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Approach to molecular subgrouping of medulloblastomas: Comparison of NanoString nCounter assay versus combination of immunohistochemistry and fluorescence in-situ hybridization in resource constrained centres

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Abstract

Introduction

Molecular classification of medulloblastomas (MB) is prognostically and therapeutically relevant and helps in better risk-stratification. Translation of this subgrouping to routine practice still remains a challenge. The most pathologist accessible techniques for molecular subgrouping include immunohistochemistry (IHC), fluorescent in-situ hybridization (FISH) and NanoString.

Objectives

(1) Molecular subgrouping of MBs by IHC and FISH, and NanoString assay (2) To compare their efficacy and cost for applicability in resource constrained centers.

Methods

Ninety-five cases of MB with adequate tissue were included. Molecular subgrouping was performed by IHC for β-catenin, GAB1 and YAP1; FISH for MYC amplification, and sequencing for CTNNB1, and by NanoString Assay on the same set of MBs. A subset of cases was subjected to 850k DNA methylation array.

Results

IHC + FISH classified MBs into 15.8% WNT, 16.8% SHH, and 67.4% non-WNT/non-SHH subgroups; with MYC amplification identified in 20.3% cases of non-WNT/non-SHH. NanoString successfully classified 91.6% MBs into 25.3% WNT, 17.2% SHH, 23% Group 3 and 34.5% Group 4. However, NanoString assay failure was seen in eight cases, all of which were > 8-years-old formalin-fixed paraffin-embedded tissue blocks. Concordant subgroup assignment was noted in 88.5% cases, while subgroup switching was seen in 11.5% cases. Both methods showed prognostic correlation. Methylation profiling performed on discordant cases revealed 1 out of 4 extra WNT identified by NanoString to be WNT, others aligned with IHC subgroups; extra SHH by NanoString turned out to be SHH by methylation.

Conclusions

Both IHC supplemented by FISH and NanoString are robust methods for molecular subgrouping, albeit with few disadvantages. IHC cannot differentiate between Groups 3 and 4, while NanoString cannot classify older-archived tumors, and is not available at most centres. Thus, both the methods complement each other and can be used in concert for high confidence allotment of molecular subgroups in clinical practice.

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Acknowledgements

The authors would like to thank the technical staff of Neuropathology laboratory for assistance in immunohistochemistry and fluorescence in-situ hybridization, and Dr. Nirupama Trehanpati, Additional Professor, Institute of Liver and Biliary Sciences for providing us with the NanoString platform. The authors are grateful to Dr. Kalaivani for helping with the statistical analysis. Kavneet Kaur is a Senior Research Associate (# 8936/A) under Scientists’ Pool Scheme, Council of Scientific and Industrial Research (CSIR).

Funding

This work was supported financially by the J. C. Bose fellowship awarded to the corresponding author Dr. Chitra Sarkar. The authors declare that the funding source did not play a role in the study design, collection, analysis or interpretation of data, writing of the report, or in the decision to submit the article for publication.

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Authors and Affiliations

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

    Kavneet Kaur, Prerana Jha, Pankaj Pathak, Vaishali Suri, Mehar Chand Sharma & Chitra Sarkar

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

    Ashish Suri

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

    Ajay Garg

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  1. Kavneet Kaur
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Correspondence to Chitra Sarkar.

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Kaur, K., Jha, P., Pathak, P. et al. Approach to molecular subgrouping of medulloblastomas: Comparison of NanoString nCounter assay versus combination of immunohistochemistry and fluorescence in-situ hybridization in resource constrained centres. J Neurooncol 143, 393–403 (2019). https://doi.org/10.1007/s11060-019-03187-y

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