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For small interfering RNA (siRNA)-based cancer therapies, we report an actively-targeted and stabilized polyion complex micelle designed to improve tumor accumulation and cancer cell uptake of siRNA following systemic administration. Improvement in micelle stability was achieved using two stabilization mechanisms; covalent disulfide cross-linking a...
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Citations
... In the complexation of polyelectrolytes, a completely different situation exists. Enthalpy exchange is usually small and, in some cases, positive except when an exothermic reaction occurs during complexation, such as the formation of hydrogen bonds between weak polyacids and weak polybases [29]. ...
... We introduced disulfide moieties within the core to allow for dynamic covalent crosslinking, stabilizing the structure while ensuring intracellular dissociation by the reductive cellular environment. Such a strategy has been proven to largely improve stability of polyplex micelles for the delivery of plasmid DNA [13][14][15][16] and siRNA, [17][18][19] resulting in extended in vivo circulation and intracellular responsiveness. We studied the extracellu-lar stability (Figure 1-I) of these different structures and how their interaction with serum proteins affects their cellular uptake (Figure 1-II). ...
Nucleic acid therapeutics (NATs), such as mRNA, small interfering RNA or antisense oligonucleotides are extremely efficient tools to modulate gene expression and tackle otherwise undruggable diseases. Spherical nucleic acids (SNAs) can efficiently deliver small NATs to cells while protecting their payload from nucleases, and have improved biodistribution and muted immune activation. Self‐assembled SNAs have emerged as nanostructures made from a single DNA‐polymer conjugate with similar favorable properties as well as small molecule encapsulation. However, because they maintain their structure by non‐covalent interactions, they might suffer from disassembly in biologically relevant conditions, especially with regard to their interaction with serum proteins. Here, we report a systematic study of the factors that govern the fate of self‐assembled SNAs. Varying the core chemistry and using stimuli‐responsive disulfide crosslinking, we show that extracellular stability upon binding with serum proteins is important for recognition by membrane receptors, triggering cellular uptake. At the same time, intracellular dissociation is required for efficient therapeutic release. Disulfide‐crosslinked SNAs combine these two properties and result in efficient and non‐toxic unaided gene silencing therapeutics. We anticipate these investigations will help the translation of promising self‐assembled structures towards in vivo gene silencing applications.
... Our lab and others have improved si-NP stability through a variety of strategies, particularly through enhancing core hydrophobic interactions in combination with canonical electrostatic interactions [17]. Shown by us and others, modification of the siRNA by conjugation with hydrophobic moieties such as palmitic acid [18,19], cholesterol [20,21], or other hydrophobes [22] increases carrier-cargo loading stability. Nanocarrier hydrophobicity can also be increased through ternary si-NP formulation strategies. ...
Gene silencing with siRNA nanoparticles (si-NPs) is promising but still clinically unrealized for inhibition of tumor driver genes. Ternary si-NPs containing siRNA, a single block NP core-forming polymer poly[(2-(dimethylamino)ethyl methacrylate)-co-(butyl methacrylate)] (DMAEMA-co-BMA, 50B), and an NP surface-forming diblock polymer 20 kDa poly(ethylene glycol)-block-50B (20kPEG-50B) have the potential to improve silencing activity in tumors due to the participation of both 50B and 20kPEG-50B in siRNA electrostatic loading and endosome disruptive activity. Functionally, single block 50B provides more potent endosomolytic activity, while 20kPEG-50B colloidally stabilizes the si-NPs. Here, we systematically explored the role of the molecular weight (MW) of the core polymer and of the core:surface polymer ratio on ternary si-NP performance. A library of ternary si-NPs was formulated with variation in the MW of the 50B polymer and in the ratio of the core and surface forming polymeric components. Increasing 50B core polymer MW and ratio improved si-NP in vitro gene silencing potency, endosome disruptive activity, and stability, but these features also correlated with cytotoxicity. Concomitant optimization of 50B size and ratio resulted in the identification of lead ternary si-NPs 50B4-DP100, 50B8-DP100, and 50B12-DP25, with potent activity and minimal toxicity. Following intravenous treatment in vivo, all lead si-NPs displayed negligible toxicological effects and enhanced pharmacokinetics and tumor gene silencing relative to more canonical binary si-NPs. Critically, a single 1 mg/kg intravenous injection of 50B8-DP100 si-NPs silenced the tumor driver gene Rictor at the protein level by 80% in an orthotopic breast tumor model. 50B8-DP100 si-NPs delivering siRictor were assessed for therapeutic efficacy in an orthotopic HCC70 mammary tumor model. This formulation significantly inhibited tumor growth compared to siControl-NP treatment. 50B8-DP100 si-NPs were also evaluated for safety and were well-tolerated following a multi-dose treatment scheme. This work provides new insight on ternary si-NP structure-function relationships and identifies core polymer optimization strategies that can yield safe si-NP formulations with potent oncogene silencing.
