Nucleic Acid Drugs

Nucleic acid medicine
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The development of an ideal delivery carrier with efficiency, specificity, as well as safety has been supported by progress in polymer chemistry and supramolecular chemistry, and accumulated experience in biological and clinical trials. Skip to main content. Advertisement Hide. Delivery of Nucleic Acid Drugs. Chapter First Online: 06 July This is a preview of subscription content, log in to check access. Marshall E Science — Google Scholar. Kaiser J Science Google Scholar. Personalised recommendations. Cite chapter How to cite? Circulation time and biodistribution of unconjugated HJs.

A Blood circulation time of Cy5. The lower panel shows a representative image of the collected organs from a mouse. C Fluorescent scan of a paraffin-embedded mouse kidney.

torrwenfiloba.tk HJs were labeled with Cy5. The color scale is rainbow red: high, blue: low. D Confocal microscopy images of a paraffin-embedded mouse kidney showing Cy5-labeled HJs in red as they pass through the kidney. The slide has been stained with DAPI to indicate nuclei and overlayed with a brightfield image. HJs administered subcutaneously S. The HJ blood levels reached a maximum approximately min post injection. Nonetheless, the overall biodistribution profile after 24 h was almost identical to that of the I.

The high stoichiometric control of the HJ allowed us to conduct a systematic investigation of the PK properties of the structure with different types and number of conjugates. Initially, HJ oligos were conjugated and assembled with variable number and sizes of PEG polymers, as PEGylation is one of the most widely used approaches for prolonging circulation times [ 35 ]. As the renal filtration cut-off is usually observed to be in the kDa range, we expected a significant change in circulation time from attaching just a single PEG20K molecule.

To test this, we injected Cy5. HJs were assembled with PEG20K at two different positions to ensure that a flexible placement was allowed. On the right side is shown a representative image of the fluorescent scan of the blood samples. D In vitro binding between HJ-Palmitoyl conjugates and human serum albumin. HJs carrying two palmitoyl groups on Q3 were preassembled in PBS and subsequently incubated with increasing amounts of albumin.

Imaging of live animals after 24 h showed no peripheral signals above background in mice injected with unfunctionalized HJs, whereas mice injected with HJs carrying one or more PEG20Ks exhibited a widely distributed signal across the body including the extremities, such as paws, tail and nose Figure S6. These combined results demonstrate that both the half-life and biodistribution of the small structure can be controlled, and that one or two PEG20K moieties is sufficient to extend circulation times in vivo.

A different strategy for half-life extension is to conjugate drugs with biodegradable fatty acids. This approach is utilized in the design of several different FDA-approved drugs, including Levemir and Victoza [ 36 , 37 ], of which the latter contains a C 16 palmitoyl group. To investigate the utility of this approach for the HJ scaffold, one module Q3 was synthesized with two palmitoyl groups at the 5'-terminus. Although the palmitoylated oligo tended to form aggregates by itself, no aggregation was observed by gel electrophoresis once it was inserted into the HJ structure Figure 4 D.

Gel shift assays with purified HSA confirmed that the palmitoylated HJ bound albumin with high binding affinity and fast kinetics Figure 4 D , similar to what has previously been observed for palmitoylated antisense oligos [ 39 ]. To analyze the effect of the palmitoylation in vivo , Cy5. As shown in Figure 4 B, the palmitoyl groups had a significant effect on the circulation half-life of the HJs, increasing it from approximately 21 min for the unconjugated HJ to more than 2 h for the HJ-Palmitoyl construct.

Importantly, after 24 h HJ-Palmitoyl was detected predominantly in the liver with only a small fraction found in the kidney, suggesting that renal clearance had to a large extent been circumvented Figure 4 C.

This is consistent with the reported PK properties of previous palmitoyl-conjugated constructs [ 40 ]. Targeted delivery in vitro. The asialoglycoprotein receptor ASGP-R is conserved in all mammals and is highly expressed on the surface of hepatocytes [ 41 ]. The functional entity is a trimer comprised of two proteins that, in the presence of calcium, exhibits high affinity for N -acetyl galactosamine GalNAc -terminated oligosaccharides [ 42 ].

The primary function of ASGP-R appears to be the clearance of glycoproteins by way of receptor-mediated endocytosis. However, more recently this receptor has also been used for targeted delivery of a variety of drugs, including oligonucleotides, to the liver in mice [ 43 , 44 ]. Using the potential to control the valency of the HJ, we investigated the effect of multimerization of the triGalNAc units on the cellular uptake in hepatocytes.

