Sex Differences in Colon Cancer Metabolism Reveal A Novel Subphenotype

Abstract

Women have a lower incidence of colorectal cancer (CRC) than men, however, they have a higher incidence of right-sided colon cancer (RCC). This is of concern as patients with RCC have the poorest clinical outcomes among all CRC patients. Aberrant metabolism is a known hallmark and therapeutic target for cancer. We propose that metabolic subphenotypes exist between CRCs due to intertumoral molecular and genomic variation, and differences in environmental milieu of the colon which vary between the sexes. Metabolomics analysis of patient colon tumors (n = 197) and normal tissues (n = 39) revealed sex-specific metabolic subphenotypes dependent on anatomic location. Tumors from women with RCC were nutrient-deplete, showing enhanced energy production to fuel asparagine synthesis and amino acid uptake. The clinical importance of our findings were further investigated in an independent data set from The Cancer Genomic Atlas, and demonstrated that high asparagine synthetase (ASNS) expression correlated with poorer survival for women. This is the first study to show a unique, nutrient-deplete metabolic subphenotype in women with RCC, with implications for tumor progression and outcomes in CRC patients.

Figure 1

Schematic illustration of the metabolomics-driven analysis strategy for the discovery of sex-related differences in colon cancer metabolism. RCC, right-sided colon cancer; LCC, left-sided colon cancer; TCGA, The Cancer Genome Atlas.

Figure 2

Sex differences in critical metabolites required for cancer cell growth. (A) ribulose-5-phosphate (R-5-P), glyceraldhyde-3-phosphate/dihydroxyacetone phosphate (G-3-P/DHAP), lactate, palmitoylcarnitine, stearoylcarnitine, and oleic acid in right-sided colon cancer (RCC). S = stage. Combination of box and violin plots display median value (center line), upper and lower quartiles (box limits), 1.5× interquartile range (black bar), and points out of interquartile range are outliers. Nonparametric Kruskal–Wallis rank sum test with pairwise Wilcoxon Mann-Whitney U test, p values adjusted for false discovery rates (FDR) (Benjamini-Hochberg). *p < 0.05, **p < 0.01, ***p < 0.001, ns. = not significant. Women RCC: Normal (n = 12), SI (n = 12), SII (n = 21), SIII (n = 17); Men RCC: Normal (n = 27), SI (n = 10), SII (n = 23), SIII (n = 15). (B) Schematic illustration of sex differences in glycolysis, pentose phosphate pathway (PPP), and fatty acid metabolism (FA Met.) in RCC. glucose (Glc), glucose-6-phosphate (Glc-6-P), fructose-6-phosphate (F-6-P), phosphoenolpyruvate (PEP). M = Men, W = Women. Up arrow indicates increased, horizontal rectangle indicates not changed in tumor compared to normal colon.

Figure 3

Increased asparagine synthesis associates with amino acid uptake in women with RCC. (A) Abundance of asparagine, adenosine monophosphate (AMP), glutamine, threonine, and serine in normal tissues and tumor tissues in stage I-III women with right-sided colon cancer (RCC) and men with RCC. Nonparametric Kruskal-Wallis rank sum test with pairwise Wilcoxon Mann-Whitney U test, p values adjusted for false discovery rate (FDR) (Benjamini-Hochberg). *p < 0.05, **p < 0.01, ***p < 0.001, ns. = not significant. Women RCC: Normal (n = 12), SI (n = 12), SII (n = 21), SIII (n = 17); Men RCC: Normal (n = 27), SI (n = 10), SII (n = 23), SIII (n = 15). (B) Pearson correlation analyses between asparagine with threonine, and serine in women with stage I-III RCC. Metabolite abundances were log2 transformed, and 95% confidence intervals (CIs) for the correlation coefficients were indicated as the shadowed area colored according to tumor stage.

Figure 4

Abundance levels of asparagine and glutamine in normal and tumor tissues. Abundance levels are shown in normal tissue, stage I tumor, stage II tumor, and stage III tumors from women with left-sided colon cancer (LCC) and men with LCC. Nonparametric Kruskal–Wallis rank sum test with pairwise Wilcoxon Mann-Whitney U test, p values adjusted for false discovery rates (FDR) (Benjamini-Hochberg). *p < 0.05, **p < 0.01, ***p < 0.001, ns. = not significant. Women LCC: Normal (n = 12), SI (n = 10), SII (n = 17), SIII (n = 18); Men LCC: Normal (n = 27), SI (n = 15), SII (n = 25), SIII (n = 14).

Figure 5

High asparagine synthetase (ASNS) expression was correlated with poorer overall survival for women with CRC. (A) Scheme of asparagine synthesis catalyzed by glutamine and ATP-dependent asparagine synthetase. Kaplan-Meier survival curve (overall survival probability) for (B) women with colon adenocarcinoma and (C) men with colon adenocarcinoma using data from The Cancer Genome Atlas (TCGA) database with low/medium and high ASNS expression levels.

Figure 6

Sex-related differences in methionine and polyamine metabolism. (A) Abundance of S-adenosylmethionine (SAM), S-adenosylhomocysteine (SAH), methionine, dimethyglycine, ornithine, spermidine, N¹-acetylspermidine, and N¹, N¹²-diacetylspermine in right-sided colon cancer (RCC). S = stage. Nonparametric Kruskal–Wallis rank sum test with pairwise Wilcoxon Mann-Whitney U test, p values adjusted for false discovery rates (FDR) (Benjamini-Hochberg). *p < 0.05, **p < 0.01, ***p < 0.001, ns. = not significant. Women RCC: Normal (n = 12), SI (n = 12), SII (n = 21), SIII (n = 17); Men RCC: Normal (n = 27), SI (n = 10), SII (n = 23), SIII (n = 15). (B) Schematic illustration of sex differences in methionine metabolism and polyamine synthesis pathways in RCC. S-adenosylmethionine (SAM), decarboxylated S-adenosylmethionine (dc.SAM), S-adenosylhomocysteine (SAH), N¹,N¹²-diacetylspermine (DAS). M = Men, W = Women, S = stage. Up arrow indicates increased, horizontal rectangle indicates not changed in tumor compared to normal colon.

Figure 7

Proposed model of RCC metabolism in women. Enhanced fatty acid metabolism produces ATP required for asparagine production, which is associated with extracellular amino acid uptake. This promotes nucleotide synthesis and activates amino acid-dependent signaling. Fatty acid (FA), mammalian target of Rapamycin (mTOR), intracellular (Intra. cell), extracellular (Extra. cell), amino acid (AA).