Sir,
We read with interest the recent publication by Heinzerling et al (2013), demonstrating intra-patient heterogeneity of BRAF mutation status between tumours in 10 of 53 (18.9%) patients. However, we have great concern that the results of the study may reflect the (less than 100%) sensitivity of the molecular techniques employed and/or an incorrect assumption that the primary melanoma was the source of the metastatic disease rather than true intra-patient BRAF heterogeneity.
Potentially, the results of the study by Heinzerling et al could have tremendous clinical importance, as accurate determination of a patient’s melanoma BRAF status is critical when planning treatment for melanoma patients with advanced stage disease. Targeting the mitogen-activated protein kinase (MAPK) pathway in patients with BRAF-mutant metastatic melanoma has vastly improved clinical outcomes; however, BRAF inhibitors may paradoxically activate the MAPK pathway in wild-type BRAF melanomas and therefore adversely affect survival if such patients are treated with BRAF inhibitors. Thus, if intra-patient melanoma BRAF heterogeneity exists and treatment decisions are made on the basis of mutation assessment of a single tumour, potentially effective treatment may not be offered in a significant proportion of patients, or alternatively, treatment may be administered that is potentially detrimental.
Although the results of the study by Heinzerling et al are in keeping with other recent reports of heterogeneity in 15% and 13.5% of patients (Colombino et al, 2012; Saint-Jean et al, 2013), two recent studies (Boursault et al, 2013; Menzies et al, 2013) demonstrated very little heterogeneity of BRAF status within metastatic melanoma patients. Several factors may have influenced the results of these studies. First, the techniques used to determine BRAF status were different in the ‘higher’ and ‘lower’ discordance studies. The latter studies used a highly sensitive and specific immunohistochemical technique (the anti-BRAFV600E VE1 antibody) that enables determination of the BRAF status in all individual cells by direct visualisation and at the same time confirmation that they are in fact tumour cells. This technique is not reliant on a certain percentage of tumour cells being present. In contrast, the former studies used molecular methods such as pyrosequencing, allele-specific PCR, and Sanger sequencing, all of which may have false-negative results when samples contain low tumour content. A recent study highlighted the problem of false-negative mutation tests by molecular techniques. Discordant BRAFV600E status was identified in 5 of 97 specimens; subsequent molecular retesting both confirmed an initial molecular misdiagnosis in 4 of the 5 cases and the greater accuracy of BRAF protein immunohistochemistry (Long et al, 2013).
Another factor that may have resulted in heterogeneity is the assumption that any given primary melanoma is the culprit tumour from which the metastatic disease was derived. Ten per cent of patients with metastatic melanoma have a history of multiple primary melanomas (Murali et al, 2012). Even in patients with a history of only a single known primary melanoma, sometimes the site of locoregional metastasis is not in keeping with the T-stage or site of the presumed primary melanoma, or it does not occur within a plausible time period, suggesting that an occult primary melanoma may have led to the metastatic disease. In this situation, close scrutiny of a patient’s clinical history is required to ensure accurate assignment of the ‘culprit’ primary melanoma (Murali et al, 2012).
Clinical responses observed in patients treated with BRAF inhibitors do not support the suggestion of intra-patient BRAF heterogeneity as all metastases have a uniform initial metabolic response to BRAF inhibition assessed using FDG-PET imaging (McArthur et al, 2012), and all resistant lesions resected from patients still contain mutant BRAF (McArthur et al, 2011; Poulikakos et al, 2011; Van Allen et al, 2013).
Further clinical studies are required to examine the issue of intra-patient discordance of BRAF. Carefully assigning primary melanomas as culprit lesions, and using accurate BRAF testing methods with adequate tumour cell content would be the requirements to underpin the data.
References
Boursault L, Haddad V, Vergier B, Cappellen D, Verdon S, Bellocq JP, Jouary T, Merlio JP (2013) Tumor homogeneity between primary and metastatic sites for BRAF status in metastatic melanoma determined by immunohistochemical and molecular testing. PLoS One 8 (8): e70826.
Colombino M, Capone M, Lissia A, Cossu A, Rubino C, De Giorgi V, Massi D, Fonsatti E, Staibano S, Nappi O, Pagani E, Casula M, Manca A, Sini M, Franco R, Botti G, Caraco C, Mozzillo N, Ascierto PA, Palmieri G (2012) BRAF/NRAS mutation frequencies among primary tumors and metastases in patients with melanoma. J Clin Oncol 30 (20): 2522–2529.
Heinzerling L, Baiter M, Kuhnapfel S, Schuler G, Keikavoussi P, Agaimy A, Kiesewetter F, Hartmann A, Schneider-Stock R (2013) Mutation landscape in melanoma patients clinical implications of heterogeneity of BRAF mutations. Br J Cancer 109: 2833–2841.
