AbstractBackgroundEarly‐onset colorectal cancer (EOCRC) has been increasing in incidence worldwide but its genomic pathogenesis is mostly undetermined. This study aimed to identify robust EOCRC‐specific gene expression patterns in non‐familial adenomatous polyposis (FAP) and non‐hereditary nonpolyposis colorectal cancer syndrome (HNPCC) EOCRC.MethodWe first performed gene expression profiling analysis using RNA sequencing of discovery cohort comprised of 49 EOCRC (age <50) and 50 late‐onset colorectal cancer (LOCRC) (age >70) specimens. To obtain robust gene expression data from this analysis, we validated differentially expressed genes (DEGs) through TCGA cohort (EOCRC:59 samples, LOCRC:229 samples) and our validation cohort (EOCRC:72 samples, LOCRC:43 samples) using real‐time RT‐PCR. After the validation of DEGs, we validated the selected gene at protein levels using Western blotting. To identify whether genomic methylation regulates the expression of a particular gene, we selected methylation sites using The Cancer Genome Atlas (TCGA) datasets and validated them by pyrosequencing in our validation cohort.ResultsThe EOCRC patients included in this study had significantly more prominent family history of cancer than the LOCRC patients (23 [46.9%] vs. 13 [26%],p= 0.050). Alanyl aminopeptidase (ANPEP) was significantly downregulated in the EOCRC tissues (FC = 1.78,p= 0.0007) and was also commonly downregulated in the TCGA cohort (FC = −1.08,p= 0.0021). Moreover, the ANPEP mRNA and protein expression levels were significantly downregulated in the EOCRC tissues of our validation cohort (p= 0.037 and 0.027). In comparisons of the normal and tumor tissues in public datasets, the ANPEP level was significantly lower in the tumor tissue in the TCGA dataset (p< 2.2 × 10−16) and GSE196006 dataset (p= 0.0005). Furthermore, the ANPEP expression level did not show a decreasing tendency at a young age in the normal colon tissue of the GTEx dataset. Lastly, the hypermethylation of cg26222247 in ANPEP was identified to be weakly associated with reduced ANPEP expression in our EOCRC cohort.ConclusionThe reduced expression of ANPEP was identified as a novel biomarker of non‐FAP and non‐HNPCC EOCRC.