TARGETING TOXINS WITH ACID-TRIGGERED HYBRID ANTIBODIES

用酸触发的混合抗体靶向毒素

• 批准号: 3194165
• 负责人: VICTOR A RASO
• 金额: $ 21.33万
• 依托单位: BOSTON BIOMEDICAL RESEARCH INSTITUTE
• 依托单位国家: 美国
• 财政年份: 1989
• 资助国家: 美国
• 起止时间: 1989-12-01 至 1994-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3194165
• 关键词: Pseudomonas; acidity /alkalinity; athymic mouse; bacterial toxins; conformation; cytoplasm; diphtheria toxin; disease /disorder model; drug design /synthesis /production; drug vehicle; exotoxins; guinea pigs; human subject; hybrid antibody; hybridomas; ligands; mesothelioma; monoclonal antibody; multiple myeloma; neoplastic cell; nonhuman therapy evaluation; plant poisons; protein sequence; thiols; toxin; transferrin receptor

The long-term objective of this project is to harness the lethal potency of plant and bacterial toxins for beneficial use in medicine. To accomplish this goal we have designed a novel mechanism which combines the accurate targeting of antibody-bound toxin to cells with a precise means of triggering its release and full activity once it has reached its intracellular destination. Certain toxic proteins such as diphtheria toxin and Pseudomonas exotoxin, undergo an acid (pH 4-5) dependent conformational change within endosomal compartments inside the cell. This denaturation is critical to the passage of toxin out of these vesicles and into the cytosol where it acts to kill the cell. The aim of this project is to generate murine and human monoclonal antibodies which will tightly bind toxin at physiological pH levels (pH 6-8) but efficiently release this toxin when it undergoes its conformational transition at pH 4-5. Such antibodies will be covalently linked to a second cell-reactive antibody or receptor ligand to form a hybrid molecule with dual specificity. This hybrid will carry and attach antibody-bound toxin to the surface of only those cells bearing chosen target sites. Toxin will be released in its lethal form when the hybrid-toxin complex is taken into cells and is exposed to the low pH in endosomes. This approach is very flexible since antibodies or ligands to virtually any membrane site can be easily coupled with the toxin-bearing antibody to form a variety of highly specific cytotoxic agents. The binding of hybrid-antibody to toxin would also block the indiscriminate attachment of toxin to cells and prevent the neutralization of hybrid-delivered toxin by any circulating endogenous antibodies. The hybrid-antibody system will be especially useful for delivering toxin fragments or genetically modified toxins with improved characteristics. A monoclonal antibody to the human transferrin receptor has been coupled to an acid-releasable monoclonal antibody directed against diphtheria toxin. This hybrid will serve to evaluate the release strategy since its receptor internalization pathway involves entry into acidified cellular compartments. Ensuing therapeutic studies with this reagent will allow us to judge its in vivo effectiveness. An athymic mouse model of early and advanced stage human malignant mesothelioma will be used to test tumoricidal action and to monitor the biodistribution of hybrid-delivered diphtheria toxin and toxin analogs.

该项目的长期目标是利用致命的效力 植物和细菌毒素可用于医学。 完成 这个目标我们设计了一种新颖的机制，该机制结合了准确 靶向抗体结合的毒素对细胞的精确均值 触发其释放和完整的活动后 细胞内目的地。 某些有毒蛋白，例如白喉毒素和假单胞菌毒素， 在内体内进行酸（pH 4-5）依赖构象变化 电池内的隔室。 这种变性对通过 这些囊泡的毒素和杀死的毒素的作用 细胞。 该项目的目的是产生鼠和人类 单克隆抗体将在生理pH下紧密结合毒素 水平（pH 6-8），但在其经历时有效释放此毒素 pH 4-5处的构象过渡。 这样的抗体将是共价的 与第二个细胞反应性抗体或受体配体链接以形成A 具有双重特异性的杂化分子。 该混合动力车将携带并附加 抗体结合的毒素仅适用于那些带有那些带有的细胞的表面 目标站点。 当毒素将以致命的形式释放 杂交毒素复合物被吸收到细胞中，并暴露于低pH中 内体。 这种方法非常灵活，因为抗体或配体几乎没有任何 膜位点很容易与含毒素抗体结合以形成 多种高度特异性的细胞毒性剂。 结合 毒素的杂交抗体也将阻止不加区分的附件 毒素向细胞毒素，并防止通过 任何循环的内源性抗体。 混合抗体系统将是 对于输送毒素片段或转基因修饰特别有用 具有改善特征的毒素。 对人转铁蛋白受体的单克隆抗体已与 针对白喉毒素的可蛋白溶质单克隆抗体。 由于其受体，该混合动力车将用于评估释放策略 内部化途径涉及进入酸化的细胞 车厢。 随后使用该试剂的治疗研究将使我们 判断其体内有效性。 早期和 晚期人类恶性间皮瘤将用于测试 肿瘤作用并监测混合分配的生物分布 白喉毒素和毒素类似物。