Hypoxia is a common feature of solid tumors and promotes resistance to apoptosis from cancer therapies that induce DNA damage. The mechanism for this resistance, however, remains unclear. Since activation of the p53 pathway plays a major role in determining whether cells undergo apoptosis in response to DNA damage, we performed a microarray analysis of p53-dependent gene expression changes in response to DNA damage combined with hypoxia. When the H460 human lung cancer cell line was treated with hypoxia and etoposide, a chemotherapy agent that induces double-stranded DNA breaks, the dominant transcriptional response was regulated by DNA damage in a p53-dependent manner. Interestingly, however, DNA damage combined with hypoxia modulated both the intensity of the p53 response and the composition of downstream target genes. For example, there was synergistic activation of known p53 target genes such as p21 and gadd45, and the unique induction of other potentially novel p53 target genes including Rad and I-Rel. In addition, analysis of repressed genes supported a model for antagonism of c-Myc signaling in hypoxia, based on the downregulation of several known c-Myc target genes and the induction of Mxi1, a c-Myc antagonist. These data suggest a hypothesis that the combination of hypoxia and DNA damage promotes resistance to therapy by eliciting a transcriptional response that favors cell cycle arrest over apoptosis.