Heme Oxygenase‐1‐Derived Carbon Monoxide Promotes Growth and Progression in Uveal Melanoma
Authors: Longhitano Lucia , Russo Giuliano , Caltabiano Rosario , Reibaldi Michele , Castracani Carlo Castruccio , Distefano Alfio , Pittalà Valeria , Salerno Loredana , Tibullo Daniele , Avola Roberto , Nicolosi Daniela , Viola Maria , Volti Giovanni Li , Barbagallo Ignazio 
Published: June 2021

Heme Oxygenase‐1 (HO‐1) catalyzes the enzymatic degradation of heme with the simultaneous release of CO, Fe2+and biliverdin. Uveal melanoma (UM) is the most common primary intraocular tumor in adults, with about 1200–1500 new cases occurring per year in the U.S. Unfortunately, no chemotherapeutic regimen or immunotherapy has been demonstrated to be effective in these patients. The aim of the present study was to characterize in vitro the role of HO‐1 and its enzymatic byproducts in UM in cancer progression. Such results were further confirmed in a series of UM human biopsies (n=80) retrieved from our database. The study was conducted according to the basic principles of the declaration oh Helsinki. Cell proliferation and colony formation capacity were evaluated by Xcelligence technology and clonogenic assay. Cells were also treated with Hemin (10 μM) and/or VP13/47 (100 μM) as potent pharmacological inducer and inhibitor of HO‐1 respectively. HO‐1 protein expression was evaluated by real time PCR and immunocytochemistry. In a separate set of experiments, we also tested the biological effects of HO activity byproducts by treating cells with CO releasing molecules (CORM‐3 and CORM‐A1 25 μM) and biliverdin (25 μM). Our results showed that hemin significantly increased (46.34±4.5‐fold increase vs untreated cells, p<0.01) HO‐1 gene and protein expression resulting in a significant increase in cell proliferation (2.3±0.3 fold increase vs untreated cells, p<0.01) and colony formation capacity (1003±20 colonies in treated cells vs 503±14 in untreated cells). Similarly, hemin treatment resulted in a significant decrease of doubling time (20.42±2.3 h vs 32.74±1.2 h, p<0.01) and this effect was abolished by concomitant treatment with VP13/47 (49.71±3.9, p<0.01). Consistently, CORMs treatments significantly decreased doubling time (13.8±2.9 h and 11.5±1.5 h for CORMA1 and CORM3 respectively, p<0.01 vs untreated cells). Our results also showed that VP13/47 reversed the proliferative effect of hemin and strongly inhibited growth when used alone. Furthermore, we observed similar proliferative effects of hemin with CORMs but not with biliverdin alone. Interestingly, CORMs treatment abolished the antiproliferative effect of VP13/47 whereas biliverdin showed no effects. The direct correlation of HO‐1 with tumor growth and progression was further confirmed in human biopsies showing that HO‐1 expression significantly correlated with the expression of p16, a well‐established biomarker of cell proliferation. In conclusion, our results showed that HO‐1 derived CO is required for UM growth and progression and the HO system may therefore represent a good candidate to be exploited as a target for new chemotherapic agents.Support or Funding InformationThis work was supported by FIR 2016–2018 from the University of Catania.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.