Maintenance of Respiration Capacity Is an Early Predictor of Drug Resistance in Cancer Cells

Inherent or acquired drug resistance is the major obstacle in cancer therapy. An individual prediction of response to cancer therapy would be highly appreciated. It has been reported that the mitochondrial function of cancer cells correlates with the treatment response in different cancer entities (Chonghaile et al., Science 2011 and Vo et al., Cell 2012).

However, as the method of BCL-2 peptide priming measurement (“BH3 profiling”) seemed to be rather complicated and error-prone in our hand we sought to determine mitochondrial function by analyzing cellular respiration. We established three different drug resistant human cancer cell lines (MV4-11, HL-60 and NCl-H82) by continuous exposure to Sorafenib, Cytarabine (Ara-C) or Etoposid (VP-16) respectively. Cellular respiration was measured as oxygen consumption rate (OCR) 24h after drug exposure employing the XF96 extracellular Flux Analyzer. Viability, reduction equivalents NADH/NADPH and mitochondrial membrane potential of the cells were concomitantly determined by flow cytometry via 4’, 6-diamidino-2-phenylindole (DAPI) exclusion, (4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and Tetramethylrhodamine ethyl ester (TMRE) staining with flow cytometric analysis, respectively.

All resistant cancer cells maintained their respiration capacity upon exposure to tyrosine kinase inhibitors, chemotherapeutics or topoisomerase inhibitors, whereas the respiration of drug sensitive cancer cells decreased significantly. Moreover, resistant MV4-11 and HL-60 cells exhibited a metabolic shift towards glycolysis after drug treatment. In contrast sensitive cancer cells showed a decline in respiration and glycolysis in a dose dependent manner. Decrease of respiration was evident at 24h post treatment, while the onset of apoptosis was measurable 48-120h later. Reduction equivalents NADH/NADPH but not mitochondrial membrane potential were diminished in sensitive cells after 24h drug exposure, but dose dependency was more distinct with OCR analysis. The increase of glycolysis in resistant cell lines was also not detectable with the MTT assay.

Our data indicate cellular metabolism and in particular respiration as an early, sensitive and reliable surrogate parameter of drug sensitivity in viable cells that might therefore be applicable in order to determine drug responsiveness in primary cancer cells in a prospective manner.