Department of Chemistry "Ugo Schiff" and CSGI, University of Florence, Via della Lastruccia 3, 50019 Sesto Fiorentino, FI, Italy.
Nanovector s.r.l., Via Livorno 60, 10144 Torino, Italy.
J Colloid Interface Sci. 2017 Dec 15;508:476-487. doi: 10.1016/j.jcis.2017.08.080. Epub 2017 Aug 24.
Transcription factor decoys (TFD) are short oligonucleotides designed to block essential genetic pathways in bacteria and defeat resistant infections. TFD protection in biological fluids and their delivery to the site of infection require formulation in appropriate delivery systems. In this work, we build on a classical phosphatidylcholine/phosphatidylethanolamine (POPC/DOPE) scaffold to design TFD-loaded cationic liposomes by combining the DNA-complexing abilities of a bolaamphiphile, (1,1'-(dodecane-1,12-diyl)-bis-(9-amino-1,2,3,4-tetrahydroacridinium) chloride (12-bis-THA), with the biocompatible cationic lipid ethyl-phosphatidylcholine (DPePC). The goal is to perform a structural study to determine the impact of the bolaamphiphile and TFD incorporation on the liposome structure, the capacity for TFD encapsulation, and the colloidal stability in saline media and cell culture environments.
The systems are characterized by means of dynamic light scattering, small-angle X-ray scattering, and ζ-potential measurements, to provide a clear picture of the liposome structure. Circular dichroism (CD) spectroscopy is used to assess the compaction of the oligonucleotide in a psi form, while steady-state fluorescence and fluorescence correlation spectroscopies give insight into the entrapment rate and distribution of the TFD in the liposomes.
We found that the combination of the two cationic species, 12-bis-THA and DPePC, allows encapsulation of 90% of the TFD. Results of CD experiments revealed that the TFD is condensed, therefore likely protected from the lytic action of serum nucleases. Finally, the systems showed colloidal stability in aqueous dispersion with ionic strength comparable to biologically relevant media.
转录因子诱饵（TFD）是设计用来阻断细菌中重要遗传途径并战胜耐药感染的短寡核苷酸。TFD 在生物体液中的保护及其递送至感染部位需要在适当的递药系统中进行制剂设计。在这项工作中，我们以经典的磷脂酰胆碱/磷脂酰乙醇胺（POPC/DOPE）支架为基础，通过将 DNA 络合能力与生物相容的阳离子脂质二棕榈酰基磷脂酰胆碱（DPePC）相结合，设计负载 TFD 的阳离子脂质体。我们采用了一种双（12-氨基-1,2,3,4-四氢吖啶）十二烷-1,12-二基二氯（12-双-THA） bola 两性离子，以 bola 两性离子和 TFD 掺入对脂质体结构、TFD 包封能力以及盐介质和细胞培养环境中的胶体稳定性的影响。
通过动态光散射、小角 X 射线散射和 ζ-电势测量对系统进行了表征，以清晰描绘脂质体结构。圆二色性（CD）光谱用于评估寡核苷酸在 psi 形式下的紧凑程度，而稳态荧光和荧光相关光谱则深入了解 TFD 在脂质体中的包封率和分布。
我们发现，两种阳离子物质 12-双-THA 和 DPePC 的结合允许 90%的 TFD 被包封。CD 实验结果表明，TFD 被浓缩，因此可能受到血清核酸酶溶胞作用的保护。最后，该系统在具有与生物相关介质相当的离子强度的水性分散体中表现出胶体稳定性。