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The tri-antennary conformation of GalNAc sugars has been shown to bind with high affinity to the ASGP-R, and we hypothesized that a multivalent display of trivalent sugars would be able to crosslink multiple receptors and therefore further enhance endocytosis. After 45 min, the cells were washed and subsequently analyzed by flow cytometry to compare cellular uptake between the different structures Figure 5 B. To ensure that the fluorescence signal detected by flow cytometry did indeed originate from internalized HJs, confocal microscopy was used to image HepG2 cells treated with these same samples Figure 5 C.

Here, a clear intracellular signal with a punctuate cytoplasmic distribution was observed, indicative of endosomal localization. Collectively, these data show that the efficient internalization via the ASGP-R receptor can be further potentiated by a multivalent display of tri-antennary ligands on the HJ. Targeted delivery in vivo. To test if the multivalency effect from conjugation of multiple triGalNacs observed in the in vitro uptake studies would also result in higher uptake in hepatocytes in vivo , we injected mice with Cy5.

Blood samples were collected at different intervals and the organs were harvested 2 h post injection. All injected samples displayed similar blood circulation half-lives Figure S7 and were essentially undetectable in the blood after 2 h. However, their biodistribution profiles were markedly different.

When comparing the three triGalNAc groups, a clear trend was observed that a higher valency of triGalNAcs resulted in stronger liver accumulation, although only the difference between a single and three triGalNAc units was statistically significant Figure 6 A. Imaging of tissue slices of the collected livers in a 2D fluorescence scanner showed that the triGalNAc-functionalized HJs were distributed evenly throughout the entire organ Figure 6 B , suggesting that uptake predominantly occurs in hepatocytes.

These data were confirmed by confocal microscopy Figure 6 C , which showed that the HJs were not confined to areas near blood vessels or in macrophages, but rather distributed evenly throughout the tissue.

Methods for Studying Nucleic Acid/Drug Interactions

Thus, we conclude that the triGalNAc-functionalized HJs are indeed taken up by hepatocytes and that the multivalency of triGalNAc groups contributes to higher uptake. Flow cytometry histograms show the cell-associated fluorescence from one replicate from each group within one experiment, and the bars show the median fluorescence from three biological replicates from two independent experiments, normalized to the HJ-Cy5 control.

C Confocal microscopy images of HepG2 cells treated with the samples shown in B. A Ex vivo biodistribution of Cy5. A representative image of the organs is shown to the right. B Fluorescent scan of an entire lobe excised from the liver. C Confocal microscopy images of a paraffin-embedded mouse liver lobe showing Cy5. The slide has been stained with DAPI to indicate nuclei and overlayed with a bright fieldimage. The assembled scaffold appears to either exist in a relaxed form with approximately equal distances between the four arms, or as a stacked, X-shaped structure, depending on the presence of divalent cations.

Each of the four arms can be functionalized and assembled in a modular fashion with, in principle, any biomolecule with an accessible chemical handle for site-specific conjugation. Due to the stabilizing effects of the modified nucleotides, the HJ oligos robustly self-assemble into uniform and stable constructs, even when functionalized with large hydrophilic or charged macromolecules on each of the four arms.

This simple covalent coupling approach, in combination with the high thermostability of the duplexes, thus allows us to create structures that are exceptionally stable with a low risk of dissociation under physiological conditions. This represents a significant improvement compared to other nucleic acid-based platforms with a potential use for drug delivery, such as doxorubicin-loaded DNA origami structures [ 46 ] that risk dissociation and drug leakage at the physiological salt concentrations and low concentrations in vivo [ 47 ].

Due to its small size and highly chemically modified nature, the HJ presented here also separates itself from similar proposed structures, such as a significantly larger DNA HJ consisting of four 50 nt long DNA strands, all modified with fluorophores for high-signal protein labeling [ 48 ]. In mice, the small non-functionalized HJ scaffold was shown to be excreted through the kidneys with a half-life of only minutes. This effect can be advantageous when rapid clearance is needed in connection with bioimaging of for instance tumors.

Nucleic Acid Drug List

“Nucleic acid medicines” utilize “nucleic acid,” which refer to substances such as DNA and RNA that control genetic information, as drugs. These allow targeting. Nucleic acid medicine. Next generation drugs using DNA and RNA With research conducted in countries around the world. Nucleic acid medicines are next.

