Abstract
BackgroundPancreatic ductal adenocarcinoma (PDAC) is an aggressive cancer with limited treatment options. Pyruvate kinase, especially the M2 isoform (PKM2), is highly expressed in PDAC cells, but its role in pancreatic cancer remains controversial. To investigate the role of pyruvate kinase in pancreatic cancer, we knocked down PKM2 individually as well as both PKM1 and PKM2 concurrently (PKM1/2) in cell lines derived from a KrasG12D/-; p53-/- pancreatic mouse model.MethodsWe used liquid chromatography tandem mass spectrometry (LC-MS/MS) to determine metabolic profiles of wildtype and PKM1/2 knockdown PDAC cells. We further used stable isotope-labeled metabolic precursors and LC-MS/MS to determine metabolic pathways upregulated in PKM1/2 knockdown cells. We then targeted metabolic pathways upregulated in PKM1/2 knockdown cells using CRISPR/Cas9 gene editing technology.ResultsPDAC cells are able to proliferate and continue to produce pyruvate despite PKM1/2 knockdown. The serine biosynthesis pathway partially contributed to pyruvate production during PKM1/2 knockdown: knockout of phosphoglycerate dehydrogenase in this pathway decreased pyruvate production from glucose. In addition, cysteine catabolism generated ~ 20% of intracellular pyruvate in PDAC cells. Other potential sources of pyruvate include the sialic acid pathway and catabolism of glutamine, serine, tryptophan, and threonine. However, these sources did not provide significant levels of pyruvate in PKM1/2 knockdown cells.ConclusionPKM1/2 knockdown does not impact the proliferation of pancreatic cancer cells. The serine biosynthesis pathway supports conversion of glucose to pyruvate during pyruvate kinase knockdown. However, direct conversion of serine to pyruvate was not observed during PKM1/2 knockdown. Investigating several alternative sources of pyruvate identified cysteine catabolism for pyruvate production during PKM1/2 knockdown. Surprisingly, we find that a large percentage of intracellular pyruvate comes from cysteine. Our results highlight the ability of PDAC cells to adaptively rewire their metabolic pathways during knockdown of a key metabolic enzyme.
Highlights
Pancreatic ductal adenocarcinoma (PDAC) is an aggressive cancer with limited treatment options
PKM2 and PKM1/2 knockdown do not decrease pancreatic cancer cell proliferation To study the role of pyruvate kinase in pancreatic cancer proliferation, we characterized PDAC cell lines (A13M21 and A13M13) derived from a KrasG12D/-; p53-/- mouse pancreatic tumor [35]
To ensure that PKL or PKR expression was not induced after PKM1/2 knockdown, we probed the expression of PKL/R in PDAC cells
Summary
Pancreatic ductal adenocarcinoma (PDAC) is an aggressive cancer with limited treatment options. Pancreatic ductal adenocarcinoma (PDAC) is a devastating disease with < 10% of patients surviving beyond 5 years after diagnosis [1] It is currently the fourth leading cause of cancer-related death in western countries and is expected to be the second leading cause by 2030 [2]. Treatment options for pancreatic cancer patients include surgical resection, radiation therapy, and/or systemic chemotherapy [3, 4]. Pancreatic cancer cells rewire metabolism to utilize a wide range of nutrients including glucose, extracellular proteins, and various amino acids to support survival and proliferation [8,9,10]. Targeting metabolic pathways upregulated in PDAC cells may be a promising direction for therapy [11]
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