Proj 3 - Targeting the Pro-tumorigenic Microenvironment

摘要/摘要 项目4的总体目标是通过以下方式发现和开发治疗耐药的外部机制 关注肿瘤相关巨噬细胞和肿瘤相关成纤维细胞的作用 肿瘤微环境(TME)中的TAFs。我们的主要假设是TAMS和TAFs合作 创造良好的致癌环境，最终导致治疗性疾病的出现 NB中的抗性和免疫逃逸。我们还假设，随着肿瘤的治疗，TME在其 组成和功能变得越来越有利于治疗耐药。这一假设是有根据的 根据我们小组发表的和初步的数据显示，TAMs和TAFs大量存在 在确诊时与复发风险高和极差相关的炎症性亚型NB中 预后。我们也有证据表明，当TAMs和TAFs暴露在肿瘤细胞中时，会刺激它们的 促肿瘤细胞因子旁分泌的增殖、存活和耐药 激活肿瘤细胞信号通路的趋化因子，如STAT3和ERK。我们的项目有三个目标。 目标1，将检查TAM和TAFs之间的合作机制，检验假设在 MYCN扩增的不产生化学诱导剂CCl-2/MCP-1的肿瘤，TAFs是 这种趋化因子。我们还将研究在共培养中产生的细胞因子和趋化因子的贡献。 TAMs、TAFs和Nb细胞及其在Nb细胞中激活的信号通路导致 扩散和生存。目标2，将检查TME环境中继发于化疗的变化 同基因小鼠NB模型(项目2)，并在通过Core获得的患者肿瘤样本中验证数据 B.通过检测长期暴露于/三苯氧胺和三氟甲烷的NB细胞的转录组的变化 他们潜在的表观遗传起源(通过项目3)，Aim 2还将识别脆弱性，以防止对 化疗或靶向治疗(项目1)。目标3，然后将这些发现转化为临床前 肿瘤模型。我们将测试最有希望的靶向TAFs、TAMs或 与化疗或免疫治疗相结合，它们在肿瘤细胞中激活的途径(项目5)， 在免疫缺陷小鼠和小鼠细胞系中使用人NB系和患者来源的异种移植 在具有免疫能力的小鼠(项目2)中，最有效的药物(S)将被建议用于早期阶段 NANT的临床试验(核心B)。因此，项目4对本项目的总体目标做出了独特的贡献 PPG通过其对TME和导致治疗的非自治机制的关注 抵抗力和免疫逃逸。

SUMMARY/ABSTRACT The overall objective of Project 4 is to discover and exploit extrinsic mechanisms of therapy resistance by focusing on the contribution of tumor-associated macrophages (TAMs) and tumor-associated fibroblasts (TAFs) in the tumor microenvironment (TME). Our overarching hypothesis is that TAMs and TAFs cooperate in creating a favorable tumorigenic environment that ultimately leads to the emergence of therapeutic resistance and immune escape in NB. We also postulate that as tumors are treated, the TME is altered in its composition and function to become increasingly favorable to therapeutic resistance. This hypothesis is based on published and preliminary data from our group demonstrating that TAMs and TAFs are abundantly present in an inflammatory subtype of NB at diagnosis associated with a high risk of recurrence and extremely poor prognosis. We also have evidence that TAMs and TAFs when exposed to tumor cells stimulate their proliferation, survival and drug-resistance via the paracrine production of pro-tumorigenic cytokines and chemokines that activate in tumor cells signaling pathways such as STAT3 and ERK. Our project has 3 aims. Aim 1, will examine mechanisms of cooperation between TAMs and TAFs, testing the hypothesis that in MYCN amplified tumors that do not produce the TAM chemoattractant CCL-2/MCP-1, TAFs are a source of this chemokine. We will also examine the contribution of cytokines and chemokines generated in co-culture of TAMs, TAFs and NB cells and the signaling pathways they activate in NB cells leading to increased proliferation and survival. Aim 2, will examine changes in the TME landscape secondary to chemotherapy in syngeneic murine NB models (with Project 2) and validate the data in patient tumor samples obtained via Core B. By examining changes in the transcriptome that occur in NB cells chronically exposed to TAM/TAF and their potential epigenetic origin (with Project 3), aim 2 will also identify vulnerabilities to prevent resistance to chemotherapy or targeted therapy (with Project 1). Aim 3, will then translate these discoveries in pre-clinical tumor models. We will test the therapeutic efficacy of the most promising agents targeting TAFs, TAMs, or pathways they activate in tumor cells in combination with chemotherapy or immunotherapy (with Project 5), using human NB lines and patient-derived xenotransplants in immunodeficient mice as well as murine cell lines in immunocompetent mice (with Project 2), The most effective agent(s) will then be proposed for early phase clinical trials to the NANT (Core B). Thus Project 4 brings a unique contribution to the overall objective of this PPG through its focus on the TME and on non-autonomous mechanisms leading towards therapeutic resistance and immune escape.