130 Novel method for non-invasive assessment of molecular markers of UV exposure in normal skin

Abstract

The use of markers of UV-induced DNA damage as a dosimeter of exposure and predictor of skin cancer risk has been proposed by multiple groups. While UV-induced mutations and adducts are well described and present in normal appearing UV-exposed epidermis, sampling normal non-lesional skin requires non-invasive methods to extract epidermal DNA for analysis. Here we demonstrate the feasibility of such an approach. Sampling was performed using a felt-tip pen soaked in a mixture of surfactants (Brij-30/DPS). In mice, we show that the epidermis can be selectively removed without scarring, with complete healing within 2 weeks. We exposed Hairless mice to low-dose solar simulated light (Newport) over a period of 3 months for a total exposure of 175 kJ/m2, a dose which normally produces tumors within 3 months. Over 2 micrograms / cm2 of DNA could be purified and exome sequenced. We noted a progressive increase in a UV-signature mutational burden over the following 2 months following the cessation of UV exposure, consistent with accumulation of mutant clones. Next we sought to assess whether this was practical to perform in human patients. Skin samples were collected from neck or shoulder (sun-exposed areas with dermatoheliosis) and lateral chest or back (minimally sun-exposed areas) with no prior preparation. Approximately 300 ng of total DNA could be isolated from an approximately 1 cm2 area of sampled skin. These samples were subjected to targeted exome sequencing using a commercially available panel (QIAseq Cancer Panel) to a depth of approximately 3500X and referenced to saliva. We identified considerable heterogeneity in multiple samples obtained from the same patient, suggesting widespread mosaicism, but also an enrichment for UV-signature mutations in key tumor suppressors including TP53 and NOTCH1/2. Our next steps are to use hypermutated areas of the genome to further refine this dosimetry method and assess its dynamic range across a larger spectrum of patients.