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| 2. |
- Bijkerk, Roel, et al.
(författare)
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In Vivo Silencing of MicroRNA-132 Reduces Blood Glucose and Improves Insulin Secretion
- 2019
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Ingår i: Nucleic acid therapeutics. - : Mary Ann Liebert Inc. - 2159-3337 .- 2159-3345. ; 29:2, s. 67-72
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Tidskriftsartikel (refereegranskat)abstract
- Dysfunctional insulin secretion is a hallmark of type 2 diabetes (T2D). Interestingly, several islet microRNAs (miRNAs) are upregulated in T2D, including miR-132. We aimed to investigate whether in vivo treatment with antagomir-132 lowers expression of miR-132 in islets thereby improving insulin secretion and lowering blood glucose. Mice injected with antagomir-132 for 24 h, had reduced expression of miR-132 expression in islets, decreased blood glucose, and increased insulin secretion. In isolated human islets treated with antagomir-132, insulin secretion from four of six donors increased. Target prediction coupled with analysis of miRNA-messenger RNA expression in human islets revealed DESI2, ARIH1, SLC25A28, DIAPH1, and FOXA1 to be targets of miR-132 that are conserved in both species. Increased expression of these targets was validated in mouse islets after antagomir-132 treatment. In conclusion, we identified a post-transcriptional role for miR-132 in insulin secretion, and demonstrated that systemic antagomir-132 treatment in mice can be used to improve insulin secretion and reduce blood glucose in vivo. Our study is a first step towards utilizing antagomirs as therapeutic agents to modulate islet miRNA levels to improve beta cell function.
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| 3. |
- Boianelli, Alessandro, et al.
(författare)
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Cross-Species Translation of Biophase Half-Life and Potency of GalNAc-Conjugated siRNAs
- 2022
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Ingår i: Nucleic Acid Therapeutics. - : Mary Ann Liebert. - 2159-3337 .- 2159-3345. ; 32:6, s. 507-512
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Tidskriftsartikel (refereegranskat)abstract
- Small interfering RNAs (siRNAs) with N-acetylgalactosamine (GalNAc) conjugation for improved liver uptake represent an emerging class of drugs to treat liver diseases. Understanding how pharmacokinetics and pharmacodynamics translate is pivotal for in vivo study design and human dose prediction. However, the literature is sparse on translational data for this modality, and pharmacokinetics in the liver is seldom measured. To overcome these difficulties, we collected time-course biomarker data for 11 GalNAc-siRNAs in various species and applied the kinetic-pharmacodynamic modeling approach to estimate the biophase (liver) half-life and the potency. Our analysis indicates that the biophase half-life is 0.6-3 weeks in mouse, 1-8 weeks in monkey, and 1.5-14 weeks in human. For individual siRNAs, the biophase half-life is 1-8 times longer in human than in mouse, and generally 1-3 times longer in human than in monkey. The analysis indicates that the siRNAs are more potent in human than in mouse and monkey.
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| 6. |
- Hernandez, Frank J, et al.
(författare)
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Degradation of nuclease-stabilized RNA oligonucleotides in Mycoplasma-contaminated cell culture media
- 2012
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Ingår i: Nucleic Acid Therapeutics. - : Mary Ann Liebert. - 2159-3337 .- 2159-3345. ; 22:1, s. 58-68
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Tidskriftsartikel (refereegranskat)abstract
- Artificial RNA reagents such as small interfering RNAs (siRNAs) and aptamers often must be chemically modified for optimal effectiveness in environments that include ribonucleases. Mycoplasmas are common bacterial contaminants of mammalian cell cultures that are known to produce ribonucleases. Here we describe the rapid degradation of nuclease-stabilized RNA oligonucleotides in a human embryonic kidney 293 (HEK) cell culture contaminated with Mycoplasma fermentans, a common species of mycoplasma. RNA with 2′-fluoro-or 2′-O-methyl-modified pyrimidines was readily degraded in conditioned media from this culture, but was stable in conditioned media from uncontaminated HEK cells. RNA completely modified with 2′-O-methyls was not degraded in the mycoplasma-contaminated media. RNA zymogram analysis of conditioned culture media and material centrifuged from the media revealed several distinct protein bands (ranging from 30 to 68kDa) capable of degrading RNA with 2′-fluoro-or 2′-O-methyl-modified pyrimidines. Finally, the mycoplasma-associated nuclease was detected in material centrifuged from the contaminated culture supernatants in as little as 15 minutes with an RNA oligo-containing 2′-O-methyl-modified pyrimidines and labeled with a 5′-fluorescein amidite (FAM) and 3′-quencher. These results suggest that mycoplasma contamination may be a critical confounding variable for cell culture experiments involving RNA-based reagents, with particular relevance for applications involving naked RNA (e.g., aptamer-siRNA chimeras). © 2012 Mary Ann Liebert, Inc.
