Arginine Metabolism Regulates Myeloid Immune Suppression in Glioblastoma

项目总结 肿瘤相关髓系细胞(TAMCs)由肿瘤相关巨噬细胞和髓系细胞组成。 衍生的抑制性细胞构成了胶质瘤中细胞浸润物的大部分。TAMC具有强大的影响力 免疫抑制，是免疫治疗成功的主要障碍。TAMC高度直通 精氨酸酶-1(Arg-1)，一种分解代谢酶，被认为可以从肿瘤微环境中耗尽精氨酸。尽管 作为免疫抑制细胞的著名标记物，这种选择的代谢原因尚不清楚。 应用RNA-SEQ、批量代谢组学和免疫组织化学方法检测小鼠脑胶质瘤模型中TAMCs的表型 碳-13精氨酸通量显示，精氨酸分解代谢的两条独立途径在世代汇合。 鸟氨酸。鸟氨酸是从头生成多胺的先决条件底物，多胺是一组含氮的化合物。 丰富的代谢物，对生物学的所有领域都具有基础性的重要性。重要的是，我们发现- 多胺产生的限制步骤鸟氨酸脱羧酶1(ODC1)在胶质瘤中显著上调 侵袭TAMCs，提示从头产生的多胺对其功能很重要。因此， 这项提案的总体目标是确定精氨酸是如何被TAMCs分解为多胺的，并 确定抑制这一代谢途径是否可以加强对胶质瘤的免疫治疗。 有趣的是，我们发现精氨酸代谢的第二条未被研究的代谢途径，即脱氧核糖核酸。 在我们的胶质瘤模型中，TAMCs优先利用鸟氨酸产生新的肌酸。第一个目标 这个项目的目的是产生基因工程小鼠(GEM)来确定新生肌酸的作用 在小鼠胶质母细胞瘤模型中产生免疫抑制的代谢。这一目标将剖析肌酸 一代人被用来作为脑瘤生存的燃料，或者被用来促进免疫抑制。 我们的初步实验表明，抑制精氨酸代谢的产物，多胺， 可扰乱TAMC免疫抑制。这表明，多胺可能是 TAMCs在脑肿瘤中的作用因此，这项建议的第二个目的是探讨 脑胶质瘤中TAMCs的多胺代谢。我们将产生独特的有条件的基因敲除动物模型 探讨该通路在脑胶质瘤TAMCs免疫抑制功能中的作用。 这个项目的第三个目标是确定这些通路的靶向是否可以与标准的 对胶质瘤的护理以促进动物存活。首先，我们将测试一种血脑屏障渗透性抑制剂是否 肌酸激酶可用于阻断胶质瘤中的TAMCs。接下来，我们将使用公认的多胺抑制剂 在试图钝化TAMC介导的胶质瘤免疫抑制中的生成和多胺摄取。最后，我们 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 神经胶质瘤的免疫治疗。

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.