Combating melanoma resistance by targeting ABL1/2

总结 尽管在过去的十年里治疗取得了进展，转移性黑色素瘤的五年生存率仍然很低 (23%). BRAF突变发生在50-75%的黑素瘤中，并且BRAF和MEK抑制剂的组合（BRAFi， MEKi）是有效的治疗方法;然而，大多数患者会迅速产生获得性耐药性。来的研究 在过去的十年中，已经确定了黑色素瘤细胞对BRAF/MEK产生耐药性的机制， 抑制剂，但到目前为止，没有FDA批准的方案，逆转BRAFi/MEKi耐药性，并增加 5年生存率。在这里，我们证明了非受体酪氨酸ABL家族的活动， 激酶在BRAFi或BRAFi/MEKi抗性和ABL 1/2驱动抗性后增强。此外，委员会认为， 用FDA批准的抗白血病药物尼洛替尼靶向ABL 1/2， （达拉非尼/曲美替尼）抗性，在体内诱导延长的肿瘤消退。基于这些令人兴奋的 研究结果，我们的建议的中心假设是ABL 1/2是激活ERK依赖性或 - 驱动BRAFi/MEKi抗性的独立途径;因此，靶向ABL 1/2可以有效克服 耐药性并最终延长难治性黑色素瘤患者的生存期。我们的长期， 翻译目标是开发一种新的方案，以提高转移性肝癌患者的5年生存率。 黑素瘤我们提出了一个全面的假设驱动的实验设计和多学科 approach.目标1将定义ABL 1/2驱动阻力的机制。为了实现我们的目标， 使用亲本/抗性黑素瘤细胞系的生物化学、分子生物学和细胞生物学方法， 药理学抑制剂/RNAi、质谱法、异种移植、GEM、PDX和同基因体内方法， 以及从临床试验（目标3）中收集的人体组织将用于检验ABL 1/2 驱动MAPK 3 K激活MEK/ERK/MYC（ERK依赖性抗性）;诱导ERBB 2/ERBB 3激活 PI 3 K/AKT（ERK非依赖性抗性）;并改变免疫细胞浸润。在目标2中，来自 亲本/抗性细胞系，使用体外和体内模型的共培养实验，以及从 动物和患者将被用于测试ABL 1/2驱动的分泌蛋白质组有助于 BRAFi/MEKi电阻。最后，Aim 3是一项多中心、无缝的I/II期临床试验，旨在测试 尼洛替尼与达拉非尼+曲美替尼（BRAFi/MEKi）联合治疗难治性患者。额外 终点包括确定推荐的II期剂量，基于II期数据的初始疗效（PFS）， PK/PD研究和初步探索性生物标志物开发。这些研究的结果不仅会增加 我们对黑色素瘤如何抵抗BRAFi/MEKi作用的理解，也将为研究黑色素瘤的治疗奠定基础。 大型多中心随机临床试验。

SUMMARY Despite treatment advances over the past decade, five-year survival rates for metastatic melanoma remain low (23%). BRAF mutations occur in 50-75% of melanoma, and a combination of BRAF and MEK inhibitors (BRAFi, MEKi) are effective therapies; however, acquired resistance rapidly develops for most patients. Research over the past decade has identified mechanisms by which melanoma cells develop resistance to BRAF/MEK inhibitors, but thus far, there are no FDA-approved regimens that reverse BRAFi/MEKi resistance and increase the 5-year survival rate. Here, we demonstrate that the activities of the ABL family of non-receptor tyrosine kinases are potentiated following BRAFi or BRAFi/MEKi resistance, and ABL1/2 drive resistance. Moreover, targeting ABL1/2 with an FDA-approved anti-leukemia drug, nilotinib, dramatically reverses BRAFi/MEKi (dabrafenib/trametinib) resistance, in vivo, inducing prolonged tumor regression. Based on these exciting findings, the central hypothesis of our proposal is that ABL1/2 are key nodes that activate ERK-dependent or -independent pathways to drive BRAFi/MEKi resistance; thus, targeting ABL1/2 may be effective for overcoming resistance and ultimately extending survival for patients with treatment-refractory melanomas. Our long-term, translational goal is to develop a novel regimen to improve the 5-year survival rate for patients with metastatic melanoma. We propose a comprehensive hypothesis-driven experimental design and multi-disciplinary approach. Aim 1 will define mechanisms by which ABL1/2 drive resistance. To achieve our objective, biochemical, molecular biological and cell biological approaches using parental/resistant melanoma cell lines, pharmacological inhibitors/RNAi, mass spectrometry, xenograft, GEM, PDX, and syngeneic in vivo approaches, as well as human tissue collected from the clinical trial (Aim 3) will be used to test the hypothesis that ABL1/2 drive MAP3K activation of MEK/ERK/MYC (ERK-dependent resistance); induce ERBB2/ERBB3 activation of PI3K/AKT (ERK-independent resistance); and alter immune cell infiltration. In Aim 2, conditioned medium from parental/resistant cell lines, co-culture experiments using in vitro and in vivo models, and serum collected from animals and patients will be used to test the prediction that the ABL1/2-driven secretome contributes to BRAFi/MEKi resistance. Finally, Aim 3 is a multi-site, seamless Phase I/II clinical trial that tests the safety of combining nilotinib with dabrafenib+trametinib (BRAFi/MEKi) for treatment-refractory patients. Additional endpoints include identifying the recommended Phase II dose, initial efficacy (PFS) based on Phase II data, PK/PD studies, and initial exploratory biomarker development. Results from these studies not only will increase our understanding for how melanomas resist the effects of BRAFi/MEKi but also will lay the groundwork for a large, multi-center, randomized clinical trial.