Disruption of cellular cholesterol transport and homeostasis as a novel mechanism of action of membrane-targeted alkylphospholipid analogues.

BACKGROUND AND
PURPOSE: Alkylphospholipid (APL) analogues constitute a new class of synthetic anti-tumour agents that act directly on cell membranes. We have previously demonstrated that hexadecylphosphocholine (HePC) alters intracellular cholesterol traffic and metabolism in HepG2 cells. We now extended our studies to analyse the effects of other clinically relevant APLs, such as edelfosine, erucylphosphocholine and perifosine on intracellular cholesterol homeostasis. EXPERIMENTAL APPROACH: Using radiolabelled substrates we determined the effect of APLs on cholesterol metabolism and cholesterol traffic from the plasma membrane to the endoplasmic reticulum (ER). Protein levels and gene expression of the main proteins involved in cholesterol homeostasis were analysed by Western blot and RT-PCR respectively. Membrane raft and non-raft fractions were isolated from HepG2 cells by a detergent-free method. KEY
RESULTS: All APLs inhibited the transport of cholesterol from the plasma membrane to the ER, which induced a significant cholesterogenic response in HepG2 cells. This response involved an increased gene expression and higher levels of several proteins related to the biosynthesis and the receptor-mediated uptake of cholesterol. Cell exposure to the APL-representative HePC enhanced the content of cholesterol mainly in the membrane raft fractions, compared with the untreated cells. CONCLUSIONS AND IMPLICATIONS: Membrane-targeted APLs exhibited a novel and common mechanism of action, through disruption of cholesterol homeostasis, which in turn affected specific lipid microdomains of cellular membranes.