Metabolic Alterations in Pancreatic Cancer Progression

Abstract

Pancreatic cancer is the third leading cause of cancer-related deaths in the USA. Pancreatic tumors are characterized by enhanced glycolytic metabolism promoted by a hypoxic tumor microenvironment and a resultant acidic milieu. The metabolic reprogramming allows cancer cells to survive hostile microenvironments. Through the analysis of the principal metabolic pathways, we identified the specific metabolites that are altered during pancreatic cancer progression in the spontaneous progression (KPC) mouse model. Genetically engineered mice exhibited metabolic alterations during PanINs formation, even before the tumor development. To account for other cells in the tumor microenvironment and to focus on metabolic adaptations concerning tumorigenic cells only, we compared the metabolic profile of KPC and orthotopic tumors with those obtained from KPC-tumor derived cell lines. We observed significant upregulation of glycolysis and the pentose phosphate pathway metabolites even at the early stages of pathogenesis. Other biosynthetic pathways also demonstrated a few common perturbations. While some of the metabolic changes in tumor cells are not detectable in orthotopic and spontaneous tumors, a significant number of tumor cell-intrinsic metabolic alterations are readily detectable in the animal models. Overall, we identified that metabolic alterations in precancerous lesions are maintained during cancer development and are largely mirrored by cancer cells in culture conditions.

Keywords: KPC mice; cancer metabolism; metabolic alterations; pancreatic cancer; precancerous lesions.

Figure 1

Pathological evaluation of the pancreas from KPC mice at different stages. (a) Scheme of the experimental design. Six mice per group with the appropriate genotype were sacrificed after 10, 15, and 25 weeks. (b) Graphs representing tumor volumes and weights at the time of necropsy Tissues volume and weight were compared by Student’s t-test. ‘*’ p < 0.05 and ‘**’ p < 0.01. # indicates the group where pancreatic tumors were found. (c) Hematoxylin and eosin stains in pancreas/tumor sections from control and KPC mice. Control mice at 10 weeks, 15 weeks, and 25 weeks showed normal acinar architecture in the pancreas with no PanIN lesions and no adenocarcinoma (top row). KPC mice pancreas showed PanIN lesions (indicated by arrowheads) at 10 and 15 weeks and invasive adenocarcinoma at 25 weeks (bottom row). (d) Masson’s trichrome staining in pancreas/tumor sections from KPC mice for evaluating fibrosis and desmoplastic reaction. Fibrosis/desmoplastic regions are stained blue. Normal periductal fibrous tissue but not significant desmoplasia can be noticed at 10 and 15 weeks in KPC mice. At 25 weeks, the invasive adenocarcinoma demonstrates tumor-associated desmoplasia.

Figure 2

Evaluation of glycolysis metabolites in pancreatic cancer models. (a) Relative glycolytic metabolite levels in 10, 15, and 25 weeks old KPC mice pancreas/tumors determined by LC-MS/MS. (b) Relative glycolytic metabolite levels in mice orthotopically implanted with a KPC cell line. (c) Relative glycolytic metabolite levels in KPC cell lines. Abbreviations: G6P, glucose-6-phosphate; FBP, fructose bisphosphate; DHAP, dihydroxyacetone phosphate; G3P, glyceraldehyde-3-phosphate; 3PG, 3-phosphoglycerate; PEP, phosphoenolpyruvate; PYR, pyruvate. Data are represented as mean ± SEM. The bar charts in (a) and (b) were compared by Student’s t-test. * p < 0.05, ** p < 0.01, and *** p < 0.001. Bar charts in (c) were compared by one-way ANOVA followed by Tukey’s post-hoc test. £ p < 0.05, # p < 0.01, and $ p < 0.001.

Figure 3

Evaluation of the pentose phosphate pathway metabolites in pancreatic cancer models. (a) Relative PPP metabolite levels in 10, 15, and 25 weeks old KPC mice. (b) Relative PPP metabolite levels in orthotopically implanted mice. (c) Relative PPP metabolite levels in KPC cell lines. Abbreviations: G6P, glucose-6-phosphate; Ru5P, ribulose-5-phosphate; R5P, ribose-5-phosphate; X5P, xylulose-5-phosphate; G3P, glyceraldehyde-3-phosphate; S7P, sedoheptulose-7-phosphate; SBP, sedoheptulose 1,7-bisphosphate; E4P, erythrose 4-phosphate. Data are represented as mean ± SEM. The bar charts in (a) and (b) were compared by Student’s t-test. * p < 0.05, ** p < 0.01, and *** p < 0.001. Bar charts in (c) were compared by one-way ANOVA followed by Tukey’s post-hoc test. £ p < 0.05, # p < 0.01, and $ p < 0.001.

Figure 4

Evaluation of the TCA cycle metabolites in pancreatic cancer models. (a) Relative TCA cycle metabolite levels in 10, 15, and 25 weeks old KPC mice. (b) Relative TCA metabolite levels in orthotopically implanted mice. (c) Relative TCA metabolite levels in KPC cell lines. Data are represented as mean ± SEM. The bar charts in (a) and (b) were compared by Student’s t-test. * p < 0.05 and ** p < 0.01. Bar charts in (c) were compared by one-way ANOVA followed by Tukey’s post-hoc test. £ p < 0.05, # p < 0.01, and $ p < 0.001.

Figure 5

Evaluation of the pyrimidine pathway in pancreatic cancer models. (a) Relative pyrimidine metabolite levels in 10, 15, and 25 weeks old KPC mice. (b) Relative pyrimidine metabolite levels in orthotopically implanted mice. (c) Relative pyrimidine metabolite levels in KPC cell lines. Abbreviations: UMP uridine 5’-monophosphate, UDP uridine 5’-diphosphate, UTP uridine-5’-triphosphate, dCMP deoxycytidine 5’-monophosphate, CTP cytidine-5’-triphosphate. Data are represented as mean ± SEM. The bar charts in (a) and (b) were compared by Student’s t test. * p < 0.05, ** p < 0.01, and ***p < 0.001. Bar chart in (c) was compared by one-way ANOVA followed by Tukey’s post-hoc test. # < 0.01 and $ < 0.001.

Figure 6

Evaluation of the urea cycle metabolites in pancreatic cancer models. (a) Relative urea cycle metabolite levels in 10, 15, and 25 weeks old KPC mice. (b) Relative urea cycle metabolite levels in orthotopically implanted mice. (c) Relative urea metabolite levels in KPC cell lines. Data are represented as mean ± SEM. The bar charts in (a) and (b) were compared by Student’s t-test. * p < 0.05, ** p < 0.01, and *** p < 0.001. Bar charts in (c) were compared by one-way ANOVA followed by Tukey’s post-hoc test. £ p < 0.05, # p < 0.01, and $ p < 0.001.