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.