Determining the accuracy of next generation sequencing based copy number variation analysis in Hereditary Breast and Ovarian Cancer

Ağaoğlu, Nihat Buğra; Unal, Busra; Doğan, Özlem Akgün; Zolfagharian, Payam; Shairfli, Pari; Karakurt, Aylin; B, Senay; Kizilboga, Tugba; Yildiz, Jale; Doğanay, Gizem Dinler; Doganay, Levent

doi:10.1080/14737159.2022.2048373

Cited by 2 publications

(3 citation statements)

References 45 publications

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“…7–10) from the results of MS assay which were caused by SNVs in the primer binding sites as revealed by TS. Similarly, SNVs located on the MLPA probe binding sites have been observed to cause false positive results ( Agaoglu et al, 2022 ). Another independent approach, such as TS, long-range PCR or fluorescent in situ hybridization is recommended to further validate the CNV-positive results derived from MS assay or MLPA.…”

Section: Discussionmentioning

confidence: 99%

A MALDI-TOF mass spectrometry-based method for detection of copy number variations in BRCA1 and BRCA2 genes

Zhou,

He,

Zhao

et al. 2024

Front. Mol. Biosci.

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Background: Identifying germline mutations in BRCA1 and BRCA2 genes (BRCAs) would benefit the carriers in multiple aspects. In addition to single-nucleotide variations and small indels, copy number variations (CNVs) is also an indispensable component of identifiable mutations in BRCAs. A sensitive, rapid and throughput-flexible method to detect CNVs would be preferred to meet the rising clinical requirements for BRCAs testing.Methods: We developed a MALDI-TOF-MS-based method (MS assay) which included three steps: first, multiplex end-point PCR followed by a single base extension reaction; second, automated analyte transfer and data acquisition; third, data analysis. We applied MS assay to detect CNVs in BRCAs in 293 Chinese patients with ovarian or pancreatic cancer. All the samples were examined by targeted next-generation sequencing (TS) simultaneously. Samples were further cross-validated by multiplex ligation-dependent probe amplification (MLPA) if the results from MS assay and TS were inconsistent. Long range PCR was then applied to identify the exact breakpoints in BRCAs.Results: MS assay introduced highly multiplexed panels to detect CNVs of BRCAs semi-quantitatively. Simplified on-board data analysis was available for MS assay and no complex bioinformatics was needed. The turnaround time of MS assay was less than 8 hours with a hands-on time of only 40 min. Compared to TS, MS assay exhibited higher sensitivity (100% vs. 75%) and was more flexible in throughput, with the reagent cost per sample remaining constant no matter how many samples were examined per assay. A total of eight CNVs in BRCAs were detected from the 293 samples, and the molecular breakpoints were successfully identified in five samples through long-range PCR followed by Sanger sequencing.Conclusion: Our results suggested that MS assay might be an effective method in primary screening for CNVs in genes such as BRCAs, especially when short turnaround time and/or high sensitivity is a top priority.

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Section: Discussionmentioning

confidence: 99%

A MALDI-TOF mass spectrometry-based method for detection of copy number variations in BRCA1 and BRCA2 genes

Zhou,

He,

Zhao

et al. 2024

Front. Mol. Biosci.

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“…FASTQ data were analyzed by SOPHiA Data‐Driven Medicine (Sophia DDM, Sophia Genetics v4.2, hg19 alignment). Copy numbers were identified by measuring the coverage levels of the desired regions along with samples within the same run 25 …”

Section: Participants Materials and Methodsmentioning

confidence: 99%

“…Copy numbers were identified by measuring the coverage levels of the desired regions along with samples within the same run. 25…”

Section: Sample Preparation Next-generation Sequencing and Bioinforma...mentioning

confidence: 99%

Genomic disparity impacts variant classification of cancer susceptibility genes in Turkish breast cancer patients

Agaoglu,

Unal,

Hayes

et al. 2024

Cancer Medicine

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ObjectiveTurkish genome is underrepresented in large genomic databases. This study aims to evaluate the effect of allele frequency in the Turkish population in determining the clinical utility of germline findings in breast cancer, including invasive lobular carcinoma (ILC), mixed invasive ductal and lobular carcinoma (IDC‐L), and ductal carcinoma (DC).MethodsTwo clinic‐based cohorts from the Umraniye Research and Training Hospital (URTH) were used in this study: a cohort consisting of 132 women with breast cancer and a non‐cancer cohort consisting of 492 participants. The evaluation of the germline landscape was performed by analysis of 27 cancer genes. The frequency and type of variants in the breast cancer cohort were compared to those in the non‐cancer cohort to investigate the effect of population genetics. The variant allele frequencies in Turkish Variome and gnomAD were statistically evaluated.ResultsThe genetic analysis identified 121 variants in the breast cancer cohort (actionable = 32, VUS = 89) and 223 variants in the non‐cancer cohort (actionable = 25, VUS = 188). The occurrence of 21 variants in both suggested a possible genetic population effect. Evaluation of allele frequency of 121 variants from the breast cancer cohort showed 22% had a significantly higher value in Turkish Variome compared to gnomAD (p < 0.0001, 95% CI) with a mean difference of 60 times (ranging from 1.37–354.4). After adjusting for variant allele frequency using the ancestry‐appropriate database, 6.7% (5/75) of VUS was reclassified to likely benign.ConclusionTo our knowledge, this is the first study of population genetic effects in breast cancer subtypes in Turkish women. Our findings underscore the need for a large genomic database representing Turkish population‐specific variants. It further highlights the significance of the ancestry‐appropriate population database for accurate variant assessment in clinical settings.

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Determining the accuracy of next generation sequencing based copy number variation analysis in Hereditary Breast and Ovarian Cancer

Cited by 2 publications

References 45 publications

A MALDI-TOF mass spectrometry-based method for detection of copy number variations in BRCA1 and BRCA2 genes

A MALDI-TOF mass spectrometry-based method for detection of copy number variations in BRCA1 and BRCA2 genes

Genomic disparity impacts variant classification of cancer susceptibility genes in Turkish breast cancer patients

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