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C11orf95-RELA fusions and upregulated NF-KB signalling characterise a subset of aggressive supratentorial ependymomas that express L1CAM and nestin

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Abstract

Ependymomas (EPN) show site specific genetic alterations and a recent DNA methylation profiling study identified nine molecular subgroups. C11orf95-RELA and YAP1 fusions characterise the RELA and YAP1 molecular subgroups, respectively, of supratentorial (ST)-EPNs. Current guidelines recommend molecular subgrou** over histological grade for accurate prognostication. Clinicopathological features of ST-EPNs in correlation with C11orf95-RELA and YAP1 fusions have been assessed in only few studies. We aimed to study these fusions in EPNs, and identify diagnostic and prognostic markers. qRT-PCR and Sanger Sequencing for the detection of C11orf95-RELA, YAP1-MAMLD1 and YAP1-FAM118B fusion transcripts, gene expression analysis for NFKB1, and immunohistochemistry for p53, MIB-1, nestin, VEGF, and L1CAM were performed. 88 EPNs (10-Grade I and 78-Grade II/III) from all sites were included. RELA fusions were unique to Grade II/III ST-EPNs, detected in 81.4% (22/27) and 18.5% (5/27) of pediatric and adult ST-EPNs respectively. ST-EPNs harbouring RELA fusions showed frequent grade III histology (81.5%), clear cell morphology (70.3%), upregulated NFKB1 expression, MIB-1 labelling indices (LI) ≥ 10% (77.8%), and immunopositivity for nestin (95.7%), VEGF (72%), L1CAM (79%), and p53 (64%). Presence of RELA fusions, L1CAM immunopositivity and MIB-1 LI ≥ 10% associated with poor outcome. L1CAM showed 81% concordance with RELA fusions. YAP1-MAMLD1 fusion was identified in a single RELA fusion negative adult anaplastic ST-EPN. RELA fusions are frequent in ST-EPNs and associate with poor outcome. L1CAM is a surrogate immunohistochemical marker. RELA fusion positive ST-EPNs strongly express nestin indicating increased stemness. Further evaluation of the interactions between NFKB and stem cell pathways is warranted.

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Acknowledgements

This work was funded by the Indian Council of Medical Research through the Senior Research Fellowship awarded to Prit Benny Malgulwar (No.3/2/3/284/2014/NCD-III) and by Neurosciences Centre, AIIMS. We would like to thank Ms. Suruchi Aggarwal from THSTI, Gurgaon for her help in analysing GEO data. We also thank Ms.Kiran Rani and Mr.Pankaj Kumar from Department of Pathology, AIIMS for their assistance in immunohistochemistry.

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    Authors and Affiliations

    1. Department of Pathology, All India Institute of Medical Sciences, Ansari Nagar, New Delhi, 110029, India

      Prit Benny Malgulwar, Aruna Nambirajan, Pankaj Pathak, Madhu Rajeshwari, Vaishali Suri, Chitra Sarkar & Mehar Chand Sharma

    2. Genomics and Molecular Medicine Unit, Institute of Genomics and Integrative Biology–Council of Scientific and Industrial Research (IGIB-CSIR), New Delhi, India

      Mohammed Faruq

    3. Department of Neurosurgery, All India Institute of Medical Sciences, New Delhi, 110029, India

      Manmohan Singh

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    1. Prit Benny Malgulwar
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    Prit Benny Malgulwar and Aruna Nambirajan have contributed equally, and are co-first authors.

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    Supplementary Figure 1: Detection of Type 1 and Type 2 C11orf95-RELA fusions in ependymomas

    : Quantitative Real Time PCR followed by Sanger sequencing used in the detection of (A) RELAFUS1 (C11orf95 exon 2: RELA exon2) and (B) RELAFUS2 (C11orf95 exon 3: RELA exon 2) fusion transcripts (JPG 285 KB)

    Supplementary Figure 2

    : NFKB1 gene expression analysis: Dot plot shows >5 fold increase in NFKB1 gene expression in RELA fusion positive supratentorial ependymomas as compared to fusion negative cases and normal brain controls (JPG 76 KB)

    Supplementary Figure 3: Kaplan Meier analysis of progression free survival

    : Among all cases included, supratentorial and infratentorial showed a trend towards inferior survival (A). Among supratentorial ependymomas, pediatric age showed an inferior outcome (B). MIB1 labelling indices ≥10% correlated with significant progression (C) while immunopositivity for nestin (D) and VEGF (E) (JPG 275 KB)

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    Supplementary material 6 (DOCX 21 KB)

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    Malgulwar, P.B., Nambirajan, A., Pathak, P. et al. C11orf95-RELA fusions and upregulated NF-KB signalling characterise a subset of aggressive supratentorial ependymomas that express L1CAM and nestin. J Neurooncol 138, 29–39 (2018). https://doi.org/10.1007/s11060-018-2767-y

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