Approximately half of metastatic cutaneous melanomas (CM) harbor a mutation in the BRAF protooncogene, upregulating the mitogen-activated protein kinase (MAPK)-pathway. The development of inhibitors targeting the MAPK pathway (MAPKi), i.e., BRAF- and MEK-inhibitors (BRAFi and MEKi), have substantially improved the survival in BRAF^V600E/K-mutated stage IV metastatic CM. However, most patients develop resistance to treatment and no predictive biomarkers exist in practice. This study aimed at discovering plasma proteome changes during treatment MAPKi in patients with metastatic (stage IV) CM. Matched plasma samples before (pre) and during treatment (trm) from 23 patients with stage IV CM, treated with BRAF-inhibitors (BRAFi) alone or BRAF- and MEK- inhibitors combined (BRAFi and MEKi), were collected and analyzed with targeted proteomics by proximity extension assays. Additionally, plasma from 9 patients treated with BRAFi and MEKi was analyzed with in-depth high-resolution isoelectric focusing liquid-chromatography mass-spectrometry proteomics. Alterations of plasma proteins involved in granzyme and interferon gamma pathways were detected in patients treated with BRAFi, and cell adhesion-, neutrophil degranulation-, and proteolysis pathways in patients treated with BRAFi and MEKi. Several proteins were associated with progression-free survival after MAPKi treatment. We show that the majority of the altered plasma proteins were traceable to BRAF^V600E-mutant metastatic CM tissue at mRNA level in 154 patients from the TCGA, further strengthening their involvement in tumoral response to treatment. This wide screen of plasma proteins unravels proteins that may serve as predictive and/or prognostic biomarkers of MAPKi treatment, opening a window of opportunity for plasma biomarker discovery in MAPKi-treatment of BRAF^V600-mutant metastatic CM.

大约一半的转移性皮肤黑色素瘤 (CM) 在BRAF原癌基因中存在突变，从而上调了丝裂原活化蛋白激酶 (MAPK) 通路。靶向 MAPK 通路 (MAPKi) 的抑制剂的开发，即 BRAF 和 MEK 抑制剂（BRAFi 和 MEKi），显着提高了BRAF ^{V600E/K的存活率}-突变的IV期转移性CM。然而，大多数患者对治疗产生抗药性，并且在实践中不存在预测性生物标志物。本研究旨在发现转移性（IV 期）CM 患者在治疗 MAPKi 期间血浆蛋白质组的变化。收集来自 23 名 IV 期 CM 患者（单独使用 BRAF 抑制剂（BRAFi）或 BRAF 和 MEK 抑制剂联合（BRAFi 和 MEKi）治疗）的匹配血浆样本，并使用靶向药物进行分析。通过邻近延伸测定进行蛋白质组学。此外，采用深入的高分辨率等电聚焦液相色谱质谱蛋白质组学分析了 9 名接受 BRAFi 和 MEKi 治疗的患者的血浆。在接受 BRAFi 治疗的患者中检测到参与颗粒酶和干扰素 γ 途径的血浆蛋白改变，以及接受 BRAFi 和 MEKi 治疗的患者的细胞粘附、中性粒细胞脱粒和蛋白水解途径。几种蛋白质与 MAPKi 治疗后的无进展生存期相关。我们表明，大多数改变的血浆蛋白可追溯到来自 TCGA 的 154 名患者的BRAF ^V600E突变转移性 CM 组织的 mRNA 水平，进一步加强了他们参与肿瘤对治疗的反应。这种广泛的血浆蛋白筛选揭示了可作为 MAPKi 治疗的预测和/或预后生物标志物的蛋白质，为在BRAF ^V600突变转移性 CM的 MAPKi 治疗中发现血浆生物标志物打开了机会之窗。