pH- SENSITIVE BIS-DETERGENTS FOR MACROMOLECULAR DELIVERY

用于大分子输送的 pH 敏感双去污剂

• 批准号: 6776804
• 负责人: MOO J CHO
• 金额: $ 20.24万
• 依托单位: UNIV OF NORTH CAROLINA CHAPEL HILL
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-05-01 至 2006-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6776804
• 关键词: acidity /alkalinity; calorimetry; chemical stability; chemical structure function; chemical synthesis; confocal scanning microscopy; crosslink; cytoplasm; cytotoxicity; detergents; drug delivery systems; electron microscopy; endocytosis; fluorescence spectrometry; hydrolysis; lipid bilayer membrane; lipid metabolism; liposomes; macromolecule; membrane permeability; micelles; protonation; technology /technique development; tissue /cell culture

DESCRIPTION (provided by applicant): Macromolecules, being unable to penetrate cell membranes readily, enter the cell via endocytosis. The contents of the endosome are eventually routed to enzyme-rich lysosomes and in all likelihood degraded. If pharmacological activity of a therapeutic macromolecule is realized only when it enters the cytosol, then a mechanism must be built within a delivery system by which these agents escape from the endosome into the cytosol. The endosomal pH drops to as low as 5.0 as the endosome matures. This provides a unique opportunity in devising such a strategy for cytosolic delivery. We have designed a series of acid-sensitive lipids that can be readily incorporated into liposomal drug carrier systems. Upon entering the cell and encountering the acidic environment of the endosome, the acid sensitive lipid breaks down to release potent detergents. Critical hypotheses being tested at this point are:(1) presence of these detergents initiates liposome-to-micelle transition that triggers catastrophic release of entrapped macromolecules within endosomes and (2) micelles then induce permeabilization of endosomal membrane to promote cytosolic transfer of macromolecules. However, it is impossible to test these hypotheses without incorporating a critical concentration of detergent into liposomal bilayers. This concentration, above which vesicle-to-micelle transition can take place is unattainably high simply because at high detergent concentrations, liposomes cannot be made. In the present study, we propose to synthesize "bis-detergents", inert double-tailed lipids that can be embedded at high concentrations in liposomal bilayers and break down only at low pH into "membrane-active" detergents. When realized, the technology developed from this study will have profound implication not only in basic biomedical research entailing cytosolic delivery of macromolecules in vitro but also in a variety of therapeutic applications in vivo, ranging from delivery of antigenic proteins for induction of cell-mediated immunity to nucleic acids for gene regulation.

描述（申请人提供）：大分子无法轻易穿透细胞膜，通过内吞作用进入细胞。内体的内容物最终被输送至富含酶的溶酶体并很可能被降解。如果治疗性大分子的药理活性只有在进入细胞质时才能实现，那么必须在递送系统内建立一种机制，通过该机制这些药物从内体逃逸到细胞质中。随着内体成熟，内体 pH 值降至 5.0。这为设计这种胞质递送策略提供了独特的机会。 我们设计了一系列酸敏感脂质，可以很容易地掺入脂质体药物载体系统中。进入细胞并遇到内体的酸性环境后，酸敏感脂质会分解并释放出有效的去污剂。此时正在测试的关键假设是：（1）这些去污剂的存在引发脂质体到胶束的转变，从而引发内体内截留的大分子的灾难性释放；（2）胶束然后诱导内体膜的透化，以促进大分子的胞质转移。然而，如果不将临界浓度的去垢剂掺入脂质体双层中，就不可能检验这些假设。这个浓度（高于该浓度可以发生囊泡到胶束的转变）是无法达到的，因为在高洗涤剂浓度下，无法制备脂质体。在本研究中，我们建议合成“双去污剂”，即惰性双尾脂质，它可以高浓度嵌入脂质体双层中，并且仅在低pH值下分解成“膜活性”去污剂。 一旦实现，这项研究开发的技术不仅将对体外大分子胞质递送的基础生物医学研究产生深远的影响，而且还将对体内的各种治疗应用产生深远的影响，包括递送抗原蛋白以诱导细胞介导的免疫转化为核酸以进行基因调控。