PURPOSE Most aggressive thyroid cancers are commonly associated with a BRAF V600E mutation. Preclinical and clinical data in BRAF V600E cancers suggest that combined BRAF and MEK inhibitor treatment results in a response, but resistance is common. One mechanism of acquired resistance is through persistent activation of tyrosine kinase (TK) signaling by alternate pathways. We hypothesized that combination therapy with BRAF and multitargeting TK inhibitors (MTKI) might be more effective in BRAF V600E thyroid cancer than in single-agent or BRAF and MEK inhibitors. EXPERIMENTAL DESIGN The combined drug activity was analyzed to predict any synergistic effect using high-throughput screening (HTS) of active drugs. We performed follow-up in vitro and in vivo studies to validate and determine the mechanism of action of synergistic drugs. RESULTS The MTKI ponatinib and the BRAF inhibitor PLX4720 showed synergistic activity by HTS. This combination significantly inhibited proliferation, colony formation, invasion, and migration in BRAF V600E thyroid cancer cell lines and downregulated pERK/MEK and c-JUN signaling pathways, and increased apoptosis. PLX4720-resistant BRAF V600E cells became sensitized to the combination treatment, with decreased proliferation at lower PLX4720 concentrations. In an orthotopic thyroid cancer mouse model, combination therapy significantly reduced tumor growth (P < 0.05), decreased the number of metastases (P < 0.05), and increased survival (P < 0.05) compared with monotherapy and vehicle control. CONCLUSIONS Combination treatment with ponatinib and PLX4720 exhibited significant synergistic anticancer activity in preclinical models of BRAF V600E thyroid cancer, in addition to overcoming PLX4720 resistance. Our results suggest this combination should be tested in clinical trials.

中文翻译：

---

使用 BRAFV600E 抑制剂 (PLX4720) 和酪氨酸激酶抑制剂 (Ponatinib) 靶向 BRAF V600E 突变癌症的组合策略。

目的 大多数侵袭性甲状腺癌通常与 BRAF V600E 突变有关。BRAF V600E 癌症的临床前和临床数据表明，BRAF 和 MEK 抑制剂联合治疗会产生反应，但耐药性很常见。获得性耐药的一种机制是通过替代途径持续激活酪氨酸激酶 (TK) 信号。我们假设与单药或 BRAF 和 MEK 抑制剂相比，BRAF 和多靶点 TK 抑制剂 (MTKI) 的联合治疗可能对 BRAF V600E 甲状腺癌更有效。实验设计 使用活性药物的高通量筛选 (HTS) 分析组合药物活性以预测任何协同效应。我们进行了体外和体内随访研究，以验证和确定协同药物的作用机制。结果 MTKI ponatinib 和 BRAF 抑制剂 PLX4720 显示出 HTS 的协同活性。这种组合显着抑制了 BRAF V600E 甲状腺癌细胞系的增殖、集落形成、侵袭和迁移，并下调了 pERK/MEK 和 c-JUN 信号通路，并增加了细胞凋亡。PLX4720 抗性 BRAF V600E 细胞对联合治疗变得敏感，在较低的 PLX4720 浓度下增殖减少。在原位甲状腺癌小鼠模型中，与单一疗法和载体对照相比，联合疗法显着减少了肿瘤生长（P < 0.05），减少了转移数量（P < 0.05），并增加了存活率（P < 0.05）。结论 除了克服 PLX4720 耐药外，普纳替尼和 PLX4720 的联合治疗在 BRAF V600E 甲状腺癌的临床前模型中表现出显着的协同抗癌活性。我们的结果表明应该在临床试验中测试这种组合。