Frequent alternative lengthening of telomeres and ATRX loss in adult NF1-associated diffuse and high-grade astrocytomas

Excerpt

Pilocytic astrocytoma (PA) is the most frequent primary brain tumor developing in patients with neurofibromatosis type 1 (NF1) syndrome. It is also known that the whole spectrum of glial neoplasia may develop in these patients, including low-grade astrocytomas, which are notably difficult to classify, and overtly diffuse gliomas, which are histologically indistinguishable from those occurring sporadically (e.g., diffuse astrocytoma, anaplastic astrocytoma, and glioblastoma) [6]. However, the specific genetic drivers of these tumors are not completely characterized in contrast to gliomas developing in a sporadic setting [1]. Prior studies have linked alterations in the alpha thalassemia/mental retardation syndrome X-linked (ATRX) or death domain-associated protein (DAXX) genes with the alternative lengthening of telomeres (ALT) phenotype in a subset of cancers [3]. ATRX alterations predominate, but ATRX and DAXX are chromatin-remodeling proteins that cooperate at repetitive regions (e.g., telomeres) to incorporate H3.3 into heterochromatin. ATRX mutations and ALT are also associated with specific molecular subgroups of brain tumors, particularly a subset of pediatric glioblastoma [7] and IDH mutant diffuse astrocytomas [1]. Here, we hypothesized that ATRX alterations and concomitant acquisition of ALT may also play a role in NF1-associated gliomas. We tested for ALT using telomere-specific FISH, and nuclear ATRX/DAXX loss through immunohistochemistry in archival paraffin sections using previously published criteria [3]. Additionally, a subset of cases that were ambiguous by FISH due to high tissue autofluorescence were evaluated using a chromogenic (CISH) method (n = 9). A total of 27 gliomas from 26 patients with NF1 syndrome were successfully tested for these markers (ATRX n = 26, ALT FISH/CISH n = 26) (clinicopathologic and molecular data are outlined in supplementary Table 1). The diagnosis of NF1 was performed using established clinical criteria [5]. ALT was present in 10 of 26 (38 %) cases and ATRX loss in 9 of 26 (35 %) cases. DAXX expression was preserved in all 19 (100 %) cases tested. Interestingly, ALT was present in 8 (of 12) diffuse/high-grade gliomas compared to only 2 (of 14) low-grade circumscribed gliomas, while ATRX loss was identified in 7 (of 12) diffuse/high-grade gliomas compared to 2 (of 14) low-grade circumscribed gliomas, a statistically significant difference (p = 0.013 and p = 0.038, respectively, two-tailed Fisher’s exact test) (Fig. 1). IDH1 (R132H) immunohistochemistry was negative in 15 (of 15) cases (6/6 ALT+/ATRX−), p53 was positive in 10 (of 17) cases (6/8 ALT+/ATRX−) and H3 p.K27M was positive in 1 (of 9) case. There was high concordance between ALT and ATRX loss (22 of 25, 88 % cases). The two ALT-positive/ATRX-preserved discordant cases consisted of a low-grade astrocytoma with peculiar SEGA-like morphology (as previously described in NF1 patients) [6] and an anaplastic pleomorphic xanthoastrocytoma (A-PXA). For these cases, we next performed combined telomere-specific FISH for ALT and ATRX immunofluorescence. While we observed no ATRX loss in the low-grade astrocytoma, the A-PXA showed ATRX mislocalization, suggesting an altered protein, which consisted of distinct punctate foci, some of which co-localized with the ultrabright telomeric foci (Fig. 1d–f). When focusing on age, combined ALT+/ATRX− was present in only 1 of 13 pediatric cases (a glioblastoma) vs. 7 of 11 adult cases (p = 0.008). The frequency was particularly high in the adult diffuse/high-grade glioma group, with all but one gliosarcoma case (7/8) demonstrating the ALT phenotype and 6/8 of cases demonstrating combined ALT+/ATRX−.

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