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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.

SUBMITTER: Fujiwara-Tani R

PROVIDER: S-EPMC9323155 | biostudies-literature | 2022 Jul

REPOSITORIES: biostudies-literature

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

Fujiwara-Tani Rina R Sasaki Takamitsu T Takagi Tadataka T Mori Shiori S Kishi Shingo S Nishiguchi Yukiko Y Ohmori Hitoshi H Fujii Kiyomu K Kuniyasu Hiroki H

International journal of molecular sciences 20220715 14

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 tre ...[more]

PMID: 35887170

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Project description:BackgroundPancreatic ductal adenocarcinoma (PDAC) is a highly lethal disease with poor prognosis, as it is difficult to predict or circumvent, and it develops chemoresistance quickly. One cellular mechanism associated with chemoresistance is alternative splicing dysfunction, a process through which nascent mRNA is spliced into different isoforms. Survivin (Baculoviral IAP Repeat-Containing Protein 5 (BIRC5)), a member of the inhibitor of apoptosis (IAP) protein family and a cell cycle-associated oncoprotein, is overexpressed in most cancers and undergoes alternative splicing (AS) to generate six different splicing isoforms.MethodsTo determine if survivin splice variants (SSV) could be involved in PDAC chemoresistance, a Gemcitabine (Gem) resistant (GR) cell line, MIA PaCa-2 GR, was created and assessed for its SSV levels and their potential association with GR. Cross-resistance was assessed in MIA-PaCa-2 GR cells to FIRINOX (5-fluorouracil (5-FU), irinotecan, and oxaliplatin). Once chemoresistance was confirmed, RT-qPCR was used to assess the expression of survivin splice variants (SSVs) in PDAC cell lines. To confirm the effect of SSVs on chemoresistance, we used siRNA to knockdown all SSVs or SSV 2β.ResultsThe MIA PaCa-2 GR cell line was 40 times more resistant to Gem and revealed increased resistance to FIRINOX (5-fluorouracil (5-FU), irinotecan, and oxaliplatin); when compared to the parental MIA-PaCa-2 cells. RT-qPCR studies revealed an 8-fold relative expression increase in SSV 2β and a 2- to 8-fold increase in the other five SSVs in the GR cells. Knockdown of all SSV or SSV 2β only, using small inhibitory RNA (siRNA), sensitized the GR cells to Gem, indicating that these SSVs play a role in PDAC chemoresistance.ConclusionThese findings provide evidence for the potential role of SSV 2β and other SSVs in innate and acquired PDAC chemoresistance. We also show that the expression of SSVs is not affected by the type of chemoresistance, therefore targeting survivin splice variants in combination with chemotherapy could benefit a wide range of patients.

Dataset Information

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

Publications

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

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