Generated networks were ordered by a score meaning significance, estimated as the ratio of the number of input probes that map to the pathway divided by the total number of pathway probes.

Significance of biological functions and canonical pathways were tested by the Fisher’s exact test p-value after application of Benjamini- Hochberg method of multiple testing correction. Significant pathways were chosen as p < 0.05, except for the significant canonical pathways in the ‘Good’ versus control experiment where a more stringent p-value (p < 0.01) was chosen to eliminate possible false-positive results due to the large number of differentially expressed probe sets. For each experiment, additional Acalabrutinib KEGG (Kyoto Encyclopedia of Genes and Genomes) pathway analysis was performed on up- or downregulated genes (corrected p-value <0.001 and a fold change of respectively >2 and <2) using GENECODIS, a web-based tool for enrichment analysis (http://​genecodis.​dacya.​ucm.​es ) using the NCBI Entrez Gene database [18]. Two statistical tests are implemented: the hypergeometric distribution and the χ2 test of independence. A stimulation-based correction approach is used to adjust for multiple testing. Results Sample selection

Based on the definition of the 2 diverse survival outcome groups and the required RIN values above 7.1, finally 7 ‘Good’ and 10 ‘Bad’ patient samples with similar pathological characteristics remained available for gene expression analysis (Table 1, Figure SPTBN5 2). The median age was 61 and 67 years, respectively. All patients had negative find more resection margins on histopathological examination. Table 1 Clinicopathological parameters of patients, with respectively good and bad outcome Category Gender Age

Location pG pT pN pM pR PNI LVI VI Postop OS DFS GOOD F 55 Head 2 2 0 0 0 1 0 1 0 156.4 156.4 GOOD M 32 Head 3 3 1 0 0 1 1 0 RCT 127.9 127.9 GOOD M 78 Head 1 3 0 0 0 0 1 0 0 71.5 71.5 GOOD M 53 Head 3 3 1 0 0 1 0 1 RCT 67.2 67.2 GOOD F 61 Head 3 3 0 0 0 1 0 1 0 56.4 56.4 GOOD F 62 Head 3 3 1 0 0 0 0 1 RCT 62.7 62.7 GOOD M 68 Tail 3 2 0 0 0 1 0 1 CT 51.5 51.5 BAD F 75 Head 3 3 0 0 0 1 0 0 0 9.4 5.2 BAD M 72 Head 2 3 1 0 0 1 1 1 CT 12.6 5.6 BAD M 52 Head 3 3 0 0 0 1 0 1 0 8.4 4.1 BAD F 78 Head 2 3 1 0 0 1 1 1 0 9.9 3.6 BAD M 59 Head 3 3 1 0 0 1 0 0 0 6.3 2.8 BAD F 51 Head 3 3 0 0 0 0 0 0 CT 19.4 6.5 BAD M 74 Tail 3 1 1 0 0 1 1 1 CT 12.3 0.5 BAD M 50 Head 2 2 1 0 0 1 1 1 CT 9.4 7.0 BAD(M) M 67 Head       1         CT 8.3 / F: female; M: male; pG: pathological tumour grade; pT: pathological tumour size; pN: pathological lymph node status; pM: pathological metastasis; pR: pathological resection margin; PNI: perineural invasion; VI: vascular invasion; LVI: lymphovascular invasion; RCT: radiochemotherapy; CT: chemotherapy; OS: overall survival; DFS: disease-free survival. Figure 2 Pathological features from ‘Good’ and ‘Bad’ patients.