Active loading of cyanine 5.5 derivatives into liposomes for deep self-quenching and their applications in deep tissue imaging

• Published in: Sensors & diagnostics Vol. 3; no. 6; pp. 128 - 138
• Main Authors: Chen, Chong-Yan, Jian, Cheng-Bang, Gao, Hua-De, Yu, Xu-En, Chang, Yuan-Chih, Leong, Shwee Khuan, Shie, Jiun-Jie, Lee, Hsien-Ming
• Format: Journal Article
• Language: English
• Published: 13.06.2024
• ISSN: 2635-0998; 2635-0998
• DOI: 10.1039/d3sd00325f

Visualizing liposome release profiles in small animals is important for evaluating the pharmacokinetic influence of vesicles. Encapsulating near-infrared (NIR) fluorescent dyes to visualize and report liposomal cargo release in vivo , which necessitates high encapsulation with deep self-quenching, is highly desirable in advanced (such as targeting or trigger-release) liposome development. However, passive loading of NIR dyes usually yields low encapsulation efficiencies (1-5%), causing significant wastage and cost-ineffectiveness while using expensive NIR fluorescent dyes. It would be highly beneficial if an active loading method, which typically has an encapsulation efficiency of nearly 100%, is developed. This research describes an active loading approach for two cyanine 5.5 (Cy5.5) derivatives. We discovered that using ammonium sucrose octasulfate (ASO) as a trapping agent allows for nearly 100% encapsulation for both Cy5.5 dyes, accompanied by the formation of nanoprecipitates inside the liposome, as evidenced by cryogenic electron microscopy. Fluorescence spectroscopy confirmed deep fluorescence self-quenching after active loading and a 60-100-fold fluorescence enhancement upon full content release via liposome rupture. Cellular uptake experiments showed that the fluorescence of Cy5.5-loaded liposomes recovered and plateaued after 9 hours of incubation with cells. In vivo fluorescence imaging (IVIS) demonstrated the same fluorescence activation in tumor-bearing mice intratumorally injected with the liposome. We believe that the developed active loading method will enable Cy5.5-loaded liposomes to be a deep tissue-compatible and cost-effective NIR fluorescence release-reporting platform. Visualizing liposome release profiles in small animals is important for evaluating the pharmacokinetic influence of vesicles.