A high therapeutic index is as important for gene-based therapies as it is for chemotherapy or radiotherapy. One approach has been transcriptional targeting through the use of tissue-specific regulatory elements. A more versatile approach would be to use a regulatory element that is controlled via a parameter common to a broad range of diseases. Ischemia is characteristic of a number of pathologies that range from vascular occlusion through to cancer. The state of low oxygen, hypoxia, triggers a transcriptional signaling pathway that is mediated by transcription factors binding to a specific enhancer, the hypoxia response element (HRE). These observations have therefore led to the use of HREs to drive gene expression in a number of target tissues from tumors to cardiac muscle. To translate these observations into a clinically useful vector system we have now assessed the potency of a number of naturally derived HREs in various configurations combined with minimal promoters. The optimal HRE has been introduced into a single transcription unit retroviral vector that can deliver regulated gene expression in response to hypoxia. An important feature of this new physiologically regulated vector is the combination of low basal expression and high-level activated expression that is on a par with that obtained with the cytomegalovirus immediate-early (CMV IE) promoter. The role of elements that stabilize mRNA in the presence of hypoxia has also been assessed. These hypoxia-regulated vectors may have utility for restricting the delivery of therapeutic proteins to tumors and ischemic sites.