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

描述（由申请人提供）：单核细胞衍生细胞的激活发生在大多数炎症部位，并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。现在已知活化的巨噬细胞是可以介导促炎（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）行为的异质细胞群体。已经表明，由M2细胞表达的细胞因子通过调节炎症反应来帮助调节M1行为。因此，假设M1和M2细胞之间的平衡在正常愈合反应中是重要的。最近，M1或M2细胞的活性增强与几种疾病状态有关，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的可塑性，包括激活的巨噬细胞的复极的证据。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解决上述疾病的基于细胞的治疗方法的关键工具。该提案的总体目标是开发M1和M2特异性肽，其可用作细胞标记物或靶向配体以将调节药物递送至活化的巨噬细胞亚群。这一目标将通过首先通过顺序阴性和阳性生物淘选从组合文库中鉴定M1-和M2-结合肽来实现。将合成潜在的M1和M2肽，并通过流式细胞术和共聚焦显微镜进行检测。然后将结合肽与量子点缀合以用作细胞特异性标记物，并在混合细胞群和组织切片中进行评估。此外，肽将与纳米颗粒缀合，并评估靶向递送至活化巨噬细胞亚群。最后，将在非人灵长类动物和人组织切片中评估这些肽对M1和M2细胞的跨种属识别。公共卫生相关性：活化的巨噬细胞亚群（M1和M2）的不平衡导致M1或M2细胞的活性升高或延长，已被假设为有助于疾病，如纤维化和动脉粥样硬化。这项工作的目标是开发M1和M2特异性肽，可用于识别，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解决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.