Cited 14 times since 2020 (8.5 per year) source: Scopus Journal of thoracic oncology : official publication of the International Association for the Study of Lung Cancer, Volume 15, Issue 6, 31 5 2020, Pages 1000-1014 Optimizing Mutation and Fusion Detection in NSCLC by Sequential DNA and RNA Sequencing. Cohen D, Hondelink LM, Solleveld-Westerink N, Uljee SM, Ruano D, Cleton-Jansen AM, von der Thüsen JH, Ramai SRS, Postmus PE, Graadt van Roggen JF, Hoppe BPC, Clahsen PC, Maas KW, Ahsmann EJM, Ten Heuvel A, Smedts F, van Rossem RN, van Wezel T

Introduction

Frequently, patients with locally advanced or metastatic NSCLC are screened for mutations and fusions. In most laboratories, molecular workup includes a multitude of tests

immunohistochemistry (ALK, ROS1, and programmed death-ligand 1 testing), DNA sequencing, in situ hybridization for fusion, and amplification detection. With the fast-emerging new drugs targeting specific fusions and exon-skipping events, this procedure harbors a growing risk of tissue exhaustion.

Methods

In this study, we evaluated the benefit of anchored, multiplexed, polymerase chain reaction-based targeted RNA sequencing (RNA next-generation sequencing [NGS]) in the identification of gene fusions and exon-skipping events in patients, in which no pathogenic driver mutation was found by DNA-based targeted cancer hotspot NGS (DNA NGS). We analyzed a cohort of stage IV NSCLC cases from both in-house and referral hospitals, consisting 38.5% cytology samples and 61.5% microdissected histology samples, mostly core needle biopsies. We compared molecular findings in a parallel workup (DNA NGS and RNA NGS, cohort 1, n = 198) with a sequential workup (DNA NGS followed by RNA NGS in selected cases, cohort 2, n = 192). We hypothesized the sequential workup to be the more efficient procedure.

Results

In both cohorts, a maximum of one oncogenic driver mutation was found per case. This is in concordance with large, whole-genome databases and suggests that it is safe to omit RNA NGS when a clear oncogenic driver is identified in DNA NGS. In addition, this reduced the number of necessary RNA NGS to only 53% of all cases. The tumors of never smokers, however, were enriched for fusions and exon-skipping events (32% versus 4% in former and current smokers, p = 0.00), and therefore benefited more often from the shorter median turnaround time of the parallel approach (15 d versus only 9 d in the parallel workup).

Conclusions

We conclude that sequentially combining DNA NGS and RNA NGS is the most efficient strategy for mutation and fusion detection in smoking-associated NSCLC, whereas for never smokers we recommend a parallel approach. This approach was shown to be feasible on small tissue samples including for cytology tests, can drastically reduce the complexity and cost of molecular workup, and also provides flexibility in the constantly evolving landscape of actionable targets in NSCLC.

J Thorac Oncol. 2020 1;15(6):1000-1014