Solid‐Phase Synthesis of Sequence‐Defined T‐, i‐, and U‐Shape Polymers for pDNA and siRNA Delivery

Schaffert, David; Troiber, Christina; Salcher, Eveline E.; Fröhlich, Thomas; Martin, Irene; Badgujar, Naresh; Dohmen, Christian; Edinger, Daniel; Kläger, Raphaela; Maiwald, G.; Farkasova, Katarina; Seeber, Silke; Jahn‐Hofmann, Kerstin; Hadwiger, Philipp; Wagner, Ernst

doi:10.1002/anie.201102165

Cited by 174 publications

(227 citation statements)

References 29 publications

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“…The preserved silencing activity of the conjugated siRNA was first shown in comparison to unmodified siRNA in a standard setting, transfecting Neuro2A/ eGFPLuc cells (Figure 3d and Figure S10) using the defined but untargeted polycationic carrier 46. 27 Results reveal that the silencing efficiency as well as cytotoxicity is not negatively affected by the attachment of the endosomolytic peptide via a disulfide linkage. Efficient and Specific Reporter Gene Silencing in Vitro.…”

Section: Articlementioning

confidence: 90%

“…Schaffert et al already showed that Stp-bearing structures bind nucleic acids very efficiently when crosslinked via disulfide bonds. 27 Thus cysteines were incorporated to enhance polyplex stability. The stabilizing effect is shown in a gel shift assay (Figure 3a), comparing the functional oligomer 356 with the analogue 420, bearing serines instead of cysteines.…”

Section: Articlementioning

confidence: 99%

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Nanosized Multifunctional Polyplexes for Receptor-Mediated SiRNA Delivery

et al. 2012

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Although our understanding of RNAi and our knowledge on designing and synthesizing active and safe siRNAs significantly increased during the past decade, targeted delivery remains the major limitation in the development of siRNA therapeutics. On one hand, practical considerations dictate robust chemistry reproducibly providing precise carrier molecules. On the other hand, the multistep delivery process requires dynamic multifunctional carriers of substantial complexity. We present a monodisperse and multifunctional carrier system, synthesized by solid phase supported chemistry, for siRNA delivery in vitro and in vivo.The sequence-defined assembly includes a precise cationic (oligoethanamino)amide core, terminated at the ends by two cysteines for bioreversible polyplex stabilization, at a defined central position attached to a monodisperse polyethylene glycol chain coupled to a terminal folic acid as cell targeting ligand. Complexation with an endosomolytic influenza peptide-siRNA conjugate results in nanosized functional polyplexes of 6 nm hydrodynamic diameter.The necessity of each functional substructure of the carrier system for a specific and efficient gene silencing was confirmed. The nanosized polyplexes showed stability in vivo, receptorspecific cell targeting, and silencing of the EG5 gene in receptor-positive tumors. The nanosized appearance of these particles can be precisely controlled by the oligomer design (from 5.8 to 8.8 nm diameter). A complete surface charge shielding together with the high stability result in good tolerability in vivo and the absence of accumulation in nontargeted tissues such as liver, lung, or spleen. Due to their small size, siRNA polyplexes are efficiently cleared by the kidney.

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Section: Articlementioning

confidence: 90%

Section: Articlementioning

confidence: 99%

Nanosized Multifunctional Polyplexes for Receptor-Mediated SiRNA Delivery

et al. 2012

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“…Studies of lipid and polymer carriers have implicated at least two common elements: chemical groups to bind small RNAs and nanoparticle (NP)-stabilizing hydrophobicity, which combine to encapsulate RNA molecules inside stable NPs and release small RNAs into the cytosol after endocytosis (35). Particular tertiary amines with optimal pK a , alkyl chains, and defined topology/ structure have been identified in effective carriers (2)(3)(4)(5)(6)(7)(8)(9)(10)(35)(36)(37).…”

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confidence: 99%

Modular degradable dendrimers enable small RNAs to extend survival in an aggressive liver cancer model

Zhou

Nguyen

Miller

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

144

135

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RNA-based cancer therapies are hindered by the lack of delivery vehicles that avoid cancer-induced organ dysfunction, which exacerbates carrier toxicity. We address this issue by reporting modular degradable dendrimers that achieve the required combination of high potency to tumors and low hepatotoxicity to provide a pronounced survival benefit in an aggressive genetic cancer model. More than 1,500 dendrimers were synthesized using sequential, orthogonal reactions where ester degradability was systematically integrated with chemically diversified cores, peripheries, and generations. A lead dendrimer, 5A2-SC8, provided a broad therapeutic window: identified as potent [EC 50 < 0.02 mg/kg siRNA against FVII (siFVII)] in doseresponse experiments, and well tolerated in separate toxicity studies in chronically ill mice bearing MYC-driven tumors (>75 mg/kg dendrimer repeated dosing). Delivery of let-7g microRNA (miRNA) mimic inhibited tumor growth and dramatically extended survival. Efficacy stemmed from a combination of a small RNA with the dendrimer's own negligible toxicity, therefore illuminating an underappreciated complication in treating cancer with RNA-based drugs.dendrimers | miRNA | cancer

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“…44 To verify the nontoxic properties also in case of protein delivery, a cell viability test was performed. This confirmed this assumption.…”

Section: ■ Results and Discussionmentioning

confidence: 99%

“…Transfection oligomer 386 was synthesized as previously reported. 44 SPDP modified proteins (1 mg) were diluted in Hepps buffer (1 mL, 0.5 M, pH 8.5) and 2-fold molar excess of oligomer 386 (compared to covalently bound linker), which was predissolved in water (50 mg/mL), was added for the modification of nlsEGFP. For the modification of β-galactosidase, a 5-fold molar excess of oligomer 386 (compared to covalently bound linker), which was predissolved in water (50 mg/mL), was used.…”

Section: ■ Experimental Sectionmentioning

confidence: 99%

Sequence Defined Disulfide-Linked Shuttle for Strongly Enhanced Intracellular Protein Delivery

2012

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Intracellular protein transduction technology is opening the door for a promising alternative to gene therapy. Techniques have to address all critical steps, like efficient cell uptake, endolysosomal escape, low toxicity, while maintaining full functional activity of the delivered protein.Here, we present the use of a chemically precise, structure defined threearm cationic oligomer carrier molecule for protein delivery. This carrier of exact and low molecular weight combines good cellular uptake with efficient endosomal escape and low toxicity. The protein cargo is covalently attached by a bioreversible disulfide linkage. Murine 3T3 fibroblasts could be transduced very efficiently with cargo nlsEGFP, which was tagged with a nuclear localization signal. We could show subcellular delivery of the nlsEGFP to the nucleus, confirming cytosolic delivery and expected subsequent subcellular trafficking. Transfection efficiency was concentration-dependent in a directly linear mode and 20-fold higher in comparison with HIV-TAT-nlsEGFP containing a functional TAT transduction domain. Furthermore, β-galactosidase as a model enzyme cargo, modified with the carrier oligomer, was transduced into neuroblastoma cells in enzymatically active form.

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Solid‐Phase Synthesis of Sequence‐Defined T‐, i‐, and U‐Shape Polymers for pDNA and siRNA Delivery

Cited by 174 publications

References 29 publications

Nanosized Multifunctional Polyplexes for Receptor-Mediated SiRNA Delivery

Nanosized Multifunctional Polyplexes for Receptor-Mediated SiRNA Delivery

Modular degradable dendrimers enable small RNAs to extend survival in an aggressive liver cancer model

Sequence Defined Disulfide-Linked Shuttle for Strongly Enhanced Intracellular Protein Delivery

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