lly substantial differences in between the oral cavity and pharynx with all the highest correlation (p 0.0001) for ANO1, TMEM22, TMEM97, TMEM116, TMEM156, and TMEM158. TMEM140, TMEM61, and TMEM43 showed no considerable variations (p 0.05). For other TMEMs statistically considerable (p 0.05) variations have been indicated for no less than two localizations, Figure 3.Figure 3. The expression levels of selected TMEMs genes rely on head and neck squamous cell carcinoma (HNSCC) localization in oral cavity (n = 316), pharynx (n = 90), and larynx (n = 116); ns: not important; p 0.05; p 0.01; p 0.001; p 0.0001.Cancers 2021, 13,8 of3.three. Expression of ANO1, TMEM156, TMEM173, and TMEM213 Correlates with Patient Survival We tested whether or not TMEMs are linked with patient survival. For each gene we measured disease-free survival (DFS) and all round survival (OS) in sufferers with high vs. low TMEM expression, Figure 4 and Table S2.Figure 4. Disease-free survival (DFS) and all round survival (OS) curves (darker lines) with 95 CI (lighter lines) of HNSCC individuals based on ANO1 or TMEM156 or TMEM173 or TMEM213 expression levels; higher and low subgroups of sufferers divided according to the mean of expression level; n–number of instances in each subgroup; pa –log-rank (Mantel ox) test, and pb –Gehan-Breslow-Wilcoxon test; HRa –Hazard Ratio-Mantel aenszel, HRb –Hazard Ratio-log-rank; CI –confidence interval; p 0.05 regarded as significant.Cancers 2021, 13,9 ofIn individuals with reduced expression of ANO1 and TMEM156 we observed substantially longer DFS (p = 0.0443, HR = two.317 with 95 CI = 1.022 to 5.254 and HR = two.415 with 95 CI = 1.066 to 5.470 and p = 0.0495, HR = two.089 with 95 CI = 0.9207 to four.741 and HR = two.124 with 95 CI = 0.9363 to four.817, respectively). Sufferers with low ANO1 displayed also longer OS (p = 0.0003; HR = 1.710 with 95 CI = 1.280 to two.284 and HR = 1.724 with 95 CI = 1.291 to two.301). In contrast, lower levels of TMEM156, TMEM173, and TMEM213 have been connected with shorter OS (p = 0.0009, HR = 0.6123 with 95 CI = 0.4585 to 0.8177 and HR = 0.6043 with 95 CI = 0.4526 to 0.8068; p = 0.0494, HR = 0.7475 with 95 CI = 0.5592 to 0.9992 and HR = 0.7490 with 95 CI = 0.5605 to 1.001 and p = 0.0212, HR = 0.7853 with 95 CI = 0.5749 to 1.073 and HR = 0.7755 with 95 CI = 0.5679 to 1.059, respectively). Other genes showed no statistical differences in DFS or OS Caspase 11 web within the two groups of sufferers, Figure S2 and Table S3. 3.4. TMEM-Correlated Genes Are Involved in Quite a few Biological Processes For further evaluation we selected the four TMEMs which correlated with patient survival (ANO1, TMEM156, TMEM173, and TMEM213) and searched for the correlations using a panel of unique genes, Figure 5A.Figure five. Variations in HNSCC phenotype according to the expression amount of ANO1, TMEM156, TMEM173, and TMEM213. (A) REACTOME pathway enrichment analysis of genes positively (R 0.three) and negatively (R -0.3) correlated with ANO1, TMEM156, TMEM173, and TMEM213. Fifteen statistically substantial (p 0.05) pathways with the highest gene count are shown; (B) Genes sets considerably (p 0.05 and FDR 0.25) Kinesin-12 Molecular Weight enriched in higher and low ANO1, TMEM156, TMEM17, and TMEM213 expression phenotypes with regard to hallmarks (H) and oncogenic (C6) gene sets determined by GSEA analysis; gray squares indicate lack of enrichment.ANO1 negatively correlated with genes related to metabolism, eukaryotic translation, and cellular response to anxiety and external stimuli; a optimistic correlatio