... Airway resistance in allergen exposure [103] Another approach to increase the stability of siRNA/carrier complex involves covalent crosslinking of the carrier and introduction of hydrophobic interactions [105]. Kataoka et al. developed a polyion complex micelle system based on Cholesterol-Conjugated siRNA (chol-siRNA) and PEG-poly(L-lysine) block-co-polymer (PEG-PLL) Modified with 1-(3-mercaptopropyl)Amidine (MPA),where the micelles were stabilized by disulfide cross-linking of the polymer and hydrophobic association of cholesterol groups [105]. ...
... Airway resistance in allergen exposure [103] Another approach to increase the stability of siRNA/carrier complex involves covalent crosslinking of the carrier and introduction of hydrophobic interactions [105]. Kataoka et al. developed a polyion complex micelle system based on Cholesterol-Conjugated siRNA (chol-siRNA) and PEG-poly(L-lysine) block-co-polymer (PEG-PLL) Modified with 1-(3-mercaptopropyl)Amidine (MPA),where the micelles were stabilized by disulfide cross-linking of the polymer and hydrophobic association of cholesterol groups [105]. siRNA was fluorescently labeled by conjugating Cy5 dye to 5′ end of the antisense strand, and the blood level of siRNA was measured by Intravital Real-Time Confocal Laser Scanning Imaging (IVRT-CLSM) [105,106]. ...
... Kataoka et al. developed a polyion complex micelle system based on Cholesterol-Conjugated siRNA (chol-siRNA) and PEG-poly(L-lysine) block-co-polymer (PEG-PLL) Modified with 1-(3-mercaptopropyl)Amidine (MPA),where the micelles were stabilized by disulfide cross-linking of the polymer and hydrophobic association of cholesterol groups [105]. siRNA was fluorescently labeled by conjugating Cy5 dye to 5′ end of the antisense strand, and the blood level of siRNA was measured by Intravital Real-Time Confocal Laser Scanning Imaging (IVRT-CLSM) [105,106]. ...
Abstract
Up regulation of cell cycle-regulating and DNA repair genes appears to have
a negative impact on recurrence-free survival in patients with papillary thyroid
cancer. Furthermore, recurrence is associated with thyroid dedifferentiation.
Most cases address local applications or diseases in the filtering organs,
reflecting remaining challenges in systemic delivery of siRNA. Small Interfering
RNA (siRNA) is a promising drug candidate, expected to have broad therapeutic
potentials toward various diseases including viral infections and cancer. With
recent advances in bio conjugate chemistry and carrier technology, several
siRNA-based drugs have advanced to clinical trials. The difficulty in siRNA
delivery is in large part due to poor circulation stability and unfavorable
pharmacokinetics and bio distribution profiles of siRNA.
In this research we describe the pharmacokinetics and bio distribution of
siRNA Nano medicines, focusing on those reported in the past 5 years, and
their pharmacological effects in selected disease models such as hepatocellular
carcinoma, liver infections, and respiratory diseases. The examples discussed
here will provide an insight into the current status of the art and unmet needs
in siRNA delivery.
