Molecular and clinicopathologic characteristics of gliomas with EP300::BCOR fusions

The BCL6 corepressor gene (BCOR) is a tumor suppressor gene located on human chromosome X that regulates cell differentiation and body structure development [2]. Alteration of BCOR has been identified in sarcomas, as well as hematopoietic and central nervous system (CNS) tumors [2]. In the case of CNS tumors, a subgroup has been identified with a distinct DNA methylation profile showing internal tandem duplication in exon 15 of the BCOR gene (BCOR-ITD) [6]. Recently, gene fusions involving BCOR and EP300 have been reported in several CNS tumors with DNA methylation profiles resembling BCOR-ITD or low-grade glioma with MYB alterations (LGG-MYB) [5, 7, 8]. A fusion involving BCORL1 has also been reported [9]. To date, only a few cases (< 10) with EP300::BCOR fusions have been described, warranting further study to understand the genomic and clinicopathologic features of these tumors relative to BCOR-ITD cases.

In this study, we have profiled tumors from 21 patients that define a single DNA methylation group predominantly harboring EP300::BCOR fusions (BCOR-fusion group), and compared them to 15 BCOR-ITD cases, along with other tumors (Table S1, Fig. 1a). The BCOR alterations in tumors from the 21 patients in the BCOR-fusion group included: documented EP300::BCOR fusions (n = 11); CREBBP::BCOR fusion (n = 1); MEAF6::CXXC5 fusion (n = 1); BCOR-Stop mutations (n = 2); and BCOR gene status undetermined but suggestive of fusion based on copy number breakpoints and methylation class (n = 6). Of note, one of the 21 patients had 2 tumors analyzed: Y691 (primary) and AX30 (recurrent), both of which showed a EP300::BCOR fusion (Table S1). RNA sequencing demonstrated fusions at EP300 exon 31 and BCOR exons 4–7, and one fusion at CREBBP exon 31 and BCOR exon 6 (Fig. S1). The DKFZ v12b6 classifier gave confidence scores > 0.9 on 13 cases of these 21 tumors, with 12 cases matching to the EP300::BCOR fusion class and 1 case (with a demonstrated EP300::BCOR fusion) matching to the BCOR-ITD class. The remaining 8 cases (scores < 0.90) were suggested as EP300::BCOR, BCOR-ITD, or other classes. Copy number variation (CNV) analysis using GISTIC2.0 [4] showed that BCOR-fusion tumors are enriched for chromosome 22q12.31 deletion (FDR-adjusted p = 5.6e-5), where EP300 is located. CNV plots showed a high frequency of amplifications near the BCOR gene on chromosome X in the BCOR-fusion tumor group (Fig. 1b). CNV breakpoints (Fig. 1c) at EP300 and BCOR were present in 18 out of 21 BCOR-fusion patients, but none of BCOR-ITD.

Fig. 1.

DNA methylation and copy number variation of BCOR-altered tumors. a UMAP analysis of BCOR-altered tumors, LGG-MYB and HGNET-MN1. b Frequency of altered CNV segments in BCOR-fusion (up) and BCOR-ITD (bottom) tumors. The chromosome segment was considered altered (amplification or deletion) if the intensity score was > 0.4 or < −0.4, respectively. c Example of chromosome breakpoints at EP300 and BCOR on sample BJ64

The median age of patients with BCOR-fusion tumors was 30 years (range 5–72), much older than the median age for BCOR-ITD tumors of 4 years (range 2–45) (Fig. 2a). The sex ratio for BCOR-fusion group was 0.6 male: female (n = 8 male, 13 female), opposite compared to BCOR-ITD cases where males predominated in our cohort as well as in a previous study [3]. In 15 of 21 patients whose tumor location was defined, 10 were supratentorial and 5 were in the posterior fossa. Initial histologic diagnoses for our 21 cases included ependymoma, ganglioglioma, astrocytoma, and glioblastoma (Table S1). Sixteen of the 21 cases with available histology showed diverse histologic appearances including oligodendroglioma-like or ependymoma-like features, some with calcifications (Fig. 2b). Elevated mitotic activity (e.g., readily identified mitotic figures) was frequent, as was tumor necrosis, but microvascular proliferation was uncommon (Fig. 2c). Recurring histologic features included ependymoma-like perivascular pseudorosettes, myxoid change and branching capillary networks. Most tumors in the BCOR-fusion group expressed glial markers (GFAP and OLIG2), and expression of BCOR was less common than in BCOR-ITD cases. The BCOR-fusion tumor patients showed median progression-free time of 45 months (Fig. 2d), which is favorable relative to the previously reported median relapse time of 20 months for BCOR-ITD tumors [3].

