Journal
JOURNAL OF DRUG TARGETING
Volume 24, Issue 6, Pages 508-519Publisher
TAYLOR & FRANCIS LTD
DOI: 10.3109/1061186X.2015.1095194
Keywords
Endocytosis; endosomal release
Categories
Funding
- EU through the European Regional Development Fund through the project Tumor-Tech, Centre of Excellence of Chemical Biology [3.2.1001.11-0008]
- Estonian Ministry of Education and Research [IUT2026]
- Estonian Science Foundation [ETF9438]
- national scholarship program Kristjan Jaak - Archimedes Foundation
- Ministry of Education and Research and Graduate School in Biomedicine and Biotechnology
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Cell-penetrating peptides (CPPs) are efficient vehicles to transport bioactive molecules into the cells. Despite numerous studies the exact mechanism by which CPPs facilitate delivery of cargo to its intracellular target is still debated. The current work presents methods that can be used for tracking CPP/pDNA complexes through endosomal transport and show the role of endosomal transport in the delivery of cargo. Separation of endosomal vesicles by differential centrifugation enables to pinpoint the localization of delivered cargo without labeling it and gives important quantitative information about pDNA trafficing in certain endosomal compartments. Single particle tracking (SPT) allows following individual CPP/cargo complex through endosomal path in live cells, using fluoresently labled cargo and green fluoresent protein expressing cells. These two different methods show similar results about tested NickFect/pDNA complexes intracellular trafficing. NF51 facilitates rapid internalization of complexes into the cells, prolongs their stay in early endosomes and promotes release to cytosol. NF1 is less capable to induce endosomal release and higher amount of complexes are routed to lysosomes for degradation. Our findings offer potential delivery vector for in vivo applications, NF51, where endosomal entrapment has been allayed. Furthermore, these methods are valuable tools to study other CPP-based delivery systems.
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