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A nCounter CNV Assay to Detect HER2 Amplification: A Correlation Study with Immunohistochemistry and In Situ Hybridization in Advanced Gastric Cancer

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Abstract

Aim

Screening amplified genes for targeted therapy with high-throughput technology is very important. The NanoString nCounter system allows multiplexed digital quantification of target molecules through the use of color-coded barcodes with the great advantage that formalin-fixed, paraffin-embedded (FFPE) tissue can be utilized.

Methods

We tested nCounter custom copy number variation (CNV) panels in 220 gastric cancer samples and evaluated the utility of this method as a screening tool for the detection of CNV using HER2. For the validation of results, we compared the nCounter results with immunohistochemistry (IHC), and we further performed in situ hybridization (ISH) in discrepant cases.

Results

The average HER2 gene copy numbers (CNs) by nCounter were 17.25, 2.0 and 2.61 for the HER2 IHC positive (3+), equivocal (2+), and negative cases, respectively. Out of the 16 IHC 3+ cases, 13 (81.3 %) were reported as HER2 CN gain (≥4). Gastric cancers with homogeneous HER2 overexpression or high tumor purity showed HER2 CN ≥10. Among the 192 cases with HER2 IHC negative and without HER2 gene amplification, 29 showed a HER2 CN ≥4 with the nCounter assay. The nCounter assay had a concordance rate of 83.4 % (kappa value, 0.35), a sensitivity of 66.7 %, a specificity of 85.2 %, a negative predictive value of 96 %, and a positive predictive value of 32.6 % compared with HER2 IHC/ISH results. Fresh frozen (FF) samples revealed a higher concordance rate (91.5 %, kappa value, 0.59) than FFPE samples (78.5 %, kappa value 0.27) and showed a high specificity (97.2 %).

Conclusion

The nCounter CNV assay is a reliable and practical method to detect high CN variations. Given the intra-tumoral HER2 heterogeneity and normal cell contamination, additional IHC and/or FISH is necessary and needs caution in interpretation, especially in FFPE tissue samples.

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    Authors and Affiliations

    1. Department of Pathology and Translational Genomics, Samsung Medical Center, Sungkyunkwan University School of Medicine, 81 Irwon-ro, Gangnam-gu, Seoul, 135-710, Korea

      Soomin Ahn & Kyoung-Mee Kim

    2. Center of Companion Diagnostics, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, Korea

      Soomin Ahn, You Jeong Lyou & Kyoung-Mee Kim

    3. Department of Pathology, Kangnam Sacred Heart Hospital, Hallym University School of Medicine, Seoul, Korea

      Mineui Hong

    4. Department of Pathology and Laboratory Medicine, Tulane University School of Medicine, New Orleans, LA, USA

      Michael Van Vrancken

    5. Department of Medicine, Division of Hematology-Oncology, Samsung Medical Center, Sungkyunkwan University School of Medicine, 81 Irwon-ro, Gangnam-gu, Seoul, 135-710, Korea

      Seung Tae Kim, Se Hoon Park, Won Ki Kang, Young Suk Park & Jeeyun Lee

    6. Biostatistics and Clinical Epidemiology Center, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, Korea

      Sin-Ho Jung & Minah Woo

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    1. Soomin Ahn
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    Correspondence to Jeeyun Lee or Kyoung-Mee Kim.

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    Conflict of interest

    SA, MH, MVV, YP, SK, SP, WK, SJ, MW, JL, and KM have declared no conflicts of interest.

    Funding

    This research was supported by a grant of the Korea Health Technology R&D project through the Korea Health Industry Development Institute (KHIDHI), funded by the Ministry of Health and Welfare, Republic of Korea (HI14C2750 to YSP, HI14C3418 to JL) and National Research Foundation of Korea (NRF) funded by the Ministry of Education, Science and Technology (NRF-2012R1A1A3015504 to KMK) and 20 by 20 project of Samsung Medical Center (GF01140111).

    Ethical approval and informed consent

    All procedures performed in studies involving human participants were in accordance with the ethical standards of the institutional and/or national research committee and with the 1964 Helsinki declaration and its later amendments or comparable ethical standards. Informed consent was obtained from all individual participants included in the study.

    Additional information

    S. Ahn and M. Hong contributed equally to this work.

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    Ahn, S., Hong, M., Van Vrancken, M. et al. A nCounter CNV Assay to Detect HER2 Amplification: A Correlation Study with Immunohistochemistry and In Situ Hybridization in Advanced Gastric Cancer. Mol Diagn Ther 20, 375–383 (2016). https://doi.org/10.1007/s40291-016-0205-4

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