The standard of care in our hospital was conventional EGFR, ALK, ROS1 sequential testing and to treat with chemotherapy if no actionable genetic mutations were detected. With the evolving precision medicine based new targeted treatment for lung cancer patients, we planned a study to develop a new NGS-based assay to identify clinically relevant 12 genes most frequently mutated in non-small cell lung cancer to help patients to benefit from precision genomics based new targeted treatment approaches and to avoid chemotherapy use.
The study population consisted of newly diagnosed or previously treated advanced non-small cell lung cancer patients. After analysis of the Formalin-Fixed Paraffin-Embedded (FFPE) blocks for tumor content, DNA and mRNA from FFPE blocks were extracted and subjected to 12 gene biomarker testing by Next Generation Sequencing (NGS) using the Ion S5 System. The test utilizes AmpliSeq technology based NGS assay. High quality nucleic acids that passed quality control checks were subjected to library preparation and analysed for relevant genomic alterations in both the DNA and RNA to simultaneously detect multiple variants, including hotspots, single-nucleotide variants, indels, copy number variants, and gene fusion as mentioned below Hotspot genes (SNVs and short indels): ALK, BRAF, EGFR, ERBB2, KRAS, MAP2K1, MET, NRAS, PIK3CA, RET and ROS1 Gene fusions: ALK, MET exon14 skipping mutation, NTRK (1,2,3), RET and ROS1 Sequencing was performed to achieve a minimum 500x depth of coverage. High quality sequencing data was then analysed using the optimized ION Torrent Suite and the ION Reporter software to accurately detect rare somatic variants. The hotspot, indels and fusions were analysed with the help of ION reporter software and variants were annotated according to ACMG and AMP guidelines.
In our study conducted over a period of 6 months, a total of 50 patients underwent 12 gene biomarker testing by NGS. The mean age of our study population was 63.3 ± 11.75 years, comprising of 60% males (n=30) and 40% females (n=20). Histopathology reported was adenocarcinoma in 98% (n=49) and squamous cell carcinoma in 2% (n=1) of patients. The mean turnover time between the analysis of FFPE blocks for tumor content and NGS testing results was around 14 ± 2 days.
Genetic mutation % of patients (n) EGFR exon 19-del and exon 21-L858R 34% (17) ALK rearrangements 6% (3) ROS1 fusion 2% (1) MET ex-14 skipping 8% (4) KRAS G12C 10% (5) ERBB2 exon 20 insertion 6% (3) BRAF V600E 6% (3) No mutations 22% (11) Co-mutations: PIK3CA gene and other genes (MET exon 14 skipping, KRAS G12C, EGFR exon 19 deletion with MYC amplification) PD-L1 Tumor proportion score % of patients (n) Precision genomics-based targeted treatment % of patients (n) Negative (<1%) 68% (34) Yes 46% (23) Low (1-49%) 24% (12) No 54% (27) High (≥50%) 08% (4) Unaffordable drug price 26% (7) No access to drugs or clinical trials 11 % (3) No Indications for targeted therapy 22% (6) No mutations 41% (11)
Our study of precision genomics based 12 gene biomarker testing by NGS as an alternative to conventional EGFR, ALK, ROS-1 testing in advanced non-small cell lung cancer patients is a cost-effective & time saving approach for targeted treatment in a tertiary care hospital.
Dr Shekar Patil, Health Care Global Enterprises Ltd. Cancer Hospital Bengaluru.
Has not received any funding.
All authors have declared no conflicts of interest.