Identification of germline cancer predisposition variants during clinical ctDNA testing.

Nawal Kassem,Santosh Philips,Bryan P Schneider,Milan Radovich,Cynthia Hunter,Leigh Anne Anne Stout

doi:10.1200/jco.2020.38.15_suppl.e15555

Abstract

e15555 Background: Next-generation sequencing (NGS) of circulating tumor DNA (ctDNA) is a non-invasive method to guide therapy selection for cancer patients. Identification of inherited germline cancer predisposition mutations that have significant implications for at-risk relatives may be missed during routine ctDNA testing. Allele frequency has the potential to enhance the likelihood that a mutation is germline; and is often reported in many NGS tests from ctDNA. Here, we report on the fidelity of allele frequency in ctDNA as a predictor for pathogenic germline variant carriage. Methods: ctDNA sequencing of patients with metastatic cancer from the Indiana University Health Precision Genomics Program was performed using the FoundationOne Liquid assay. All variants detected by the ctDNA assay report were considered. All patients also had germline testing information and pathogenicity of germline variants were determined using ClinVar. Germline variants with conflicting interpretations were manually reviewed to determine pathogenicity. Comparisons between ctDNA results with known germline status were performed. Results: Of 91 previously identified germline cancer predisposition variants, 36 (40%) were also identified by ctDNA analysis. All germline variants that were tested for in the ctDNA assay (n = 36, 100%) were identified. When detected, the allele frequencies of detected germline variants in the ctDNA ranged from 39-87.6% with an average of 52.1%. Conversely, 111 of 160 (69%) variants identified by ctDNA analysis with allele frequency between 40-60% in a cancer predisposition gene were found to be germline in origin (regardless of pathogenicity). Variants in the BRCA2, BRCA1, and CDH1 genes were most likely to be germline in origin (26/27 [96%], 20/22 [91%], 13/15 [87%], respectively). Variants in the TP53 and APC genes were least likely to be germline in origin (9/36 [25%] and 1/6 [17%], respectively). There was an 85% (95/111) concordance in actionability between the somatic testing lab and ClinVar germline classifications. Of the 16 discordant variants, 100% were determined to be actionable by the somatic testing lab but not actionable in ClinVar. Conclusions: ctDNA allele frequency can alter the likelihood that a variant is germline. Importantly, however, this testing is far from comprehensive and should not be used as a replacement for germline testing. Variants with allele frequency between 40-60% in cancer predisposition genes should induce a high level of suspicion for germline status.

Highlights

Tumor generation sequencing (NGS) has become commonplace in routine clinical practice for patients with a variety of malignancies to help identify potential drug targets
We report the concordance between the variant allele frequency (VAF) from circulating tumor DNA (ctDNA) analysis and germline carrier status utilizing two patient cohorts sequenced at the Indiana University Health Precision Genomics program
Of patients seen at the IU Health Precision Genomics Program, a total of 156 pathogenic germline variants were identified (CONSORT diagram 1). ctDNA results using FoundationOne Liquid were available for 86 patients (91 variants)

Summary

Introduction

Tumor generation sequencing (NGS) has become commonplace in routine clinical practice for patients with a variety of malignancies to help identify potential drug targets. While traditionally established risk factors (e.g. age of diagnosis, family history, and tumor characteristics) have prompted the consideration for germline testing [9], more recently, expert consensus guidelines have considered specific findings on tumor NGS. The current NCCN guidelines recommend referral for appropriate germline genetic testing any time a variant is identified through tumor NGS that would have clinical implications if the variant was determined to be pathogenic and germline in origin [10]. Tumor NGS is designed to identify drug targets which has implications for both the variant coverage and the definition of pathogenicity. Tumor NGS tests are designed to uncover variants that would have therapeutic implications and these variants are not in complete concordance to those that would confer risk of disease [11]. It is important to note that some pathogenic germline variants are missed by tumor panels due to the variant type including structural rearrangements and/or a variant being present in a pseudogene region [12, 13]

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Results

Conclusion