pH- SENSITIVE BIS-DETERGENTS FOR MACROMOLECULAR DELIVERY

性状（由申请方提供）：大分子不能轻易穿透细胞膜，通过胞吞作用进入细胞。内体的内容物最终被路由到富含酶的溶酶体，并且很可能被降解。如果治疗性大分子的药理学活性仅在其进入胞质溶胶时实现，则必须在递送系统内建立机制，通过该机制这些药剂从内体逃逸到胞质溶胶中。随着内体成熟，内体pH下降至低至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，可以很容易地纳入脂质体药物载体系统。一旦进入细胞并遇到内体的酸性环境，酸敏感性脂质就会分解以释放有效的去污剂。在这一点上测试的关键假设是：（1）这些去污剂的存在引发脂质体到胶束的转变，引发内体内捕获的大分子的灾难性释放，以及（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污剂掺入脂质体双层中，就不可能检验这些假设。高于该浓度，囊泡至胶束的转变可以发生，该浓度是无法达到的高浓度，这仅仅是因为在高去污剂浓度下，不能制备脂质体。在本研究中，我们建议合成“双洗涤剂”，惰性双尾脂质，可以嵌入在高浓度的脂质体双层和分解，只有在低pH值成“膜活性”洗涤剂。 当实现时，从本研究开发的技术将具有深远的影响，不仅在基础生物医学研究中需要在体外的大分子的胞质递送，而且在体内的各种治疗应用中，从用于诱导细胞介导的免疫的抗原蛋白的递送到用于基因调控的核酸。

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.