The superhelical properties of nuclear DNA from malignant cells of human origin (leukemic cell lines Reh and D-562, mesothelioma cell line, and peripheral blood monocytes from acute monocytic leukemia) were investigated in neutral sucrose gradients with ethidium bromide. As compared with the universal superhelical density of nuclear DNA from normal cells, the DNA released from malignant cells showed a substantially higher negative superhelix content, equivalent to an increased deficiency in right-handed DNA duplex turns. In regard to analogous results obtained from other malignant cell systems, the extremely underwound structural state of nuclear DNA appears to be a feature common to malignant cells.