Cancer Res Treat.  2022 Jan;54(1):30-39. 10.4143/crt.2021.218.

Comparison of the Data of a Next-Generation Sequencing Panel from K-MASTER Project with That of Orthogonal Methods for Detecting Targetable Genetic Alterations

• Choi YJ
• Choi JY
• Kim JW
• Lim AR
• Lee Y
• Chang WJ
• Lee S
• Sung JS
• Chung HJ
• Lee JW
• Kang EJ
• Kim JS
• Lim T
• Kim HS
• Kim YJ
• Ahn MS
• Kim YS
• Park JH
• Lim S
• Cho SS
• Cho JH
• Shin SW
• Park KH
• Kim YH

• KMID: 2524585
• DOI: http://doi.org/10.4143/crt.2021.218

Abstract

Purpose
K-MASTER project is a Korean national precision medicine platform that screened actionable mutations by analyzing next-generation sequencing (NGS) of solid tumor patients. We compared gene analyses between NGS panel from the K-MASTER project and orthogonal methods.
Materials and Methods
Colorectal, breast, non–small cell lung, and gastric cancer patients were included. We compared NGS results from K-MASTER projects with those of non-NGS orthogonal methods (KRAS, NRAS, and BRAF mutations in colorectal cancer [CRC]; epidermal growth factor receptor [EGFR], anaplastic lymphoma kinase [ALK] fusion, and reactive oxygen species 1 [ROS1] fusion in non–small cell lung cancer [NSCLC], and Erb-B2 receptor tyrosine kinase 2 (ERBB2) positivity in breast and gastric cancers).
Results
In the CRC cohort (n=225), the sensitivity and specificity of NGS were 87.4% and 79.3% (KRAS); 88.9% and 98.9% (NRAS); and 77.8% and 100.0% (BRAF), respectively. In the NSCLC cohort (n=109), the sensitivity and specificity of NGS for EGFR were 86.2% and 97.5%, respectively. The concordance rate for ALK fusion was 100%, but ROS1 fusion was positive in only one of three cases that were positive in orthogonal tests. In the breast cancer cohort (n=260), ERBB2 amplification was detected in 45 by NGS. Compared with orthogonal methods that integrated immunohistochemistry and in situ hybridization, sensitivity and specificity were 53.7% and 99.4%, respectively. In the gastric cancer cohort (n=64), ERBB2 amplification was detected in six by NGS. Compared with orthogonal methods, sensitivity and specificity were 62.5% and 98.2%, respectively.
Conclusion
The results of the K-MASTER NGS panel and orthogonal methods showed a different degree of agreement for each genetic alteration, but generally showed a high agreement rate.

Keyword

High-throughput nucleotide sequencing; Pathology; Molecular; Precision medicine; Targetable gene alteration

Reference

References

1. Collins FS, Varmus H. A new initiative on precision medicine. N Engl J Med. 2015; 372:793–5.

2. Kummar S, Williams PM, Lih CJ, Polley EC, Chen AP, Rubinstein LV, et al. Application of molecular profiling in clinical trials for advanced metastatic cancers. J Natl Cancer Inst. 2015; 107:djv003.

3. Allegra CJ, Rumble RB, Hamilton SR, Mangu PB, Roach N, Hantel A, et al. Extended RAS gene mutation testing in metastatic colorectal carcinoma to predict response to anti-epidermal growth factor receptor monoclonal antibody therapy: American Society of Clinical Oncology provisional clinical opinion update 2015. J Clin Oncol. 2016; 34:179–85.

4. Amado RG, Wolf M, Peeters M, Van Cutsem E, Siena S, Freeman DJ, et al. Wild-type KRAS is required for panitumumab efficacy in patients with metastatic colorectal cancer. J Clin Oncol. 2008; 26:1626–34.

5. MacConaill LE. Existing and emerging technologies for tumor genomic profiling. J Clin Oncol. 2013; 31:1815–24.

6. Shao D, Lin Y, Liu J, Wan L, Liu Z, Cheng S, et al. A targeted next-generation sequencing method for identifying clinically relevant mutation profiles in lung adenocarcinoma. Sci Rep. 2016; 6:22338.

7. Hagemann IS, Devarakonda S, Lockwood CM, Spencer DH, Guebert K, Bredemeyer AJ, et al. Clinical next-generation sequencing in patients with non-small cell lung cancer. Cancer. 2015; 121:631–9.

8. Lee Y, Lee S, Sung JS, Chung HJ, Lim AR, Kim JW, et al. Clinical application of targeted deep sequencing in metastatic colorectal cancer patients: actionable genomic alteration in K-MASTER Project. Cancer Res Treat. 2021; 53:123–30.

9. Cha YJ, Lee JS, Kim HR, Lim SM, Cho BC, Lee CY, et al. Screening of ROS1 rearrangements in lung adenocarcinoma by immunohistochemistry and comparison with ALK rearrangements. PLoS One. 2014; 9:e103333.

10. Sung JS, Chong HY, Kwon NJ, Kim HM, Lee JW, Kim B, et al. Detection of somatic variants and EGFR mutations in cell-free DNA from non-small cell lung cancer patients by ultra-deep sequencing using the ion ampliseq cancer hotspot panel and droplet digital polymerase chain reaction. Oncotarget. 2017; 8:106901–12.

