Efficient siRNA delivery and gene silencing using a lipopolypeptide hybrid vector mediated by a caveolae-mediated and temperature-dependent endocytic pathway

• Published in: Journal of nanobiotechnology 2019-01, Vol.17 (1), p.11-11, Article 11
• Main Authors: Kasai, Hironori, Inoue, Kenji, Imamura, Kentaro, Yuvienco, Carlo, Montclare, Jin K, Yamano, Seiichi
• Format: Article
• Language: English
• Subjects: Actin; Carcinoma; Caveolae; Cell Line; Charged particles; Chlorpromazine; Clathrin; Cyclodextrins; Cysteine; Cystine; Cytotoxicity; Deoxyribonucleic acid; DNA; Drug dosages; Efficiency; Endocytosis; Endocytosis - drug effects; Expression vectors; Feasibility studies; Gene delivery; Gene expression; Gene silencing; Gene Silencing - drug effects; Gene therapy; Gene Transfer Techniques; Genetic engineering; Histidine; HIV; Human immunodeficiency virus; Humans; Hypotheses; Infection; Inhibitors; Lipids; Lipopolysaccharides; Methods; Muscle proteins; Nanoparticles; Non-viral vector; Peptides; Peptides - chemistry; Physiological aspects; Plasmids; Polyethyleneimine; Properties; Proteins; Reagents; Ribonucleic acid; RNA; RNA interference; RNA, Small Interfering - administration & dosage; siRNA; Small interfering RNA; Squamous cell carcinoma; Tat protein; Temperature dependence; Toxicity; Transfection; Vectors (Biology); β-Cyclodextrin
• ISSN: 1477-3155; 1477-3155
• DOI: 10.1186/s12951-019-0444-8

We developed a non-viral vector, a combination of HIV-1 Tat peptide modified with histidine and cysteine (mTat) and polyethylenimine, jetPEI (PEI), displaying the high efficiency of plasmid DNA transfection with little toxicity. Since the highest efficiency of INTERFERin (INT), a cationic amphiphilic lipid-based reagent, for small interfering RNA (siRNA) transfection among six commercial reagents was shown, we hypothesized that combining mTat/PEI with INT would improve transfection efficiency of siRNA delivery. To elucidate the efficacy of the hybrid vector for siRNA silencing, β-actin expression was measured after siRNA β-actin was transfected with mTat/PEI/INT or other vectors in HSC-3 human oral squamous carcinoma cells. mTat/PEI/INT/siRNA produced significant improvement in transfection efficiency with little cytotoxicity compared to other vectors and achieved ≈ 100% knockdown of β-actin expression compared to non-treated cells. The electric charge of mTat/PEI/INT/siRNA was significantly higher than INT/siRNA. The particle size of mTat/PEI/INT/siRNA was significantly smaller than INT/siRNA. Filipin III and β-cyclodextrin, an inhibitor of caveolae-mediated endocytosis, significantly inhibited mTat/PEI/INT/siRNA transfection, while chlorpromazine, an inhibitor of clathrin-mediated endocytosis, did not inhibit mTat/PEI/INT/siRNA transfection. Furthermore, the transfection efficiency of mTat/PEI/INT at 4 °C was significantly lower than 37 °C. These findings demonstrated the feasibility of using mTat/PEI/INT as a potentially attractive non-viral vector for siRNA delivery.