Abstract Body: Skin chronically exposed to sunlight contains numerous p53 mutant patches which contain TP53
mutations and which are considered early potential precursors of skin cancer. Initially, in this study, whole exome
sequencing (WES) of AKs (n=5) was conducted and showed that the dysplastic cells within AKs have a median
mutation burden of 34.5 mutations/Mb, which is greater than that seen in many highly fatal malignancies. Comparison
of these results with the COSMIC database and WES results from cSCCs led to selection of 18 cancer-related genes
for next generation sequencing in p53 mutant patches (n=23). Targeted sequencing of these genes was also
conducted on actinic keratoses (AK, n=32), Bowen’s disease (BD, n=37) and squamous cell cancers (cSCC) which
had arisen from these precancers (n=15). 13 of the 18 genes were mutated within the p53 mutant patches, with these
genes also mutated in the AKs and BDs, as well as in cSCCs. The presence of clonal non-silent and/or synonymous
mutations confirmed the clonal nature of the p53 mutant patches, however, all TP53 mutations and many other
genetic mutations were subclonal in these lesions. Tracking shared mutations from the same skin site in one subject
suggested evolution along a common pathway from a shared precursor cell pool before the development of four
separate p53 mutant patches. In cases where histological evidence suggested that cSCCs had arisen from an
adjacent AK and/or BD, multiple shared mutations in the cancer and precancerous lesion confirmed the interpretation
of previous clinical observations that human cSCCs can arise from AKs and BDs. Our results demonstrate that p53
mutant patches in sun-exposed skin contain multiple mutations in cancer-related genes, which are also mutated in
later precancerous lesions and cSCCs. Furthermore, our data provides evidence for clonal evolution of mutations
within p53 mutant patches in normal skin in a similar manner to that encountered in established cancer.

Multiple cancer-related genes are mutated in p53 mutant patches in human skin.