Topical 5-fluorouracil more effectively eliminates epidermal clonal mutations than photodynamic therapy in a mouse model of early UV carcinogenesis

Summary: Abstract Body: Field treatment with topical 5-fluorouracil cream (5FU) and photodynamic therapy (PDT) is frequently used to treat actinic keratoses and early skin cancers. Previous studies by our group demonstrated that PDT effectively reduces the burden of epidermal clonal mutations, particularly those with variant allele fractions (VAFs) below 1%, in both human and mouse models. In this study, we expanded our investigation to include SKH1 mice treated with 5FU. Four weeks after the field treatment, 131 epidermal samples were collected from 9 SKH1 hairless mice treated by PDT (n=5) and 5FU (n=4) following 8 weeks of chronic 3x/week UVB exposure (120mJ/cm2). A total of 66 samples were from the treated side (38 PDT and 28 5FU-treated), and 75 were untreated samples from the same animals, serving as controls. Ultradeep sequencing (>20,000X) was performed using a custom-targeted panel capturing 251 kb of mouse genomic regions commonly mutated by UV exposure. Of the 25,973 mutations, 87% displayed a UV signature pattern. Both PDT and 5FU treatments significantly reduced mutation burdens in the treated skin (PDT: p<0.01, 5FU: p<0.0001), with 5FU showing a markedly stronger effect compared to PDT (median mutations per sample: Control = 246, PDT = 173, 5FU = 66). This overall difference was primarily driven by low-VAF (<1%) mutations (median mutations per sample: Control = 237, PDT = 167.5, 5FU = 51, p<0.0001) rather than high-VAF (≥1%) mutations (median mutations per sample: Control = 7, PDT = 3.5, 5FU = 3.5). Although the magnitude of the difference is likely model and dose-specific, the current study provides the first objective and quantitative comparison of the early field effect of PDT and 5FU treatments. Our findings indicate that clonal mutations, particularly low-VAF mutations, may help assess early field treatment response and help design treatments to reduce skin cancer risk more effectively. Gyorgy Paragh¹, Mitsuko Murakami¹, Megan Fitzgerald¹, Li Yan¹, Jonathan Li¹, Prashant Singh¹, Barbara Foster¹, Wendy J. Huss¹, Lei Wei¹ 1. Roswell Park Comprehensive Cancer Center, Buffalo, NY, United States. UV Biology/Injury and Non-melanoma Cancers