Abstract 4602: Exome sequencing of UKLS lung cancer CT screened early stage cancers
Authors: Hyde Russell , Davies Michael , Ledson Martin , Holemans John A. , Page Richard D. , Gosney John , Baldwin David R. , Devaraj Anand , Hansell David M. , Duffy Stephen W. , Field John K. 
Published: August 2015

Abstract
INTRODUCTION
Developments in both low-dose CT-screening within populations at high risk of lung cancer and sequencing analysis of lung tumours have improved the prospects for early detection and clinical management of lung cancer. Such approaches potentially could provide biological insight into the early stages of lung cancer development, which until now have been refractory to genomic analysis, since lung tumours are rarely identified at a presymptomatic timepoint.
BRIEF DESCRIPTION OF PERTINENT EXPERIMENTAL PROCEDURES
Individuals with at least a 5% risk of developing lung cancer within 5 years (according to the LLPv2 lung cancer risk score) were recruited to the UK Lung low dose CT Screening (UKLS) trial. Ten of the early stage NSCLC UKLS tumours were prepared for exome sequencing. Tumour DNA was extracted from FFPE material and DNA was also extracted from matched blood samples. Tumour and blood libraries were prepared using an Agilent SureSelect exome-capture kit and sequenced to 100x and 50x, depth respectively, on the Illumina HiSeq platform (Oxford Gene Technology, UK). Sequence data were aligned against the GRCh37 human reference and matched samples were subject to local realignment in pairs. Somatic variants were identified using a combination of EBCall, MuTect and VarScan2 and were post-filtered to remove false positive variants (alignment artifacts and FFPE-hypersensitive sites) by comparison between samples.
TCGA (The Cancer Genome Atlas) lung adenocarcinoma and squamous cell carcinoma samples from smokers and former smokers (who had complete data for a minimal set of clinical and epidemiological variables) were compared to the UKLS samples described above. To ensure comparable genomic regions were studied, both the UKLS and TCGA somatic variants were restricted to protein-coding regions covered by the Agilent SureSelect array but disjoint from simple-repetitive regions.
SUMMARY OF THE NEW, UNPUBLISHED DATA
Despite detection at an early stage, somatic mutations were identified in all CT-detected tumours, including non-synonymous variants in known driver genes. Between 71 and 471 variants were detected within non-repetitive protein-coding regions, which is comparable to, if somewhat lower than, the number of variants in the same regions in the TCGA ever-smoker samples (p = 0.17; Kruskal-Wallis). The results suggest that the majority of somatic variants exist in lung tumours prior to the evolution of symptoms and are consistent with recent studies of lung cancer heterogeneity wherein the majority of mutations occur prior to subclonal branching.
STATEMENT OF CONCLUSIONS
Somatic variations in the host genome are readily detected in FFPE material from pre-symptomatic lung tumours. Mutational burden is comparable to lung adenocarcinoma and squamous carcinoma from mature symptomatic individuals providing further evidence for the long latency of lung tumour development.
Citation Format: Russell Hyde, Michael Davies, Martin Ledson, John A. Holemans, Richard D. Page, John Gosney, David R. Baldwin, Anand Devaraj, David M. Hansell, Stephen W. Duffy, John K. Field. Exome sequencing of UKLS lung cancer CT screened early stage cancers. [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 4602. doi:10.1158/1538-7445.AM2015-4602