Abstract
Oxythiamine (OT), an analogue of anti-metabolite, can suppress the nonoxidative synthesis of ribose and induce cell apoptosis by causing a G1 phase arrest in vitro and in vivo. However, the molecular mechanism remains unclear yet. In the present study, a quantitative proteomic analysis using the modified SILAC method (mSILAC) was performed to determine the effect of metabolic inhibition on dynamic changes of protein expression in MIA PaCa-2 cancer cells treated with OT at various doses (0 μM, 5 μM, 50 μM and 500 μM) and time points (0 h, 12 h and 48 h). A total of 52 differential proteins in MIA PaCa-2 cells treated with OT were identified, including 14 phosphorylated proteins. Based on the dynamic expression pattern, these proteins were categorized in three clusters, straight down-regulation (cluster 1, 37% of total proteins), upright “V” shape expression pattern (cluster 2, 47.8% total), and downright “V” shape pattern (cluster 3, 15.2% total). Among them, Annexin A1 expression was significantly down-regulated by OT treatment in time-dependent manner, while no change of this protein was observed in OT dose-dependent fashion. Pathway analysis suggested that inhibition of transketolase resulted in changes of multiple cellular signaling pathways associated with cell apoptosis. The temporal expression patterns of proteins revealed that OT altered dynamics of protein expression in time-dependent fashion by suppressing phosphor kinase expression, resulting in cancer cell apoptosis. Results from this study suggest that interference of single metabolic enzyme activity altered multiple cellular signaling pathways.
Highlights
It has been known for decades that most tumor cells and tissues enhanced glucose metabolism by glycolysis [1,2]
Oxythiamine caused protein expression in a dose-dependent manner Using MTT assay, we determined the toxicity of OT to MIA PaCa-2 cells and found that the IC50 of OT for MIA PaCa-2 is 14.95 μM (Additional file 1: Figure S1)
A number of our previous studies have shown that inhibition of activity of either transketolase in the pentose phosphate cycle, or glycogen phosphorylase causes cell cycle arrest leading to cancer cell apoptosis [17,23,47,48]
Summary
It has been known for decades that most tumor cells and tissues enhanced glucose metabolism by glycolysis [1,2]. Its causal relationship with cancer cell proliferation is still unclear, the phenomenon has been developed a reliable technique for detecting and classifying tumors by fluorodeoxyglucose positron emission tomography (FDG-PET) [3,4] In recent years, this metabolic alteration of malignant cells has been observed in OT is a thiamine antagonist and inhibits transketolase (TK) which is an enzyme of the pentose phosphate pathway in animals. The exactly molecular mechanism is not clear, it has been accepted that the decreased biological macromolecular synthesis can inhibit cell proliferation and induces cell apoptosis These features of metabolism are used for cancer therapeutic approach known as “metabolic therapy” [19,20]
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