Analysis of stress-induced metabolic response in UM-UC-3 cells exposed to the APIM-containing peptide drug ATX-101
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The proliferating cell nuclear antigen (PCNA) is an essential scaffold protein central that is indispensable in several of the cellular repair processes through its numerous protein interaction partners. Under cellular stress, like genotoxic stress caused by chemotherapeutic agents, there is a stress switch, which leads PCNA to interact with cell repair proteins through the AlkB homologue 2 PCNA-interacting motif (APIM). By successfully blocking the interactions between PCNA and the DNA-repair proteins using APIM-containing peptides, the cell will not initiate DNA repair and instead undergo cell arrest or apoptosis and increase the chemotherapeutic effect. The APIM-containing peptide ATX-101 appear to influence signaling pathways regulating the primary metabolism in cells. Therefore, a stress-response experiment was performed on the bladder cancer cell line Um-Uc-3 when exposed to ATX-101 both alone and in combination with the chemotherapeutic agent cisplatin to assess metabolic changes. An additive effect was observed for cell apoptosis when ATX-101 was added to the cisplatin. Any effects on selected metabolite levels 24 h after treatment were analyzed by applying three targeted mass spectrometric methods. Furthermore, the glucose, and glutamine consumption and lactate production was assessed using enzymatic assays.No evident changes were found for metabolites in the amino acid and organic acid pool between the different treatment groups. However, indications of changed metabolic levels of the phosphorylated metabolites were found between all treatment groups. Several of the metabolites considerably altered were regulators or intermediates in the glycolytic pathway, and implied that the APIM-peptide had an effect on the primary metabolites involved in these metabolic processes in UM-UC-3 cells, both alone and in combination with cisplatin. Further, the energy charge was reduced in samples receiving ATX-101 and cisplatin in combination, and not in the other groups. All stress exposed samples showed an increased glucose consumption, the changes was highest in the samples treated with cisplatin alone or in combination with ATX-101. Altogether, the data clearly states that an effect arose under APIM-peptide exposure of Um-Uc-3 cells, and that APIM-mediated PCNA-interaction plays a role in the regulation of the primary carbon metabolism. Still, much remains to be discovered before the role of the APIM-mediated PCNA-interactions are fully understood.