However, for drug delivery purposes, a sufficiently long circulatory half-life is vital to ensure maximum exposure of the payload to its target. To accommodate both potential applications, we investigated the possibility of adjusting this circulation time by conjugating various PEG or palmitoyl groups. By attaching one or more PEG modules, the circulation time of the HJ was significantly increased, and the HJ remained detectable in the blood after 24 h.

However, in spite of its wide use and popularity, there has in recent years been an increased focus on the disadvantages of the use of PEG [ 50 , 51 ]. One of the primary concerns is the presence of antibodies that specifically recognize and bind to PEG, including both pre-existing and treatment-induced antibodies [ 50 , 52 ]. In addition, while PEG prolongs blood circulation and presumably increases bioavailability at the target tissue, heavy PEGylation can lead to shielding of the targeting ligands on the structure. Using PEG for PK enhancement is in this way a balancing act between achieving optimal circulation time and effective target recognition.

We therefore also investigated an alternative strategy by attaching biodegradable fatty acids to the HJ.

Delivery of Nucleic Acid Drugs

These are known to associate non-covalently with fatty acid binding sites on circulating HSA and take advantage of its extraordinarily long half-life [ 53 ]. We synthesized a HJ module carrying two palmitoyl groups in its 5' end. The palmitoyl module was shown to facilitate efficient binding to HSA in vitro and afforded significant PK enhancement by extending the half-life of the HJ from approximately 20 min to more than two hours. In this way, both the half-life and biodistribution of the HJ in circulation can be tightly controlled by conjugation of PEG or palmitoyl groups, thereby providing a method to fine-tune its PK properties for diagnostic and therapeutic applications.

It is likely that such PK optimization could translate into higher drug efficacy and consequently reduce the required drug dose. This could thereby allow for the delivery of cytotoxic drugs with low half-lives and narrow therapeutic windows with a reduced risk of adverse effects. In addition, the modular nature of the HJ provides a unique opportunity to explore different combinations of PK enhancers without the need for extensive resynthesis to identify the best suited configurations for a particular clinical application.

The ability to mediate active and specific tissue targeting is an important property for any drug delivery platform intended for personalized and theranostic medicine. To investigate the potential of the HJ for cell-specific uptake, we functionalized the scaffold with triGalNAcs with precise stoichiometry. This led to pronounced hepatocyte-specific uptake of the HJ in HepG2 cells and also fast liver accumulation in vivo.

The increase in valency of the triGalNAc targeting ligands also led to an increasing hepatocyte internalization both in vitro and in vivo, demonstrating that although the trivalent GalNAc moiety is an effective targeting ligand by itself, its targeting potential can be further enhanced by a multivalent display of two or more triGalNAcs. This approach could thereby add further potency to sugar-based delivery approaches including the triGalNAc-conjugated siRNA systems developed by Alnylam [ 54 ] and the triGalNAc-conjugated antimiR system from Regulus [ 55 ].

Furthermore, this indicates a potential of the HJ to improve the effect of receptor-mediated targeting with low-affinity systems. In clinical development of specific targeting ligands, such as peptides derived from phage display, many candidates are discarded from further development due to insufficient affinity for receptors under in vivo conditions. A controlled multivalent display of such candidates could enhance the utility of these ligands and produce effective targeted carriers for imaging modalities or therapeutic drugs.

For many applications, this precludes the need for post-assembly purification. This feature of the HJ therefore represents a significant improvement compared to for example nanoparticles or DNA origami where the functionalized substructures do not associate in a ratio and must be purified post assembly to avoid off-target effects. Additionally, this unique property opens up the possibility for rapid on-site combination of imaging agents or drugs targeted to selected disease conditions, based on larger libraries of differently functionalized oligonucleotide modules.

Bill Marshall

The current report focuses on the structural characterization of the HJ scaffold and on establishing the most vital properties required for a safe, stable and versatile delivery platform for imaging agents and drugs. The system was shown to be useful for molecular targeting with an adjustable half-life and could specifically deliver fluorophores for optical imaging to the target tissues. For clinical translation of this, attachment of radioactive compounds for positron emission tomography PET imaging or magnetic contrast for magnetic resonance imaging MRI is desired, and this work is therefore currently on-going.

