Bechet Denise, Gielen Gerrit G H, Korshunov Andrey, Pfister Stefan M, Rousso Caterina, Faury Damien, Fiset Pierre-Olivier, Benlimane Naciba, Lewis Peter W, Lu Chao, David Allis C, Kieran Mark W, Ligon Keith L, Pietsch Torsten, Ellezam Benjamin, Albrecht Steffen, Jabado Nada
Departments of Experimental Medicine and of Human Genetics, McGill University, 4060 Ste Catherine West, PT239, Montreal, QC, H3Z2Z3, Canada.
Acta Neuropathol. 2014 Nov;128(5):733-41. doi: 10.1007/s00401-014-1337-4. Epub 2014 Sep 9.
Studies in pediatric high-grade astrocytomas (HGA) by our group and others have uncovered recurrent somatic mutations affecting highly conserved residues in histone 3 (H3) variants. One of these mutations leads to analogous p.Lys27Met (K27M) mutations in both H3.3 and H3.1 variants, is associated with rapid fatal outcome, and occurs specifically in HGA of the midline in children and young adults. This includes diffuse intrinsic pontine gliomas (80 %) and thalamic or spinal HGA (>90 %), which are surgically challenging locations with often limited tumor material available and critical need for specific histopathological markers. Here, we analyzed formalin-fixed paraffin-embedded tissues from 143 pediatric HGA and 297 other primary brain tumors or normal brain. Immunohistochemical staining for H3K27M was compared to tumor genotype, and also compared to H3 tri-methylated lysine 27 (H3K27me3) staining, previously shown to be drastically decreased in samples carrying this mutation. There was a 100 % concordance between genotype and immunohistochemical analysis of H3K27M in tumor samples. Mutant H3K27M was expressed in the majority of tumor cells, indicating limited intra-tumor heterogeneity for this specific mutation within the limits of our dataset. Both H3.1 and H3.3K27M mutants were recognized by this antibody while non-neoplastic elements, such as endothelial and vascular smooth muscle cells or lymphocytes, did not stain. H3K27me3 immunoreactivity was largely mutually exclusive with H3K27M positivity. These results demonstrate that mutant H3K27M can be specifically identified with high specificity and sensitivity using an H3K27M antibody and immunohistochemistry. Use of this antibody in the clinical setting will prove very useful for diagnosis, especially in the context of small biopsies in challenging midline tumors and will help orient care in the context of the extremely poor prognosis associated with this mutation.
我们团队及其他团队针对儿童高级别星形细胞瘤（HGA）开展的研究发现，组蛋白3（H3）变体中影响高度保守残基的体细胞突变会反复出现。其中一种突变在H3.3和H3.1变体中均导致类似的p.Lys27Met（K27M）突变，与快速致命的结局相关，且特别发生于儿童和青年的中线HGA中。这包括弥漫性固有脑桥胶质瘤（80%）以及丘脑或脊髓HGA（>90%），这些部位手术难度大，通常可获取的肿瘤组织有限，且迫切需要特定的组织病理学标志物。在此，我们分析了143例儿童HGA以及297例其他原发性脑肿瘤或正常脑的福尔马林固定石蜡包埋组织。将H3K27M的免疫组织化学染色与肿瘤基因型进行比较，并与先前显示在携带该突变的样本中显著减少的H3三甲基化赖氨酸27（H3K27me3）染色进行比较。肿瘤样本中H3K27M的基因型与免疫组织化学分析之间的一致性为100%。突变型H3K27M在大多数肿瘤细胞中表达，表明在我们数据集的范围内，该特定突变的肿瘤内异质性有限。该抗体可识别H3.1和H3.3K27M突变体，而诸如内皮细胞、血管平滑肌细胞或淋巴细胞等非肿瘤成分不着色。H3K27me3免疫反应性与H3K27M阳性在很大程度上相互排斥。这些结果表明，使用H3K27M抗体和免疫组织化学可高特异性和高灵敏度地特异性识别突变型H3K27M。在临床环境中使用该抗体将对诊断非常有用，尤其是在具有挑战性的中线肿瘤的小活检情况下，并且将有助于在与该突变相关的极差预后背景下指导治疗。
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