The antltumour antibiotic actinomycin D normally binds to DNA by intercalation at sequences containing the CpG step, but In the presence of daunomycin It has been reported to Interact with poly(dA-dT). This observation has neither been confirmed nor explained. Here we have used a photoreactlve 7-azido derivative of actinomycin to study the effect of daunomycin on its binding to three DNA fragments. Daunomycin did indeed alter the binding of actinomycin to the DNA, such that the antibiotic was displaced from Its primary GpC sites onto secondary sites in the DNA, though not to AT regions especially. These findings suggest a possible scientific explanation for the Increased toxldty seen during combination chemotherapy with these two drugs. © 1994 Oxford University Press.