Abstract Background The PTEN tumour suppressor gene and PIK3CA proto-oncogene encode proteins which contribute to regulation and propagation of signal transduction through the PI3K/AKT signalling pathway. This study investigates the prevalence of loss of PTEN expression and mutations in both PTEN and PIK3CA in colorectal cancers (CRC) and their associations with tumour clinicopathological features, lifestyle factors and dietary consumptions. Methods 186 adenocarcinomas and 16 adenomas from the EPIC Norfolk study were tested for PTEN and PIK3CA mutations by DNA sequencing and PTEN expression changes by immunohistochemistry. Dietary and lifestyle data were collected prospectively using seven day food diaries and lifestyle questionnaires. Results Mutations in exons 7 and 8 of PTEN were observed in 2.2% of CRC and PTEN loss of expression was identified in 34.9% CRC. Negative PTEN expression was associated with lower blood low-density lipoprotein concentrations (p = 0.05). PIK3CA mutations were observed in 7% of cancers and were more frequent in CRCs in females (p = 0.04). Analysis of dietary intakes demonstrated no link between PTEN expression status and any specific dietary factor. PTEN expression negative, proximal CRC were of more advanced Dukes' stage (p = 0.02) and poor differentiation (p < 0.01). Testing of the prevalence of PIK3CA mutations and loss of PTEN expression demonstrated that these two events were independent (p = 0.55). Conclusion These data demonstrated the frequent occurrence (34.9%) of PTEN loss of expression in colorectal cancers, for which gene mutations do not appear to be the main cause. Furthermore, dietary factors are not associated with loss of PTEN expression. PTEN expression negative CRC were not homogenous, as proximal cancers were associated with a more advanced Dukes' stage and poor differentiation, whereas distal cancers were associated with earlier Dukes' stage.