Abstract 5230: Circulating tumor DNA as a ‘liquid biopsy’ in head and neck cancer
Authors: Perdomo Sandra , Anantharaman Devasena , Mckay James , Brennan Paul 
Published: August 2015

Abstract
In cancer patients, a larger fraction of cell-free DNA is of tumor origin, released in circulating blood as a consequence of cell death and tissue trauma, ctDNA carries information on tumor-related genetic and epi¬genetic changes. Given the utility as a noninvasive marker ctDNA is commonly referred to as “liquid biopsy”. Some studies have correlated the abundance of ctDNA with tumor size and stage, with as much as 10% of ctDNA in advanced-stage cancers but as low as 0.1-1% in premalignant or early stage disease. Only one published study has explored the detection of ctDNA in Head and neck tumors in advanced cancers detecting mutations in 60% of the cases.
Our primary goal was to provide a comprehensive evaluation of the presence of ctDNA in circulating blood fractions from a series of HNSCC cases at early and late stages. We also evaluated the extent to which measures of ctDNA in blood samples correlate with similar findings from paired tumor tissue and correlate with outcome. 48 cases (including plasma, tumor and germline samples) diagnosed as head and neck SCC were selected from a multi center case- control study (ARCAGE), conducted between 2002-2005 involving 10 countries across Europe. Stage distribution was as following: 40% stage IV, 23% stage II, 19% stage III, 15% stage I, 3% no Info. Additionally, 65 controls matched by center, age, sex, smoking and alcohol status were included to assess a threshold of the background noise after sequencing.
Targeted sequencing was performed independently on both germline DNA and the tumor material using the Ion Proton™ System at an average depth of 250X. Sequencing included the entire coding region of 14 of the most frequently mutated genes in HNSCC: TP53, NOTCH1, CDKN2A, CASP8, PIK3CA, HRAS, FAT1, MLL2, NSD1, FBXW7, RB1, TP63, IRF6 and PTEN. Plasma samples were sequenced using an amplicon based approach. All 65 controls were also sequenced for all mutations detected. Plasma samples were sequenced at an average depth of 10000X. Mutation calls were based on a poisson distribution to distinguish true variants from background noise. Indels were manually inspected using IGV. Statistical tests were carried out in R statistical programming environment. 80% of tumor samples sequenced harboured a somatic mutation in at least one of the 14 genes, for a total 54 somatic mutations. 50% of these mutations including 2 indels were also detected in the corresponding plasma samples, allelic frequencies ranged from 0.005 to 0.30; cases were mostly stage IV although mutations were also detected in four early stage cases. Overall survival was lower for cases harboring TP53 mutations in both tumor and plasma.
It is feasible to detect frequent mutations in circulating free DNA in both early and late stage HNSCC. Mutations detected in both tumor and plasma correlated with poor survival especially in oropharyngeal cases demonstrating that ctDNA might be included as a potential biomarker for patient monitoring and outcome prediction.
Citation Format: Sandra Perdomo, Devasena Anantharaman, James Mckay, Paul Brennan. Circulating tumor DNA as a ‘liquid biopsy’ in head and neck cancer. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 5230. doi:10.1158/1538-7445.AM2015-5230