Background and Objectives
The use of ultra-sensitive diagnostic tests to detect clinically actionable somatic alterations within the gene encoding the epidermal growth factor receptor (
EGFR
) within circulating cell-free DNA is an important first step in determining the eligibility of patients with non-small cell lung cancer to receive tyrosine kinase inhibitors.
Methods
We present the clinical validation (accuracy, sensitivity, and specificity) of a highly sensitive OncoBEAM
TM
EGFR
V2 test, which we compare to a custom next-generation sequencing assay, for the treatment of patients with non-small cell lung cancer with
EGFR
tyrosine kinase inhibitor therapies. The OncoBEAM
TM
digital-polymerase chain reaction method detects 36 different
EGFR
alterations in circulating cell-free DNA, whereas the next-generation sequencing assay covers major solid tumor oncodrivers. Of the 540 samples analyzed with the OncoBEAM
TM
EGFR
V2 test, 42.4% of patients had undergone molecular testing at diagnosis (
N
= 229/540) and 57.7% of patients during disease progression (
N
= 311/540).
Results
The sensitivity and specificity were measured for this BEAMing assay. The number of mutant beads and mutant allelic fraction were measured for each
EGFR
alteration and the level of detection was established at 0.1% for a median of 2861 genome equivalent (GE) in each reaction using HD780 horizon control DNA, as well as by an internal quality reference standard. Approximately 10%, 27%, and 63% of the 540 samples contained < 1500 GE, a range of 1500–3000 GE, and > 3000 GE, which corresponded to a maximal assay sensitivity of 2.0%, 0.5–0.1%, and 0.1–0.05% mutant allelic fraction, respectively. In a routine hospital setting, 11.4% of non-small cell lung cancer tumors were positive at diagnosis for
EGFR
alterations, while 43.7% samples harbored
EGFR
mutations at progression, among which 40.3% expressed
EGFR
resistance mutations after first-line tyrosine kinase inhibitor treatment with first- and second-generation drugs.
Conclusions
The OncoBEAM
TM
EGFR
V2 is a sensitive, robust, and accurate assay that delivers reproducible results. Next-generation sequencing and BEAMing technologies act complementarily in the routine molecular screening. We show that using a next-generation sequencing assay, despite its lower sensitivity, enables the identification of rare
EGFR
alterations or resistance mechanisms (mutation, deletion, insertion, and copy number variation) to orient first- and second-line treatments.
Supplementary ...