Improved Gene Fusion Detection in Childhood Cancer Diagnostics Using RNA Sequencing.

Jayne Y Hehir-Kwa,Eric Strengman,Marco J Koudijs,Arjan Buijs,Ellen Van De Geer,Jasper Van Der Lugt,Mariëtte E G Kranendonk,Philip Lijnzaad,Frank C P Holstege,Marc Van Tuil,Valerie De Haas,Bastiaan B J Tops,Patrick Kemmeren,Wendy De Leng,Eugene T P Verwiel,Lennart A Kester,Laura S Hiemcke-Jiwa

doi:10.1200/po.20.00504

Jayne Y Hehir-Kwa, Eric Strengman + Show 15 more

Open Access

https://doi.org/10.1200/po.20.00504

Copy DOI

Abstract

PURPOSEGene fusions play a significant role in cancer etiology, making their detection crucial for accurate diagnosis, prognosis, and determining therapeutic targets. Current diagnostic methods largely focus on either targeted or low-resolution genome-wide techniques, which may be unable to capture rare events or both fusion partners. We investigate if RNA sequencing can overcome current limitations with traditional diagnostic techniques to identify gene fusion events.METHODSWe first performed RNA sequencing on a validation cohort of 24 samples with a known gene fusion event, after which a prospective pan-pediatric cancer cohort (n = 244) was tested by RNA sequencing in parallel to existing diagnostic procedures. This cohort included hematologic malignancies, tumors of the CNS, solid tumors, and suspected neoplastic samples. All samples were processed in the routine diagnostic workflow and analyzed for gene fusions using standard-of-care methods and RNA sequencing.RESULTSWe identified a clinically relevant gene fusion in 83 of 244 cases in the prospective cohort. Sixty fusions were detected by both routine diagnostic techniques and RNA sequencing, and one fusion was detected only in routine diagnostics, but an additional 24 fusions were detected solely by RNA sequencing. RNA sequencing, therefore, increased the diagnostic yield by 38%-39%. In addition, RNA sequencing identified both gene partners involved in the gene fusion, in contrast to most routine techniques. For two patients, the newly identified fusion by RNA sequencing resulted in treatment with targeted agents.CONCLUSIONWe show that RNA sequencing is sufficiently robust for gene fusion detection in routine diagnostics of childhood cancers and can make a difference in treatment decisions.

Highlights

Chromosomal rearrangements in the genomes of tumor cells can lead to the formation of chimeric transcripts or gene fusions
We first performed RNA sequencing on a validation cohort of 24 samples with a known gene fusion event, after which a prospective pan-pediatric cancer cohort (n = 244) was tested by RNA sequencing in parallel to existing diagnostic procedures
Sixty fusions were detected by both routine diagnostic techniques and RNA sequencing, and one fusion was detected only in routine diagnostics, but an additional 24 fusions were detected solely by RNA sequencing

Summary

Introduction

Chromosomal rearrangements in the genomes of tumor cells can lead to the formation of chimeric transcripts or gene fusions. Gene fusion detection in a diagnostic setting poses several challenges: (1) specific gene fusions may be rare,[7-9] (2) breakpoints may be atypical,[10-13] and (3) fusion partners may be promiscuous.[14]. Both reverse transcriptase polymerase chain reaction (RT-PCR)– based assays and fluorescence in situ hybridization (FISH) assays using break-apart probes are a sensitive, but time-consuming method to detect chromosomal rearrangements. For most cancer samples, multiple methods are performed in parallel to detect clinically relevant gene fusions. Most of these challenges and technical limitations are mitigated by targeted RNA

Methods

Results

Discussion

Conclusion