Colorectal cancer develops in a multi-step manner from normal epithelium, through a pre-malignant lesion (so-called adenoma), into a malignant lesion (carcinoma), which invades surrounding tissues and eventually can spread systemically (metastasis). It is estimated that only about 5% of adenomas do progress to a carcinoma. The present study aimed to unravel the biology of adenoma to carcinoma progression by mRNA expression profiling, and to identify candidate biomarkers for adenomas that are truly at high risk of progression. Genome-wide mRNA expression profiles were obtained from a series of 37 colorectal adenomas and 31 colorectal carcinomas using oligonucleotide microarrays. Differentially expressed genes were validated in an independent colorectal gene expression data set. Gene Set Enrichment Analysis (GSEA) was used to identify altered expression of sets of genes associated with specific biological processes, in order to better understand the biology of colorectal adenoma to carcinoma progression. mRNA expression of 248 genes was significantly different, of which 96 were upregulated and 152 downregulated in carcinomas compared to adenomas. Classification of adenomas and carcinomas using the expression of these genes showed to be very accurate, also when tested in an independent expression data set. Gene-sets associated with ageing (which is related to senescence) and chromosomal instability were upregulated, and a gene-set associated with fatty acid metabolism was downregulated in carcinomas compared to adenomas. Moreover, gene-sets associated with chromosomal location revealed chromosome 4q22 loss and chromosome 20q gain of gene-set expression as being relevant in this progression. These data are consistent with the notion that adenomas and carcinomas are distinct biological entities. Disruption of specific biological processes like senescence (ageing), maintenance of chromosomal instability and altered metabolism, are key factors in the progression from adenoma to carcinoma.