Combined multimodal ctDNA analysis and radiological imaging for tumor surveillance in Non-small cell lung cancer

Highlights

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  High concordance rate in variant calls from tumor biopsies and the corresponding liquid biopsy-derived DNA sample.

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  ctDNA sequencing allows for unbiased characterization of molecular tumor architecture and reveal the presence of otherwise undetectable resistance-mediating tumor cell clones.

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  Serial ctDNA analysis allows for monitoring tumor dynamics but limited by tumor cell clonal molecular heterogeneity.

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  cfDNA methylation analysis using methylation-sensitive restriction enzyme-coupled ddPCR can complement cfDNA mutation scoring to broaden the spectrum of eligible patients for molecular surveillance.

Abstract

Background

Radiology is the current standard for monitoring treatment responses in lung cancer. Limited sensitivity, exposure to ionizing radiations and related sequelae constitute some of its major limitation. Non-invasive and highly sensitive methods for early detection of treatment failures and resistance-associated disease progression would have additional clinical utility.

Methods

We analyzed serially collected plasma and paired tumor samples from lung cancer patients (61 with stage IV, 48 with stages I-III disease) and 61 healthy samples by means of next-generation sequencing, radiological imaging and droplet digital polymerase chain reaction (ddPCR) mutation and methylation assays.

Results

A 62% variant concordance between tumor-reported and circulating-free DNA (cfDNA) sequencing was observed between baseline liquid and tissue biopsies in stage IV patients. Interestingly, ctDNA sequencing allowed for the identification of resistance-mediating p.T790M mutations in baseline plasma samples for which no such mutation was observed in the corresponding tissue. Serial circulating tumor DNA (ctDNA) mutation analysis by means of ddPCR revealed a general decrease in ctDNA loads between baseline and first reassessment. Additionally, serial ctDNA analyses only recapitulated computed tomography (CT) -monitored tumor dynamics of some, but not all lesions within the same patient. To complement ctDNA variant analysis we devised a ctDNA methylation assay ( ^methcfDNA) based on methylation-sensitive restriction enzymes. cfDNA methylation showed and area under the curve (AUC) of > 0.90 in early and late stage cases. A decrease in ^methcfDNA between baseline and first reassessment was reflected by a decrease in CT-derive tumor surface area, irrespective of tumor mutational status.

Conclusion

Taken together, our data support the use of cfDNA sequencing for unbiased characterization of the molecular tumor architecture, highlights the impact of tumor architectural heterogeneity on ctDNA-based tumor surveillance and the added value of complementary approaches such as cfDNA methylation for early detection and monitoring