Keywords: RNA interference; Small interfering RNA; Delivery;
Pharmacokinetics; Biodistribution; Cancer
... When sulfhydryl groups are introduced into hydrophilic block polymer with polylysine segment, under oxidation condition, sulfhydryl groups are oxidized to form disulfide bond, thereby cross-linking polylysine segment and forming core cross-linked micelle (Miyata et al., 2004;Vachutinsky et al., 2011;Oe et al., 2014;Takeda et al., 2017). Core cross-linked micelles show enhanced resistance to the shear stress of blood circulation. ...
Natural amino acids and their derivatives are excellent building blocks of polymers for various biomedical applications owing to the non-toxicity, biocompatibility, and ease of multifunctionalization. In the present review, we summarized the common approaches to designing and constructing functional polymeric micelles based on basic amino acids including lysine, histidine, and arginine and highlighted their applications as drug carriers for cancer therapy. Different polypeptide architectures including linear polypeptides and dendrimers were developed for efficient drug loading and delivery. Besides, polylysine- and polyhistidine-based micelles could enable pH-responsive drug release, and polyarginine can realize enhanced membrane penetration and gas therapy by generating metabolites of nitric oxide (NO). It is worth mentioning that according to the structural or functional characteristics of basic amino acids and their derivatives, key points for designing functional micelles with excellent drug delivery efficiency are importantly elaborated in order to pave the way for exploring micelles based on basic amino acids.
... After intravenous injection into mice, the same formulation also improved accumulation in both tumor mass and tumor-associated blood vessels. These systems were successively improved by functionalizing the copolymer with dithiobispropionimidate (DTBP) and by encapsulating cholesterol-conjugated siRNA (Chlo-RNA) [112]. Chemical physical characterizations confirm that the Chol moiety did not alter the micelle formation behavior. ...
... cRGD-PEG-b-(PLL-IM) c(RGDyK) HeLa-luc SiRNA [111] cRGD-PEG-PLL(MPA) c(RGDyK) HeLa-Luc SiRNA [112] RGD-PEG-p(Lys)-p(Leu) RGD MDA-MB-231-MCF7 DCPT [113] Tat-p(Lys)-p(Leu) YGRKKRRQRRR HELA DOX [114] c-RGD-PEG-P(Glu) c(RGDyK)-RAD U87MG glioblastoma DACHPt [116] cRGD-PEG-P(Glu) c(RGDyK) LNM DACHPt [117] cRGD-PEG-P(Glu) c(RGDyK) HNSCC-CSC CDDP [118] PLG-g-Ve/PEG cRGD B16F10 DTX-CDDP [119] cRGD-PEG-PLA PEG-PGA)-API c(RGDyK) B16F10 DOX [120] cRGD-PEG-PDEARGDPEGPAsp c(RGDfC) C6 Cd-Te (QD) [121] cRGD PEG-b-PBLA c(RGDfC) U87MG Epi [122] PEG-GPLGVRGDG-P(BLA-co-Asp) GPLGVRGDG HT1080 DOX [123] cRGD-PEG-b-PAPA c(RGDyK) B16F10 DTX [124] cRGD-Lipep-Ms c(RGDyK) HCT-116 MMAE [126] c(RGD)-PF127 c(RGDyK) MDR KBv DOX-PTX [127] c(RGD)-PF127 c(RGDyK) U87MG DOX-PTX [128] Pf127-D4-(AdA) 2 (C18) 2 LARLLT HepG2 - [130] cRGDfK cRGDfK B16 t Quercitin [131] cRGDfK cRGDfK B16F10 Anlotinib [132] CCK8-DSPE-PEG 2000 DYMGWMDF -Gd [133] (C18) 2 DTPAGlu-Gd RGDBiot-StreptBiot-PEG62-b-PMLABe73 c(RGDyK) HepaRG DiD DiR [138] cRGD-PEG-PTMC c(RGDyK) U87MG PTX [140] c-RGD-PEG-SS-PTX c(RGDyK) SGC7901 PTX [141] iRGD-PEG-SS-CPT CRGDRGPDC U87MG CPT-IR780 [142] RGD-PEG-H 9 -DEVD-TPE RGD 4T1 DOX-TPE [143] OCT-PEG-b-PGu(DA-TPE) fCFwLTCT-ol LCC-18-NCH-446 c ETO [144] Tat-PEG-PBM YGRKKRRQRRR HT1080, MCF-7 MDA-MB-231 DOX [145] Tat-PEG1k-PHHD-GA-P5kCV AYGRKKRRQRRR SMCC 7721 DOX [146] ...
