Defining Metabolic Rewiring in Lung Squamous Cell Carcinoma

Abstract

Metabolomics based on untargeted flow infusion electrospray ionization high-resolution mass spectrometry (FIE-HRMS) can provide a snap-shot of metabolism in living cells. Lung Squamous Cell Carcinoma (SCC) is one of the predominant subtypes of Non-Small Cell Lung Cancers (NSCLCs), which usually shows a poor prognosis. We analysed lung SCC samples and matched histologically normal lung tissues from eight patients. Metabolites were profiled by FIE-HRMS and assessed using t-test and principal component analysis (PCA). Differentially accumulating metabolites were mapped to pathways using the mummichog algorithm in R, and biologically meaningful patterns were indicated by Metabolite Set Enrichment Analysis (MSEA). We identified metabolic rewiring networks, including the suppression of the oxidative pentose pathway and found that the normal tricarboxylic acid (TCA) cycle were decoupled from increases in glycolysis and glutamine reductive carboxylation. Well-established associated effects on nucleotide, amino acid and thiol metabolism were also seen. Novel aspects in SCC tissue were increased in Vitamin B complex cofactors, serotonin and a reduction of γ-aminobutyric acid (GABA). Our results show the value of FIE-HRMS as a high throughput screening method that could be exploited in clinical contexts.

Keywords: flow infusion electrospray ionization high resolution mass spectrometry; lung squamous cell carcinoma; pathways; untargeted metabolites.

Figure 1

Principal component analysis (PCA) score plot between the selected PCs. The explained variances are shown in brackets, (A) is the negative ionization mode with a variation of 22.8% and (B) is the positive ionization mode with a variation of 18%, and the areas in red and green have 95% of confidence. The PCA shows distinction between both groups, normal tissue and cancer tissue.

Figure 2

Hierarchical clustering analyses of negative ionization mode (A) and positive ionization mode (B) with top 50 differentially accumulated metabolites.

Figure 3

Significantly enriched pathways in SCC tumour tissue in (A) negative and (B) positive ionisation modes.

Figure 4

Joint-pathway analysis through an unbiased analyses linked our results to KEGG disease interaction focusing on lung cancer.

Figure 5

Altered nucleotide metabolism is a major feature of metabolism in lung SCC samples. Identified nucleotide metabolites for each sample were extracted from FIE-MS derived matrix and assessed by (A) principal component analyses and (B) hierarchical cluster analysis. (C) Area under the curve (AUC) and associated box and whisker plots of the main discriminatory nucleotide metabolites.

Figure 6

Anaerobic glycolysis is used to sustain glutamine reductive carboxylation. NAD(P)H electron transfer flux can sustain such as lipid synthesis. Red colours represent cancer tissue and green colours represent normal tissue.