Biochemical studies of transient intermediates in relation to chemical carcinogenesis

Many chemical carcinogens must be metabolized to chemically reactive transient species before they can exert their full toxic action on mammalian cells. In general, this metabolic activation is performed by NADPH-dependent enzymes in the endoplasmic reticulum; the NADPH-cytochrome P-450 electron-transport chain is very important in this respect. Biochemical studies on the chemical reactivities of such transient intermediates require the application of various fast-reaction and free-radical techniques: the use of such techniques is illustrated by reference to the metabolism of carbon tetrachloride. CCl4 is metabolized by liver endoplasmic reticulum in the presence of NADPH to a highly reactive product, probably CCl3; this activation of CCl4 results in covalent binding of CCl3 and lipid peroxidation. The steady-state concentration of CCl3 is too low to be measured directly by e.s.r. spectroscopy but radical species can be accumulated with spin-trap techniques. The CCl3 radical can be generated by pulse radiolysis and the ensuing reactions with biologically important neighbouring species can be followed in the microsecond range by kinetic spectroscopy. The results point to the high reactivity of CCl3 and its restriction to a microenvironment within the endoplasmic reticulum. Highly reactive electrophilic radicals (e.g. CCl3) can initiate lipid peroxidation in biomembranes and this is associated with changes in polyunsaturated fatty acids and in membrane fluidity. The results are discussed in relation to carcinogen activation, to free-radical-mediated reactions in biomembranes, and to the general thesis that the production of reactive aldehydes by lipid peroxidation may act as a 'coarse control' of cell division.