With the incorporation of such clinically relevant imaging agents, the HJ could provide a flexible platform for highly detailed multi-modal diagnostics of cancer. Furthermore, with the planned production of modules containing different classes of bioactive cargos, including cytotoxic drugs and nucleic acid-based therapeutics, more complex multifunctional HJs could be assembled for applications in cancer, viral- inflammatory- and metabolic diseases. A and J.

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Gottesman MM. Mechanisms of cancer drug resistance. Annu Rev Med. Chemotherapy and the war on cancer. Nat Rev Cancer.

Bozic I, Nowak MA. Resisting Resistance. Annu Rev Cancer Biol. Translating cancer research into targeted therapeutics.

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User Name Password Sign In. Agonists of TLRs 7, 8, and 9 are being studied as therapeutic agents for various diseases, including cancers, allergies, asthma, and infectious diseases, and also as adjuvants with vaccines. To overcome problems related to safety and the cost of production resulting from the fully chemical modified ODN, we developed a non-chemically modified decoy ODN, the ribbon-type decoy, by ligation of the extremities of two single phosphodiester strands. The requirement for these drugs to use cellular machinery for their mechanism of action limits the amount of chemical modification that can be deployed since specific proteins in the cell need to recognize the molecule. This study is the first to demonstrate antisense-mediated protein suppression in patients with a neurodegenerative disease. Nature Reviews Neurology 15 ,

Diamantis N, Banerji U. Antibody-drug conjugates—an emerging class of cancer treatment. Br J Cancer. Strategies and challenges for the next generation of antibody-drug conjugates. Nat Rev Drug Discov. Combinatorial drug therapy for cancer in the post-genomic era. Nat Biotechnol. Nanomaterials for theranostics: recent advances and future challenges. Chem Rev. Adv Drug Deliv Rev. Imaging and drug delivery using theranostic nanoparticles.

Biocompatibility of engineered nanoparticles for drug delivery. J Control release. Engineered nanoparticles for drug delivery in cancer therapy. Angew Chemie Int Ed. Principles in the design of ligand-targeted cancer therapeutics and imaging agents. A logic-gated nanorobot for targeted transport of molecular payloads. Science DNA nanomaterials for preclinical imaging and drug delivery.

J Control Release. DNA nanotechnology for cancer therapy. Okholm AH, Kjems J. DNA nanovehicles and the biological barriers. A DNA nanorobot functions as a cancer therapeutic in response to a molecular trigger in vivo. Adv Mater. Aptamer chemistry. Bramsen JB, Kjems J. Development of therapeutic-grade small interfering RNAs by chemical engineering.

Front Genet. Pharmacokinetics, biodistribution and cell uptake of antisense oligonucleotides. Wengel J. Synthesis of 3 '-C-and 4 '-C-branched oligodeoxynucleotides and the development of locked nucleic acid LNA. Acc Chem Res. Vester B, Wengel J. Potent and nontoxic antisense oligonucleotides containing locked nucleic acids. Proc Natl Acad Sci. A large-scale chemical modification screen identifies design rules to generate siRNAs with high activity, high stability and low toxicity. Nucleic Acids Res. PLoS One. Synthesis of abasic locked nucleic acid and two seco-LNA derivatives and evaluation of their hybridization properties compared with their more flexible DNA counterparts.

J Org Chem. Org Biomol Chem. Ho PS. Structure of the Holliday junction: applications beyond recombination. Biochem Soc Trans. Petersen M, Wengel J. LNA: a versatile tool for therapeutics and genomics. Trends Biotechnol. Lilley DMJ. Structures of helical junctions in nucleic acids.

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Q Rev Biophys. PEGylation as a strategy for improving nanoparticle-based drug and gene delivery. The mechanism of protraction of insulin detemir, a long-acting, acylated analog of human insulin. Pharm Res. Clin Pharmacokinet. The major histocompatibility complex-related Fc receptor for IgG FcRn binds albumin and prolongs its lifespan. J Exp Med. Mol Ther. Albumin-based drug delivery: harnessing nature to cure disease.

Mol Cell Ther. Human Hepatic Lectin. J Biol Chem. J Mol Biol. Design and synthesis of novel N-acetylgalactosamine-terminated glycolipids for targeting of lipoproteins to the hepatic asialoglycoprotein receptor. J Med Chem. Targeted delivery of antisense oligonucleotides to hepatocytes using triantennary N-acetyl galactosamine improves potency fold in mice. The role of metal ions in the conformation of the four-way DNA junction.

EMBO J.