DNase I hypersensitivity regions correlate with genetic regulatory loci and binding sites for sequence-specific DNA-binding proteins. We present data supporting the presence of novel DNase 1 hypersensitive sites (which we have designated sites VI–IX) in both the body of the human c-myc gene downstream from exon 2 and the 3 ′-flanking region of the c-myc gene in HL-60 cells. All of these novel DH sites are markedly decreased when HL-60 cells are treated with either dimethyl sulfoxide (DMSO) or retinoic acid. Moreover, a similar pattern of DNase I hypersensitive sites in this region of c-myc was present in MCF-7 human breast cancer cells growing in culture. Our results suggest a potential role for these sites in transcriptional regulation of the human c-myc gene.