Multifunctional lipids based on a molecular switch for the improved delivery of nucleic acids

50年前，当DNA的结构刚刚被发现时，谁能想到将基因用作药物是可能的？由于整个科学界的协同努力，这个概念已经从长凳上走到了床边，现在市场上有两种基因产品。然而，这些新疗法仍处于初级阶段，需要优化。通过使用精心设计的载体来提高它们的效率是可能的，这些载体能够携带遗传物质穿过它们需要跨越的每一个生物屏障。特别是，进入细胞是一个重大的挑战，因为这些亲水性核酸在被降解为溶酶体之前，必须穿过富含脂肪的质膜和/或内体膜。设计一种能够在特定刺激下破坏细胞膜稳定的脂类载体将有助于核酸到达靶细胞的细胞质。这项研究计划专注于多功能脂类的设计和评价，具有DNA结合亲和力和pH触发的膜失稳特性。DNA结合亲和力将由可变的阳离子头基提供。PH敏感性将基于以前开发的分子钳单元的使用，这些单元已经显示了适合内体逃逸的pH范围。这项研究计划将分三个阶段进行。在第一阶段，将合成和表征一个由20种脂类组成的文库。根据它们的pH触发的释放和对不同核酸的亲和力，将应用选择过程来选择最佳的6个候选者。第二阶段将确认膜的不稳定特性，并评估转染率和细胞毒性。两个最好的候选者将进入第三阶段，在那里细胞内核酸的输送将被成像。最后，在小鼠身上进行的概念验证将评估这些具有生物响应性的脂质纳米颗粒在提高体内遗传物质输送效率方面的潜力。

Who could have envisaged it would be possible to use a gene as a drug 50 years ago, when the structure of DNA has just been discovered? Thanks to synergistic efforts of a whole scientific community, the concept has gone from bench to bedside, and two gene products are now available on the market. However, these new therapeutics are still in their first stages and require optimization. It is possible to improve their efficiency by the use of carefully engineered vectors, able to carry the genetic material through each of the biological barriers they need to cross. In particular, getting inside the cell is a major challenge, because these hydrophilic nucleic acids have to cross the lipid-rich plasmatic and/or endosomal membrane, before being degraded into lysosomes. Designing a lipidic vector able to destabilize the membrane upon a specific stimulus would facilitate nucleic acids to reach the cytoplasm of the targeted cell. This research program is focused on the design and evaluation of multifunctional lipids, bearing DNA binding affinity and pH-triggered membrane destabilization properties. The DNA binding affinity will be provided by variable cationic headgroups. The pH-sensitivity will be based on the use of previously developed molecular tweezer units, which have shown the suitable pH range for endosomal escape. This research program will process in three phases. In phase I, a library of 20 lipids will be synthesized and characterized. A selection process, based on their pH-triggered release and affinity for diverse nucleic acids, will be applied to select the best 6 candidates. The phase II will confirm the membrane-destabilizing properties and assess the transfection efficiency and cytotoxicity. The two best candidates will access to phase III, where the cytosolic delivery of nucleic acids will be imaged. Finally, a proof of concept in mice will evaluate the potential of these bioresponsive lipid nanoparticles to improve the efficiency of genetic material delivery in vivo.