Oops, you're using an old version of your browser so some of the features on this page may not be displaying properly.

MINIMAL Requirements: Google Chrome 24+Mozilla Firefox 20+Internet Explorer 11Opera 15–18Apple Safari 7SeaMonkey 2.15-2.23

Poster Display session 3

2436 - Development and Validation of an RNA-Seq Assay for Gene Fusions Detection in Formalin-Fixed Paraffin-Embedded Samples

Date

30 Sep 2019

Session

Poster Display session 3

Topics

Translational Research

Tumour Site

Presenters

Hua Dong

Citation

Annals of Oncology (2019) 30 (suppl_5): v574-v584. 10.1093/annonc/mdz257

Authors

H. Peng1, C. Wang2, Y. Guo3, B. Li3, C. Chen2, L. Xiong2, H. Dong3, F. Li3, L. Tian4, Q. Xu2

Author affiliations

  • 1 -, The First People's Hospital of Yunnan Province, 00 - Yunnan/CN
  • 2 The R&d Center, 3DMed Inc, 200120 - Shanghai/CN
  • 3 Bioinformatics, 3D medicines Inc., 201114 - Shanghai/CN
  • 4 Department Of Thoracic Surgery, The First Affiliated Hospital of Chongqing Medical University, Chongqing/CN

Resources

Login to access the resources on OncologyPRO.

If you do not have an ESMO account, please create one for free.

Abstract 2436

Background

RNA sequencing (RNA-Seq) assay has been widely used for transcript level gene quantification and fusion detection for research use in fresh frozen samples. Clinical use of RNA is complicated by the common use of formalin-fixed paraffin-embedded (FFPE) tissue storage, which can cause low yield and RNA degradation. We evaluate the feasibility, quality, and analytical performance of RNA-Seq on clinical FFPE tumor samples for gene fusion detection.

Methods

Total RNA was extracted from FFPE tumor samples and/or adjacent normal samples. Ribosomal RNA depletion, cDNA synthesis, and library preparation were used to prepare next-generation sequencing (NGS) libraries that were sequenced on Illumina HiSeq X instrument. Sequencing data were analyzed and annotated with an in-house developed pipeline. A set of experimental and data quality control parameters were set up.

Results

The assay identified all the positive fusions from RNA reference material with 15 NTRK fusions spiked-in and ALK, RET, NTRK1 and FGFR3 fusions from 5 positive cell lines. The assay Limit of Detection was tested by diluting RNA from ALK fusion positive cell H2228C to fusion-negative cell line. Gene fusions were generally detectable down to 10% dilutions for all fusion types and as little as 5% for some fusion types. RNA-Seq assay detected 10 of 12 gene fusions detected by DNA based NGS assay, for a sensitivity of 83%. No false-positive gene fusions were identified in 28 tumor specimens that were negative for fusions, for a specificity of 100%. The assay also identified 6 novel fusions in 3 tumor specimens, which had been confirmed by RT-PCR and Sanger sequencing. Good intra-assay and inter assay reproducibility was observed with complete concordance for the presence or absence of gene fusions in 3 samples and 6 replicates. We observed 81% success rate on whole transcriptome RNA-Seq process for more than 100 FFPE samples.

Conclusions

RNA-Seq assay can help identify gene fusions in patients with cancer, which is a good supplement for DNA based NGS assay, especially for novel fusion detection or fusion breakpoints which are hard to design probes for DNA samples. These patients may in turn benefit from approved and investigational fusion related targeted therapies.

Clinical trial identification

Editorial acknowledgement

Legal entity responsible for the study

3DMed Inc.

Funding

3DMed Inc.

Disclosure

H. Dong: Full / Part-time employment: 3DMed Inc. C. Wang: Full / Part-time employment: 3DMed Inc. Q. Xu: Full / Part-time employment: 3DMed Inc. Y. Guo: Full / Part-time employment: 3DMed Inc. Y. Chen: Full / Part-time employment: 3DMed Inc. B. Li: Full / Part-time employment: 3DMed Inc. S. Liu: Full / Part-time employment: 3DMed Inc. C. Chen: Full / Part-time employment: 3DMed Inc. L. Xiong: Full / Part-time employment: 3DMed Inc. F. Li: Full / Part-time employment: 3DMed Inc. All other authors have declared no conflicts of interest.

This site uses cookies. Some of these cookies are essential, while others help us improve your experience by providing insights into how the site is being used.

For more detailed information on the cookies we use, please check our Privacy Policy.

Customise settings
  • Necessary cookies enable core functionality. The website cannot function properly without these cookies, and you can only disable them by changing your browser preferences.