thiamine and its derivatives in a manner that reflects its dynamic response to genetic modification and pathophysiological stimuli is essential. In this work, we developed a mass spectrometry based-analytical method to track metabolites derived from stable isotope tracers for a better understanding of the metabolic fate of thiamine in cancer cells. This method used ion-pair reversed phase liquid chromatography to simultaneously quantify underivatized thiamine, thiamine monophosphate (TMP) and thiamine pyrophosphate (TPP) in cells. Hexylamine was used as an ion-pairing agent. The method was successfully validated for accuracy, precision and selectivity in accordance with U.S. FDA guidance. Furthermore, the method was then applied for the determination of thiamine and its derivatives with stable isotope labeling to explore the metabolic fate of intracellular thiamine in cancer cells. The finding shows that thiamine is rapidly converted to TPP however, the TPP does not return to thiamine. It appears that TPP may be utilized for other purposes rather than simply being an enzyme cofactor, suggesting unexplored roles for thiamine in cancer.
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•Hexylamine used for ion-pair chromatography and amprolium as the internal standard•Method validated following US FDA guidelines•Method applied to explore the metabolic fate of thiamine in cancer cells