Analogues of staurosporine were synthesized and their ability to inhibit protein kinases was examined. Staurosporine is a potent but non‐selective inhibitor of in vitro protein kinase C (PKC) activity (IC₅₀ 6.0 _nm). The derivative CGP 41 251 had reduced PKC activity with an IC₅₀ of 50 nm but showed a high degree of selectivity when assayed for inhibition of cyclic AMP‐dependent protein kinase (IC₅₀, 2.4 μm). 56 kinase (IC₅₀ 5.0 μm). and tyrosine‐kinase‐specific activity of epidermal growth factor receptor (IC₅₀ 3.0 μm). Staurosporine and CGP 41 251 exerted growth inhibition in the human bladder carcinoma line T‐24, human promyelocytic leukemia line HL‐60 and bovine corneal endothelial cells at concentrations which correlated well with in vitro PKC inhibition. In addition, both compounds inhibited the release of H₂O₂ from human monocytes pre‐treated with 12‐O‐tetradecanoyl‐phorbol‐13‐acetate at non‐toxic concentrations. In vivo anti‐tumor activity was examined in T‐24 human bladder carcinoma xenografts in athymic nude mice. Tumor growth inhibition tests revealed significant anti‐tumor activity (2_p < 0.001) at 1/10 of the maximum tolerated doses for both compounds. By contrast, a closely related derivative of staurosporine (CGP 42 700) was inactive at concentrations of over 100 μm in all in vitro enzyme and anti‐proliferative assays as well as in animal tumor models. Our data suggest an association between PKC inhibition and anti‐proliferative and anti‐tumor activity.