Arginine Metabolism Regulates Myeloid Immune Suppression in Glioblastoma

项目摘要 肿瘤相关髓样细胞（TAMCs），由肿瘤相关巨噬细胞和髓样-巨噬细胞组成。 衍生的抑制细胞构成胶质瘤中的大部分细胞浸润。TAMCs可以有效地 免疫抑制剂，并代表成功免疫治疗的主要障碍。TAMCs高表达 精氨酸酶-1（Arg-1），一种被认为从肿瘤微环境中消耗精氨酸的分解代谢酶。尽管 作为免疫抑制细胞的公知标志物，这种选择的代谢原因尚不清楚。 使用RNA-seq、整体代谢组学和生物信息学检查鼠神经胶质瘤模型中的TAMC表型 碳-13精氨酸通量揭示了精氨酸催化剂的两个独立途径在生成 鸟氨酸。鸟氨酸是从头生成多胺的必要底物，多胺是一组氮- 丰富的代谢产物，对生物学的所有领域都具有重要的基础性意义。更重要的是，我们发现- 多胺生成的限制性步骤，鸟氨酸脱羧酶1（ODC 1），被胶质瘤显著上调 浸润TAMCS，表明从头多胺的产生对其功能很重要。因此 本提案的总体目标是确定精氨酸如何被TAMC分解代谢为多胺， 确定抑制这种代谢途径是否可以增强神经胶质瘤的免疫治疗。 有趣的是，我们发现精氨酸代谢的第二个未研究的代谢途径，即去- 在我们的神经胶质瘤模型中，由鸟氨酸重新产生肌酸被TAMC优先利用。第一个目标 该项目的目的是产生基因工程小鼠（GEM），以确定从头肌酸的作用， 在胶质母细胞瘤小鼠模型中，代谢产生免疫抑制。这一目标将解剖，如果肌酸 生成被用作在脑肿瘤中生存的燃料，或被用于促进免疫抑制。 我们的初步实验表明，抑制精氨酸代谢产物多胺， 可以扰乱TAMC的免疫抑制。这表明，多胺可能是关键的代谢产物， TAMCs在脑肿瘤中的功能。因此，本建议的第二个目的是探讨以下方面的重要性： 胶质瘤中TAMCs的多胺代谢。我们将产生独特的条件性基因敲除动物模型， 探讨该通路在胶质瘤中TAMCs免疫抑制功能中的重要性。 该项目的第三个目的是确定这些途径的靶向是否可以与标准的 来提高动物的存活率。首先，我们将测试血脑屏障渗透性抑制剂是否 肌酸激酶可用于阻止胶质瘤中的TAMC。接下来，我们将使用成熟的多胺抑制剂 产生和多胺摄取，试图钝化胶质瘤中TAMC介导的免疫抑制。最后我们 我将服用上述临床抑制剂，并确定它们是否可用于增强检查点 胶质瘤免疫治疗

PROJECT SUMMARY Tumor-associated myeloid cells (TAMCs), which consist of tumor associated macrophages and myeloid- derived suppressor cells, make up a majority of cellular infiltrates in glioma. TAMCs are potently immunosuppressive, and represent a major barrier to successful immunotherapy. TAMCs highly express arginase-1 (Arg-1), a catabolic enzyme thought to deplete arginine from the tumor microenvironment. Despite being a well-known marker of immunosuppressive cells, the metabolic reasons for this choice are not clear. Examination of TAMCs phenotype in murine glioma models using RNA-seq, bulk metabolomics, and carbon-13 arginine flux revealed that two separate pathways of arginine catabolism converge on the generation of ornithine. Ornithine is the prerequisite substrate for the de-novo generation of polyamines, a group of nitrogen- rich metabolites with foundational importance to all domains of biology. Importantly, we found that the rate- limiting step of polyamine generation, ornithine decarboxylase 1 (ODC1), is dramatically upregulated by glioma infiltrating TAMCS, suggesting de-novo polyamine generation is important for their function. Therefore, the overall goal of this proposal is to determine how arginine is catabolized into polyamines by TAMCs, and to determine if inhibition of this metabolic pathway can enhance immunotherapy for glioma. Interestingly, we discovered that a second, unstudied metabolic pathway of arginine metabolism, the de- novo generation of creatine from ornithine, is preferentially utilized by TAMCs in our glioma models. The first aim of this project is to generate genetically engineered mice (GEM) to determine the role of the de-novo creatine metabolism in generating immunosuppression in mouse models of glioblastoma. This aim will dissect if creatine generation is used as a fuel to survive in brain tumors, or is being used to promote immunosuppression. Our preliminary experiments suggest that inhibition of the products of arginine metabolism, polyamines, can perturb TAMC immune suppression. This suggests that polyamines may be critical metabolites for the functions of TAMCs in brain tumors. Therefore, the second aim of this proposal is to probe the importance of polyamine metabolism by TAMCs in glioma. We will generate unique conditional knockout animal models to probe the importance of this pathway in the immunosuppressive functions of TAMCs in glioma. The third aim of this project is to determine if targeting of these pathways can be combined with standard of care for glioma to promote animal survival. First, we will test if a blood-brain-barrier permeable inhibitor of creatine kinase can be used to stymie TAMCs in glioma. Next, we will use well established inhibitors of polyamine generation and polyamine uptake in attempts to blunt TAMC mediated immunosuppression in glioma. Lastly, we will take the clinical inhibitors described above, and determine if they can be used to potentiate checkpoint immunotherapy for glioma.