Gene fusions and oncogenic mutations in MLH1 deficient and BRAFV600E wild-type colorectal cancers

Iiris Ukkola,Veli Kairisto,Caj Haglund,Jenni Tuominen,Markku Kallajoki,Soili Kytölä,Mia Kero,Pirjo Nummela,Ari Ristimäki,Päivi Peltomäki,Hanna Tammio

doi:10.1007/s00428-022-03302-x

Abstract

Gene fusions can act as oncogenic drivers and offer targets for cancer therapy. Since fusions are rare in colorectal cancer (CRC), their universal screening seems impractical. Our aim was to investigate gene fusions in 62 CRC cases with deficient MLH1 (dMLH1) and BRAFV600E wild-type (wt) status from a consecutive real-life series of 2079 CRCs. First, gene fusions were analysed using a novel FusionPlex Lung v2 RNA–based next-generation sequencing (NGS) panel, and these results were compared to a novel Idylla GeneFusion assay and pan-TRK immunohistochemistry (IHC). NGS detected seven (7/62, 11%) NTRK1 fusions (TPM3::NTRK1, PLEKHA6::NTRK1 and LMNA::NTRK1, each in two cases, and IRF2BP2::NTRK1 in one case). In addition, two ALK, four RET and seven BRAF fusions were identified. Idylla detected seven NTRK1 expression imbalances, in line with the NGS results (overall agreement 100%). Furthermore, Idylla detected the two NGS–identified ALK rearrangements as one specific ALK fusion and one ALK expression imbalance, whilst only two of the four RET fusions were discovered. However, Idylla detected several expression imbalances of ALK (n = 7) and RET (n = 1) that were found to be fusion negative with the NGS. Pan-TRK IHC showed clearly detectable, fusion partner-dependent staining patterns in the seven NTRK1 fusion cases. Overall agreement for pan-TRK antibody clone EPR17341 was 98% and for A7H6R 100% when compared to the NGS. Of the 62 CRCs, 43 were MLH1 promoter hypermethylated (MLH1ph) and 39 were RASwt. All fusion cases were both MLH1ph and RASwt. Our results show that kinase fusions (20/30, 67%) and most importantly targetable NTRK1 fusions (7/30, 23%) are frequent in CRCs with dMLH1/BRAFV600Ewt/MLH1ph/RASwt. NGS was the most comprehensive method in finding the fusions, of which a subset can be screened by Idylla or IHC, provided that the result is confirmed by NGS.

Highlights

Universal screening for mismatch repair deficiency amongst colorectal cancer (CRC) patients has been recommended to facilitate identification of Lynch syndrome (LS) and to direct optimal oncological treatment of those cases presenting a sporadic microsatellite unstable tumour [1]
In case of deficient MLH1 IHC, the dMMR screening algorithm leads to BRAFV600E mutation–targeted IHC testing to identify potential LS patients, i.e. immunonegative and BRAFV600Ewt, to be further tested for MLH1 promoter hypermethylation (MLH1ph)
We screened 2079 CRC resection specimens for deficient MLH1 (dMLH1) and BRAFV600Ewt using IHC and identified 64 such cases of which 62 were available for this study

Summary

Introduction

Universal screening for mismatch repair deficiency (dMMR) amongst colorectal cancer (CRC) patients has been recommended to facilitate identification of Lynch syndrome (LS) and to direct optimal oncological treatment of those cases presenting a sporadic microsatellite unstable tumour [1]. Besides DNA-repair deficiency phenotype in CRC, gene fusions that act as oncogenic drivers offer targets for cancer therapy. To this end, larotrectinib became the first and entrectinib the second tumour agnostic, i.e. ‘histology-independent’, cancer treatment approved by EMA (2019 and 2020, respectively) in patients whose solid tumours display a neurotrophic tyrosine receptor kinase (NTRK) gene fusion and are advanced, have spread to other parts of the body or are not amenable to surgery, and who have no satisfactory alternative treatments [2]. Recent studies have recognised an enrichment of NTRK fusions in a subset of CRCs presenting dMMR due to loss of MLH1 gene expression, BRAFV600E wild-type (wt), MLH1 promoter hypermethylation (MLH1ph) and RASwt [4–7]

Objectives

Methods

Results

Conclusion