Disseminated diffuse midline gliomas, H3K27-altered mimicking diffuse leptomeningeal glioneuronal tumors: a diagnostical challenge!

Diffuse midline gliomas (DMG) are divided into four subtypes depending on their molecular characteristics, and/or location: DMG, H3.3 K27-mutant; DMG, H3.1 or H3.2 K27—mutant; DMG, H3-wildtype, with EZHIP overexpression and DMG, EGFR-altered [1]. Leptomeningeal dissemination at diagnosis has been variably reported depending on the series (up to 42%) [2]. Very little genetic and epigenetic data is available for those disseminated cases, with one case harboring a concomitant FGFR1 mutation [3] and another a 1p deletion [4]. Consequently, their relationship with diffuse leptomeningeal glioneuronal tumors (DLGNT), remains unclarified.

Herein, we describe the histopathological, neuroradiological and molecular (including DNA-methylation profiling) features of three initially disseminated H3K27-altered tumors with glioneuronal features including two cases with an associated MAPK pathway alteration.

The cases concerned three females, aged 14, 13 and 40-year-old (see Additional file 1: Table S1). At the initial diagnosis, in all cases, the tumors were disseminated with supra-tentorial and infra-tentorial leptomeningeal infiltration. An intraparenchymal mono-thalamic involvement was observed in cases 1 and 2; case 3 did not present any intraparenchymal involvement, until the end of the follow-up (Fig. 1). A leptomeningeal biopsy was performed in all cases. Histopathologically, all tumors presented a glioneuronal immunophenotype, and, one of them also had numerous microcalcifications (Fig. 2 and Additional file 2: Table S2). A 1p deletion was evidenced in case 1 and therefore a diagnosis of DLGNT was suggested (Fig. 2D). NGS sequencing showed a FGFR1 N546K mutation (case 1), a BRAF V600E mutation (case 2) and a H3F3A K27M mutation (case 3). The DNA-methylation profiling classified cases 1 and 3 as DMG, H3K27-altered, subtype H3K27M/EZHIP overexpressing (calibrated scores 0.99 and 0.82 respectively) and case 2 as DMG H27K27-altered, subtype EGFR-altered (calibrated score 0.95) (Additional file 3: Fig. S1). Complementary analyses found a loss of H3K27me3 (in all cases), an EZHIP overexpression (cases 1 and 2) (Fig. 2), but no EGFR alteration (all exons were tested by whole exome sequencing, and an amplification was ruled out by FISH analyses) was evidenced. Case 1 received several lines of chemotherapy and craniospinal radiation therapy but passed away 16 months after the initial diagnosis, whereas the case 2, treated by chemotherapy and targeted anti-BRAF therapy, is still alive with a stable disease, 7 months after the initial diagnosis. The patient 3 received chemotherapy and craniospinal irradiation but died 4 months after the diagnosis.

Fig. 1

Radiological features. Case #1 A Axial FLAIR brain MRI shows a hyperintense infiltrative lesion of the right thalamus extended to the right lateral ventricle. B Axial contrast-enhanced T1-weighted brain MRI shows a heterogeneous enhancement after gadolinium injection. C Axial T2-weighted brain MRI shows other nodular FLAIR hyperintensities of the cerebellum (arrows). D Sagittal T2-weighted spine MRI shows a hyperintense peripheral lesion of the spinal cord (arrow). Case #2 E Axial FLAIR-weighted brain MRI shows a hyperintense lesion of the right thalamus extended to the third ventricle and the right hippocampus. F Axial contrast-enhanced T1-weighted brain MRI shows a heterogeneous enhancement of this lesion. G Axial contrast-enhanced T1-weighted brain MRI shows an intraventricular localization in the fourth ventricle (arrow). H Sagittal contrast-enhanced T1-weighted spine MRI shows multiple spinal leptomeningeal lesions. Case #3 I Sagittal T2-weighted spine MRI shows a thoracic hyperintense leptomeningeal lesion. J Sagittal T2-weighted lumbar MRI shows multiple lumbar intradural lesions, attached to nerve roots and in the lower end of the dural sac. K Sagittal and L Axial contrast-enhanced T1-weighted lumbar MRI show an enhancement of these lesions. FLAIR: Fluid Attenuated Inversion Recovery

