Understanding and controlling the cellular fate of fluorine-modified biologics

项目总结 有机氟化合物具有诱人的化学、药理和生物特性，具有 使他们能够在生物制药和生物材料的设计上进行范式转变。这个 将氟原子引入氨基酸和核酸中，开辟了一个广阔的新的化学图景 它可以改变多肽、蛋白质和DNA的折叠、稳定性、齐聚倾向和生物活性。 然而，尽管显示了良好的性能，但在生物支架中添加氟的影响是 很难预测。此外，越来越多的证据表明，全氟化合物混杂地吸附到 许多细胞的基本组成成分--包括脂质、蛋白质和DNA--引发了许多 生物效应。PI的实验室最近发现的这些效应之一是有机氟分子能够 将蛋白质和DNA直接组装成含氟微域，然后相分离成含氟液体，而无需 使生物变性。PI最近利用了这些新特性来实现超声引导 三维组织中的氟化蛋白质。根据这些初步研究结果，建议的 研究计划将从机械上探索有机氟化合物如何影响结构和 吸附的蛋白质和DNA的功能，并利用这些见解来指导新的超分子设计 组装好的生物材料。我们的主要假设是有机氟化合物以非共价吸附 到蛋白质和DNA，引导它们分离成富氟相，进而改变它们的低聚物 组装、细胞命运和生物活性。为了测试这个断言，我们将扩展我们的全氟化合物(PFC) 文库包括各种具有碱性/酸性官能团和杂环部分的分子。我们将使用 这个文库用于建立结构-活性关系，控制全氟碳化合物吸附蛋白质的能力，以及 研究PFC络合如何改变蛋白质细胞摄取、细胞内转运和生物活性。在……里面 同时，我们将利用这个文库来研究PFC-DNA相互作用的分子机制，并研究 PFC络合如何改变暴露细胞的DNA稳定性和代谢动态平衡。总而言之，这些研究 将建立对PFC如何与蛋白质和DNA相互作用的全面机制理解，并将 允许我们合理地设计利用这种不寻常的组装现象和阶段的氟生物技术- 出现的分离属性。例如，我们将创建超声波敏感的氟纳米乳剂 装载PFC修饰的核糖核蛋白(RNPs)，以实现在肾脏组织中进行成像引导的基因编辑。 这项研究的成功将推动PFC作为一种新的分子基序来控制蛋白质和DNA的使用 组装，以及开发的方法应用于发现新的试剂细胞内转导 含氟生物大分子。最终，提高关于有机氟化合物如何相互作用的知识 蛋白质和DNA及其对细胞的影响，将指导新的PFC启用技术的合理设计 具有用于药物发现和纳米医学应用的理想的功能特性。

PROJECT SUMMARY Organofluorine compounds possess attractive chemical, pharmacological and biological properties that have allowed them to make paradigm shifts in the design of biopharmaceuticals and biologic materials. The introduction of fluorine atoms into amino acids and nucleic acids opens a vast new chemical landscape with which to alter the folding, stability, oligomerization propensity and bioactivity of peptides, proteins and DNA. However, although shown to impart favorable properties, the impact of adding fluorine into biologic scaffolds is rarely predictable. Further, increasing evidence suggests perfluorinated compounds promiscuously adsorb to many of the fundamental building blocks of cells - including lipids, proteins and DNA - to elicit a plurality of bioeffects. One of these effects, recently discovered by the PI’s lab, is the ability of organofluorine molecules to direct protein and DNA assembly into fluorous microdomains that phase separate into fluorinated liquids without denaturing the biologic. The PI has recently exploited these emergent properties to enable ultrasound-guidance of fluorinated proteins in three-dimensional tissues. Building upon these preliminary findings, the proposed research program will mechanistically explore how organofluorine compounds influence the structure and function of adsorbed proteins and DNA and use these insights to guide the design of new supramolecular assembled biomaterials. Our overarching hypothesis is that organofluorine compounds non-covalently adsorb to proteins and DNA to direct their separation into fluorine-rich phases, which in turn alters their oligomeric assembly, cellular fate and bioactivity. To test this assertion, we will expand our perfluorinated compound (PFC) library to include a diversity of molecules with basic/acidic functionalities and heterocyclic moieties. We will use this library to establish structure-activity relationships governing the ability of PFCs to adsorb proteins, and investigate how PFC complexation alters protein cellular uptake, intracellular trafficking and bioactivity. In parallel, we will use this library to study the molecular mechanisms mediating PFC-DNA interactions and examine how PFC complexation alters DNA stability and metabolic homeostasis in exposed cells. Together, these studies will establish a comprehensive mechanistic understanding of how PFCs interact with proteins and DNA and will allow us to rationally design fluorous biotechnologies that exploit the unusual assembly phenomenon and phase- separation properties that emerge. As an example, we will create ultrasound-sensitive fluorine nanoemulsions loaded with PFC-modified ribonucleoproteins (RNPs) to enable imaging-guided gene editing in kidney tissue. Success of this research will advance the use of PFCs as a new molecular motif to control protein and DNA assembly, and the methods developed applied to discover new reagents for intracellular transduction of fluorinated biomacromolecules. Ultimately, advancing knowledge on how organofluorine compounds interact with proteins and DNA, and its effects on cells, will guide the rational design of new PFC enabled technologies with desirable functional properties for drug discovery and nanomedicine applications.