Photodynamic therapy mediates the oxygen-independent activation of hypoxia-inducible factor 1α

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Abstract
Photodynamic therapy (PDT) induces the expression of the hypoxia-inducible factor 1α (HIF-1α) subunit of the HIF-1 transcription factor and its target genes in vitro and in vivo. PDT also induces the expression of the enzyme cyclooxygenase-2 and its metabolite, prostaglandin E2 (PGE2). PGE2 and hypoxia act independently and synergistically to increase HIF-1α accumulation and nuclear translocation. To examine the expression of HIF-1 target genes in response to PDT-mediated oxidative stress and PGE2 under normoxic conditions, we established EMT6 cells transfected with a plasmid consisting of a hypoxia response element promoter and a downstream gene encoding for green fluorescent protein (GFP). To examine the temporal kinetics of HIF-1α nuclear translocation in response to PDT, we transfected a second line of EMT6 cells with a GFP-tagged HIF-1α fusion vector. Cell monolayers were incubated with 1 μg mL−1 Photofrin for 24 h and irradiated with fluences of 1, 2.5, and 5 J cm−2. Direct measurement of oxygen concentration during irradiation confirmed that cells remained well oxygenated. Cells were also exposed to 1 and 10 μmol/L PGE2 for 3 h. In normoxic conditions, Photofrin, PDT, and PGE2 treatment activated HIF-1α and induced its nuclear translocation. Maximal Photofrin-PDT–mediated HIF-1α activation was intermediate in magnitude between that induced by PGE2 and that by the hypoxia mimic cobalt chloride. This work establishes that PDT induces significant activation of the HIF-1α pathway in the absence of hypoxia and supports the interpretation that the induction of HIF-1 target genes by PDT may be mediated, at least in part, by the prostaglandin pathway. [Mol Cancer Ther 2006;5(12):3268–74]