Actinomycin (Act) analogs, differing in the chemical substitution(s) made at various positions in either their pentapeptide chain(s) or chromophore ring, were evaluated for their antitumor activity in mice bearing either Ridgway osteogenic sarcoma (ROS) or P388 leukemia. Of the analogs tested against advanced (2--3-g) ROS tumors, azetomicin I and Act III caused therapeutic responses which, although variable, were nevertheless indicative of antitumor activities greater than was found using Act D. Several other analogs, Act C2, 2-N-(gamma-hydroxypropyl)-Act D, Act X0delta, and azetomicin II, displayed antitumor activity in ROS-bearing mice which varied, in different experiments, from comparable to superior to that achieved using Act D. Additionally, Act Pip1beta and 3'-(4-cisCl-Pro)-Act were comparable to, and Act-2-hydroxy-C3 inferior to, Act D in activity against ROS. Both azetomicin I and II were as effective as Act D in mice bearing P388 leukemia. Moreover, a subline of P388 that is resistant to Act D was cross-resistant to both azetomicin I and II.