A series of new substituted 7-phenyl-3H-pyrrolo[3,2-f]quinolin-9-ones were synthesized and evaluated for their antiproliferative activity. The most active derivatives showed high selectivity against human leukemia cell lines and potently inhibited their growth, with GI(50) values in the nanomolar range. The active compounds strongly blocked tubulin assembly and colchicine binding to tubulin. Their activities were equal to or greater than that of the reference compound combretastatin A-4. Flow cytometry studies showed that the two most active compounds arrested Jurkat cells in the G(2)/M cell-cycle phase in a concentration-dependent manner. This effect was associated with apoptosis, mitochondrial depolarization, generation of reactive oxygen species, activation of caspase-3, and cleavage of the enzyme poly(ADP-ribose) polymerase.