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RNAi as New Class of Nanomedicines
Published in Dan Peer, Handbook of Harnessing Biomaterials in Nanomedicine, 2021
Monika Dominska, Derek M. Dykxhoorn
Pioneering preclinical studies have shown that potent silencing responses could be produced in mammalian cells through the introduction of specific siRNAs, which have been designed to target genes involved in a variety of pathogenic processes, including viral infections, tumor and metastases formation, and neurodegeneration [35, 48, 49, 121, 143, 147, 146]. Two basic strategies are employed to trigger the RNAi pathway for targeted gene knockdown: (1) delivery of synthetic siRNAs to the cytoplasm of cells and (2) expression from DNA-based vectors of short hairpin RNAs (shRNAs) that are recognized and processed by Dicer into active siRNAs [32, 33]. shRNAs are highly effective and frequently used experimentally for gene function analysis where permanent knockdown of a gene is desirable. However, stable over-expression of shRNAs in vivo has, in some cases, been associated with cytotoxic effects resulting from outcompeting endogenous molecules for limiting amounts of RNAi pathway components [6, 55]. In addition, toxicities associated with the integration of retroviral-based vectors, a commonly used approach for the stable delivery of shRNAs to many cell types, into the host cell genome have raised concerns about the safety of these delivery systems [113]. Once integrated, these vectors are maintained for the life span of the target cells and, presumably, retain siRNA expression even after their therapeutic utility has been exhausted. In contrast, the delivery of chemically synthesized siRNAs can efficiently silence gene expression without altering the host genetic material. The transient nature of the silencing response mediated by synthetic siRNAs means that repeated doses of the siRNA may be needed to maintain robust silencing [98, 135]. As a result of this transient silencing phenotype, the use of synthetic siRNAs allows for the gene-silencing treatment to be tailored to the specific clinical needs, thereby, providing a greater range of therapeutic options. This can be helpful if the target sequence needs to be altered (for example, siRNAs targeting sequences that are subject to mutation and selection like that seen during the replication of HIV), as well as if changing the dosage or discontinuing the treatment is desired due to untoward side effects or the achievement of a clinical end point. This chapter will focus on the delivery of synthetic siRNAs as mediators of the therapeutic gene silencing.
Lipid-based liquid crystalline films and solutions for the delivery of cargo to cells
Published in Liquid Crystals Reviews, 2019
Marilyn Porras-Gomez, Cecilia Leal
The delivery of nucleic acids into the cytosol has become a growing field in biomedicine [9,22,24,27,35,42,44–51]. siRNA is a class of double stranded short (ribo)nucleic acids (21-23 nucleotides long) capable of achieving gene knockdown (i.e. silenced or decreased gene expression) in eukaryotic cells. The clinical potential of siRNA in diseases such as cancer, neurodegenerative disorders and viral infectious diseases has been amply shown [52]. DNA depends upon entering the nucleus to initiate the transfection process. In contrast, siRNA does not require entering the nucleus: once it enters the cytosol, it is incorporated into a protein complex that finds and destroys target messenger RNA, which further propagates gene silencing [53,54]. Naked siRNA has shown silencing efficiency but is prone to enzymatic degradation and its negative charge prevents it to easily enter the cytosol. The design of non-cytotoxic carriers that deliver siRNA in a localised manner [49] and provide further specificity to gene silencing has become a critical challenge for the development of better therapies. Various lipid-nucleic acid complexes have been developed in order to address some of these issues and in this paper we describe a few examples [10,15,21,22,42,43,51,55,56].
Preparation and characterization of novel albumin-sericin nanoparticles as siRNA delivery vehicle for laryngeal cancer treatment
Published in Preparative Biochemistry and Biotechnology, 2019
Eda Yalcin, Goknur Kara, Ekin Celik, Ferda Alpaslan Pinarli, Guleser Saylam, Ceren Sucularli, Serhat Ozturk, Esin Yilmaz, Omer Bayir, Mehmet Hakan Korkmaz, Emir Baki Denkbas
RNA interference (RNAi) has become a widely used powerful tool in gene functional analyses and therapeutic applications since it has first emerged as a post-transcriptional gene silencing strategy in 1998.[1,2] This process provides sequence-specific degradation of the target mRNA and hence inhibits translation of the target protein via the key role of siRNA. In this mechanism, the double-stranded RNAs (dsRNAs) are cleaved by the enzyme called Dicer and divided into fragments of 21–23 nucleotides which are called siRNAs. These siRNAs, consisting of a passenger strand and a guide strand, bind to the RNA-Induced Silencing Complex (RISC). Subsequently, the guide strand is complementarily directed to the target mRNA by the activity of Argonaute-2 enzyme to cleave between the 10th and 11th bases of the mRNA.[3] siRNAs can be also produced synthetically with unique sequences that are highly specific to the target. siRNAs have been studied commonly for cancer treatment owing to their high potency for gene silencing.[4] Despite this great potential, a number of drawbacks must be considered for its therapeutic use. The difficulty in delivery of the siRNA molecule to the targeted area resulting from its rapid enzymatic degradation and insufficient cellular uptake is the major obstacle.[4] Therefore, an efficient carrier is still a need in siRNA therapies, and lately, nanoparticles have come forward as favorable delivery systems for safe siRNA transportation and accomplishment of effective gene knockdown in targeted cells.
Block catiomer with flexible cationic segment enhances complexation with siRNA and the delivery performance in vitro
Published in Science and Technology of Advanced Materials, 2021
Wenqian Yang, Takuya Miyazaki, Pengwen Chen, Taehun Hong, Mitsuru Naito, Yuji Miyahara, Akira Matsumoto, Kazunori Kataoka, Kanjiro Miyata, Horacio Cabral
Herein, we aimed to gain insight on the effect of the flexibility of the cationic segment of the block catiomers on the complexation with siRNA. Thus, to discriminate the contribution of the flexibility of the polycation, we used two corresponding block copolymers bearing polycation segments with different flexibility and PEG, as follows: PEG-PLL with the relatively rigid amide backbone, and PEG-poly(glycidyl butylamine) (PEG-PGBA) having a polyether backbone, which presents ether bonds with low rotational barrier [19,20]. The block catiomer systems have similar PEG length and weight fraction, and equivalent polycation length, i.e. 40 units (Figure 1). Moreover, the pendant primary amines serving as the cationic moieties have the same distance to the block backbone, i.e. a (CH2)4 spacer. In addition, we have previously found that PEG-PLL and PEG-PGBA have comparable degree of hydration as determined by differential scanning calorimetry (DSC) [21]. Besides, since the ionic strength and temperature affect the formation of polyion complexes [22], in our study, we have fixed these parameters for relating the differences in complexation to the polymers. The PICs resulting from mixing siRNA and PEG-PLL, or PEG-PGBA, were compared by analyzing the polymer complexation, the size and morphology of the PIC, and the PICs stability. Moreover, molecular dynamics (MD) simulations were applied to gain understanding on the siRNA-polycation interactions. In addition, the gene knockdown ability was evaluated in cancer cells to determine the in vitro delivery efficiency. Our results indicate that the flexibility of the cationic block could be an important parameter affecting PIC stability and the ensuing delivery efficiency, opening new opportunities for enhanced siRNA delivery.