octaarginine and Neoplasms

Nanomedicine promises to play an important role in next generation therapy, including Nucleic acid/Gene therapy. To accomplish this, innovative nanotechnologies will be needed to support nanomedicine by controlling not only the biodistribution but also the intracellular trafficking of macromolecules such as RNA/DNA. A multifunctional envelope-type nano device (MEND) was developed to meet this requirement. We herein provide an update regarding the functions of the MEND system focusing on the introduction of different functional biomaterials that enhance efficiency. The octaarginine (R8) peptide enhances cellular uptake and controls intracellular trafficking to induce synergism in transgene expression. The R8 was also used for developing a MITO-Porter system for mitochondrial targeting. The function of the MITO-Porter system was extended by developing a mitochondrial reporter gene for mitochondrial gene therapy. For efficient in vivo gene delivery, new pH-sensitive lipids have been introduced to achieve controlled biodistribution and to enhance endosomal escape. For example, the CL4H6 lipid exerts a more efficient in vivo gene silencing than that of ONPATTRO

Topics: Animals; Biocompatible Materials; Drug Delivery Systems; Genetic Therapy; Humans; Immunotherapy; Nanomedicine; Nanotechnology; Neoplasms; Nucleic Acids; Oligopeptides

Cell-penetrating peptides (CPPs) are promising delivery vehicles. These short peptides can transport wide range of cargos into cells, although their usage has often limitations. One of them is the endosomatic internalisation and thus the vesicular entrapment. Modifications which increases the direct delivery into the cytosol is highly researched area. Among the oligoarginines the longer ones (n > 6) show efficient internalisation and they are well-known members of CPPs. Herein, we describe the modification of tetra- and hexaarginine with (4-((4-(dimethylamino)phenyl)azo)benzoyl) (Dabcyl) group. This chromophore, which is often used in FRET system increased the internalisation of both peptides, and its effect was more outstanding in case of hexaarginine. The modified hexaarginine may enter into cells more effectively than octaarginine, and showed diffuse distribution besides vesicular transport already at low concentration. The attachment of Dabcyl group not only increases the cellular uptake of the cell-penetrating peptides but it may affect the mechanism of their internalisation. Their conjugates with antitumor drugs were studied on different cells and showed antitumor activity.

Topics: Antineoplastic Agents; Cations; Cell Proliferation; Cell-Penetrating Peptides; Humans; Neoplasms; Oligopeptides; p-Dimethylaminoazobenzene; Peptides; Tumor Cells, Cultured

Efficiently delivering plasmid DNA (pDNA) to the spleen is particularly significant for DNA immunization. However, increasing the efficiency of gene expression in spleen cells for achieving a therapeutic effect remains a serious challenge. An ideal spleen-targeted system should avoid liver uptake and should efficiently transfect specific functional spleen cells. Here, we report on pDNA nanocarriers with enhanced transfection in spleen cells driven by synergism between an octaarginine (R8) peptide and YSK05; a pH-responsive ionizable lipid. A double-coating design is essential for enhancing spleen selective transfection which is significantly affected by the total amount of lipid and the composition of the outer coat. The optimized R8/YSK system shows a high gene expression in the spleen with a high spleen/liver ratio and a surprising ability to target spleen B cells. Compared to other organs, the high spleen activity cannot be explained based on the amount of pDNA delivered to each organ, indicating that the system is extremely efficient in transfecting spleen cells. The system can be used in cancer immunization where a strong anti-tumor effect was observed in mice immunized with the R8/YSK system encapsulating antigen-encoding pDNA. The R8/YSK system holds great promise for future applications in the field of DNA vaccination.

Topics: Animals; B-Lymphocytes; DNA; Female; Gene Transfer Techniques; Humans; Hydrogen-Ion Concentration; Immunization; Lipids; Liver; Male; Mice, Inbred C57BL; Mice, Inbred ICR; Nanocapsules; Neoplasms; Oligopeptides; Piperidines; Plasmids; Spleen; Transfection

Topics: A549 Cells; Acrylamides; Cell Line, Tumor; Cytoplasm; Endosomes; Female; Fluorescein-5-isothiocyanate; Gene Expression Regulation, Neoplastic; Gene Silencing; Gene Transfer Techniques; Humans; Microscopy, Confocal; Neoplasms; Oligopeptides; Ovarian Neoplasms; Peptides; Polymers; Real-Time Polymerase Chain Reaction; RNA; RNA Interference; RNA, Small Interfering; Surface Properties

We previously developed octaarginine (R8)-modified lipid envelope-type nanoparticles for siRNA delivery (R8-MEND). Herein, we report on their ex vivo siRNA delivery to primary mouse bone marrow-derived dendritic cells (BMDCs) for potential use as a cancer vaccine. Quantitative imaging analysis of the intracellular trafficking of siRNA revealed that the dissociation process, as well as the rate of endosomal escape limits the siRNA efficiency of the prototype R8-MEND, prepared by the hydration method (R8-MEND(hydo)). Successful endosomal escape was achieved by using a pH-dependent fusogenic peptide (GALA) modified on a lipid mixture that was optimized for endosomal fusion. Furthermore, a modified protocol for the preparation of nanoparticles, mixing the siRNA/STR-R8 complex and small unilamellar vesicles (R8/GALA-MEND(SUV)), results in a more homogenous, smaller particle size, and results in a more efficient intracellular dissociation. Gene knockdown of the suppressor of cytokine signaling 1 (SOCS1), a negative-feedback regulator of the immune response in BMDCs resulted in an enhanced phosphorylation of STAT1, and the production of proinflammatory cytokines. Moreover, SOCS1-silenced BMDCs were more potent in suppressing tumor growth. Collectively, these results show that siRNA loaded in R8/GALA-MEND(SUV) efficiently suppresses endogenous gene expression and consequently enhances dendritic cell-based vaccine potency in vivo.

Topics: Animals; Cancer Vaccines; Dendritic Cells; Endosomes; Female; Gene Knockdown Techniques; Genetic Therapy; HeLa Cells; Humans; Mice; Mice, Inbred C57BL; Nanoparticles; Neoplasms; Oligopeptides; RNA, Small Interfering; Suppressor of Cytokine Signaling Proteins