Precision medicine is based on innovative administration methods of active principles. Drug delivery on tissue of interest allows improving the therapeutic index and reducing the side effects. Active targeting by means of drug-encapsulated micelles decorated with targeting bioactive moieties represents a new frontier. Between the bioactive moieties, peptides, for their versatility, easy synthesis and immunogenicity, can be selected to direct a drug toward a considerable number of molecular targets overexpressed on both cancer vasculature and cancer cells. Moreover, short peptide sequences can facilitate cellular intake. This review focuses on micelles achieved by self-assembling or mixing peptide-grafted surfactants or peptide-decorated amphiphilic copolymers. Nanovectors loaded with hydrophobic or hydrophilic cytotoxic drugs or with gene silence sequences and externally functionalized with natural or synthetic peptides are described based on their formulation and in vitro and in vivo behaviors.
... However, almost all current siRNA vectors (e.g. lipoplexes, 4 polymers, 14 inorganic nanoparticles, 15 cell penetrating peptide, 16 and micelles 17 ) are positively charged, and so are their siRNA complexes. Although cationic carriers facilitate their condensation of negatively charged siRNA, and also enable endosomal escape, cationic carriers can interact with negative charged serum components, form unstable complexes in the circulation system, and cause off-target effect. ...
Despite the recent successes in siRNA therapeutics, targeted delivery beyond the liver remains the major hurdle for the widespread application of siRNA in vivo. Current cationic liposome or polymer-based delivery agents are restricted to the liver and suffer from off-target effect, poor clearance, low serum stability, and high toxicity. In this study, we have genetically engineered a non-cationic tumor-targeted universal siRNA nanocarrier. This protein nanocarrier consists of three function domains: dsRNA binding domain (dsRBD) (from human protein kinase R) for any siRNA binding, 18-histidines for endosome escape, and two RGD peptides at N-and C-termini for targeting tumor and tumor neovasculature. We showed that cloned dual-RGD-dsRBD-18his (dual-RGD) protein protects siRNA against RNases, induces effective siRNA endosomal escape, specific targets on integrin αvβ3 expressing cells in vitro, and homes siRNA to tumor in vivo. The delivered siRNA leads target gene knockdown in the cell lines and tumor xenografts with low toxicity. This multifunctional, biomimetic, charge-neutral siRNA carrier is biodegradable, low toxic, suitable for mass production by fermentation, and serum stable, holding great potential to provide a widely applicable siRNA carrier for tumor-targeted siRNA delivery.
... Furthermore, the RNAi therapeutics in blood circulation, leakage from blood to tumor tissues as well as distribution in tumor tissues can be monitored in a real-time manner with intravital confocal laser scanning microscopy (IVRTCLSM). For instance, the Cy5-siRNAincorporated polymeric micelles and naked siRNA (Cy5-siRNA) could be traced in blood circulation to study their pharmacokinetics in vivo, to investigate their entry from blood to tumor tissues, as well as their distribution in tumor tissues by utilizing IVRTCLSM (Figure 4b,c,d) [110][111][112]. ...
The RNA interference (RNAi) technique is a new modality for cancer therapy, and several candidates are being tested clinically. Nanotheranostics is a rapidly growing field combining disease diagnosis and therapy, which ultimately may add in the development of ‘personalized medicine’. Technologies on theranostic nanomedicines has been discussed. We designed and developed bioresponsive and fluorescent hyaluronic acid-iodixanol nanogels (HAI-NGs) for targeted X-ray computed tomography (CT) imaging and chemotherapy of MCF-7 human breast tumors. HAI-NGs were obtained with a small size of ca. 90 nm, bright green fluorescence and high serum stability from hyaluronic acid-cystamine-tetrazole and reductively degradable polyiodixanol-methacrylate via nanoprecipitation and a photo-click crosslinking reaction. This chapter presents an over view of the current status of translating the RNAi cancer therapeutics in the clinic, a brief description of the biological barriers in drug delivery, and the roles of imaging in aspects of administration route, systemic circulation, and cellular barriers for the clinical translation of RNAi cancer therapeutics, and with partial content for discussing the safety concerns. Finally, we focus on imaging-guided delivery of RNAi therapeutics in preclinical development, including the basic principles of different imaging modalities, and their advantages and limitations for biological imaging. With growing number of RNAi therapeutics entering the clinic, various imaging methods will play an important role in facilitating the translation of RNAi cancer therapeutics from bench to bedside.