Fig. 2

Histopathological and molecular features. Case #1 A A glial proliferation with oligo-like features and one microcalcification (HPS, magnification × 400). B Diffuse expression of Olig2 (magnification × 400). C Diffuse synaptophysin immunoreactivity without true neuropil islands (magnification × 400). D 1p deletion by FISH analysis (green signal for 1q25 and orange signal for 1p36, magnification × 400). E EZHIP overexpression in all tumor cells (magnification × 400). Case #2 F A glial proliferation with astrocytic features (magnification × 400). G Diffuse expression of Olig2 (magnification × 400). H Diffuse synaptophysin immunoreactivity without true neuropil islands (magnification × 400). I Loss of the trimethylation H3K27me3 in tumor cells (magnification × 400). J EZHIP overexpression in all tumor cells (magnification × 400). Case #3 K A high-grade glial proliferation with several mitoses and necrosis (magnification × 400). L Immunoreactivity for neurofilament in a subset of tumor cells (magnification × 400). N Loss of the trimethylation H3K27me3 in tumor cells (magnification × 400). O H3K27M immunopositivity in all tumor cells (magnification × 400). Black scale bars represent 50 μm

DLGNTs are glioneuronal tumors molecularly defined by a chromosome arm 1p deletion and a MAPK pathway alterations [1]. Contrary to what their name suggest, they can present a parenchymal component, which can include a thalamic location, with or without leptomeningeal involvement [5]. The already published H3K27M-mutant cases with a disseminated radiological presentation (including a case with a 1p deletion) raises the question of a potential overlap between DLGNT and DMG [3, 4, 6]. However, those cases did not have DNA-methylation analysis, and their relationship to DMG, H3 K27–altered remains open in the last version of the World Health Organization classification [1]. Herein, we present three initially disseminated leptomeningeal tumors, including one case with a 1p deletion and two with BRAF/FGFR1 mutations, classified as DMG using DNA-methylation profiling. Like patients with DMG-H3K27 mutant with concomitant BRAF or FGFR1 mutation, the two current disseminated cases H3K27-altered (one with EZHIP overexpression) with a MAPK mutation were older than classical DMG and histologically presented a glioneuronal immunophenotype and /or microcalcifications [7, 8]. The case 2, classified as DMG, H3K27-altered (EGFR-mutant) proven by DNA-methylation analysis, represents the first example of a disseminated presentation of this typically bithalamic tumor type [9]. Another particularity of this case was its having a BRAF V600E mutation without an EGFR alteration (as 20%, 8/40 of all published cases), representing the second example of this discrepancy between genetic and epigenetic results (the first being reported as unilateral thalamic) [9]. Gliomas with concomitant mutations of H3K27M and BRAF/FGFR1 are supposed to be associated with a better prognosis than other DMG, H3K27-altered according to some publications [7, 8]. As a result, it can be suggested that these molecular alterations (MAPK and H3K27M/EZHIP alterations) confer a different biological behavior, with a metastatic phenotype and/ or a slower local progression ultimately allowing the development of disseminated lesions. Arguing for this hypothesis, a previously published monothalamic tumor classified as ganglioglioma, H3K27M- and BRAF V600E-mutant presented secondary leptomeningeal dissemination 7 years after the initial diagnosis [10]. Further data is needed to understand this disseminated phenotype in detail.

In summary, we showed that despite the histopathological and molecular overlaps with DLGNT, DMG, H3K27-altered may be found to have, in exceptional cases, an initial disseminated radiological presentation.