Gemcitabine Resistance in Pancreatic Ductal Carcinoma Cell Lines Stems from Reprogramming of Energy Metabolism

Abstract

Pancreatic ductal adenocarcinoma (PDAC) is associated with poor prognosis because it is often detected at an advanced stage, and drug resistance interferes with treatment. However, the mechanism underlying drug resistance in PDAC remains unclear. Here, we investigated metabolic changes between a parental PDAC cell line and a gemcitabine (GEM)-resistant PDAC cell line. We established a GEM-resistant cell line, MIA-G, from MIA-PaCa-2 parental (MIA-P) cells using continuous therapeutic-dose GEM treatment. MIA-G cells were also more resistant to 5-fluorouracil in comparison to MIA-P cells. Metabolic flux analysis showed a higher oxygen consumption rate (OCR) in MIA-G cells than in MIA-P cells. Notably, OCR was suppressed by GEM treatment only in MIA-G cells. GEM treatment increased mitochondrial membrane potential and mitochondrial reactive oxygen species (ROS) in MIA-P cells, but not in MIA-G cells. Glutamine uptake and peroxidase levels were elevated in MIA-G cells. The antioxidants N-acetyl-L-cysteine and vitamin C increased the sensitivity to GEM in both cell lines. In MIA-G cells, the expression of the mitochondrial transcription factor A also decreased. Furthermore, rotenone reduced the sensitivity of MIA-P cells to GEM. These findings suggest that the suppression of oxidative phosphorylation contributes to GEM resistance by reducing ROS production. Our study provides a new approach for reducing GEM resistance in PDAC.

Increased mitochondria are responsible for the acquisition of gemcitabine resistance in pancreatic cancer cell lines

“…This mitochondrial number increase in the GEM‐resistant cell lines likely enhances OXPHOS. Furthermore, OXPHOS capacity was clearly improved, not only in basal respiration but also in terms of reserve capacity and maximal respiration (Figure 2B), which contrasted with Fujiwara et al's results 28 . The reason for the discrepancy is unknown; however, it is possible that the experimental conditions may play a role, because of the large discrepancies in the OCR baselines.…”

“…In pancreatic cancer stem cells, the enhancement of OXPHOS with increased mitochondria and high sensitivity to OXPHOS inhibition, and the antitumor effect of metformin, an OXPHOS inhibitor, have also been reported 26,27,34 . However, Fujiwara et al reported that, although the basal respiration of a GEM‐resistant cell line was high, GEM treatment reduced TFAM expression and OXPHOS, resulting in reduced ROS production and contributing to GEM resistance 28 …”

“…26,27,34 However, Fujiwara et al reported that, although the basal respiration of a GEM-resistant cell line was high, GEM treatment reduced TFAM expression and OXPHOS, resulting in reduced ROS production and contributing to GEM resistance. 28 mtDNA-encoded genes are essential for mitochondrial OX-PHOS. 47,48 In this study, no change was noted in respiratory chain complex expression in the resistant strains (Figure 3A), suggesting that no pathogenic mtDNA mutations had occurred.…”

“…23,24 Mitochondrial dynamics associated with anticancer drug exposure and acquisition of resistance have been reported to increase the number of mitochondria in cisplatin-resistant ovarian cancer. 25 In pancreatic cancer, increased mitochondria in cancer stem cells 26,27 and decreased TFAM expression in GEM-resistant cells 28 have been reported; however, few reports have evaluated mitochondrial dynamics before and after GEM-resistance acquisition by GEM-sensitive pancreatic cancer cell lines.…”

Lauric Acid Overcomes Hypoxia-Induced Gemcitabine Chemoresistance in Pancreatic Ductal Adenocarcinoma

“…Cell growth was assessed using the 3-(4,5-Dimethylthiazol-2-yl)-5-(3-Carboxymethoxyphenyl)-2-(4-Sulfophenyl)-2 H-tetrazolium (MTS) assay as previously described [ 50 ]. MTS assays were performed using the CellTiter 96 Aqueous One Solution Cell Proliferation Assay kit (Promega Biosciences, Inc., San Luis Obispo, CA, USA).…”