... Redox potential responsiveness [98] 20min 40 Extracellular pH responsiveness [120] 5 h 150 ...
Recent progress in RNA biology has broadened the scope of therapeutic targets of RNA drugs for cancer therapy. However, RNA drugs, typically small interfering RNAs (siRNAs), are rapidly degraded by RNases and filtrated in the kidney, thereby requiring a delivery
vehicle for efficient transport to the target cells. To date, various delivery formulations have been developed from cationic lipids, polymers, and/or inorganic nanoparticles for systemic delivery of siRNA to solid tumors. This research article describes the current
status of clinical trials related to siRNA-based cancer therapy, as well as the remaining issues that need to be overcome to establish a successful therapy. It, then introduces various promising design strategies of delivery vehicles for stable and targeted siRNA delivery, including the prospects for future design. The current major strategies to design delivery vehicles for systemic siRNA delivery involve the construction of multimolecular assemblies from more than dozens of monomer components, including siRNA. The success of RNAi-based cancer therapy is closely associated with tumor biology as well
as architecture of delivery vehicles. Tumor cell plasticity evokes a resistance mechanism against clinical treatments, and cancer stem cells are gradually being identified as the root of cancer recurrence. New target RNA genes should be discovered to increase apoptosis in cancer cells and simultaneously reduce side effects in normal and healthy cells. Multidisciplinary research studies will guide the development of highly effective and safer RNAi-based drugs in clinical trials.
... A study reported that cRGD ligand enabled increased cellular uptake and in vitro gene silencing of cholesterol-conjugated siRNA/PEG-PLL micelleplexes, in α v β 5 integrin receptor-expressing HeLa-Luc cells (Oe et al., 2014). Conjugation with cholesterol conferred additional stability to the micelleplexes. ...
... Conjugation with cholesterol conferred additional stability to the micelleplexes. Further in vivo results corroborated the active-targeted and stabilized features of the micelleplexes, not only with regards to improved tumor accumulation but also concerning gene knockdown efficiency of the delivered Luc siRNA (siLuc) to mice bearing subcutaneous HeLa-Luc tumors (Oe et al., 2014). A new potential strategy for lung cancer treatment was based on co-delivery of doxorubicin and Bcl-2 siRNA by mPEG-PLGA-PLL micelleplexes decorated by epidermal growth factor (EGF) (Zhang et al., 2016a). ...
Cell-targeted nucleic acid (NA) therapeutics, either DNA or RNA, have experienced considerable attention regarding their potential applications in disease management. Their therapeutic potential as selective, safe and multispecific biomolecules is principally hindered by their instability in biological fluids and deficient cellular uptake, urgently calling for intelligent design strategies such as loading in effective nanosystems. Therefore, NA nanosystems which bypass biological hurdles and are capable of safeguarding the NA payload have been extensively explored. Micelleplexes consist of tailored and multifunctional micelle-like nanoassemblies of negatively-charged NAs and cationic blocks, thus ensuring efficient NAs protection, transportation, cellular transfection and intracellular trafficking. Herein, we review the biomedical applications of stable micelleplexes as robust and smart NA delivery nanosystems focusing on the fine-tuning of their properties toward stimuli-responsiveness and the nanosystem's versatility to accommodate distinct ligands for selective tissue-targeting purposes. Subsequent optimization regarding the nanosafety and regulatory considerations will also be discussed toward potential future clinical translation and thus paving the way to next-generation micelleplexes-based NA therapeutics. Micelleplexes as multifunctional micellar nucleic acid/polymer nanocomplexes for targeted delivery of diagnostic and therapeutic agents to cells, evidencing broad applications in biomedicine.
