Comparison of Fluorescence In Situ Hybridization, Next-Generation Sequencing, and DNA Methylation Microarray for Copy...

Gene-level and chromosomal copy number variation (CNV) assessments are critical in the integrated diagnosis of gliomas. Whereas fluorescence in situ hybridization (FISH) has been traditionally employed for CNV detection, emerging technologies such as next-generation sequencing (NGS) and DNA methylation microarray (DMM) are available in clinical practice. Nevertheless, the comparative performance of these 3 assays and the concordance of them remain unclear. A retrospective cohort study comprising 104 patients diagnosed with gliomas was conducted at our hospital. We systematically compared FISH, NGS, and DMM for detecting the following 6 CNV-related diagnostic or prognostic parameters: epidermal growth factor receptor (EGFR), cyclin-dependent kinase inhibitor 2A/B (CDKN2A/B), 1p, 19q, chromosome 7, and chromosome 10. All the 3 methods showed high consistency in EGFR assessment; however, FISH demonstrated relatively low concordance with NGS/DMM in detecting other parameters. In contrast, NGS and DMM exhibited strong concordance in the assessment of all the 6 parameters. Notably, discordant cases were associated with high-grade gliomas (grade 3/4; P < .05) and a high fraction of genome altered (P < .01), indicating high malignancy and genomic instability of discordant cases. This study elucidated the discrepancies and limitations of conventional FISH compared with NGS/DMM in CNV assessments. The discrepancies were associated with high-grade gliomas and genomic instability. We propose a process with recommendations on methods, highlighting the importance of integrated multiplatform assays in accurate clinical diagnosis.