Olaparib (LynparzaTM) was recently approved by the European Medicine Agency (EMA) for the treatment of recurrent, platinum-sensitive, high-grade serous ovarian carcinoma (OC). The detection of a somatic or germline mutation in the BRCA1 or BRCA2 genes is a requirement for treatment with this drug. While the EMA did not specify the origin of a mutation, the American Food and Drug Administration (FDA) recommend treatment with Olaparib only for germline mutations. CeGaT offers genetic diagnostics for both purposes, and recommends the comparison of both tumor and normal tissues for the most accurate and informative analysis.
BRCA1 and BRCA2 are genes that play an important role in DNA-repair mechanisms. While 17% of patients with ovarian carcinoma have a germline mutation in BRCA1 or BRCA2, an additional 6% have somatic mutations that are restricted to the tumor tissue. Sporadic OCs are also often due to germline mutations, thus all patients for whom Olaparib treatment is an option should be informed about the implications of germline mutations and should attend Genetic Counseling. The detection of a germline mutation means an increased risk for descendants and relatives to have the same mutation and therefore an enhanced risk of disease.
Due to the heterogeneity of a tumor, the comparative analysis of tumor and normal tissue is important to obtain results with highest sensitivity and to ensure a complete analysis of the genetic mutations. We therefore routinely examine tumor tissue and normal tissue in parallel. CeGaT offers four options for the analysis of mutations involved in ovarian carcinoma. While options 1, 2 and 3 are restricted to the analysis of the gene regions of BRCA1 and BRCA2, option 4 includes the analysis of 649 cancer-relevant genes (incl. BRCA1 and BRCA2), in which mutations known to contribute to tumor development have been documented. Option 4 supports a treatment decision based on a comprehensive characterization of the tumor. In addition to genetic analysis using modern sequencing technologies, our service includes the evaluation and interpretation of detected variants by a team of experts in bioinformatics and human genetics.
Option 1: BRCA1 and BRCA2 analysis in tumor tissue only
In option 1, only mutations in the tumor are analyzed, no differentiation between germline and somatic mutations can be made. We do not recommend this option, but will perform it when it is explicitly requested by a patient.
Option 2: BRCA1 and BRCA2 analysis in blood only
In option 2, only germline mutations are analyzed. This option is relevant if the tumor of the patient is not available.
Option 3: BRCA1 and BRCA2 analysis in tumor tissue and normal tissue (blood)
In option 3, a comprehensive analysis of somatic and germline mutations in the genes BRCA1 and BRCA2 is carried out. The comparison of tumor and normal DNA allows the most sensitive and specific analysis to be performed.
Option 4: Somatic Tumor Panel
In option 4, a comprehensive analysis of somatic and germline mutations in 649 cancer-relevant genes is performed. In addition to the estimation of a therapeutic response to Olaparib, additional therapeutic options can be identified.
When samples arrive at the laboratory, tumors may be macro-dissected to ensure high tumor content in the DNA sample. After DNA isolation, enrichment of the coding regions and neighboring intronic regions (+/- 10 bp) is performed using an amplicon strategy. High-throughput sequencing is performed using the Illumina HiSeq platform. In addition to sequencing, multiplex ligation-dependent probe amplification (MLPA) for the gene regions of BRCA1 and BRCA2 is carried out to analyze these genes for deletions or duplications in the germ line. For option 4 there is also the possibility to examine the tumor tissue for deletions and duplications using array comparative genomic hybridization (aCGH).
Sequencing reads are mapped to the reference genome (hg19), variants are detected (SNPs, SNVs, small deletions and insertions), and the variants are annotated using internal and external databases. Variants found in the tumor are compared to the reference genome (options 1 and 2) for the identification of mutations. In options 3 and 4, the variants are compared between a patient’s own tumor and normal tissue which ensures the clear differentiation between germline and somatic mutations. A medical report is issued by our team of experienced scientists and geneticists.
Tumor tissue (FFPE or frozen) or tumor DNA (1 μg DNA)
DNA or blood of the patient (5 μg DNA or 5-10 ml EDTA-blood)
CeGaT order form including the written consent of the patient
10-12 working days for options 1-3
20 working days for option 4
Recommendations from the Breast Cancer Consortium
The genetic analysis of the genes BRCA1 and BRCA2 is indicated in the following family situations, as the probability of having a mutation in BRCA1 or BRCA2 is over 10%.
At least two women with breast cancer, at least one of which was younger than 51 years when the first tumor appeared
At least one woman with breast cancer and one woman with ovarian cancer, or one woman with both breast and ovarian cancers
At least two women with ovarian cancer
At least one woman with breast cancer in both breasts, who was younger than 51 years when the first tumor appeared
At least one woman with breast cancer, who was younger than 36 years when the first tumor appeared
At least one man with breast cancer and one additional person with breast or ovarian cancer
At least three women with breast cancer, independent of the age at which the first tumor appeared