Inhibition of hypoxia-inducible factor 1 with acriflavine sensitizes hypoxic tumor cells to photodynamic therapy with zinc phthalocyanine-encapsulating cationic liposomes

Photodynamic therapy (PDT) is a tumor treatment modality in which a tumorlocalized photosensitizer is excited with light, which results in local production of reactive oxygen species, destruction of tumor vasculature, tumor hypoxia, tumor cell death, and induction of an anti-tumor immune response. However, pre-existing tumor hypoxia may desensitize tumors to PDT by activating the hypoxia-inducible factor 1 (HIF-1) survival pathway. Therefore, we hypothesized that inhibition of HIF-1 with acriflavine (ACF) would exacerbate cell death in human epidermoid carcinoma (A431) cells. PDT of A431 tumor cells was performed using newly developed and optimized PEGylated cationic liposomes containing the photosensitizer zinc phthalocyanine (ZnPC). Molecular docking revealed that ACF binds to the dimerization domain of HIF-1 alpha, and confocal microscopy confirmed translocation of ACF from the cytosol to the nucleus under hypoxia. HIF-1 was stabilized in hypoxic, but not normoxic, A431 cells following PDT. Inhibition of HIF-1 with ACF increased the extent of PDT-induced cell death under hypoxic conditions and reduced the expression of the HIF-1 target genes VEGF, PTGS2, and EDN1. Moreover, co-encapsulation of ACF in the aqueous core of ZnPC-containing liposomes yielded an adjuvant effect on PDT efficacy that was comparable to non-encapsulated ACF. In conclusion, HIF-1 contributes to A431 tumor cell survival following PDT with liposomal ZnPC. Inhibition of HIF-1 with free or liposomal ACF improves PDT efficacy.