Combining Bacterial Glycosylation Tools and Nanotechnology to Optimize Siglec Ligands for the Treatment of Allergy and Anaphylaxis

项目总结 在美国，过敏是慢性疾病的第六大常见原因，但有效的治疗方案 仍然是有限的。最近发现的细胞特异性抑制性免疫受体，如唾液酸结合 免疫球蛋白型凝集素(Siglec)受体是抑制这些有害物质的有希望的治疗靶点。 免疫反应，但一直很难靶向，因为它们专门结合含有唾液酸的 碳水化合物，或多聚糖。使用两种新开发的工具：细菌多糖构建平台 构建可修饰的Siglec受体葡聚糖配体和可变化的葡聚糖展示纳米凝胶 多糖密度，我们的目标是产生优化的Siglec受体配体，用于治疗 治疗过敏和预防过敏反应的候选人。我们将使用Siglec 3(CD33)作为模型 为了验证这种方法，因为它在肥大细胞和嗜碱性粒细胞上都有表达，两种细胞类型与 过敏和过敏反应。在第一个目标中，我们将产生一个化学修饰的葡聚糖变异体文库 是候选的Siglec受体配体，用于评估与ELISA的高亲和力相互作用。然后我们将装载很高的货物 不同密度的纳米凝胶上的亲和葡聚糖配体，适用于致敏的原代人皮肤来源的肥大 并评估肥大细胞活性的指标，包括颗粒成分、表面标志物和 细胞因子的释放。在第二个目标中，我们将用一个新的工具来补充这个领域，以描述 Siglec受体。将糖基化机制在工程大肠杆菌中的基因组掺入将产生 产生可修饰唾液酸糖蛋白的稳定菌株，这些唾液酸糖蛋白是候选Siglec配体。最终，使用 细菌糖工程，我们将开发一个可定制的平台来生成Siglec受体配体， 可以很容易地移植到其他实验室，并广泛应用于其他凝集素的表征。一旦完成， 建议的目标不仅将产生能够高亲和力Siglec 3(CD33)结合的治疗候选， 还提供了一种新的方法和工具，可用于确定其他Siglec的候选治疗方案 受体和凝集素。这个项目跨越了糖生物学、细胞生物学、纳米材料，并使用了 疾病。因此，它将提供丰富的培训经验，并得到研究导师的支持和协同 来自西北大学、合成生物学中心、医学中心的专业知识和充足的支持 科学家培训计划和跨学科生物科学研究生计划。除了……之外 提供强大的训练体验，这也将提高靶向治疗的潜力 Siglec受体用于治疗过敏和预防过敏反应，以及其他相关疾病。

PROJECT SUMMARY Allergies are the 6th most common cause of chronic illness in the US, but effective treatment options remain limited. Recently discovered, cell-specific, inhibitory immune receptors, such as sialic acid-binding immunoglobulin-type lectin (Siglec) receptors, are promising therapeutic targets to suppress these unwanted immune responses, but have been difficult to target because they specifically bind sialic-acid containing carbohydrates, or glycans. Using two newly developed tools: a bacterial glycan-building platform for constructing modifiable Siglec receptor glycan ligands and a glycan-displaying nanogel capable of varying glycan density, we aim to generate optimized Siglec receptor ligands towards the development of therapeutic candidates for the treatment of allergy and prevention of anaphylaxis. We will use Siglec 3 (CD33) as a model to validate this approach since it is expressed on both mast cells and basophils, two cell types implicated in allergy and anaphylaxis. In the first aim, we will generate a library of chemically modified glycan variants that are candidate Siglec receptor ligands to assess for high affinity interactions with ELISA. We will then load high affinity glycan ligands on nanogels at varying densities, apply to sensitized primary human skin-derived mast cells, and assess metrics of mast cell activity including release of granule components, surface markers, and cytokine release. In the second aim, we will supplement the field with a new tool for the characterization of Siglec receptors. Genomic incorporation of glycosylation machinery in engineered Escherichia coli will yield a stable strain for generating modifiable sialoglycoproteins that are candidate Siglec ligands. Ultimately, using bacterial glycoengineering, we will develop a customizable platform for generating Siglec receptor ligands that can be readily ported to other labs and broadly applied for the characterization of other lectins. Once complete, the proposed aims will not only yield therapeutic candidates capable of high affinity Siglec 3 (CD33) binding, but also a new approach and tool that can be applied to identify therapeutic candidates for other Siglec receptors and lectins. This project spans glycobiology, cell biology, nanomaterials, and using allergy models of disease. As such, it will provide a rich training experience bolstered by research mentors with synergistic expertise and ample support from Northwestern University, the Center for Synthetic Biology, the Medical Scientist Training Program, and the Interdisciplinary Biological Sciences Graduate Program. In addition to providing a robust training experience, this will work will also advance the therapeutic potential of targeting Siglec receptors for the treatment of allergy and prevention of anaphylaxis, among other relevant diseases.