The mechanism of HAMLET-induced cell death - cellular signalling, oncogenes and clinical perspectives

Despite recent advances in cancer treatment, truly innovative approaches are required to move beyond the modest benefits achieved to date. HAMLET is a human protein-lipid complex originally discovered in breast milk able to kill a wide range of tumour cells while leaving healthy, differentiated cells unaffected. The aim of this thesis was to identify mechanisms dictating HAMLET sensitivity and to define the events that lead to cell death in response to HAMLET. In Paper I we show that the properties sensitising cells to HAMLET coincide with the ‘’Hallmarks of cancer’’. Using a combination of shRNA screens, proteomic and metabolomic technology, we identified the c-Myc oncogene and hexokinase as essential determinants of HAMLET sensitivity. HAMLET sensitivity was modified by the glycolytic state of tumor cells and HAMLET induced a metabolic paralysis in tumour cells. In paper II the initiating events for tumour cell death are elucidated. By rapid activation of a single ion channel HAMLET disturbs the flux of cations in tumour cells and subsequent cellular responses are shown to rely on this mechanism for their initiation. Ion fluxes activated p38 MAPK signalling, which was found to be crucially involved in tumour cell death. Paper III identifies oleate as the functional ligand in HAMLET formation and shows that oleate actively contributes to the induction of the cellular response to HAMLET. Both protein and lipid are needed for HAMLET’s tumoricidal effect, however. Finally, in paper IV we identify HAMLET as a new therapeutic agent in colon cancer. HAMLET caused a significant reduction in tumour numbers as well as mortality in mice carrying a human APC mutation. In tumours surviving HAMLET challenge a reduction in onco-protein expression was detected as well as an increase in expression of glycolic enzymes. By long-term prophylaxis, prevention of tumour development was achieved