Abstract The effect of N-methyl- N-nitrosourea (MNU) on the cell cycle, DNA synthesis and chromosomal sensitivity of cultivated diploid fibroblasts and fibroblasts with trisomy 21 was investigated in vitro. With the exception of the inhibition of G 2, Down's cells proved to be more sensitive than diploid cells with respect to the decrease of the mitotic and labelling index, the inhibition of the progression of cells through the early and middle S and the frequency of induced chromosomal aberrations. The chromosomal sensitivity was dependent on the position of cells in the cell cycle during treatment with MNU. If treated during late S no differences concerning the S block and aberration frequencies were found between diploid and Down's cells. However, if MNU treatment took place in the middle and early S, Down's cells were more sensitive. The higher aberration frequencies in Down's cells resulted from elevated levels of chromatid breaks, multiple fragmentations and chromatid translocations. Possible reasons for the increased sensitivity of Down's cells are discussed.