Cancer Therapeutics that Anchor Proteins to Membranes

描述（由申请人提供）：多药耐药（MDR）癌症仍然是治愈性癌症化疗的主要障碍。MDR癌细胞与典型肿瘤细胞的不同之处在于显著上调了几种因子的产生，包括药物转运蛋白P-糖蛋白、胆固醇结合蛋白小窝蛋白和细胞质膜的脂筏微区组分。这些筏结构域富含胆固醇和鞘脂，并在信号转导过程中发挥关键作用。MDR癌症的质膜的独特组成可能使选择性化疗靶向这些癌症。在最后一个资助周期中，合成了新的小分子，其包含与蛋白质配体如生物素共价连接的胆固醇胺。用合成的生物素-胆固醇胺配体（配体#1）和蛋白质链霉亲和素（SA）处理癌细胞系可有效地将SA靶向脂筏，导致这种蛋白质-配体复合物的快速网格蛋白介导的内吞作用。这种新的系统模拟霍乱毒素对细胞的渗透，霍乱毒素结合脂筏中的神经节苷脂GM 1。该项目基于以下假设：配体#1将调节与癌细胞系中的内体激活的毒素柔红霉素和外毒素相关的SA的内吞作用。由于配体#1结合脂筏，将在体外和体内在鼠癌症模型中研究SA连接的毒素向富含脂筏的MDR癌细胞的选择性递送。还将评估配体#1在增强融合至神经肽Y的Satoxins的内吞作用方面的有效性，所述神经肽Y靶向神经母细胞瘤细胞上的特异性受体。这种通过靶向具有小分子的脂筏来增强表面受体的内吞作用的新方法可以解决非内化肿瘤抗原的免疫治疗中的主要问题。还将测试配体调节的SA向抗原呈递细胞（APC）的递送将刺激免疫应答的假设。新型免疫刺激剂将通过将SA与卵清蛋白抗原融合、用配体#1调节APC中的内吞作用以及分析T细胞活化来研究。这种方法可以在分子水平上控制免疫刺激，并为疫苗开发提供新的工具。还将研究通过配体#1将细胞内抗生物素蛋白融合蛋白募集至质膜以条件性调节细胞生长和死亡

DESCRIPTION (provided by applicant): Multidrug resistant (MDR) cancer remains the primary impediment to curative cancer chemotherapy. MDR cancer cells differ from typical tumor cells by dramatically upregulating production of several factors including the drug transporter P-glycoprotein, the cholesterol binding protein caveolin, and components of lipid raft microdomains of cellular plasma membranes. These raft domains are enriched in cholesterol and sphingolipids and play key roles in signal transduction processes. The distinct composition of plasma membranes of MDR cancers may enable selective chemotherapy targeting these cancers. During the last grant cycle, novel small molecules were synthesized that comprise cholesterylamine covalently linked to protein ligands such as biotin. These compounds bind lipid rafts in plasma membranes of cancer cells.Treatment of cancer cell lines with a synthetic biotin-cholesterylamine ligand (ligand #1) and the protein Streptavidin (SA) efficiently targets SA to lipid rafts, resulting in rapid clathrin-mediated endocytosis of this protein-ligand complex. This novel system mimics penetration of cells by Cholera toxin, which binds ganglioside GM1 in lipid rafts. This project is based on the hypothesis that ligand #1 will regulate endocytosis of SA linked to endosome-activated toxins daunorubicin and exotoxin in cancer cell lines. Since ligand #1 binds lipid rafts, selective delivery of SA-linked toxins to lipid raft-rich MDR cancer cells will be investigated in vitro and in vivo in murine cancer models. The effectiveness of ligand #1 at enhancing endocytosis of Satoxins fused to neuropeptide Y, which targets specific receptors on neuroblastoma cells, will also be evaluated. This novel approach directed at enhancing endocytosis of surface receptors by targeting to lipid rafts with small molecules could address the major problem in immunotherapy of non-internalized tumor antigens. The hypothesis that ligand-regulated delivery of SA to antigen presenting cells (APCs) will stimulate immune responses will also be tested. Novel immunostimulants will be investigated by fusing SA to the ovalbumin antigen, regulating endocytosis in APCs with ligand #1, and analyzing T-cell activation. This approach could control immunostimulation at the molecular level and yield novel tools for vaccine development. Recruitment of intracellular avidin fusion proteins to plasma membranes by ligand #1 will also be studied in an effort to conditionally regulate cellular growth and death