POLYMERIC IMMUNOGLOBULIN RECEPTOR TARGETING OF AIRWAYS

描述(申请人提供)：聚合免疫球蛋白受体(PlgR)在呼吸道上皮细胞和粘膜下腺的浆液细胞中表达。它的正常功能是与基底外侧表面的聚合免疫球蛋白结合，并将它们转移到根尖表面(管腔)，在那里它们被释放，仍然作为分泌性免疫球蛋白与受体的一部分(现在称为分泌成分，或SC)结合。它特别适合于通过上皮转移大量货物，它表达在呼吸道中的细胞类型中，囊性纤维化跨膜传导调节因子也在其中表达。因此，从治疗的角度来看，该受体有两个候选目的--第一，将基因从基底外侧途径输送到呼吸道上皮细胞；第二，将治疗药物从血液输送到管腔。已经制备了针对该受体的抗体，并应用了二次筛选，要求它们也与分泌型免疫球蛋白反应(即结合在天然配体使用的位置以外的位置)。这些抗体分为两类。其中一类人经历了快速的细胞穿透。另一种快速摄取，但以膜结合的核周小泡形式保留在细胞内。这一建议是基于这样一种假设，即“快速跨细胞”抗体将产生更好的跨细胞治疗性载体，而“细胞滞留”抗体将更有利于基因转移，因为它们将使其货物在细胞内保留更长时间，从而允许逃逸和核进入。单链抗体将从这两类抗体的代表中制备出来，并比较它们从致密DNA的非病毒基因转移载体中促进基因转移的能力，以及它们作为融合蛋白将潜在的治疗分子1-抗胰蛋白酶运送到气道腔的能力。测试系统将是两个极化的细胞系：在培养中转染人plgR的细胞；在气液界面生长的表达plgR的人呼吸道上皮细胞；以及由CC-10启动子驱动的转人plgR的小鼠，因此该受体仅在呼吸道上皮细胞中表达。如果研究成功，囊性纤维化和其他呼吸道疾病的新疗法就可以在这个基础上开发出来。

DESCRIPTION (provided by applicant): The polymeric immunoglobulin receptor (plgR) is expressed in airway epithelial cells and in the serous cells of the submucosal glands. Its normal function is to bind to polymeric immunoglobulins on the basolateral surface and transfer them to the apical surface (the lumen) where they are released, still bound to a portion of the receptor (now called secretory component, or SC) as secretory immunoglobulins. It is specifically adapted to transfer large amounts of cargo across the epithelium, and it is expressed in the cell types in the airway in which the cystic fibrosis transmembrane conductance regulator is also expressed. Thus, from the therapeutic perspective, this receptor is a candidate for two purposes - one, to deliver genes to airway epithelium from the basolateral approach, and two, to ferry therapeutics across the airway from the blood to the lumen. Antibodies have been prepared against this receptor, and a secondary screen requiring them to react with secretory immunoglobulin as well (that is, to bind at a site other than that used by the natural ligand) was applied. These antibodies fall into two categories. One class undergoes rapid transcytosis. The other undergoes rapid uptake, but is retained within the cell in membrane-bounded perinuclear vesicles. This proposal is based on the hypothesis that the "rapid transcytosis" antibodies will produce superior transcytotic therapeutic carriers, whereas the "cell retention" antibodies will be superior for gene transfer, for they will keep their cargo in the cells longer, allowing for escape and nuclear entry. Single chain Fvs will be prepared from representatives of both classes of antibodies and compared for their ability to enhance gene transfer from nonviral gene transfer vectors of compacted DNA, and for their ability to transport, as a fusion protein, a potential therapeutic molecule, 1-antitrypsin, into the airway lumen. Test systems will be both polarized cell lines in culture transfected with the human plgR; human airway epithelial cells grown at the air-liquid interface, which express plgR; and mice transgenic for the human plgR driven by the CC-10 promoter, so that the receptor is expressed only in airway epithelium. If the studies are successful, new therapeutics for cystic fibrosis and other airway diseases can be developed from this base.