Background Most colon cancers have aberrant Wnt/β-catenin signalling and are a major therapeutic challenge. HAMLET (human alpha-lactalbumin made lethal to tumour cells) is the first member of a new family of tumouricidal, unfolded, protein-lipid complexes consisting of partially unfolded α-lactalbumin and oleic acid. HAMLET exhibits broad antitumour activity and kills many types of tumour cells in vitro; this tumouricidal activity is maintained in vivo. Methods We investigated if HAMLET can be used for colon cancer therapy and prevention. Findings Peroral HAMLET treatment reduced tumour progression and mortality in APC(Min/+) mice carrying mutations relevant to human colorectal tumours. HAMLET treatment reduced nuclear β-catenin and related tumour markers. Gene expression analysis of treated tumours showed inhibition of Wnt signalling and a shift to a more differentiated phenotype. In APC−mutated colon cancer cells, HAMLET altered β-catenin integrity and localisation through an ion-channel-dependent pathway, defining a novel mechanism controlling Wnt signalling. Tumour development was also significantly prevented by supplying HAMLET in the drinking water from the time of weaning. Interpretation We have identified HAMLET as a potential new and efficient therapeutic and prophylactic modality against colon cancer.