MYB-NFIB gene fusions identified in archival adenoid cystic carcinoma tissue employing NanoString analysis: an exploratory study

John B Mcintyre,Roderick H W Simpson,Jenny J Ko,Desiree Hao,Angela M Y Chan,Jodi Siever,Harold Y Lau

doi:10.1186/s13000-019-0855-8

Abstract

BackgroundAdenoid cystic carcinoma (ACC) is a slow growing salivary gland malignancy that is molecularly characterized by t(6:9)(q22–23;p23–24) translocations which predominantly result in MYB-NFIB gene fusions in nearly half of tumours. Detection of MYB-NFIB transcripts is typically performed with fresh ACC tissue using conventional RT-PCR fragment analysis or FISH techniques, which are prone to failure when only archival formalin fixed paraffin embedded (FFPE) tissue is available. The purpose of this pilot study was to evaluate the utility of NanoString probe technology for the detection of MYB-NFIB transcripts in archival ACC tissue.MethodsA NanoString probeset panel was designed targeting the junctions of three currently annotated MYB-NFIB fusion genes as well as 5′/3′ MYB probesets designed to detect MYB gene expression imbalance. RNA isolated from twenty-five archival ACC specimens was profiled and analyzed. RT-qPCR and sequencing were performed to confirm NanoString results. MYB protein expression was analyzed by immunohistochemistry.ResultsOf the 25 samples analyzed, 11/25 (44%) expressed a high degree of MYB 5′/3′ imbalance and five of these samples were positive for at least one specific MYB-NFIB variant in our panel. MYB-NFIB variant detection on NanoString analysis was confirmed by direct cDNA sequencing. No clinical correlations were found to be associated with MYB fusion status.ConclusionWe conclude that the application of NanoString digital probe counting technology is well suited for the detection and quantification of MYB-NFIB fusion transcripts in archival ACC specimens.

Highlights

Adenoid cystic carcinoma (ACC) is a slow growing salivary gland malignancy that is molecularly characterized by t(6:9)(q22–23;p23–24) translocations which predominantly result in MYB-NFIB gene fusions in nearly half of tumours
MYB 5′-3′ gene expression imbalance detected by NanoString NanoString gene expression profiling was performed on 25 unique ACC tumours and 12 normal matched tissues using a custom-designed MYB fusion gene panel containing
The putative MYB fusion-positive ACCs had MYB 5′:MYB 3′ ratio values ranging from 4.84–91.04 whereas putative fusionnegative ACCs had significantly lower ratios ranging from 0.64–0.88 (Fig. 1)

Summary

Introduction

Adenoid cystic carcinoma (ACC) is a slow growing salivary gland malignancy that is molecularly characterized by t(6:9)(q22–23;p23–24) translocations which predominantly result in MYB-NFIB gene fusions in nearly half of tumours. Detection of MYB-NFIB transcripts is typically performed with fresh ACC tissue using conventional RT-PCR fragment analysis or FISH techniques, which are prone to failure when only archival formalin fixed paraffin embedded (FFPE) tissue is available. The purpose of this pilot study was to evaluate the utility of NanoString probe technology for the detection of MYB-NFIB transcripts in archival ACC tissue. Persson et al were the first to report the presence of chimeric MYB-NFIB fusion transcripts in both breast and head and neck ACCs but total absence in non-ACC tumours [6] They and others were able to identify 14 MYB-NFIB transcript variants by conventional RT-PCR methods [6] [7].

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