Interactions of Indolo[3,2-b]carbazoles and Related Polycyclic Aromatic Hydrocarbons with Specific Binding Sites for 2,3,7,8-tetrachlorodibenzo-p-dioxin in Rat Liver

In the present study we have investigated the capacity of various compounds sterically related to indolo[3,2-b]carbazole to inhibit specific 2,3,7,8-tetrachloro[1,6-3H]dibenzo-p-dioxin binding in rat liver cytosol, as analyzed by electrofocusing in polyacrylamide gels. When the two nitrogen atoms of indolo[3,2-b]carbazole (IC50 = 3.6 nM) were replaced with sulfur atoms, the affinity for the specific binding sites (IC50 = 3.3 nM) was similar to that of the parent compound, whereas the affinity decreased when the two nitrogen atoms were replaced with oxygen atoms (IC50 = 29 nM). Substitution with methyl groups at positions 5 and 11 (on the nitrogens) of indolo[3,2-b]carbazole resulted in increased affinity (IC50 = 1.2 nM), compared with that of the parent compound, whereas dimethylation at the 4,10- or 2,8-positions decreased the affinity (IC50 = 19 nM and IC50 > 150 nM, respectively). Substitution at positions 5 and 11 of indolo[3,2-b]carbazole with substituents larger than methyl, as in 5,11-diethylindolo[3,2-b]carbazole (IC50 = 8.9 nM), diacetylindolo[3,2-b]carbazole (IC50 = 11.2 nM), 5,11-dibutylindolo[3,2-b]carbazole (IC50 > 150 nM), and 5,11-di(N,N-dimethylaminoethyl)indolo[3,2-b]carbazole (IC50 > 1500 nM), also decreased the affinity. Introduction of oxygen in, or hydroxylation of, the middle ring of indolo[3,2-b]carbazole, giving indolo[3,2-b]carbazole-6,12-quinone (IC50 > 150 nM) or 6,12-dihydroxyindolo[3,2-b]carbazole (IC50 > 1500 nM), respectively, also lowered the affinity. We calculated the Gibbs free energy of solvation of the analogue isoquino[3,4-b]phenanthridine (IC50 = 137 nM), relative to that of dibenz[a,h]anthracene (IC50 = 2.5 nM), in water to be -6 kcal/mol by free energy perturbation, which indicates that the most important explanation for the observed difference in binding affinity is the smaller difference in relative free energy of binding at the binding sites, compared with the Gibbs free energy of solvation of the two compounds.