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| 8. |
- Järver, Peter, et al.
(författare)
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Peptide nanoparticle delivery of charge-neutral splice-switching morpholino oligonucleotides.
- 2015
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Ingår i: Nucleic Acid Therapeutics. - : Mary Ann Liebert Inc. - 2159-3337 .- 2159-3345. ; 25:2, s. 65-77
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Tidskriftsartikel (refereegranskat)abstract
- Oligonucleotide analogs have provided novel therapeutics targeting various disorders. However, their poor cellular uptake remains a major obstacle for their clinical development. Negatively charged oligonucleotides, such as 2'-O-Methyl RNA and locked nucleic acids have in recent years been delivered successfully into cells through complex formation with cationic polymers, peptides, liposomes, or similar nanoparticle delivery systems. However, due to the lack of electrostatic interactions, this promising delivery method has been unsuccessful to date using charge-neutral oligonucleotide analogs. We show here that lipid-functionalized cell-penetrating peptides can be efficiently exploited for cellular transfection of the charge-neutral oligonucleotide analog phosphorodiamidate morpholino. The lipopeptides form complexes with splice-switching phosphorodiamidate morpholino oligonucleotide and can be delivered into clinically relevant cell lines that are otherwise difficult to transfect while retaining biological activity. To our knowledge, this is the first study to show delivery through complex formation of biologically active charge-neutral oligonucleotides by cationic peptides.
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| 9. |
- Muñoz-Alarcón, Andrés, et al.
(författare)
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Novel Efficient Cell-Penetrating, Peptide-Mediated Strategy for Enhancing Telomerase Inhibitor Oligonucleotides
- 2015
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Ingår i: Nucleic Acid Therapeutics. - : Mary Ann Liebert Inc. - 2159-3337 .- 2159-3345. ; 25:6, s. 306-310
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Tidskriftsartikel (refereegranskat)abstract
- At present, there are several therapeutic approaches for targeting telomerase in tumors. One in particular, currently undergoing clinical trials, is based on synthetic lipid-modified oligonucleotide antagonists aimed at inhibiting the ribonucleoprotein subunit of human telomerase. However, while enabling efficient uptake, the lipid modifications reduce the potency of the therapeutic oligonucleotides compared to nonmodified oligonucleotides. Moreover, lipid modification may increase oligonucleotide accumulation in the liver causing undesirable hepatotoxicity. Noncovalent complexation strategies for cell-penetrating peptide (CPP)-mediated delivery present an option to circumvent the need for potency-reducing modifications, while allowing for a highly efficient uptake, and could significantly improve the efficiency of telomerase-targeting cancer therapeutics. Delivery of a nonlipidated locked nucleic acid/2-O-methyl mixmer significantly inhibits the telomerase activity in treated HeLa cells. The inhibitory effect was further improved through addition of a CPP. Furthermore, calculated IC50-values for the oligonucleotide delivered by CPPs into HeLa cells are more than 20 times lower than telomerase inhibitor Imetelstat, currently undergoing clinical trials. These results emphasize the potential of CPP-mediated delivery of future pharmaceuticals and provide means by which to enhance an already promising therapeutic strategy for cancer treatment.
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