Heme Oxygenase Promotes B-Raf-dependent Melanosphere Formation.

Melanoma is the fifth most common malignancy in the US. While the overall 5-year survival rate has improved in the past decade, the 5-year survival rate for melanoma patients that are diagnosed with metastatic disease is only 23%, due in part to acquired drug resistance to both targeted therapies (i.e., B-Raf inhibitors) and immunotherapies, underscoring the need for novel therapeutic targets. Biosynthesis and degradation of heme, an iron-bound protoporphyrin molecule utilized by a wide variety of metabolic processes, is tightly regulated. Two closely related enzymes, heme oxygenase-1 (HMOX1) and heme oxygenase-2 (HMOX2), degrade free heme to produce carbon monoxide, Fe2+ and biliverdin. HMOX1 expression is controlled via the transcriptional activator, NFE2L2, and the transcriptional repressor, Bach1. Transcription of HMOX1 and other NFE2L2-dependent genes is increased in response to electrophilic and reactive oxygen species (ROS). Many tumor-derived cell lines have elevated levels of NFE2L2 and elevated expression of NFE2L2-dependent genes contributes to tumor growth and acquired resistance to therapies. We utilized an in vitro melanosphere assay that correlates with melanoma tumorigenesis in vivo to report a novel role for heme oxygenase activity in melanosphere formation by human melanoma-derived cell lines. Transcriptional induction of HMOX1 through derepression of Bach1 with cobalt protoporphyrin (CoPP) or transcriptional activation of HMOX2 by oncogenic B-RafV600E results in increased melanosphere formation. Genetic ablation of HMOX1 diminishes melanosphere formation. Further, inhibition of heme oxygenase activity with tin protoporphyrin markedly reduces melanosphere formation driven by either Bach1 derepression or B-RafV600E expression. Global transcriptome analysis implicates genes involved in focal adhesion and extracellular matrix interactions in melanosphere formation. These results suggest heme oxygenase activity as a novel target for drug development. Importantly, current therapeutic metalloporphoryins (e.g., SnMP, SnPP) may be repurposed for the treatment of metastatic melanoma.