16. Evaluating the Safety of Transdermal Deferoxamine Patch in Oncological Settings

PURPOSE: Transdermal deferoxamine (DFO) patch (TauTona Group) represents a novel experimental therapeutic modality with the potential to mitigate radiation-induced dermal injury in breast cancer patients undergoing radiotherapy. Nonetheless, the safety profile of DFO patch within an oncological context has not been extensively explored. Consequently, this study aims to investigate the effect of DFO on cancer cell biology using a xenograft murine breast cancer model. METHODS: Human-derived breast cancer cells (MCF7-Luc2; ATCC) were engrafted into the mammary gland of 8-week-old immunodeficient female mice (NCG) to create the xenograft model. An estrogen pellet was inserted subcutaneously three days before breast cancer cells injection to support tumor growth. The in vivo tumor growth was evaluated weekly using luciferin injection and LAGO bioluminescence imaging system. Once tumor reached bioluminescence intensity ~1x108, DFO patches were applied daily for 30 days. Non-DFO treated mice served as control. The changes in mice body weight were recorded weekly. Harvested skin and tumor samples were preserved in 10% formalin and embedded in paraffin for immunofluorescence (IF) staining, and in All Protect Reagent for protein extraction. IF staining of Ki-67 and CD31 was performed on 4 μm tumor sections to evaluate cell proliferation and tissue vascularization, respectively. Stained slides were photographed on Zeiss confocal or Keyence microscope. ImageJ software was used to calculate the number of Ki-67 positive cells and endothelial cell density. Western blot analysis on skin and tumor tissues was performed to quantify HIF-1α (Hypoxia-inducible factor 1-alpha) protein expression. Statistical analysis was performed using a student t-test and GraphPad Prism software. p-val < 0.05 was considered significant. RESULTS: Western blot analysis revealed an increase in HIF-1α protein level in mice skin treated with DFO, confirming effective DFO-treatment. Simultaneously, bioluminiscence assay demonstrated no significant differences in relative tumor growth over the 30-day study period between the treated and control groups (FC=0.8, p-val=0.392). The quantitative analysis of Ki-67 positive cells in tumor tissue showed no significant difference in cell proliferation between DFO-treated and control tumors (FC=0.8, p-val=0.131). Tumor vascularization, presented as a percentage area of CD31 positive signal, was not affected by topical DFO treatment (FC=0.7, p-val=0.132). Notably, mice body weight monitoring throughout the treatment period revealed a trend towards greater overall mass loss in DFO-treated group compared to controls (FC=0.9, p=0.011). CONCLUSION: This study reinforces the importance of verifying the oncological safety of radioprotective interventions in vivo conditions. Our findings suggest that the ability of DFO to exert regenerative and pro-vasculogenic effects is limited to superficial tissues. Thus, topical DFO-treatment most likely exposes minimal risk of potentially accelerating the growth of residual cancer cells in deeper tissues, as indicated by no significant difference in tumor growth, cancer cell proliferation, and vascularization in DFO-treated mice compared to control. Additional studies are necessary to investigate the effect of DFO-treatment on the efficacy of radiation therapy.