9 Liquid biopsy in small cell lung cancer (SCLC)

Introduction Analysis of circulating cell-free DNA (cfDNA) and circulating tumour cells (CTCs) obtained from the peripheral blood of patients with cancer is a powerful and minimally invasive approach that allows the reconstruction of tumour genomes, with potential as a companion diagnostic for both patient stratification and monitoring. Here, we use cfDNA next-generation sequencing (NGS) analysis to assess and compare circulating tumour DNA (ctDNA) profiles in patients with SCLC. Material and methods NGS libraries were prepared from cfDNA isolated from the pre-treatment peripheral blood samples from 69 patients and 32 cancer-free controls. Libraries were subjected to wholegenome sequencing (WGS) to establish genome wide copy number aberrations (CNA) as well as targeted mutation analysis of 110 genes frequently altered in SCLC. In addition CellSearch, an epitope dependent enrichment platform was used to enumerate CTCs from a parallel blood sample. Results and discussions CNA patterns revealed SCLC associated changes including losses on chromosomes 3 p, 5q and 17 p and gains on chromosome 3q and 5 p as well as amplification of MYC in 21/69 (30%) and SOX2 in 36/69 (52%) as well losses on FHIT in 40/69 (58%), RASSFI in 38/69 (55%) and RB1 in 24/69 (35%) patients. CNA metrics enabled detection of tumour associated changes in 58/64 (84%) patients, Targeted NGS detected tumour associated mutations in 59/63 (94%) with TP53 mutations detected in 50 patients (85% of patients with any detectable ctDNA). CNA and somatic mutations were detected in both limited stage (LS-SCLC, confined to 1 hemithorax) and extensive stage (ES-SCLC, with distant metastases) with statistically significant differences seen for CNA metrics and variant allele frequencies of mutations consistent with higher levels of ctDNA in ES-SCLC. However, no significant differences between ES and LS were observed in the mutation patterns with respect to DNA damage repair, RAS and PI3K and transcriptional regulation pathways between the two stages of SCLC. There was considerable overlap between the detection of ctDNA and CTC counts, with cfDNA readouts detecting tumour related changes in 93% and CTCs detected only in 78% of patients. Conclusion We have established sensitive methods for detecting ctDNA in SCLC and combined with CTC enumeration we have an effective liquid biopsy applicable to 98% of patients in this cohort. Future work will involve utilising this optimised NGS approach in an independent cohort of patients to correlate cfDNA metrics with clinical outcome.