Long GV, Wilmott JS, Capper D, Preusser M, Zhang YE, Thompson JF, Kefford RF, von Deimling A, Scolyer RA (2013) Immunohistochemistry is highly sensitive and specific for the detection of V600E BRAF mutation in melanoma. Am J Surg Pathol 37 (1): 61–65.
McArthur GA, Puzanov I, Amaravadi R, Ribas A, Chapman P, Kim KB, Sosman JA, Lee RJ, Nolop K, Flaherty KT, Callahan J, Hicks RJ (2012) Marked, homogeneous, and early [18F]fluorodeoxyglucose-positron emission tomography responses to vemurafenib in BRAF-mutant advanced melanoma. J Clin Oncol 30 (14): 1628–1634.
McArthur GA, Ribas A, Chapman PB, Flaherty KT, Kim KB, Puzanov I, Nathanson KL, Lee RJ, Koehler A, Spleiss O, Bollag G, Wu W, Trunzer K, Sosman JA (2011) Molecular analyses from a phase I trial of vemurafenib to study mechanism of action (MOA) and resistance in repeated biopsies from BRAF mutation-positive metastatic melanoma patients (pts). J Clin Oncol 29 (Suppl 15): abstract 8502.
Menzies AM, Lum T, Wilmott JS, Hyman J, Kefford RF, Thompson JF, O'Toole S, Long GV, Scolyer RA (2013) Intrapatient homogeneity of BRAFV600E expression in melanoma. Am J Surg Pathol e-pub ahead of print 12 December 2013 doi:10.1097/PAS.0000000000000136.
Murali R, Brown PT, Kefford RF, Scolyer RA, Thompson JF, Atkins MB, Long GV (2012) Number of primary melanomas is an independent predictor of survival in patients with metastatic melanoma. Cancer 118 (18): 4519–4529.
Poulikakos PI, Persaud Y, Janakiraman M, Kong X, Ng C, Moriceau G, Shi H, Atefi M, Titz B, Gabay MT, Salton M, Dahlman KB, Tadi M, Wargo JA, Flaherty KT, Kelley MC, Misteli T, Chapman PB, Sosman JA, Graeber TG, Ribas A, Lo RS, Rosen N, Solit DB (2011) RAF inhibitor resistance is mediated by dimerization of aberrantly spliced BRAF(V600E). Nature 480 (7377): 387–390.
Saint-Jean M, Quéreux G, Nguyen J-M, Peuvrel L, Brocard A, Vallée A, Knol A-C, Khammari A, Denis MG, Dréno B (2013) Is a single BRAF wild-type test sufficient to exclude melanoma patients from vemurafenib therapy? J Invest Dermatol e-pub ahead of print 11 September 2013; doi:10.1038/jid.2013.378.
Van Allen EM, Wagle N, Sucker A, Treacy DJ, Johannessen CM, Goetz EM, Place CS, Taylor-Weiner A, Whittaker S, Kryukov GV, Hodis E, Rosenberg M, McKenna A, Cibulskis K, Farlow D, Zimmer L, Hillen U, Gutzmer R, Goldinger SM, Ugurel S, Gogas HJ, Egberts F, Berking C, Trefzer U, Loquai C, Weide B, Hassel JC, Gabriel SB, Carter SL, Getz G, Garraway LA, Schadendorf D (2013) The genetic landscape of clinical resistance to RAF inhibition in metastatic melanoma. Cancer Discov e-pub ahead of print 22 November 2013.
Acknowledgements
This work is supported by Program Grant 633004 of the National Health and Medical Research Council of Australia (NHMRC), Translational Research Program Grant 10/TPG/1-02 of the Cancer Institute NSW. GVL and RAS are funded by the Cancer Institute New South Wales and NHMRC Fellowship programmes. The funding bodies had no role in the opinions expressed in the letter.
Author information
Authors and Affiliations
Corresponding author
Ethics declarations
Competing interests
AMM has received honoraria from Roche and travel support from Roche and GlaxoSmithKline (GSK). JSW declares no conflict of interest. GVL has been a consultant for Roche, Bristol-Myers Squibb, GSK and Novartis, and has received honoraria and travel support from Roche. RAS has been a consultant for Roche and GSK, and has received honoraria from Abbott Molecular.
Rights and permissions
This work is licensed under the Creative Commons Attribution-NonCommercial-Share Alike 3.0 Unported License. To view a copy of this license, visit http://creativecommons.org/licenses/by-nc-sa/3.0/
About this article
Cite this article
Menzies, A., Wilmott, J., Long, G. et al. Intra-patient heterogeneity of BRAF mutation status: fact or fiction?. Br J Cancer 111, 1678–1679 (2014). https://doi.org/10.1038/bjc.2013.796
Published:
Issue Date:
DOI: https://doi.org/10.1038/bjc.2013.796
This article is cited by
-
Reply: Intra-patient heterogeneity of BRAF mutation status: fact or fiction?
British Journal of Cancer (2014)