Overcoming Pancreatic Cancer Radioresistance: Novel Therapeutic Approaches Exploiting Metabolic Reprogramming

Introduction: Pancreatic tumors are resistant to multiples agents including radiation therapy. Remarkably, these tumors rarely shrink, even after exposure to ablative doses of ionizing radiation (IR); the reason for this resistance is not understood. We hypothesize that pancreatic cancer cells survive IR by rewiring their metabolism. These metabolic changes potentially promote DNA repair and the synthesis of anti-oxidative factors. An understanding of which metabolic pathways are activated may open new therapeutic horizons.

Methods: We established a novel model of radioresistance by exposing Panc-1, AsPC-1 and BxPC3 pancreatic cancer cells to fractionated irradiation. The metabolic profile was investigated through nutrient deprivation, exposure to metabolic inhibitors and Seahorse analysis. Intracellular metabolites levels were assessed by Liquid Chromatography-Mass Spectrometry and gene expression profiling was performed using Nanostring technology.

Results and discussion: The subclones were found to be resistant to irradiation; for instance, at 6 Gy survival was 19.4 % vs 6.2 % (p=0.001) for radioresistant / parental Panc-1 cells, 29 % vs 12 % (p=0.0002) for radioresistant / parental AsPC-1 cells and 29.5 % vs 10.7 % (p=0.004) for radioresistant / parental BxPC3 cells. We found that radioresistant Panc-1 and AsPC-1 subclones (RR-Panc-1 and RR-AsPC-1) were more sensitive to glutamine deprivation than their parental counterparts. Moreover RR-Panc-1 and RR-AsPC-1 cells were twofold more sensitive to Bis-2-(5-phenylacetamido-1,3,4-thiadiazol-2-yl)ethyl sulfide - a selective inhibitor of glutaminase- than parental cells. In addition, RR-Panc-1 and RR-AsPC-1 cells were six- and three-fold more sensitive to 6-aminonicotinamide, an inhibitor of the pentose-phosphate pathway which generates the ultimate reducing molecule NADPH and the last precursor for nucleotides synthesis. Radioresistant cells produced low levels of acetyl-CoA, aspartate and succinic acid, indicating that the tricarboxylic acid cycle (TCA) rate was slowed down. Moreover, these cells displayed reduced oxygen consumption rate, pointing out a decrease in mitochondrial respiration. Finally, radioresistant subclones transcriptionally expressed higher levels of pyruvate dehydrogenase kinase which inhibits the transformation of pyruvate to acetyl-CoA.

Conclusion: These findings suggest that radioresistant cells are dependent on glutamine and shift their metabolism from the TCA cycle and oxidative phosphorylation, potentially to the pentose-phosphate pathway to generate nucleotides and reducing power. The discovery that radioresistant cells are more sensitive to metabolic inhibitors was unexpected and without precedent. Ongoing experiments are investigating how the metabolic signature of radioresistance may be exploited for therapeutic intent. A clinical trial is planned.