Novel Targeting Ligands for Sub-populations of monocyte-derived cells

描述（由申请人提供）：单核细胞源性细胞的激活发生在大多数炎症部位，通过清除异物和向其他免疫细胞发出信号，作为免疫系统的第一道防线。目前已知活化的巨噬细胞是一种异质性细胞群，可以介导促炎（经典活化的巨噬细胞，或M1细胞）或促愈合（替代活化的巨噬细胞，或M2细胞）行为。研究表明，M2细胞表达的细胞因子通过调节炎症反应来帮助调节M1的行为。因此，假设M1和M2细胞之间的平衡在正常愈合反应中很重要。最近，M1或M2细胞活性升高与多种疾病状态相关，包括纤维化、动脉粥样硬化和肿瘤进展。有趣的是，最近的研究表明，即使在分化的巨噬细胞中也存在可塑性，包括活化的巨噬细胞复极化的证据。因此，识别利用和指导单核细胞衍生细胞的特定亚群的生物工程技术是开发针对上述疾病的基于细胞的治疗方法的关键工具。本提案的总体目标是开发M1和m2特异性肽，这些肽可以用作细胞标记物或靶向配体，以向活化的巨噬细胞亚群递送调节药物。这一目标将首先通过顺序的阴性和阳性生物筛选从组合文库中鉴定M1和m2结合肽来实现。潜在的M1-和m2 -肽将被合成并通过流式细胞术和共聚焦显微镜检测。然后将结合肽与量子点结合，作为细胞特异性标记物，并在混合细胞群和组织切片中进行评估。此外，多肽将与纳米颗粒结合，并评估靶向递送到活化巨噬细胞亚群的能力。最后，这些肽对M1和M2细胞的跨物种识别将在非人灵长类动物和人类组织切片中进行评估。公共卫生相关性：活化巨噬细胞亚群（M1和M2）的不平衡导致M1或M2细胞活性升高或延长，这被认为是导致纤维化和动脉粥样硬化等疾病的原因。这项工作的目标是开发M1和m2特异性肽，可用于鉴定、纯化或靶向这些巨噬细胞亚群。识别、利用和指导单核细胞衍生细胞的特定亚群的生物工程技术是开发针对上述疾病的基于细胞的治疗方法的关键工具。

DESCRIPTION (provided by applicant): Activation of monocyte-derived cells occurs at most sites of inflammation and serves as a first line of defense in the immune system by removing foreign objects and by signaling to other immune cells. Activated macrophages are now known to be heterogeneous populations of cells that can mediate both pro-inflammatory (classically-activated macrophage, or M1 cells) or pro-healing (alternatively-activated macrophage, or M2 cells) behavior. It has been shown that cytokines expressed by M2 cells help to regulate M1 behavior by moderating the inflammatory response. Thus, the balance between M1 and M2 cells is hypothesized to be important in the normal healing response. Recently, heightened activity from M1 or M2 cells has been associated with several disease states, including fibrosis, atherosclerosis, and tumor progression. Interestingly, recent studies indicate plasticity even in differentiated macrophage cells, including evidence of repolarization of activated macrophage. Bioengineering technologies that identify harness and direct specific subpopulations of monocyte-derived cells are therefore critical tools in the development of cell-based therapy approaches addressing the aforementioned diseases. The overall goal of this proposal is to develop M1- and M2-specific peptides that can be used as cell markers or as targeting ligands to deliver modulating drugs to sub-populations of activated macrophages. This goal will be achieved by first identifying M1- and M2-binding peptides from combinatorial libraries by sequential negative and positive biopanning. Potential M1- and M2-peptides will be synthesized and tested by flow cytometry and confocal microscopy. Binding peptides will then be conjugated to quantum dots to serve as cell specific markers and assessed in mixed cell populations and in tissue sections. In addition, peptides will be conjugated to nanoparticles and evaluated for targeted delivery to sub-populations of activated macrophages. Finally, the cross-species recognition of M1 and M2 cells by these peptides will be assessed in non-human primate and human tissue sections. Public Health Relevance: An imbalance in subpopulations (M1 and M2) of activated macrophage cells resulting in heightened or prolonged activity from either M1 or M2 cells has been hypothesized to contribute toward diseases such as fibrosis and atherosclerosis. The goal of this work is to develop M1- and M2-specific peptides that can be used to identify, purify, or target these macrophage subpopulations. Bioengineering technologies that identify, harness and direct specific subpopulations of monocyte-derived cells are critical tools in the development of cell-based therapy approaches addressing the aforementioned diseases.