THE USE OF A 3 COMPARTMENT IN-VITRO MODEL TO INVESTIGATE THE ROLE OF HEPATIC DRUG-METABOLISM IN DRUG-INDUCED BLOOD DYSCRASIAS

1 N-hydroxylation is thought to be an essential step in the haemotoxic ity of dapsone (DDS). To investigate both metabolism-dependent and cel l-selective drug toxicity in vitro we have developed a three-compartme nt system in which an hepatic drug metabolizing system is contained wi thin a central compartment separated by semi-permeable membranes from compartments containing mononuclear leucocytes (MNL) and red blood cel ls (RBC). 2 Metabolism of dapsone (100 mum) by rat liver microsomes re sulted in toxicity to RBC cells (47.3 +/- 2.1% methaemoglobin), but th ere was no significant toxicity toward MNL (3.7 +/- 1.3% cell death) c ompared with control values (1.6 +/- 0.9%). However, when RBC were rep laced with buffer in the third compartment there was significantly gre ater (P < 0.001) white cell toxicity (17.6 +/- 0.6% cell death), demon strating the protection of MNL by RBC. Metabolism of dapsone by human liver microsomes again resulted in RBC toxicity (12.5 +/- 3.3% methaem oglobin) but no significant MNL toxicity (2.9 +/- 0.8% cell death). Re placement of RBC resulted in a significant (P < 0.001) increase in MNL toxicity (6.5 +/- 0.7% cell death). Addition of synthetic dapsone hyd roxylamine (30 mum) in the absence of a metabolizing system and with n o RBC in the third compartment resulted in significant (P < 0.001) tox icity toward MNL (43.36 +/- 5.82% cell death) compared with control (1 .8 +/- 1.1%). The presence of RBC in the third compartment resulted in a significant (P < 0.001) decrease in MNL toxicity (17.6 +/- 2.2% cel l death), with 40.1 +/- 3.7% methaemoglobin in the RBC. 3 Like dapsone , procainamide and hydralazine both undergo bioactivation at a nitroge n centre, but are not toxic to red cells in vivo. Both drugs were bioa ctivated by rat liver microsomes to species which were toxic toward MN L in the three compartment model (13.2 +/- 1.3 and 13.3 +/- 2.0% respe ctively), but non-toxic toward RBC. Replacement of red cells with buff er had no significant effect on the toxicity of either compound toward s MNL. No bioactivation could be detected in the presence of human liv er microsomes. 4 Primaquine, which is associated with a high incidence of red cell toxicity in vivo, was apparently non-toxic to either red or white cells in the three compartment system. However, in incubation s without membranes, in which the target cells were not separated from the drug activating system, primaquine was metabolized to a species t oxic toward both MNL and RBC by both rat (13.7 +/- 1.4% cell death, 3. 2 +/- 0.3% methaemoglobin) and human liver microsomes (7.6 +/- 1.3% ce ll death, 3.2 +/- 0.2% methaemoglobin). This is consistent with the hy pothesis that highly reactive and short-lived metabolites are responsi ble for toxicity associated with primaquine.