The associations between OS, RFS and clinicopathologic factors in LGG
Patients’ age, tumor weight, histological type, histologic grade, IDH1 mutation and targeted therapy were significantly associated with OS in the TCGA cohort (P<0.05 for all cases, student’s t test or Fisher exact test, Table1). Moreover, IDH1 mutation was significantly associated with favourable RFS in the TCGA cohort (P<0.05, Fisher exact test, Table1). The remaining factors did not show significant association with OS in the TCGA cohort (P > 0.05 for all cases, student’s t test or Fisher exact test, Table1). The CGGA cohort was utilized to validate the associations of clinicopathologic factors with survival in LGG patients. Histologic grade, IDH1 mutation and 1p19q codeletion were significantly correlated with patients’ OS in the CGGA cohort (P < 0.05, Fisher exact test, supplementary Table3). While, histologic grade and chemotherapy were significantly associated with RFS in the LGG dataset (P < 0.05 for all cases, Fisher exact test, student’s t test, supplementary Table3).
The associations between clinicopathologic factors and SLC7 family gene expression in LGG
A linear regression model was utilized to investigate the associations between clinicopathologic factors and mRNA expression values of SLC7 family genes. Patients’ age was positively correlated with SLC7A2, SLC7A3, SLC7A11 and SLC7A14 expression and negatively corelated with SLC7A1 expression. Tumour weight was positively correlated with SLC7A2 expression and negatively corelated with SLC7A5, SLC7A6 and SLC7A9 expression. Histological type showed negative correlation with SLC7A3, SLC7A7, SLC7A9 and positive correlation with SLC7A1, SLC7A4, SLC7A5 and SLC7A14 expression. Histologic grade was negatively associated with SLC7A2, SLC7A4, SLC7A11,SLC7A14 and positively associated with SLC7A3, SLC7A5, SLC7A6 and SLC7A7 expression. IDH1 mutation was positively associated with SLC7A1, SLC7A14 and negatively associated with SLC7A2, SLC7A3, SLC7A4, SLC7A7, SLC7A8, SLC7A9, SLC7A10 and SLC7A11 expression. TP53 mutation was positively associated with SLC7A7, SLC7A9 and negatively associated with SLC7A1, SLC7A4, SLC7A5, SLC7A8, SLC7A10, SLC7A11 and SLC7A14 expression. Radiation therapy exhibited negative correlation with SLC7A1, SLC7A14 and positive correlation with SLC7A7, SLC7A9 expression. Targeted molecular therapy was negatively corelated with SLC7A11 expression (P values< 0.05 for all cases, Table2). In line with the results above, many SLC7 genes were significantly correlated with clinical factors in the CGGA dataset, with detailed results presented in the supplementary table4.
Diagnostic values of SLC7 family genes
Of the 12 SLC7 family genes, SLC7A4, SLC7A9, SLC7A10 expression was significantly down-regulated, while SLC7A1, SLC7A2, SLC7A3, SLC7A5, SLC7A6, SLC7A7, SLC7A11 expression was significantly up-regulated in LGG tissues in comparison with normal brain tissues (P values< 0.05 for all cases, student t test, Figure1A). ROC curves were constructed to further explore the diagnostic values of the ten genes. SLC7A5, SLC7A7, SLC7A10 particularly exhibited good performance in differentiating glioma tissues from normal brain tissues, with AUC values > 0.80 for all cases (Figure1B). All the results suggest these three genes might serve as diagnostic biomarkers in LGG.
The co-expression patterns and protein-protein interactions of SLC7 family genes
The expression of SLC7 family genes was highly correlated, of the 11 SLC7 family members, SLC7A1 expression was significantly correlated with all SLC7 family genes.SLC7A4, SLC7A6 SLC7A7, SLC7A9 expression showed significant correlation with the expression of other eight family members (P <0.05 for all cases, pearson correlation, Figure2A). The PPI network of SLC7 family genes consisted of 12 nodes and 12 edges, with a median node degree of 2. The PPI network exhibited more interactions than expected (PPI enrichment p-value<0.0001, Figure2B). The co-expression patterns and PPI networks suggested that the SLC7 family genes were co-expressed at the mRNA level and exhibited extensive homology at the protein level.
