We have previously reported tumour-selective killing by the sigma (σ) receptor ligand rimcazole. We now report that rimcazole elevates hypoxia inducible factor-1α (HIF-1α) protein levels under normoxic conditions in colorectal (HCT-116) and mammary carcinoma (MDA MB 231) cells but fails to induce HIF-1α in normal fibroblasts or mammary epithelial cells. Combining the σ-1 agonist (+)-pentazocine with rimcazole substantially reduces the accumulation of HIF-1α, confirming that the effect is mediated at least partly by antagonism of σ-1 sites. HIF-1α knockdown by RNA interference attenuates rimcazole-induced cell death in both cell types. Thus, the induction of HIF-1α by rimcazole contributes to tumour cell killing. In a comparison of HCT-116p53+/+ and HCT-116p53−/− cells, HIF-1α levels are consistently higher after rimcazole treatment in HCT-116p53+/+ cells. Furthermore, although rimcazole kills HCT-116p53−/− cells, it has a more potent apoptosis-inducing effect in HCT-116p53+/+ cells. This suggests that the presence of functional p53 protein may enhance death induction by rimcazole in part through greater induction of HIF-1α. p53 is not required, however, for the rimcazole-induced engagement of HIF-1α in proapoptotic mode as HIF-1α knockdown attenuates rimcazole-induced death to comparable extents in p53 mutant and wild-type cell systems. Knowledge of HIF-1α involvement may assist the re-profiling of rimcazole and other σ ligands as cancer therapeutics.