A.B. Madhankumar

Abstract 4436: The down-regulation of ferritin heavy chain as an adjuvant therapy in human glioma.

Proceedings: AACR 104th Annual Meeting 2013; Apr 6-10, 2013; Washington, DC Increased iron requirements and heightened iron metabolism is among the key characteristics of highly proliferative cancer cells. As the major iron storage protein, ferritin expression is elevated in cancer cells and we have previously shown that ferritin down-regulation in human glioblastoma cells increases their sensitivity to chemotherapeutic agent both in vitro and in vivo. In this study, we report that the sensitization of H-ferritin down-regulation in glioma cells against radiation and suggest the potential of H-ferritin down-regulation as an adjuvant therapy in human glioma. Down-regulation of H-ferritin was performed in human glioma cell line U251 through a nanotechnology-based transfection platform and effective knockdown efficiency was achieved. A transient decrease in transferrin receptor level was also observed, suggesting a release of intracellular iron with loss of ferritin. This iron release in turn produced intracellular oxidative stress, demonstrated by protein oxidation damage, as well as a decrease in the stability of hypoxia-inducible factors (HIFs), which is an indicator of radioresistance. In U251 cells, exposure to radiation resulted in protein oxidation which could be exacerbated by H-ferritin down-regulation. Previously we have demonstrated a DNA protection role for H-ferritin. Here we report that the down-regulation of H-ferritin is associated with increased DNA vulnerability when cells were exposed to ionizing radiation. Moreover, the activation of DNA repair mechanisms was impaired when H-ferritin was absent. The in vitro data suggest a synergistic effect of radiation and H-ferritin down-regulation. Additionally, we expand our research into CD133-positive glioma stem cells (GSCs), which are notoriously resistant to anti-cancer treatment. Down-regulation of H-ferritin in these cells inhibited cell proliferation in vitro. With an intracranial glioma model established by GSC implantation, we demonstrated that the survival was significantly prolonged by H-ferritin siRNA transfection through intravenous injection, in tumor bearing mice treated with first-line drug Temodar. This study supports the potential of H-ferritin siRNA as an adjuvant therapy in glioma treatment. Citation Format: Min Pang, A.B. Madhankumar, Becky Webb, Xiaoli Liu, James R. Connor. The down-regulation of ferritin heavy chain as an adjuvant therapy in human glioma. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 4436. doi:10.1158/1538-7445.AM2013-4436

Abstract 1471: Role of ferritin heavy chain in therapeutic sensitivity of human glioblastoma cells

Proceedings: AACR 103rd Annual Meeting 2012‐‐ Mar 31‐Apr 4, 2012; Chicago, IL Elevated iron requirements and heightened iron metabolism is key to the survival and proliferation of cancer cells. As one of the major iron regulatory proteins, ferritin is elevated in cancer cells and we have previously shown that decreasing ferritin expression in human glioblastoma cells will increase the sensitivity to BCNU and radiation both in vitro and in vivo. In this study, we investigate the underlying mechanism of this finding. Through a nanotechnology-based transfection platform, the human glioma cell line U251 was transfected with H-ferritin siRNA and an effective knockdown efficiency of protein was achieved within 48 hours. H-ferritin down-regulation in glioma cells was associated with a transient decrease in transferrin receptor, suggesting a release of intracellular iron with loss of ferritin. This iron release in turn generated intracellular oxidative stress, which was demonstrated by protein oxidation. The stability of hypoxia-inducible factors (HIFs), which is an indicator of radioresistance, was decreased in glioma cells with H-ferritin down-regulation. This observation is consistent with the increased sensitivity to treatment. Exposure to radiation resulted in increased protein oxidation which could be exacerbated by H-ferritin down-regulation. Thus there is a synergistic effect of radiation and H-ferritin down-regulation; possibly also including the destabilization of HIFs. Additionally, we have previously demonstrated a DNA protection role for H-ferritin. Down-regulation of H-ferritin is associated with increased DNA vulnerability when cells were exposed to ionizing radiation. Moreover, the activation of DNA repair mechanisms was impaired when H-ferritin was absent. Finally, we expanded the vulnerability studies to include the first-line chemotherapeutic agent temozolomide and found that H-ferritin down-regulation increased the sensitivity of glioma cells to this DNA alkylating agent. This study suggests that the iron management protein, H-ferritin, has a cytoprotective role in human glioma cells, which renders them more resistant to conventional therapies. Therefore down-regulation of H-ferritin via nanoliposomes delivering siRNA is a potential adjuvant therapy in glioma treatment. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1471. doi:1538-7445.AM2012-1471