Utility of Serial cfDNA NGS for Prospective Genomic Analysis of Patients on a Phase I Basket Study

Abstract

Purpose: Cell-free DNA (cfDNA) analysis offers a noninvasive means to access the tumor genome. Despite limited sensitivity of broad-panel sequencing for detecting low-frequency mutations in cfDNA, it may enable more comprehensive genomic characterization in patients with sufficiently high disease burden. We investigated the utility of large-panel cfDNA sequencing in patients enrolled to a Phase I AKT1-mutant solid tumor basket study.

Methods: Patients had AKT1 E17K-mutant solid tumors and were treated on the multicenter basket study (ClinicalTrials.gov identifier: NCT01226316) of capivasertib, an AKT inhibitor. Serial plasma samples were prospectively collected and sequenced using exon-capture next-generation sequencing (NGS) analysis of 410 genes (Memorial Sloan Kettering [MSK]-Integrated Molecular Profiling of Actionable Cancer Target [IMPACT]) and allele-specific droplet digital polymerase chain reaction (ddPCR) for AKT1 E17K. Tumor DNA (tDNA) NGS (MSK-IMPACT) was also performed on available pretreatment tissue biopsy specimens.

Results: Among 25 patients, pretreatment plasma samples were sequenced to an average coverage of 504×. Somatic mutations were called in 20/25 (80%), with mutant allele fractions highly concordant with ddPCR of AKT1 E17K (r 2 = 0.976). Among 17 of 20 cfDNA-positive patients with available tDNA for comparison, mutational concordance was acceptable, with 82% of recurrent mutations shared between tissue and plasma. cfDNA NGS captured additional tumor heterogeneity, identifying mutations not observed in tDNA in 38% of patients, and revealed oncogenic mutations in patients without available baseline tDNA. Longitudinal cfDNA NGS (n = 98 samples) revealed distinct patterns of clonal dynamics in response to therapy.

Conclusion: Large gene panel cfDNA NGS is feasible for patients with high disease burden and is concordant with single-analyte approaches, providing a robust alternative to ddPCR with greater breadth. cfDNA NGS can identify heterogeneity and potentially biologically informative and clinically relevant alterations.

© 2021 by American Society of Clinical Oncology.

Figures

FIG 1.

AKT1-mutant allele fraction (MAF) concordance in cell-free DNA between Droplet Digital polymerase chain reaction (ddPCR) and next-generation sequencing (NGS).

FIG 2.

Patterns of clonal dynamics in longitudinal cell-free DNA samples. (A) Concordant: mutant allele fraction (MAF) of each variant rises together in this patient with ovarian cancer. (B) Discordant: as KRAS MAF rises, MAFs of the other variants are declining in this patient with colorectal cancer.

FIG A1.

Consort diagram.

FIG A2.

Time between tumor tissue and cfDNA acquisition. Time between pretreatment tumor tissue biopsies and cfDNA collection did not correlate with discordance between mutation calls in tumor tissue versus cfDNA. cfDNA, cell-free DNA.