Efficacy of Vemurafenib in a Trametinib-Resistant Stage IV Melanoma Patient−−Letter

Clinical Cancer Letter to the Editor Research

Efﬁcacy of Vemurafenib in a Trametinib-Resistant Stage IV Melanoma Patient—Letter

Mathias Bernhardt1,2, Elias Orouji1,2, Lionel Larribere1,2, Christoffer Gebhardt1,2, and Jochen Utikal1,2

Targeted treatment of BRAF gene–mutated melanoma resistance to BRAF and MEK inhibition treatment have been with BRAF and MEK inhibitors has prolonged progres- described, most of which involve reactivation of down- sion-free and overall survival (1, 2). Several mechanisms of stream MAP–ERK kinase/extracellular signal–regulated

AB Before trametinib treatment

Trametinib resistant Figure 1. A, parafﬁn-embedded biopsies of melanoma metastases taken before (top) and after treatment with trametinib (bottom). CDERK activity S100B expression is low in 3 Melanoma dedifferentiated trametinib- Phospho p44 cells resistant tumor cells. B, melanoma Phospho p42 before cell line established from a 2 trametinib trametinib-resistant subcutaneous p44 treatment metastasis. C, strong pERK activity

p42 pERK/ERK 1 measured by Western blot analysis in trametinib-resistant tumor cells (in comparison with treatment- GAPDH naïve BRAF V600E–mutated Trametinib resistant HT144 melanoma cells). D, FISH melanoma shows no gain of BRAF gene copy cells numbers after trametinib treatment (green signals, BRAF gene locus (7q34); orange signals, HT-144 cells HT-144 cells trametinib res. trametinib res. centromeric 7 reference probe). E, Melanoma cells Melanoma cells efﬁcacy of vemurafenib in EFHT-144 melanoma cells trametinib-resistant melanoma cells similar to naïve V600E- mutated HT144 melanoma cells in 100 Melanoma cells the AlamarBlue cell proliferation trametinib res. assay. F, Western blot analysis 50

control showing a decrease of pERK phosphoERK expression after

% Viable cells to 0 DMSO vemurafenib administration in 0 50 100 150 1 nmol/L ERK trametinib-resistant melanoma 10 nmol/L cells. DMSO, dimethyl sulfoxide; Melanoma cells trametinib res. 100 nmol/L GAPDH GAPDH, glyceraldehyde-3- 1000 nmol/L phosphate dehydrogenase. Vemurafenib

100 DMSO

50 control

% Viable cells to 0

0 50 100 150 Vemurafenib100 nmol/L Vemurafenib 1,000 nmol/L

Authors' Afﬁliations: 1Skin Cancer Unit, German Cancer Research Center Corresponding Author: Jochen Utikal, Skin Cancer Unit, German Cancer (DKFZ), Heidelberg and 2Department of Dermatology, Venereology and Research Center (DKFZ) Theodor-Kutzer-Ufer 1-3 Mannheim 68135, Allergology, University Medical Center Mannheim, Ruprecht-Karl Univer- Germany. Phone: 49-621-383-4461; Fax: 49-621-383-3815; E-mail: sity of Heidelberg, Mannheim, Germany [email protected] Note: Supplementary data for this article are available at Clinical Cancer doi: 10.1158/1078-0432.CCR-13-2349 Research Online (http://clincancerres.aacrjournals.org/). Ó2014 American Association for Cancer Research.

2498 Clin Cancer Res; 20(9) May 1, 2014

kinase (MEK/ERK) signaling (3). Thus, BRAF-inhibitor cells could be seen, suggesting other transcriptional resistance mechanisms would be expected to confer resis- mechanisms explaining the trametinib resistance. tance to downstream MEK inhibitor (4). The outcome of In search for a potential treatment option for our an upstream inhibition of the signaling pathway is elusive patient, we tested viable melanoma cells (Fig. 1B) in vitro in which an MEK inhibitor is followed by a BRAF inhib- for potential targeted therapies. We could detect still a itor (5). strong inhibition of cell growth and pERK levels with Here, we report on a 43-year-old male Caucasian the BRAF inhibitor vemurafenib (Fig. 1E and F). This patient with BRAF V600E–mutated stage IV melanoma prompted us to treat our patient with vemurafenib 480 who was treated successfully for 10 months with the MEK mg twice per day. One week after induction of vemur- inhibitor trametinib. Under this treatment, metastases in afenib treatment, the symptoms of the patient ameliora- lung, lymph nodes, and subcutaneous tissue were regres- ted (ECOG2 improved to ECOG0) and tumor masses sive. However, tumor cells became resistant indicated by regressed (Supplementary Fig. S1). Unfortunately, treat- fast progressive disease with new brain and dediffe- ment with vemurafenib had to be interrupted after rentiated subcutaneous metastases (Fig. 1A). A cell line 1monthduetograde3tachycardiaandhearttoxicity established from trametinib-resistant melanoma cells and the patient died 2 months later. (Fig. 1B) showed high levels of ERK and perk, suggesting This case shows that upstream sequential treatment with still a strong activity of the BRAF/MEK/ERK signaling a BRAF-inhibitor might be still efficient in MEK inhibitor– pathway (Fig. 1C). We could not detect mutations in the resistant patients with melanoma. Besides the sequential coding sequence of MEK1 (MEK2 was not transcription- administration shown here, intermittent administration ally expressed) that may inhibit the binding-site of tra- also should be studied further. metinib and thus might explain the activation of the pathway. To exclude amplification of gene copy numbers Disclosure of Potential Conflicts of Interest of genes involved in the BRAF/MEK/ERK signaling path- J. Utikal is consultant/advisory board member for and has received way resulting in a higher transcriptional activity, we speakers bureau honoraria from Roche and MSD. No potential conflicts of performed FISH experiments with gene locus-specific interest were disclosed by the other authors. probes (Fig. 1D and Supplementary Table S1). However, Received August 28, 2013; revised October 23, 2013; accepted November no amplification of these gene loci in trametinib-resistant 18, 2013; published online May 1, 2014.

References 1. Flaherty KT, Robert C, Hersey P, Nathan P, Garbe C, Milhem M, et al. noma successfully rechallenged after progression. Clin Cancer Res Improved survival with MEK inhibition in BRAF-mutated melanoma. 2013;19:5749–57. N Engl J Med 2012;367:107–14. 4. Kim KB, Kefford R, Pavlick AC, Infante JR, Ribas A, Sosman JA, et al. 2. Flaherty KT, Infante JR, Daud A, Gonzalez R, Kefford RF, Sosman J, Phase II study of the MEK1/MEK2 inhibitor Trametinib in patients with et al. Combined BRAF and MEK inhibition in melanoma with BRAF V600 metastatic BRAF-mutant cutaneous melanoma previously treated with mutations. N Engl J Med 2012;367:1694–703. or without a BRAF inhibitor. J Clin Oncol 2013;31:482–9. 3. Romano E, Pradervand S, Paillusson A, Weber J, Harshman K, Mueh- 5. Goldinger SM, Murer C, Stieger P, Dummer R. Upstream MAPK path- lethaler K, et al. Identiﬁcation of multiple mechanisms of resistance to way inhibition: MEK inhibitor followed by a BRAF inhibitor in advanced vemurafenib in a patient with BRAFV600E-mutated cutaneous mela- melanoma patients. Eur J Cancer 2014;50:406–10.

www.aacrjournals.org Clin Cancer Res; 20(9) May 1, 2014 2499

Mathias Bernhardt, Elias Orouji, Lionel Larribere, et al.

Clin Cancer Res 2014;20:2498-2499.

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