Figure 2. Challenges in optimizing a ctDNA panel based on the diversity of HNSCC molecular alterations.

A) Top ten most recurrent molecular alterations in primary untreated disease as define by the HNSCC TCGA sequencing set (N = 530). Several of these genes are tumor suppressors from long, multi-exon genes. B) Top ten most recurrent copy number alterations from HNSCC TCGA data and the total genes in each region are indicated. C) Representation of regions that are required to be covered if ctDNA analysis focuses on tumor suppressors using FBXW7 schematic as an example (all exons would need coverage based on the low recurrence of hot spot alterations). Boxes represent exons, with blue boxes representing untranslated regions and black regions indicating coding sequences. Vertical lines indicate site of called non-synonymous single nucleotide variant and only the recurrent mutations were annotated. D) As in (C), with representative regions of interest in the HRAS oncogene highlighting recurrent mutation sites presented in schematic as shown suggesting the majority of HRAS mutant patients would be detected with probes covering G12, G13 and Q61.