Identification of Incidental Germline Mutations in Patients With Advanced Solid Tumors Who Underwent Cell-Free Circulating Tumor DNA Sequencing

Abstract

Purpose: To determine the potential for detection of incidental germline cancer predisposition mutations through cell-free DNA (cfDNA) analyses in patients who underwent solid tumor somatic mutation evaluation.

Patients and methods: Data were evaluated from 10,888 unselected patients with advanced (stage III/IV) cancer who underwent Guardant360 testing between November 2015 and December 2016. The main outcome was prevalence of putative germline mutations identified among 16 actionable hereditary cancer predisposition genes.

Results: More than 50 cancer types were studied, including lung (41%), breast (19%), colorectal (8%), prostate (6%), pancreatic (3%), and ovarian (2%). Average patient age was 63.5 years (range, 18 to 95 years); 43% were male. One hundred and fifty-six individuals (1.4%) had suspected hereditary cancer mutations in 11 genes. Putative germline mutations were more frequent in individuals younger than 50 years versus those 50 years and older (3.0% v 1.2%, respectively; P < .001). Highest yields of putative germline findings were in patients with ovarian (8.13%), prostate (3.46%), pancreatic (3.34%), and breast (2.2%) cancer. Putative germline mutation identification was consistent among 12 individuals with multiple samples. Patients with circulating tumor DNA copy number variation and/or reversion mutations suggestive of functional loss of the wild-type allele in the tumor DNA also are described.

Conclusion: Detection of putative germline mutations from cfDNA is feasible across multiple genes and cancer types without prior mutation knowledge. Many mutations were found in cancers without clear guidelines for hereditary cancer genetic counseling/testing. Given the clinical significance of identifying hereditary cancer predisposition for patients and their families as well as targetable germline alterations such as in BRCA1 or BRCA2, research on the best way to validate and return potential germline results from cfDNA analysis to clinicians and patients is needed.

Fig 1.

Somatic mutation and putative pathogenic germline mutant allele fractions (MAFs) for all 156 individuals with putative germline mutations arranged by lowest to highest somatic MAF. This included 169 samples in total; 144 individuals had one sample evaluated, and 12 had more than one sample evaluated (25 samples total; Appendix Tables A1 and A2). Putative germline mutations (pathogenic germline) had MAFs close to 50%, whereas somatic mutations had lower MAFs, as expected. Copy number variation corrections account for MAFs greater than 60%. Some samples did not have somatic mutations.

Fig 2.

Distribution of mutant allele fractions (MAFs) of putative germline mutations. The majority of mutations had MAFs close to 50%, as expected. Mutations with MAFs less than 40% were not evaluated; those with MAFs greater than 60% were included if concomitant locus copy number loss was identified (see Figs 3 and 4).

Fig 3.

Relationship between mutant allele fractions (MAFs) of identified putative germline mutations and common single nucleotide polymorphisms. The putative germline MAFs are plotted against, and in agreement with, genomic copy number (CN) measured using corresponding single nucleotide polymorphisms at the same allele. Mutations under 40% MAF were not evaluated.

Fig 4.

Three example genetic profiles of patients with secondary tumor mutations. The putative germline mutations identified are presented with any respective BRCA1 or BRCA2 circulating tumor findings. (A) Patient with a BRCA2 gene deletion and concomitant putative germline single nucleotide variant (SNV). (B) Patient with both germline and somatic BRCA2 SNVs. (C) Patient with a reversion of the germline SNV in the circulating tumor DNA with an allele frequency of 2.15%, potentially explaining the patient’s reported resistance to platinum-based chemotherapy (the wild-type allele presumed lost because of a BRCA1 gene deletion). chr, chromosome; ID, identifier; MAF, mutant allele fractions.