Inhibition of heme oxygenase-1 increases responsiveness of melanoma cells to ALA-based photodynamic therapy

Abstract

Based on the observation that 5-aminolevulinic acid (ALA) induces the expression of heme oxygenase-1 (HO-1) in cultured melanoma cells, the role of HO-1 on the effectiveness of 5-aminolevulinic acid-based photodynamic therapy (ALA-PDT) was examined. Transcriptional activation of the HO-1 gene is considered to be an adaptive response to oxidative and cellular stress and confers a protective capacity against cell and tissue injury, which could affect the responsiveness to ALA-PDT. A time-dependent accumulation (0-16 h) of protoporphyrin IX (PPIX) within melanoma cells was seen after incubation with ALA (0.5 mM ALA). Over the same time interval, a significant increase (up to 25-fold) in HO-1 protein expression was observed. Thus, the production and degradation of PPIX (via heme by HO-1) were simultaneously enhanced, leading to a reduced intracellular concentration of the photodynamically active substance PPIX. Diminishing HO-1 activity by the HO-1 inhibitor tin protoporphyrin IX (SnPPIX) significantly enhanced the formation of PPIX up to 1.8 fold. A further strong increase in HO-1 protein expression (up to 128-fold) was seen after ALA-PDT treatment. Induction of HO-1 is an essential step in the 'rescue response' of tumor cells. The pharmacological inhibition of HO-1 activity by SnPPIX leads to a considerable increase in the sensitivity of tumor cells to ALA-PDT treatment. At low radiation doses (0.42 J/cm2), the percentages of death cells increased significantly from 7.3±1.3% to 43.7±6.4%. This effect could be further intensified by cellular depletion of HO-1 mRNA by siRNA. The combination of pharmacological inactivation of HO-1 with gene silencing led to an increase in the death rate of up to 54.1±8.6%. The results presented indicate that HO-1 can play a protective role against ALA-PDT mediated cytotoxicity so that a specific inhibition of HO-1 activity and/or expression might be used to increase the efficacy of ALA-based photodynamic therapy.