Overcoming resistance in BCR-ABL-rearranged acute lymphoblastic leukemia

项目总结/摘要 BCR-ABL融合是急性淋巴细胞白血病最常见的分子亚型（BCR-ABL+ ALL），但在历史上预后不良。加入酪氨酸激酶抑制剂（TKI）， 将ABL催化结构域结合到一线治疗方案中可以诱导>90%的患者完全缓解，但 几乎所有的都会复发，典型的是带有不利于药物结合的耐药突变。这表明复发 仍然对ABL激酶活性上瘾。相比之下，新型IV型抑制剂结合ABL肉豆蔻酸酯位点， 变构调节BCR-ABL功能。我们假设催化和变构TKI的组合 可以防止交叉耐药克隆的出现，并治愈一部分未经治疗的BCR-ABL+ ALL。我们 已经建立了一个由16个BCR-ABL+ ALL患者来源的异种移植物（PDX）模型组成的多样化小组， 可以进行对照和充分把握度的临床前试验，以评估体内 联合阻断，开发预测性生物标志物，并建立体内获得性耐药模型。我们 还开发了一种表征单个肿瘤细胞治疗敏感性的方法， 适用于微小残留病（MRD）标本。具体来说，我们使用基于微处理器的平台 被称为悬浮微通道谐振器（SMR），通过测量药物敏感性的变化， 具有毫微微克范围灵敏度的离体暴露于靶向抑制剂的单个细胞的漂浮质量。这些 然后在SMR下游收集细胞用于单细胞RNA-Seq（scRNA-seq）以确定转录水平。 调节对治疗方法的差异反应的程序和细胞状态。为了验证我们的假设 联合ABL阻断具有治疗潜力，并满足对快速和稳健方法的迫切需求 为了表征MRD中单个肿瘤标本的灵敏度，我们提出了以下具体目标： (1)测试联合催化和变构BCR-ABL抑制是否可以治愈一些BCR-ABL+ ALL PDX 模型任何在联合治疗中复发的模型将被询问突变、转录（两者都是）， 编码和非编码RNA），以及获得性体内抗性的蛋白质组学相关性。(2)确定方法， 直接克服体内MRD内的治疗耐药性。我们将使用SMR来鉴定抗性亚克隆 并应用scRNA-seq来定义转录程序和介导耐药性的细胞状态， 其中大体积肿瘤测定是不敏感的。确定可通过联合ABL阻滞治愈的患者 将是变革性的，正如结合PML-RAR靶向与砷和ATRA已被证明是在 早幼粒细胞白血病申请人Mark Murakami博士概述了5年职业发展计划， 成为转化白血病生物学的独立研究者。他召集了一群杰出的人 导师，顾问和合作者，他们的专业知识涵盖了从板凳到床边的翻译。 丹娜-法伯癌症研究所是完成他的科学和职业目标的理想环境， 其杰出的研究社区和培训独立的医生科学家的记录。

Project Summary/Abstract The BCR-ABL fusion defines the most common molecular subtype of acute lymphoblastic leukemia (BCR-ABL+ ALL) in adults but historically conferred a poor prognosis. Incorporation of tyrosine kinase inhibitors (TKIs) that bind the ABL catalytic domain into frontline regimens can induce complete remissions in >90% of patients, but nearly all will relapse, typically with resistance mutations that disfavor drug binding. This suggests that relapses remain addicted to ABL kinase activity. In contrast, novel type IV inhibitors bind the ABL myristate site and allosterically modulate BCR-ABL function. We hypothesize that the combination of catalytic and allosteric TKIs can prevent the emergence of cross-resistant clones and cure a subset of treatment-naïve BCR-ABL+ ALLs. We have established a diverse panel of 16 patient-derived xenograft (PDX) models of BCR-ABL+ ALL in which we can perform controlled and adequately powered pre-clinical trials to assess heterogeneity of response to in vivo combined blockade, develop predictive biomarkers, and establish models of acquired in vivo resistance. We have also developed an approach for characterizing therapeutic sensitivity in individual tumor cells that is amenable to minimal residual disease (MRD) specimens. Specifically, we use a microcantilever-based platform known as the suspended microchannel resonator (SMR) to assay drug sensitivity by measuring changes in the buoyant mass of individual cells exposed to targeted inhibitors ex vivo with femtogram-range sensitivity. These cells are then collected downstream of the SMR for single-cell RNA-Seq (scRNA-seq) to define transcriptional programs and cell states that modulate differential response to therapeutics. To test our hypothesis that combined ABL blockade has curative potential, and to meet the pressing need for rapid and robust approaches to characterize sensitivity within individual tumor specimens at MRD, we propose the following Specific Aims: (1) Test whether combined catalytic and allosteric BCR-ABL inhibition can cure some BCR-ABL+ ALL PDX models. Any models that relapse on combination therapy will be interrogated for mutational, transcriptional (both coding and non-coding RNA), and proteomic correlates of acquired in vivo resistance. (2) Define approaches to overcome therapeutic resistance directly within MRD in vivo. We will use the SMR to identify resistant subclones within MRD and apply scRNA-seq to define transcriptional programs and cell states mediating resistance for which bulk tumor assays would be insensitive. Defining patients who could be cured with combined ABL blockade would be transformative, just as combined PML-RAR targeting with arsenic and ATRA has proven to be in promyelocytic leukemias. The applicant Dr. Mark Murakami has outlined a 5-year career development plan to become an independent investigator in translational leukemia biology. He has assembled a distinguished group of mentors, advisors, and collaborators whose expertise spans the continuum of bench to bedside translation. The Dana-Farber Cancer Institute is the ideal environment for completion of his scientific and career goals given its outstanding research community and record of training independent physician-scientists.