J Mol Diagn. 2017 Sep;19(5):682-696. doi: 10.1016/j.jmoldx.2017.05.006. Epub 2017 Aug 9.

Validation of a Targeted RNA Sequencing Assay for Kinase Fusion Detection in Solid Tumors.

Reeser JW¹, Martin D¹, Miya J¹, Kautto EA¹, Lyon E¹, Zhu E¹, Wing MR¹, Smith A¹, Reeder M¹, Samorodnitsky E¹, Parks H¹, Naik KR¹, Gozgit J², Nowacki N³, Davies KD⁴, Varella-Garcia M⁴, Yu L⁵, Freud AG³, Coleman J³, Aisner DL⁴, Roychowdhury S⁶.

Author information

1: Comprehensive Cancer Center, The Ohio State University, Columbus, Ohio.
2: Ariad Pharmaceuticals, Cambridge, Massachusetts.
3: Department of Pathology, The Ohio State University, Columbus, Ohio.
4: Department of Pathology, University of Colorado, Anschutz Medical Campus, Denver, Colorado.
5: Department of Biomedical Informatics, The Ohio State University, Columbus, Ohio.
6: Comprehensive Cancer Center, The Ohio State University, Columbus, Ohio; Department of Internal Medicine, Division of Medical Oncology, The Ohio State University, Columbus, Ohio. Electronic address: sameek.roychowdhury@osumc.edu.

Abstract

Kinase gene fusions are important drivers of oncogenic transformation and can be inhibited with targeted therapies. Clinical grade diagnostics using RNA sequencing to detect gene rearrangements in solid tumors are limited, and the few that are available require prior knowledge of fusion break points. To address this, we have analytically validated a targeted RNA sequencing assay (OSU-SpARKFuse) for fusion detection that interrogates complete transcripts from 93 kinase and transcription factor genes. From a total of 74 positive and 36 negative control samples, OSU-SpARKFuse had 93.3% sensitivity and 100% specificity for fusion detection. Assessment of repeatability and reproducibility revealed 96.3% and 94.4% concordance between intrarun and interrun technical replicates, respectively. Application of this assay on prospective patient samples uncovered OLFM4 as a novel RET fusion partner in a small-bowel cancer and led to the discovery of a KLK2-FGFR2 fusion in a patient with prostate cancer who subsequently underwent treatment with a pan-fibroblast growth factor receptor inhibitor. Beyond fusion detection, OSU-SpARKFuse has built-in capabilities for discovery research, including gene expression analysis, detection of single-nucleotide variants, and identification of alternative splicing events.