The GO term and KEGG pathway enrichment analysis
We performed GO term and KEGG pathway enrichment analysis for 12 SLC7 family genes and found the 12 genes were significantly enriched in 50 GO terms and 1 KEGG pathway (protein digestion and absorption). The top five GO terms showing the highest enrichment for SLC7 family genes were amino acid transmembrane transport, carboxylic acid transmembrane transport, organic acid transmembrane transport, amino acid transport and L-alpha-amino acid transmembrane transport (adjusted P value < 0.05 for all cases, supplementary Table5).
Overall survival analysesin LGG
Kaplan-Meier survival analysis showed significant differences in patients’ OS between the high and low expression groups for SLC7A3, SLC7A6, SLC7A7, SLC7A10, SLC7A1, SLC7A4, SLC7A8, SLC7A11 and SLC7A14 in the TCGA cohort (P <0.05 for all cases, log rank test, Figure3 and supplementary table6). Univariate analysis showed that elevated SLC7A7 and SLC7A10 expression levels were significantly associated with increased mortality, while increased SLC7A1, SLC7A4, SLC7A8, SLC7A11 and SLC7A14 expression levels were significantly associated with reduced mortality (P<0.05 for all cases, supplementary table6). Then multivariate analysis was performed between patients’ OS and SLC7 family gene expression levels, the mortality-associated factors, including patients’ age, tumour weight, histological type, histologic grade and IDH1 mutation. Multivariate analysis confirmed that increased SLC7A7 expression was associated with increased mortality (P≤0.001, Odd ratio [OR]:2.66, 95% Confidence interval [CI]: 1.56–4.6, supplementary table6). While, increased SLC7A4, SLC7A8, SLC7A11 and SLC7A14 expression was significantly associated with reduced mortality (P=0.02, OR:0.38, 95% CI: 0.16–0.81; P≤0.001, OR:0.44, 95% CI: 0.26–0.77; P=0.03, OR:0.54, 95% CI: 0.31–0.95; P≤0.001, OR:0.38, 95% CI: 0.21–0.67, respectively, supplementary table6).
Validation of overall survival analyses in LGG
Kaplan-Meier survival analysis confirmed that high SLC7A7 expression was associated with inferior prognosis, while high expression levels of SLC7A4 and SLC7A14 were associated with favourable prognosis in the CGGA cohort (P <0.05 for all cases, log rank test, supplementary table7). Univariate analysis showed that increased SLC7A7 expression, decreased SLC7A4 and SLC7A14 expression were significantly associated with increased mortality (P<0.05 for all cases, supplementary Table7). Then multivariate analysis was applied between patients’ OS and the mortality-associated features as well as SLC7A4, SLC7A7and SLC7A14 expression levels. Multivariate analysis confirmed that increased SLC7A7 expression was associated with increased mortality following the adjustment of survival-related clinical features (P=0.02, OR:1.45, 95% CI: 1.05–2.01, supplementary Table7).
Relapse-free survival analyses
Kaplan-Meier RFS analysis showed that high SLC7A1, SLC7A8, SLC7A11 and SLC7A14 expression levels were associated with favourable RFS, whereas, high SLC7A3 and SLC7A7 expression levels were indicative of poor RFS in the TCGA cohort (P <0.05 for all cases, log rank test, Figure4 and supplementary table8). Univariate and multivariate analysis exhibited that increased SLC7A1, SLC7A8, SLC7A11, and SLC7A14 expression levels were associated with favourable RFS, while, increased SLC7A7 expression levels were associated with poor RFS (P <0.05 for all cases, supplementary table6). In order to validate the findings above, we analysed the associations of RFS and SLC7 family member expression in the CGGA cohort. The Kaplan-Meier analysis together with univariate and multivariate analysis confirmed that increased SLC7A11 expression was associated with decreased RFS (P <0.05 for all cases, supplementary Table9).