Although RAF monomer inhibitors (type I.5, BRAF(V600)) are clinically approved for the treatment of BRAF^V600-mutant melanoma, they are ineffective in non-BRAF^V600 mutant cells^1,2,3. Belvarafenib is a potent and selective RAF dimer (type II) inhibitor that exhibits clinical activity in patients with BRAF^V600E- and NRAS-mutant melanomas. Here we report the first-in-human phase I study investigating the maximum tolerated dose, and assessing the safety and preliminary efficacy of belvarafenib in BRAF^V600E- and RAS-mutated advanced solid tumours (NCT02405065, NCT03118817). By generating belvarafenib-resistant NRAS-mutant melanoma cells and analysing circulating tumour DNA from patients treated with belvarafenib, we identified new recurrent mutations in ARAF within the kinase domain. ARAF mutants conferred resistance to belvarafenib in both a dimer- and a kinase activity-dependent manner. Belvarafenib induced ARAF mutant dimers, and dimers containing mutant ARAF were active in the presence of inhibitor. ARAF mutations may serve as a general resistance mechanism for RAF dimer inhibitors as the mutants exhibit reduced sensitivity to a panel of type II RAF inhibitors. The combination of RAF plus MEK inhibition may be used to delay ARAF-driven resistance and suggests a rational combination for clinical use. Together, our findings reveal specific and compensatory functions for the ARAF isoform and implicate ARAF mutations as a driver of resistance to RAF dimer inhibitors.

尽管 RAF 单体抑制剂（I.5 型，BRAF(V600)）在临床上被批准用于治疗BRAF ^V600突变黑色素瘤，但它们对非BRAF ^V600突变细胞无效^1,2,3。Belvarafenib 是一种有效的选择性 RAF 二聚体（II型）抑制剂，在BRAF ^V600E和NRAS突变黑色素瘤患者中表现出临床活性。在这里，我们报告了研究最大耐受剂量的首次人体 I 期研究，并评估了贝伐拉非尼在BRAF ^V600E和RAS中的安全性和初步疗效-突变的晚期实体瘤（NCT02405065，NCT03118817）。通过生成对贝伐拉非尼耐药的NRAS突变黑色素瘤细胞并分析来自接受贝伐非尼治疗的患者的循环肿瘤 DNA，我们在激酶结构域内发现了ARAF中新的复发突变。ARAF 突变体以二聚体和激酶活性依赖性方式赋予对贝伐非尼的抗性。Belvarafenib 诱导 ARAF 突变二聚体，含有突变 ARAF 的二聚体在抑制剂存在下具有活性。ARAF突变可作为 RAF 二聚体抑制剂的一般耐药机制，因为突变体对一组 II 型 RAF 抑制剂的敏感性降低。RAF 加 MEK 抑制的组合可用于延迟ARAF-驱动的耐药性，并建议临床使用的合理组合。总之，我们的研究结果揭示了 ARAF 亚型的特异性和补偿功能，并暗示ARAF突变是对 RAF 二聚体抑制剂耐药的驱动因素。