Bishydroxamic acids are effective inducers of differentiation in murine erythroleukemia cells. Flexible analogs of suberic acid bisdimethylamide are approximately 100 times as active as the parent compound or hexamethylenebisacetamide. They also induce differentiation of human promyelocytic leukemia cells (HL-60) and a subclone of human colon carcinoma cells (HT-29-U4). Some rigid bishydroxamic acids with benzene rings in the spacers are even more active toward murine erythroleukemia cells but show curious biological differences. In contrast to the flexible molecules, those with benzene spacers show poor activity toward HL-60 cells; they also have different geometric requirements, and they are not additive with hexamethylenebisacetamide in their effect. It is likely that rigid bishydroxamic acids, with a benzene ring spacer, induce differentiation by a different mechanism in spite of their chemical resemblance to the flexible bisamide and bishydroxamic acid inducers.