Normal Skin Accrues High Number of Mutations Associated with Cancer
News May 22, 2015
They examined 234 skin biopsies from four cancer-free individuals aged 55 to 73 who were undergoing routine surgery to remove excess eyelid skin that was obscuring vision. Next, for each biopsy, they ran a targeted next-generation sequencing panel of 74 genes, including genes known to play a role in skin cancers, genes involved in a wide range of cancers, and genes whose function are unknown but are frequently mutated in skin samples that were taken from the COSMIC database.
"With this technology, we can now peer into the first steps a cell takes to become cancerous," Peter Campbell, a corresponding author from the Wellcome Trust Sanger Institute, said in a statement.
The researchers chose to look at eyelid skin because it has a high level of sun exposure. They used a panel targeting the coding regions of 74 cancer-associated genes and sequenced each of the 234 biopsies to 500x coverage on the Illumina HiSeq. For one sample they sequenced the whole genome to 147x coverage.
In total, they found 3,760 mutations, corresponding to about 140 per square centimeter of skin. Between 18 percent and 32 percent of skin cells carried at least one mutation in the targeted genes.
Interestingly, the mutational pattern "closely matched that expected for ultraviolet light exposure and that seen in skin cancers," the authors wrote. For instance, there were high rates of cytosine to thymine mutations and high rates of CC to TT changes, "consistent with the known chemistry of sunlight-induced damage to DNA."
Next, the researchers looked at clonality and positive selection, and found that six genes had an excess of protein-altering base substitutions than would be expected by chance, and five of those also had an excess of indels or di-nucleotide substitutions.
The NOTCH1 gene was the most frequently mutated gene and NOTCH2 and NOTCH3 also had an excess of mutations. In total, NOTCH mutations were found in around 25 percent of cells. The NOTCH receptors are key regulators and frequently mutated in epithelial cancers and lymphoid malignancies. "The density of positively selected driver mutations was surprisingly high" in the NOTCH genes, the authors wrote, estimated to be about 83 clones per square centimeter.
Aside from NOTCH mutations, there was also an excess of mutations to the tumor suppressor genes FAT1 and TP53, in about 3 percent to 5 percent of cells.
For one sample that contained a massive clone with a known driver mutation in TP53 and an activating mutation in FGFR3 that spanned six adjacent biopsies, the researchers performed whole-genome sequencing. They identified nearly 74,000 base substitutions and 2,248 indels with a mutational signature corresponding to UV light exposure.
Despite the similarities seen between the normal skin samples and cancerous samples, the authors caution that there are still huge differences, particularly in the frequency of driver mutations. For the four normal samples the researchers evaluated, there were approximately .27 driver mutations per cell. But genomic dat from actual skin cancer genomes has found about 2.7 driver mutations per cell.
"The burden of mutations observed is high but almost certainly none of these clones would have developed into skin cancer," Iñigo Martincorena, first author from the Sanger Institute, said in a statement.
Campbell added that the study showed evidence that clonal selection for cancer-associated mutations "give cells a boost" that "increases the pool of cells waiting for the next mutation to push them even further," but what is not known is how many of these events are needed before cells become fully cancerous.