11. Yang SR, Bouhlal Y, De La Vega FM, Ballard M, Kuo CJ, Vilborg A, et al. Integrated genomic characterization of ERBB2/HER2 alterations in invasive breast carcinoma: a focus on unusual FISH groups. Mod Pathol. 2020; 33:1546–56.

12. Jing C, Mao X, Wang Z, Sun K, Ma R, Wu J, et al. Nextgeneration sequencing based detection of EGFR, KRAS, BRAF, NRAS, PIK3CA, Her2 and TP53 mutations in patients with nonsmall cell lung cancer. Mol Med Rep. 2018; 18:2191–7.

13. Arsenic R, Treue D, Lehmann A, Hummel M, Dietel M, Denkert C, et al. Comparison of targeted next-generation sequencing and Sanger sequencing for the detection of PIK3CA mutations in breast cancer. BMC Clin Pathol. 2015; 15:20.

14. Sakai K, Tsurutani J, Yamanaka T, Yoneshige A, Ito A, Togashi Y, et al. Extended RAS and BRAF mutation analysis using next-generation sequencing. PLoS One. 2015; 10:e0121891.

15. Park E, Shim HS. Detection of targetable genetic alterations in Korean lung cancer patients: a comparison study of single-gene assays and targeted next-generation sequencing. Cancer Res Treat. 2020; 52:543–51.

16. Franczak C, Kandathil SM, Gilson P, Husson M, Rouyer M, Demange J, et al. Uncommon mutational profiles of metastatic colorectal cancer detected during routine genotyping using next generation sequencing. Sci Rep. 2019; 9:7083.

17. Nambiar M, Raghavan SC. How does DNA break during chromosomal translocations? Nucleic Acids Res. 2011; 39:5813–25.

18. Beadling C, Wald AI, Warrick A, Neff TL, Zhong S, Nikiforov YE, et al. A multiplexed amplicon approach for detecting gene fusions by next-generation sequencing. J Mol Diagn. 2016; 18:165–75.

19. Cheng DT, Mitchell TN, Zehir A, Shah RH, Benayed R, Syed A, et al. Memorial Sloan Kettering-Integrated Mutation Profiling of Actionable Cancer Targets (MSK-IMPACT): a hybridization capture-based next-generation sequencing clinical assay for solid tumor nolecular oncology. J Mol Diagn. 2015; 17:251–64.

20. Davies KD, Le AT, Sheren J, Nijmeh H, Gowan K, Jones KL, et al. Comparison of molecular testing modalities for detection of ROS1 rearrangements in a cohort of positive patient samples. J Thorac Oncol. 2018; 13:1474–82.

21. Nong L, Zhang Z, Xiong Y, Zheng Y, Li X, Li D, et al. Comparison of next-generation sequencing and immunohistochemistry analysis for targeted therapy-related genomic status in lung cancer patients. J Thorac Dis. 2019; 11:4992–5003.

22. Wolff AC, Hammond ME, Allison KH, Harvey BE, Mangu PB, Bartlett JMS, et al. Human epidermal growth factor receptor 2 testing in breast cancer: American Society of Clinical Oncology/College of American Pathologists clinical practice guideline focused update. J Clin Oncol. 2018; 36:2105–22.

23. Krumm N, Sudmant PH, Ko A, O’Roak BJ, Malig M, Coe BP, et al. Copy number variation detection and genotyping from exome sequence data. Genome Res. 2012; 22:1525–32.

24. Amarasinghe KC, Li J, Halgamuge SK. CoNVEX: copy number variation estimation in exome sequencing data using HMM. BMC Bioinformatics. 2013; 14(Suppl 2):S2.

25. Grasso C, Butler T, Rhodes K, Quist M, Neff TL, Moore S, et al. Assessing copy number alterations in targeted, amplicon-based next-generation sequencing data. J Mol Diagn. 2015; 17:53–63.

26. Ross DS, Zehir A, Cheng DT, Benayed R, Nafa K, Hechtman JF, et al. Next-generation assessment of human epidermal growth factor receptor 2 (ERBB2) amplification status: clinical validation in the context of a hybrid capture-based, comprehensive solid tumor genomic profiling assay. J Mol Diagn. 2017; 19:244–54.

27. Niu D, Li L, Yu Y, Zang W, Li Z, Zhou L, et al. Evaluation of next generation sequencing for detecting HER2 copy number in breast and gastric cancers. Pathol Oncol Res. 2020; 26:2577–85.

28. Hosonaga M, Arima Y, Sampetrean O, Komura D, Koya I, Sasaki T, et al. HER2 heterogeneity is associated with poor survival in HER2-positive breast cancer. Int J Mol Sci. 2018; 19:2158.

29. Wakatsuki T, Yamamoto N, Sano T, Chin K, Kawachi H, Takahari D, et al. Clinical impact of intratumoral HER2 heterogeneity on trastuzumab efficacy in patients with HER2-positive gastric cancer. J Gastroenterol. 2018; 53:1186–95.

30. McGranahan N, Swanton C. Clonal heterogeneity and tumor evolution: past, present, and the future. Cell. 2017; 168:613–28.