DDDR-61. Ferroportin-dependent iron efflux as a novel mechanism of gallium maltolate in glioblastoma

Abstract BACKGROUND Gallium maltolate (GaM) is a metallocompound being evaluated in clinical trials for the treatment of glioblastoma (GBM). Gallium (Ga) blocks tumor iron-dependent processes including cellular iron uptake, ribonucleotide reductase activity, and mitochondrial function. GaM inhibits GBM growth and prolongs survival in animals with orthotopically implanted GBM xenografts. Ga’s action on cellular iron trafficking and efflux by ferroportin, the iron exporter, is unknown. Therefore, we examined the interaction between GaM and ferroportin to determine whether ferroportin impacts GBM sensitivity to GaM. METHODS Ferroportin expression was quantified by western blot in human GBM cells (D54) and a ferroportin Tet-on inducible cell line (IEC-6). GaM’s effect on cellular iron uptake and efflux was measured by 55Fe pulse-chase experiments. Cell proliferation was measured by Incucyte assay. RESULTS GaM (15 and 30μM) increased ferroportin expression in D54 cells at 24, 48, and 72h consistent with a disruption of the iron pool which regulates ferroportin and iron efflux. In contrast, GaM 60μM at 72h decreased ferroportin expression (p=0.0288) suggesting a compensatory downregulation by cells to retain iron. In D54 cells, GaM produced iron efflux within hours (p=0.0195). In IEC-6 cells, GaM produced an iron efflux only when ferroportin expression was induced (p=0.0221). IEC-6 cell growth rate was reduced in ferroportin-on cells relative to ferroportin-off (Δtime to reach 50% confluence = 49.9h). The decrease in cell growth with upregulated ferroportin was partially restored by hepcidin, a ferroportin inhibitor. Overexpression of ferroportin increased the antiproliferative action of GaM by 2.5-fold. CONCLUSIONS Our results indicate GaM contributes to intracellular iron depletion by increasing ferroportin-mediated iron efflux. Changes in ferroportin appear to modulate cell sensitivity to GaM, suggesting that ferroportin plays a functional role in GaM’s antiproliferative mechanism of action. Ongoing studies of GaM sensitivity and Ga export in ferroportin-inducible GBM cells will also be presented.