Abstract
In the treatment of metastatic melanoma, a highly therapy-refractory cancer, alkylating agents are used and, for the subgroup of BRAFV600E cancers, the B-Raf inhibitor vemurafenib. Although vemurafenib is initially beneficial, development of drug resistance occurs leading to tumor relapse, which necessitates the requirement for combined or sequential therapy with other drugs, including genotoxic alkylating agents. This leads to the question whether vemurafenib and alkylating agents act synergistically and whether chronic vemurafenib treatment alters the melanoma cell response to alkylating agents. Here we show that a) BRAFV600E melanoma cells are killed by vemurafenib, driving apoptosis, b) BRAFV600E melanoma cells are neither more resistant nor sensitive to temozolomide/fotemustine than non-mutant cells, c) combined treatment with vemurafenib plus temozolomide or fotemustine has an additive effect on cell kill, d) acquired vemurafenib resistance of BRAFV600E melanoma cells does not affect MGMT, MSH2, MSH6, PMS2 and MLH1, nor does it affect the resistance to temozolomide and fotemustine, e) metastatic melanoma biopsies obtained from patients prior to and after vemurafenib treatment did not show a change in the MGMT promoter methylation status and MGMT expression level. The data suggest that consecutive treatment with vemurafenib and alkylating drugs is a reasonable strategy for metastatic melanoma treatment.
Highlights
Malignant melanoma is a highly therapy-refractory cancer, contributing significantly to the worldwide cancer-related mortality [1]
O6MeG needs processing by the DNA mismatch repair (MMR) proteins MSH2, MSH6, PMS2 and MLH1, which converts it during replication into DNA double-strand breaks (DSB) that trigger apoptosis [7] and senescence [8]
We addressed the following questions. a) Does simultaneous treatment of melanoma cells with vemurafenib and TMZ or FM provoke synergistic cell kill? b) Does chronic treatment with vemurafenib cause vemurafenib resistance in vitro and is this accompanied by a change in methylguanine-DNA methyltransferase (MGMT) activity? c) Are vemurafenib resistant BRAFV600E melanoma cells still responsive to TMZ or FM? d) Does vemurafenib treatment change the MGMT promoter methylation status of melanoma tumors in vivo? Our data did not reveal a synergistic effect for both drugs, but encourage a sequential application as vemurafenib resistant cells did not display a change in the MGMT status and retained the killing response towards TMZ and FM
Summary
Malignant melanoma is a highly therapy-refractory cancer, contributing significantly to the worldwide cancer-related mortality [1]. DTIC needs metabolic activation by cytochrome P450 [4] whereas TMZ decomposes spontaneously [5] both giving rise to the DNA reactive methylating species 5-(3-methyltriazen-1-yl)imidazole4-carboximide (MTIC). The main killing DNA lesion induced by DTIC and TMZ in tumor cells is O6methylguanine (O6MeG) [6]. The damage induces autophagy, which in glioma cells counteracts the www.impactjournals.com/oncotarget killing response to TMZ [9]. In contrast to DTIC and TMZ, chloroethylating agents such as lomustine, nimustine, carmustine and fotemustine (FM) induce O6-chloroethylguanine (O6ClEtG) in the DNA, which is the principal critical cytotoxic DNA damage. FM is used as a second line therapeutic in melanoma therapy [11], notably for the treatment of brain metastases [12, 13]
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