Trajectory and Architecture of Microenvironment-Mediated Resistance in AML

项目摘要/摘要：项目2 急性髓系白血病(AML)患者的预后在过去40年中几乎没有改善。这在很大程度上是因为 与获得性耐药和难治性疾病的发展有关。因此，迫切需要新的 在残留的AML细胞或难治性克隆细胞引发疾病复发之前将其作为目标的策略。肿瘤 微环境是恶性进展和治疗耐药发生的关键因素。双性恋- 癌细胞和微环境细胞之间的定向通讯比 最初感觉到的。因此，减少复发的一个策略是针对由生长因子介导的信号转导 由骨髓微环境中的基质或免疫细胞分泌，在 促进白血病细胞存活、发展抗药性和免疫逃避。因此，长久以来- 该项目的学期目标是解开事件和细胞异质性的全面网络 以上下文相关的方式促进获得性耐药性，以确定新的治疗方法。作为一部分 在我们之前的U54 DRSN资助中，基于组学的分析发现，350名原发AML患者样本中 分泌的细胞因子和免疫因子可能与耐药有关。因此，我们假设 在AML中，骨髓小生境的重新编程调节药物反应并驱动获得性耐药。 对这些机制的全面了解将导致新的联合疗法的确定。 我们将集中在AML的5个基本治疗靶点：BCL2，Flt3，JAK2，MEK，和表观遗传途径和 将在以下三个很好地整合的目标下解决这一假设。在目标1中，我们将绘制微环境图 单细胞治疗急性髓系白血病药物反应特征的信号和肿瘤串扰机制 骨髓生态位背景下的基因表达和表观遗传学分析。肿瘤微环境交叉- 将在目标2中进一步探讨Talk，重点是单核细胞和基质细胞以及在NK和T细胞的背景下 用共培养平台进行CRISPR/Cas筛选，获得AIM 3细胞。已确定的目标/路径将 被验证以充分确定获得性耐药的微环境机制。这些结果将建立 治疗应激反应中信号串扰的预测模型和确定联合治疗 克服获得性抵抗力，特别是在微环境方面。我们将测试这些措施的效果 应用多参数流技术研究抑制剂对细胞活性、细胞异质性、分化和靶向抑制的影响 细胞计数和免疫荧光分析。我们将把这些调查结果与项目1结合起来，并确定优先顺序 将在项目3中测试目标的翻译影响。总体而言，阐明 微环境驱动的AML细胞药物反应和存活信号将有助于识别新的易驯化的 克服急性髓细胞白血病获得性耐药的联合治疗的靶点。

PROJECT SUMMARY/ABSTRACT: Project 2 Outcomes for acute myeloid leukemia (AML) patients have improved little over four decades. This is largely due to the development of acquired drug resistance and refractory disease. Thus, there is an urgent need for new strategies to target residual AML cells or refractory clones before they trigger disease relapse. The tumor microenvironment is a key factor in the development of malignant progression and therapy resistance. The bi- directional communication between cancer cells and microenvironmental cells is much more complex than initially perceived. Thus, one strategy to reduce relapse is to target the signaling mediated by growth factors secreted by stromal or immune cells within the bone marrow microenvironment that play a critical role in promoting leukemia cell survival, development of drug resistance, and immune evasion. Therefore, the long- term goal of this project is to deconvolute the comprehensive network of events and cellular heterogeneity that contribute to acquired resistance in a context dependent manner to identify new therapeutic approaches. As part of our prior U54 DRSN funding, omics-based analysis of 350 primary AML patient samples found that multiple secreted cytokines and immune factors may contribute to drug resistance. Therefore, we hypothesize that reprogramming of the bone marrow niche modulates drug response and drives acquired resistance in AML. Comprehensive understanding of these mechanisms will lead to the identification of new combination therapies. We will focus on 5 essential therapeutic targets in AML: BCL2, FLT3, JAK2, MEK, and epigenetic pathways and will address this hypothesis following three well integrated aims. In Aim 1, we will map microenvironmental signatures and tumor cross-talk mechanisms against drug response features of AML by performing single cell gene expression and epigenetic analyses in the context of bone marrow niche. Tumor microenvironment cross- talk will be further explored in Aim 2 focusing on monocytes and stromal cells and in the context of NK and T cells in Aim 3 by performing CRISPR/Cas screening with co-culture platforms. Identified targets/pathways will be validated to fully define microenvironmental mechanisms of acquired drug resistance. These results will build predictive models of signaling crosstalk in response to therapeutic stress and identify combination therapies to overcome acquired resistance, particularly in the context of the microenvironment. We will test the effect of these inhibitors on cell viability, cellular heterogeneity, differentiation, and target inhibition using multi-parametric flow cytometry and immunofluorescence analysis. We will integrate these findings with Project 1 and prioritized targets will be tested for their translational impact in Project 3. Overall, elucidating the influence of microenvironment-driven signaling on drug response and survival of AML cells will help identify novel tractable targets for combination therapies to overcome acquired drug resistance in AML.