Fig. 2.

Age, sex, histopathology, immunohistochemistry results. a Patient age and sex in the two BCOR-altered tumor types. b Histopathological features of BCOR-fusion gliomas. From top left: calcifications can be a prominent feature (Y290). Some tumors show delicate ‘chicken-wire’ vasculature (AN98, BL98). An ependymoma-like histological pattern is predominant in some tumors (BE54, BM60, BJ64). Tumors can show a microcystic, vacuolated appearance, sometimes with myxoid change (AY30, X353, BK22, CH20). Tumor necrosis is common (CC93, Y691, Y844). Some tumors have, at least focally, an oligodendroglioma-like appearance (CB22, Y624). c Histological (top six) and immunohistochemistry (BCOR, OLIG2, GFAP) results along with patient sex (female: black, male: grey) and age group (Child; ≤ 18 years old: black, Adult; > 18: grey). Histomorphology/immunohistochemistry results indicated as black (Yes/Positive) versus grey (No/Negative), and white indicates result not available. Tumor class was indicated at the bottom row. d Progression-free survival curve of patients with BCOR-fusion tumors

In summary, we report 21 cases in a defined methylation class composed predominantly of BCOR-fusion-positive tumors. Compared to BCOR-ITD tumors, the BCOR-fusion group was more frequently observed in adults, and a lower proportion of cases were BCOR immunopositive. Most tumors in the BCOR-fusion group were positive for glial markers (warranting the provisional designation as glioma with BCOR fusion) and matched to the EP300::BCOR fusion class, with occasional cases suggesting BCOR-ITD or other classes. Genomic alterations in tumors of this group include EP300::BCOR fusion (or CREBBP::BCOR and MEAF6::CXXC5 fusion), BCOR stop mutation, and characteristic CNV breakpoints at the EP300 and BCOR loci on chromosomes 22 and X. Of interest, fusions in MEAF6 have also been described as an alternative fusion in other tumor types also known to harbor BCOR fusions, including endometrial stromal sarcoma [1], consistent with our MEAF6::CXXC5 fusion-positive case embedded within the methylation class enriched for BCOR fusions. The mechanism by which inactivating BCOR mutations result in a similar methylation class with BCOR-fusion cases is not known, however, we note that inactivating BCOR mutations are well recognized in neuroepithelial tumors. The original work describing the methylation class of BCOR-altered neuroepithelial tumors (predominantly ITD cases) included 2 cases that harbored BCOR-inactivating mutations, suggesting complex relationships between the different types of alterations involving BCOR and its role in the epigenome and tumorigenesis in neuroepithelial tumors [6]. Taking the histological, immunohistochemical, and molecular characteristics together, tumors in the BCOR-fusion group are distinguished from BCOR-ITD tumors based on the typical patient age (adult versus pediatric), glial differentiation (positive GFAP/OLIG2 expression in most of the studied cases in the fusion-positive group) and different mechanisms of BCOR alteration (predominately BCOR fusions versus ITD). While more experience is required for definitive conclusions, the evidence so far suggests that BCOR fusion-positive gliomas are enriched in the adolescent-young adult (AYA) group, but like other AYA tumor types, can occur outside this range. The histopathology of these tumors can mimic other entities; indeed, one of the samples in the TCGA-LGG cohort, initially labeled as oligodendroglioma, was found to be a fusion-positive BCOR-EP300 glioma in a previous study. We note that the BCOR-fusion group and ITD group have a partially overlapping methylation profile, one case with a documented EP300::BCOR fusion but which matched to the BCOR-ITD class on v12b6 of the classifier, illustrating the importance of orthogonal data (e.g., fusion, breakpoint, or BCOR ITD) in the diagnostic assessment of this group of tumors that match to the larger family of BCOR-altered tumors. Further investigations of the various types of BCOR alteration (ITD, fusions with EP300 and other partners, as well as inactivating mutations) and their role in CNS tumor development are needed to precisely characterize the relationship of BCOR alterations with tumor classification and clinical behavior.

Data availability

Processed methylation results and raw data are available at the Gene Expression Omnibus (GEO) repository under the accession number GSE211156.