Altered Iron-Mediator Expression Identifies Chemotherapeutic Effects of Deferoxamine on Head and Neck Squamous Cell Carcinoma
Alexis Donneys, MD, MS, Chitra Subramanian, PhD, Jeremy Lynn, BS, Kevin Urlaub, BS, Halil Safak Uygur, MD, Keshav Lalchandani, BS, Kevin Kovatch, MD, Alexandra Luby, BS, Lauren Buchman, Noah S. Nelson, BS, Matthew Knudson, Mark S. Cohen, MD, Steven R. Buchman, MD.
University of Michigan, Ann Arbor, MI, USA.
PURPOSE: We have previously validated the pre-clinical use of deferoxamine (DFO) as an osteogenic stimulant capable of preventing non-unions in irradiated mandible fractures. DFO indirectly bolsters bone formation by stimulating angiogenesis and restoring diminished vascularity within irradiated tissues. Despite this promise, there have been concerns regarding the use of DFO to treat osseous defects in head and neck cancer survivors due to its remarkable ability to induce angiogenesis and promote tissue vascularization. Because of its iron chelating mechanism of action, we subsequently demonstrated that DFO actually elicits toxic, anti-tumorigenic actions on MDA-1986 head and neck squamous cell carcinoma (HNSCC) in vitro and in vivo. Aside from iron's critical metabolic functions, it is also crucial for the survival of rapidly dividing cancer cells, and its removal intensely inhibits tumor formation. In essence, DFO kills cancer cells by depleting iron stores that these rapidly dividing cells require, while protecting angiogenic mechanisms and stimulating the normal growth of blood vessels in injured tissues.
To further investigate this line of inquiry, we surveyed the Alabama HNSCC patient survivorship TGCA database for iron related genes and found that Transferrin Receptor Protein (TFRC) and Ferritin Heavy Chain-1 (FTH-1) were significantly upregulated in HNSCC, suggesting that dysregulation of iron mediators play a role in HNSCC development. Based on these observations, we posited that DFO exposure would alter HNSCC iron-mediator expression.
METHODS: We analyzed human MDA-1986 and UMSCC 108 HNSCC cells (HPV + and - cells, respectively) with and without DFO treatment (50 and 100ÁM) with Western Blot analysis for TFRC and FTH-1. DFO treated cells were also compared to non-treated controls and positive controls (5Gy radiation [XRT]). All experiments were conducted in triplicate. Subsequently, we developed in vivo tagged buccal xenografts in Nu/Nu mice. Mice were randomly assigned to control, XRT, and DFO groups (n=5/group). XRT mice received 3 fractionated doses of 3 Gy over 10 days. DFO mice received 5 peritumoral injections of DFO (100ÁM each). Tumor volumes were measured every third day (ANOVA, p > 0.05). After 12 weeks, excised tumors were analyzed with western blot and qPCR.
RESULTS: In vitro we found that both cell lines downregulated TFRC in response to 100ÁM DFO, FTH-1 was downregulated in HPV- cells, but was decreased throughout all groups in HPV+ cells (untreated and treated). In vivo, we observed a significant decrease in tumor volume upon addition of DFO and downregulation of FTH-1 was observed and validated with western blot and qPCR in DFO treated tumors.
CONCLUSION: These results suggest that DFO is not only safe to use in HNSCC, but it may offer a promising chemotherapeutic approach for oncologic management. Additionally, its chemotherapeutic properties can be gauged utilizing unique signature gene patterns of iron mediator expression as biomarkers within varying sub-types of HNSCC. Due to the heterogeneous nature of HNSCC, further studies examining the effects of DFO in a variety of HNSCC subtypes are warranted.
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