Name: BRAF Mutation Analysis
Test Name: BRAF Mutation Analysis
Health Link Test Code: HCBRAF
CPT Code(s): 81210
Methodology: Real-Time PCR followed by Direct Sequencing
Mutation of the proto-oncogene BRAF occurs in many human cancers, including virtually all hairy cell leukemias (HCL), 27-70% of melanomas, 36-53% of papillary thyroid carcinomas (PTC), 30% of ovarian cancers, 5-22% of colorectal cancers (CRC) and 5-10% of gastrointestinal stromal tumors (GIST) that are negative for KIT and PDGFRA mutations. While the predominant BRAF mutation is c.1799 T>A in exon 15 that results in substitution of glutamate for valine at amino acid 600 (Val600Glu; V600E), there are other mutations of all types (missense substitutions, insertions/deletions and nonsense mutations) that have been described either directly involving or proximal to codon 600. BRAF V600E, and other proximal mutations with similar biological consequences, result in constant activation of BRAF and the MEK/ERK pathway. Consideration of BRAF mutation status should be given when determining appropriate therapy and in differential diagnoses. In CRC, this mutation is a negative predictor of response to Cetuximab (Erbitux®) and Panitumab (Vectibix®). Additionally, BRAF V600E mutations are found in sporadic microsatellite instability high (MSI-H) cases of CRC, but not in hereditary non-polyposis colorectal cancer (HNPCC). BRAF mutation testing may also aid in the diagnosis of papillary thyroid carcinoma as benign thyroid neoplasms are not associated with BRAF mutations. The presence of the V600E BRAF mutation, and others within or proximal to codon 600, may also predict response to Vemurafenib (Zelboraf®) and other BRAF V600E targeted therapies.
Days Performed: Mon, Wed.
In-Lab Turnaround Time: 10 days.
Specimen: Formalin-fixed, paraffin embedded tissue, fine needle aspirates (FNA), and Bone marrow aspirate.
Collection Container: Lavender top
Batched runs begin on Monday or first working day of each week, and Wednesday.
Provider must contact patient’s insurance about coverage for this molecular test and then counsel the patient on out-of-pocket costs.
For Unity and WI-MA patients, documented prior authorization will need to be present in Health Link or sent to laboratory before test will be collected and/or performed.
For all other managed care plans, referral must be placed in Health Link in order for testing on tissue or bone marrow specimens to be performed at UW Health; and patients should be referred to their primary care physician for collection of blood specimens.
For any other insurers, providers must check with insurer before ordering testing.
Collection Volume: 1 mL
Pediatric Collection Volume: 0.6 mL
Sample Analyzed: Tissue
Three slides each containing 5 microns (uM) of FFPE tissue should be sent. Second slide should be H&E stained with the tumor circled. Please indicate percent tumor on Intra-Lab Send-Out Form.
Fine Needle Aspirates (FNA) and Bone Marrow Aspirate have also been validated for this method. Bone Marrow Aspirate do not centrifuge.
If an add-on order is needed, please contact UWHC Surgical Pathology at (608)263-8443. A Surgical Pathology Tissue Examination Request form will need to be completed and faxed to (608)262-7174.
In order to ensure that molecular testing is conducted on a representative sample and a direct correlation of molecular results with morphologic findings is possible, a pathologist selects the specimen(s) and, whenever possible, identifies the suspicious or neoplastic cells submitted for molecular testing.
Outreach Specimen Processing:
Transport with a cold pack. Avoid excessive heat.
Transport at room temperature or with a cold pack. Avoid excessive heat.
Specimens processed in alternative fixatives.
This test is validated to qualitatively detect the human BRAF Val600Glu (V600E) mutation and other mutations within or proximal to codon 600, including, but not limited to V600K, V600R, V600D and small insertions/deletions, in DNA extracted from formalin-fixed, paraffin-embedded (FFPE) tissue, fresh/frozen tissue, fine needle aspirates, selected cells from peripheral blood and bone marrow specimens. The analytical sensitivity of the assay is estimated to be 5%. Consequently, BRAF codon 600 mutations present in cells at less than 5% may not be detected. Additionally, BRAF mutations that are not within or proximal to codon 600 will not be detected. Detection of mutations in or proximal to BRAF codon 600 depends on other numerous factors including, but not limited to, the integrity and heterogenous nature of tissue/tumor samples, the presence/absence of PCR inhibitors and/or the presence/absence of other mutations and/or DNA sequence polymorphisms in BRAF affecting the assay. In addition to not ruling out the presence of other mutations in BRAF, this assay also does not exclude the possibility of mutations in other components of the EGFR signaling cascade. Test results should not be used as the only criterion to form a clinical conclusion, but should be interpreted in the context of all clinical findings, tumor sampling and laboratory data.
A written interpretive report is provided by the laboratory.
Interpretation Type: Expected Results
A “Not Detected” result does not preclude the presence of a mutation in the 600 codon of BRAF since analytical detection depends on numerous factors such as: the heterogenous nature of the tissue sample and ~5% analytical sensitivity of this assay, sample integrity and the absence of PCR inhibitors.
A “Detected” result indicates the presence of a mutation. A “Not Detected” result does not rule out the presence of a BRAF mutation. Inadequate specimen collection, processing and storage may invalidate test results. Test results should not be used as the only criterion to form a clinical conclusion but should be interpreted in the context of all clinical findings, tumor sampling and laboratory data.
The performance characteristics of this test were validated by UWHC Clinical Laboratories. The U.S. Food and Drug Administration (FDA) has not approved or cleared this test; however, FDA approval or clearance is currently not required for clinical use of this test. The results are not intended to be used as the sole means for clinical diagnosis or patient management decisions. The UWHC Clinical Laboratories is authorized under Clinical Laboratory Improvement Amendments (CLIA) to perform high-complexity testing.
A professional fee is associated with this test.