Related Experiment Video
Updated: Aug 12, 2025
Integration of Wet and Dry Bench Processes Optimizes Targeted Next-generation Sequencing of Low-quality and Low-quantity Tumor Biopsies
Published on: April 11, 2016
Validation of multi-gene panel next-generation sequencing for the detection of BRCA mutation in formalin-fixed,
Eun Taeg Kim1, Ha Eun Jeong1, Hyung Joon Yoon1
1Department of Obstetrics and Gynecology, Pusan National University School of Medicine, Biomedical Research Institute, Pusan National University Hospital, Busan, South Korea.
Objective:
The therapeutic effect of poly (ADP-ribose) polymerase (PARP) inhibitors in patients with epithelial ovarian cancer (EOC) with somatic BRCA mutations is consistent with that observed in patients with germline BRCA mutations, indicating the importance of detecting both germline and somatic BRCA mutations concurrently. We compared the efficacy of multi-gene panel next generation sequencing (NGS) in EOC patients' formalin-fixed, paraffin-embedded (FFPE) tissue to that of conventional Sanger sequencing in blood samples.
Materials And Methods:
This study included 48 patients with EOC, and both blood Sanger sequencing and FFPE tissue NGS were conducted in all of them. Clinical and pathological data were reviewed, including age at diagnosis, histology, and stage. Blood Sanger sequencing was performed using peripheral blood leukocytes. The target regions of 90 cancer-related genes were identified using FFPE tissue.
Results:
The median age of patients was 56.1 years, with serous carcinoma (n = 40, 83.3%) and stage III (n = 37, 77.1%) being the most common histology and International Federation of Gynecology and Obstetrics (FIGO) stage, respectively. FFPE tissue NGS identified ten pathogenic variants, including all eight pathogenic variants identified by blood Sanger sequencing and two additional pathogenic variants. Furthermore, FFPE tissue NGS identified 19 variants of uncertain significance (VUS), including all ten VUS identified by blood Sanger sequencing and nine additional VUS.
Conclusion:
The FFPE tissue multi-gene panel NGS had 100% sensitivity for detecting BRCA germline mutations and could detect additional somatic mutations. Furthermore, performing FFPE tissue multi-gene panel NGS followed by blood Sanger sequencing sequentially may help differentiate germline from somatic BRCA mutations for genetic counseling.
Insights
Next-generation sequencing (NGS) on formalin-fixed, paraffin-embedded (FFPE) tissue effectively detects BRCA mutations in epithelial ovarian cancer (EOC). This method identifies both germline and somatic mutations, aiding in genetic counseling for PARP inhibitor therapy.
Area of Science:
- Oncology
- Genetics
- Molecular Diagnostics
Background:
- Poly (ADP-ribose) polymerase (PARP) inhibitors show therapeutic efficacy in epithelial ovarian cancer (EOC) patients with somatic BRCA mutations, similar to those with germline BRCA mutations.
- Accurate detection of both germline and somatic BRCA mutations is crucial for optimizing PARP inhibitor treatment strategies in EOC.
- Conventional Sanger sequencing in blood samples has limitations in comprehensively identifying all relevant BRCA mutations.
Purpose of the Study:
- To compare the efficacy of multi-gene panel next-generation sequencing (NGS) using formalin-fixed, paraffin-embedded (FFPE) EOC tissue against conventional Sanger sequencing in blood.
- To evaluate the capability of FFPE tissue NGS in detecting both germline and somatic BRCA mutations concurrently.
- To assess the utility of FFPE tissue NGS in conjunction with blood Sanger sequencing for differentiating mutation types for genetic counseling.
Main Methods:
- Included 48 EOC patients, with both blood Sanger sequencing and FFPE tissue NGS performed on all participants.
- Clinical and pathological data, including age, histology, and FIGO stage, were reviewed.
- FFPE tissue NGS targeted 90 cancer-related genes, while blood Sanger sequencing analyzed peripheral blood leukocytes.
Main Results:
- FFPE tissue NGS identified all pathogenic variants found by blood Sanger sequencing, plus two additional pathogenic variants.
- FFPE tissue NGS detected 19 variants of uncertain significance (VUS), including all 10 identified by blood Sanger sequencing and nine novel VUS.
- The study demonstrated FFPE tissue NGS's superior sensitivity in detecting a broader spectrum of BRCA-related genetic alterations.
Conclusions:
- Multi-gene panel NGS on FFPE tissue demonstrated 100% sensitivity for detecting BRCA germline mutations and identified additional somatic mutations.
- Sequential testing, starting with FFPE tissue NGS followed by blood Sanger sequencing, can effectively differentiate germline from somatic BRCA mutations.
- This approach enhances diagnostic accuracy and supports informed genetic counseling for EOC patients undergoing PARP inhibitor therapy.
More Related Videos
10:27Testing Targeted Therapies in Cancer using Structural DNA Alteration Analysis and Patient-Derived Xenografts
Published on: July 25, 2020
08:15gDNA Enrichment by a Transposase-based Technology for NGS Analysis of the Whole Sequence of BRCA1, BRCA2, and 9 Genes Involved in DNA Damage Repair
Published on: October 6, 2014