Genomic analysis demonstrates that histologically-defined astroblastomas are molecularly heterogeneous and that tumors with MN1 rearrangement exhibit the most favorable prognosis

Abstract

Astroblastoma (AB) is a rare CNS tumor demonstrating abundant astroblastomatous pseudorosettes. Its molecular features have not been comprehensively studied and its status as a tumor entity is controversial. We analyzed a cohort of 27 histologically-defined ABs using DNA methylation profiling, copy number analysis, FISH and site-directed sequencing. Most cases demonstrated mutually exclusive MN1 rearrangements (n = 10) or BRAF^V600E mutations (n = 7). Two additional cases harbored RELA rearrangements. Other cases lacked these specific genetic alterations (n = 8). By DNA methylation profiling, tumors with MN1 or RELA rearrangement clustered with high-grade neuroepithelial tumor with MN1 alteration (HGNET-MN1) and RELA-fusion ependymoma, respectively. In contrast, BRAF^V600E-mutant tumors grouped with pleomorphic xanthoastrocytoma (PXA). Six additional tumors clustered with either supratentorial pilocytic astrocytoma and ganglioglioma (LGG-PA/GG-ST), normal or reactive cerebrum, or with no defined DNA methylation class. While certain histologic features favored one genetic group over another, no group could be reliably distinguished by histopathology alone. Survival analysis between genetic AB subtypes was limited by sample size, but showed that MN1-rearranged AB tumors were characterized by better overall survival compared to other genetic subtypes, in fact, significantly better than BRAF^V600E-mutant tumors (P = 0.013). Our data confirm that histologically-defined ABs are molecularly heterogeneous and do not represent a single entity. They rather encompass several low- to higher-grade glial tumors including neuroepithelial tumors with MN1 rearrangement, PXA-like tumors, RELA ependymomas, and possibly yet uncharacterized lesions. Genetic subtyping of tumors exhibiting AB histology, particularly determination of MN1 and BRAF^V600E status, is necessary for important prognostic and possible treatment implications.

Fig. 1

Histologically-defined ABs do not represent a homogeneous molecular group by DNA methylation profiling. The top 10,000 differentially methylated probes were used to perform unsupervised hierarchical clustering (a) or t-SNE analysis (b) of ABs and a reference group of tumors from selected DNA methylation classes from the Capper et al. dataset [5]

Fig. 2

Orthogonal validation of molecular classification of ABs. Orthogonal testing including DNA methylation profiling, BRAF^V600E sequencing, and MN1 and RELA FISH was performed on ABs with sufficient material. Supervised analysis was performed using the random forest methylation class prediction algorithm version 11b2 (www.molecularneuropathology.org). Recurrent tumors are indicated by “r”

Fig. 3

Chromosomal copy number variations in AB molecular groups compared to reference tumors. Copy number frequency plots were constructed using copy number profiles of reference tumors and ABs grouping with (a) HGNET-MN1, (b) PXA, and (c) EPN-RELA DNA methylation classes

Fig. 4

AB tumor histology. a–d MN1-rearranged tumors: (a) Case C10 showing mild vascular sclerosis (arrows). This image depicts recurrent tumor. b Case 10. The original lesion was highly sclerotic. c Case C3 showed thin tapering process with expanded endfeet in some areas (shown) and stouter clear cells in other areas. Mild vascular sclerosis is again noted. d Case C16 demonstrating fibrillary areas and vascular sclerosis. e–g BRAF^V600E-positive cases: (e) Case C12 with mild vascular sclerosis. f Case C11. A pseudonuclear inclusion is present (arrow). g Case C6. h–j Cases without known driver mutations: (h) Case 22 showing monopolar columnar-like cells and a pseudonuclear inclusion (arrow). i Case C14. An eosinophilic granular body is shown in the inset. j Case C19 demonstrating mitotic activity (arrow), free hyaline bodies (arrowhead), intracellular hyaline bodies (lower inset), and multinucleate cells (upper inset). k–l C11orf95-RELA tumors: (k) Case C1, low-grade C11orf95-RELA lesion showing AB histology and mild vascular sclerosis (arrow). l Case C30 demonstrating ependymal-like pseudorosettes and clear signet ring-like cells (inset)

Fig. 5

MN1-rearranged ABs show significantly better survival compared to BRAF^V600E-mutant ABs. The initial numbers of patients in each group are indicated in the key. The two RELA/EPEN tumors were not included in the analysis