Opioids promote tumor angiogenesis in mammary malignancies, but the underlying signaling mechanism is largely unknown. The current study investigated the hypothesis that stimulation of -opioid receptors (DOR) in breast cancer (BCa) cells activates the hypoxia-inducible factor 1 (HIF-1), which triggers synthesis and release of diverse angiogenic factors. Immunoblotting revealed that incubation of human MCF-7 and T47D breast cancer cells with the DOR agonist d-Ala,d-Leu-enkephalin (DADLE) resulted in a transient accumulation and thus activation of HIF-1 DADLE-induced HIF-1 activation preceded PI3K/Akt stimulation and was blocked by the DOR antagonist naltrindole and naloxone, pertussis toxin, different phosphoinositide 3-kinase (PI3K) inhibitors, and the Akt inhibitor Akti-1/2. Whereas DADLE exposure had no effect on the expression and secretion of vascular endothelial growth factor (VEGF) in BCa cells, an increased abundance of cyclooxygenase-2 (COX-2) and release of prostaglandin E2 (PGE) was detected. DADLE-induced COX-2 expression was also observed in three-dimensional cultured MCF-7 cells and impaired by PI3K/Akt inhibitors and the HIF-1 inhibitor echinomycin. Supernatant from DADLE-treated MCF-7 cells triggered sprouting of endothelial (END) cells, which was blocked when MCF-7 cells were pretreated with echinomycin or the COX-2 inhibitor celecoxib. Also no sprouting was observed when END cells were exposed to the PGE receptor antagonist PF-04418948. The findings together indicate that DOR stimulation in BCa cells leads to PI3K/Akt-dependent HIF-1 activation and COX-2 expression, which trigger END cell sprouting by paracrine activation of PGE receptors. These findings provide a potential mechanism of opioid-driven tumor angiogenesis and thus therapeutic targets to combat the tumor-angiogenic opioid effect. SIGNIFICANCE STATEMENT: Opioids are indispensable analgesics for treating cancer-related pain. However, opioids were found to promote tumor growth and metastasis, which questions the use of these potent pain-relieving drugs in cancer patients. Enhanced tumor vascularization after opioid treatment implies that tumor progression results from angiogenic opioid effects. Thus, understanding the signaling mechanism of opioid-driven tumor angiogenesis helps to identify therapeutic targets to combat these undesired tumor effects. The present study reveals that stimulation of δ-opioid receptors in breast cancer cells leads to an activation of HIF-1α and expression of COX-2 via PI3K/Akt stimulation, which results in a paracrine activation of vascular endothelial cells by prostaglandin E receptors.