TERT gene rearrangement in chordomas and comparison to other TERT-rearranged solid tumors
Introduction
Chordomas are rare, slow-growing neoplasms thought to arise from the fetal notochord [1]. Histologically, cords and lobules of cells separated by fibrous septa, often with extensive myxoid stroma, and the “physaliferous” tumor cells are notable for abundant, bubbly and eosinophillic cytoplasm. This unique morphology may be related to their genetics; recurrent mutations have been reported in the LYST gene, which encodes for a lysosomal trafficking regulator [2]. Mutations in the LYST gene have been associated with Chediak-Higashi syndrome, where the mutations result in defective lysosomal biogenesis and enlarged organelles in cytotoxic T-cells [3]. A similar phenomenon may be happening in chordoma cells, which are also notable for abundant vacuoles, which have been characterized as lysosome-related organelles [4]. Possible role for these vacuoles in chemo-resistance has been hypothesized, while they have also been hypothesized as potential therapy targets [4]. Immunohistochemically, the tumors are generally immunoreactive for the S100 protein expression, with brachyury being commonly employed as a diagnostic marker. While brachyury expression is seen in nearly all cases of chordomas and their pro-oncogenic tissue notochord, duplication of the TBXT gene (T-box transcription factor T, encoding brachyury) has been reported to be present in only 3/11 (27%) chordoma cases, suggesting different mechanisms are likely responsible for the brachyury expression in other chordomas. In the rare, familial chordomas focal germline tandem TBXT gene duplication has been reported [5]. Despite these interesting genotype-phenotype correlations, the mechanism of tumorigenesis in chordoma is poorly understood. Loss of CDKN2A (encoding p16(INK4A) and p14(ARF) proteins) and mutations in the PI3K/AKT signalling pathway have been reported in a subset of chordomas, but a large proportion of chordomas remain without clear driver mutations [2].
Telomere length stabilization is an important hallmark of cancer cells, crucial for supporting continued cell division [6]. Telomeres are not necessarily long, and telomere lengths are heterogeneous across the different cancers [7]. Telomerase activation and alternative lengthening of telomeres (ALT) are the two key mechanisms in telomere maintenance. The TERT gene encodes a component of the telomerase complex, namely the telomerase reverse transcriptase [8]. The TERT gene is transcriptionally silent in most non-neoplastic tissues, while aberrant telomerase reactivation is commonly observed, perhaps in up to 90% of human cancers [9]. Telomerase reactivation may be associated with point mutations in the TERT gene (including TERT promoter mutations such as C228T and C250T), TERT gene rearrangements (including TERT fusion transcripts) and TERT DNA copy number gains [7, [10], [11], [12], [13]]. TERT promoter mutation analysis and immunohistochemistry for ATRX (loss of which is associated with telomerase dysfunction) are important components of diffuse glioma workup [14, 15]. However, the TERT gene poses a challenge for molecular pathology laboratories due to the GC-rich nature of the region, and the TERT promoter is not included in many commercial and non-commercial cancer sequencing panels. The TERT gene is also not routinely included in a number of fusion transcript assays.
In this study we identified a case of chordoma with a novel TERT gene fusion gene by RNA sequencing. We screened two cohorts of chordomas by fluorescent in-situ hybridization (FISH), and 2/55 cases were found to harbor TERT rearrangement. Furthermore, we assessed 1,913 non-chordoma, solid tumors by RNA sequencing to assess the frequency of TERT fusions, and we compared the molecular features of TERT-rearranged tumors.
Section snippets
Patient selection and tissue microarrays (TMAs)
This study was performed with IRB (University of Pennsylvania, protocol number 834162; Thomas Jefferson University Hospital IRB 20D.235). The discovery cohort (including the index case) comprised 19 cases of chordomas, where the pathology diagnoses were confirmed based on the histomorphology and brachyury IHC positivity (along with S100 protein results when available) (see supplementary data, case 1-23). The discovery cohort TMA was constructed with one-to-six 1.0 mm diameter cores, depending
TERT gene rearrangement in chordomas
In our index chordoma, in-frame fusion transcripts were detected in one case of skull base chordoma from a 60-year old (at diagnosis) female patient. The fusion occurred between exon 5 of RPH3AL (transcript ID NM_001190411.1) and exon 2 of TERT (NM_001193376.1) (Fig. 1A). This fusion met our numerous quality control metrics, including the number of reads (49 reads), number of unique RNA start sites (20 sites), and the average unique start sites per control transcript (140.38). As this fusion
Discussion
With our discovery of TERT gene rearrangements in chordomas, we expand our knowledge of the mutational landscape of chordomas. By FISH, we observed TERT rearrangement in 2/55 (3.6%) of chordomas. By RNA sequencing, TERT fusion transcripts were observed in 1/4 chordomas examined. We suspect that one case of FISH-RNA sequencing discordance (study ID #5) may be related to TERT exon 3 fusions being missed by our RNA sequencing assay, which is limited to fusions involving TERT exon 2. By IHC, we
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