Unsaturated glycoceramide as novel platform for mucosal vaccine and drug delivery

描述（由申请人提供）：本提案的目的是测试我们最近关于上皮细胞中脂质运输的发现是否可以转化为临床应用。具体来说，我们将测试使用“短或不饱和”的神经酰胺为基础的脂质分子载体，以提供治疗肽或疫苗佐剂跨粘膜上皮屏障的效用。粘液表面代表了宿主组织与环境仅通过精细但高效的单层柱状上皮细胞分离的广阔区域，所述柱状上皮细胞通过蛋白质甚至小肽不可渗透的紧密连接连接。到目前为止，缺乏合理和有效的方法来绕过这一障碍，阻碍了大多数治疗性蛋白质的应用，口服或鼻腔给药和粘膜疫苗。在我们对霍乱毒素的糖脂受体神经节苷脂GM 1的生物学研究过程中，我们最近发现神经酰胺（脂质）结构域决定了GM 1在上皮细胞中的运输。当顶部施用时，含有“短”C12：0或“扭结链”不饱和C16：1脂肪酸（GM 1 short/unsat）的GM 1-神经酰胺进入共同/再循环内体。在这里，它们被分选用于运输到各种细胞内目的地，并进入到基底外侧细胞表面的“跨胞吞”途径。相反，具有完全饱和脂肪酸链（C16：0或更长）的GM 1-神经酰胺（GM 1 long/sat）被转运到晚期内体和溶酶体进行降解。在这个探索性项目中，我们将测试这一基本发现是否可以用于生物活性肽或蛋白质佐剂的经上皮递送，这两者都具有临床应用。我们将把含有不饱和“扭结”或饱和脂肪酸的GM 1分子连接到治疗性肽激素胰高血糖素样肽-1（GLP 1），其作用是调节血糖（Aim 1）;并连接到TLR 5激动剂FliC（沙门氏菌鞭毛蛋白），我们在这里使用它来模拟粘膜疫苗佐剂（Aim 2）。这些研究将测试一种用于跨粘膜表面转运治疗/疫苗分子的新平台。非常需要治疗性肽和蛋白质的非胃肠外递送。非常需要改进的粘膜疫苗策略来保护免受病原体的侵害，其中绝大多数病原体通过粘膜表面侵入。 公共卫生相关性：本申请的目的是开发一种允许口服或鼻腔施用治疗性蛋白质和疫苗的方法。通常，这些蛋白质不能被吸收，必须通过注射递送;严重限制了治疗应用。

DESCRIPTION (provided by applicant): The goal of this proposal is to test if our recent discovery on lipid trafficking in epithelial cells can be translated to clinical applications. Specifically, we will test the utility of using "short or unsaturated" ceramide-based lipids as molecular carriers to deliver therapeutic peptides or vaccine adjuvants across mucosal epithelial barriers. Mucosal surfaces represent vast areas where host tissues are separated from the environment only by a delicate but highly effective single layer of columnar epithelial cells, joined by tight junctions that are impermeable to proteins and even small peptides. So far, the lack of rational and efficient methods to circumvent this barrier has prevented the application of most therapeutic proteins for oral or nasal drug delivery and for mucosal vaccines. In the course of our studies on the biology of the glycolipid receptor for cholera toxin, ganglioside GM1, we recently discovered that the structure of the ceramide (lipid) domain dictates GM1 trafficking in epithelial cells. When applied apically, GM1-ceramides containing "short" C12:0 or "kinked chain" unsaturated C16:1 fatty acids (GM1short/unsat) enter the common/recycling endosome. Here, they are sorted for transport to various intracellular destinations and into the "transcytotic" pathway to the basolateral cell surface. In contrast, the GM1-ceramides with fully saturated fatty acid chains (C16:0 or longer) (GM1 long/sat) are instead transported to the late endosome and lysosome for degradation. In this exploratory project, we will test whether this basic discovery can be harnessed for transepithelial delivery of a bioactive peptide or protein adjuvant, both of which have clinical applications. We will link GM1 molecules containing unsaturated "kinked" or saturated fatty acids to the therapeutic peptide hormone glucagon-like peptide-1 (GLP1), which acts to regulate blood sugar (Aim 1); and to the TLR5-agonist FliC (Salmonella flagellin), which we use here to model a mucosal vaccine adjuvant (Aim 2). These studies will test a novel platform for transport of therapeutic/vaccine molecules across mucosal surfaces. There is great need for non-parental delivery of therapeutic peptides and proteins. Improved mucosal vaccine strategies are greatly needed for protection against pathogens, the vast majority of which invade via mucosal surfaces. PUBLIC HEALTH RELEVANCE: The goal of this application is to develop a way to allow for oral or nasal administration of therapeutic proteins and vaccines. Normally such proteins are not absorbed and must be delivered by injection; severely limiting therapeutic applications.