BRAF Mutation Testing - Melanoma testing for Biomarkers and the many challenges

Malignant melanoma is the deadliest form of skin cancer. Fast-growing, melanoma is best beaten with an early and accurate diagnosis followed by the right treatment tailored to the patient based on the identification of a specific biomarker. Biomarkers can predict therapeutic response to treatment. An example of a biomarker is a genetic mutation in a tumor. One of the biomarkers commonly found in melanoma is the BRAF gene mutation, in which V600E is the most common mutation.1 

Recent advances in the biomarker-based treatment of melanoma include the targeted treatments vemurafenib and dabrafenib, which target tumors with the BRAF mutation. When combined with mitogen-activated protein kinase (MEK) inhibitors, vemurafenib and dabrafenib have prolonged progression-free survival and overall survival rates in patients with the BRAF mutation.2-7   

Melanoma deriving from a BRAF mutation is more aggressive than other forms of melanoma. It typically affects younger patients, is more likely to metastasize to the brain, and is associated with shorter overall survival in patients with advanced-stage melanoma.8-10  Therefore, once a diagnosis has been made, it is imperative that biomarker testing immediately ensue to confirm the presence of the BRAF mutation in order to initiate targeted treatment, which will yield a therapeutic response far superior to standard care.  

Diagnostic tests play a crucial role in identifying patients and, ultimately, predicting a therapeutic response and optimizing treatment selection. Tests for determining BRAF mutation status include pharma-produced DNA-based companion diagnostics (CDx kits) and DNA- and protein-based tests developed by laboratories, called laboratory developed tests (LDTs).11  However, just because a test exists does not mean it will be used, run on the optimal platform, or interpreted correctly—all of which may lead to inaccurate results, missed patients, and suboptimal treatment. 

The setting of melanoma presents additional challenges to understanding tumor mutational status because of the common occurrence of tumor heterogeneity and discordance in BRAF mutational status.11  When a tumor is heterogenous, not all the cells within the tumor will express the BRAF mutation. This means patients with BRAF mutations could possibly be missed because the tissue sample biopsied did not contain cells expressing the BRAF mutation, whereas a second sample or a larger sample may. Alternatively, the biomarker test used may not have been sensitive enough to detect the low frequency of the BRAF mutations in the tissue sample. For example, traditional Sanger sequencing is a form of genetic testing that is limited to detecting substitutions and small deletions and insertions of DNA, and provides only 20% sensitivity.12  This may lead to sample rejection or patients being missed, as revealed in a poster recently presented at the European Congress of Pathology.13  In contrast, Next Generation Sequencing (NGS) offers complete mutational coverage and excellent sensitivity, and, if used, is much more likely to pick up on the BRAF mutation even when expressed with low frequency on a particular tumor tissue sample.  

Additionally, BRAF mutational status may change within the same patient, so biomarker testing should not stop at diagnosis. For example, patients who initially presented negative for the BRAF mutation may develop it once the melanoma has metastasized.11  Patients may develop primary or acquired resistance to both immune and molecularly targeted agents, which necessitates on-treatment biomarker monitoring that may predict the likelihood of treatment failure and disease relapse.14   

References 

1. Bradish JR, Cheng L. Molecular pathology of malignant melanoma: changing the clinical practice paradigm toward a personalized approach. Hum Pathol. 2014;45:1315–1326. 
2. Long GV, Weber JS, Infante JR et al. Overall survival and durable responses in patients with BRAF V600-mutant metastatic melanoma receiving dabrafenib combined with trametinib. J Clin Oncol. 2016;34:871–878. 
3. Long GV, Stroyakovskiy D, Gogas H et al. Dabrafenib and trametinib versus dabrafenib and placebo for Val600 BRAF-mutant melanoma: a multicentre, double-blind, phase 3 randomised controlled trial. Lancet. 2015;386:444–451.  
4. Robert C, Karaszewska B, Schachter J et al. Improved overall survival in melanoma with combined dabrafenib and trametinib. N Engl J Med. 2015;372:30–39.  
5. Ascierto PA, McArthur GA, Dréno B et al. Cobimetinib combined with vemurafenib in advanced BRAF(V600)-mutant melanoma (coBRIM): updated efficacy results from a randomised, double-blind, phase 3 trial. Lancet Oncol. 2016;17:1248–1260.  
6. McArthur GA, Chapman PB, Robert C et al. Safety and efficacy of vemurafenib in BRAF(V600E) and BRAF(V600K) mutation-positive melanoma (BRIM-3): extended follow-up of a phase 3, randomised, open-label study. Lancet Oncol. 2014;15:323–332.  
7. Ugurel S, Rohmel J, Ascierto PA et al. Survival of patients with advanced metastatic melanoma: the impact of novel therapies. Eur J Cancer. 2016;53:125–134. 
8. Long GV, Menzies AM, Nagrial AM et al. Prognostic and clinicopathologic associations of oncogenic BRAF in metastatic melanoma. J Clin Oncol. 2011;29:1239–1246.  
9. Ribas A, Flaherty KT. BRAF targeted therapy changes the treatment paradigm in melanoma. Nat Rev Clin Oncol. 2011;24:426–433.  
10. Hugdahl E, Kalvenes MB, Puntervoll HE et al. BRAF-V600E expression in primary nodular melanoma is associated with aggressive tumour features and reduced survival. Br J Cancer. 2016;114:801–808. 
11. Cheng L, Lopez-Beltran A, Massari F, MacLennan GT, Montironi R. Molecular testing for BRAF mutations to inform melanoma treatment decisions: a move toward precision medicine. Mod Pathol. 2018;31(1):24-38.  
12. MacConaill LE. Existing and emerging technologies for tumor genomic profiling. J Clin Oncol. 2013;31(15):1815–1824. 
13. Hartmann A, Stöhr R, Berger I, et al. BRAF mutation testing in melanoma: a study including Austria, Germany and UK, highlighting concordance for current technologies, and potential requirement of more sensitive technologies in future applications. Poster presented at: European Congress of Pathology; September 11, 2018; https://www.diaceutics.com/wp-content/uploads/2018/09/braf-mutation-testing-in-melanoma.pdf. Accessed May 5, 2019 
14. Tarhini A, Kudchadkar RR. Predictive and on-treatment monitoring biomarkers in advanced melanoma: Moving toward personalized medicine. Cancer Treat Rev. 2018;71:8-18.