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Keywords = phosphoryl guanidine group

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14 pages, 2162 KB  
Article
Chemical Modifications Influence the Number of siRNA Molecules Adsorbed on Gold Nanoparticles and the Efficiency of Downregulation of a Target Protein
by Anna V. Epanchintseva, Julia E. Poletaeva, Anton S. Dome, Ilya S. Dovydenko, Inna A. Pyshnaya and Elena I. Ryabchikova
Nanomaterials 2022, 12(24), 4450; https://doi.org/10.3390/nano12244450 - 14 Dec 2022
Cited by 6 | Viewed by 2314
Abstract
Small interfering RNAs (siRNAs) are a powerful tool for specific suppression of protein synthesis in the cell, and this determines the attractiveness of siRNAs as a drug. Low resistance of siRNA to nucleases and inability to enter into target cells are the most [...] Read more.
Small interfering RNAs (siRNAs) are a powerful tool for specific suppression of protein synthesis in the cell, and this determines the attractiveness of siRNAs as a drug. Low resistance of siRNA to nucleases and inability to enter into target cells are the most crucial issues in developing siRNA-based therapy. To face this challenge, we designed multilayer nanoconstruct (MLNC) with AuNP core bearing chemically modified siRNAs. We applied chemical modifications 2′-OMe and 2′-F substitutions as well as their combinations with phosphoryl guanidine group in the internucleotide phosphate. The effect of modification on the efficiency of siRNA loading into nanocarriers was examined. The introduction of the internucleotide modifications into at least one of the strands raised the efficiency of siRNA adsorption on the surface of gold core. We also tested the stability of modified siRNA adsorbed on gold core in the presence of serum. Based on loading efficiency and stability, MLNCs with the most siRNA effective cargo were selected, and they showed an increase in biological activity compared to control MLNCs. Our study demonstrated the effect of chemical modifications of siRNA on its binding to the AuNP-based carrier, which directly affects the efficiency of target protein expression inhibition. Full article
(This article belongs to the Special Issue Advances in Nanoscale Materials in Biomedicine)
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16 pages, 3745 KB  
Article
An Influence of Modification with Phosphoryl Guanidine Combined with a 2′-O-Methyl or 2′-Fluoro Group on the Small-Interfering-RNA Effect
by Anna S. Pavlova, Kristina I. Yakovleva, Anna V. Epanchitseva, Maxim S. Kupryushkin, Inna A. Pyshnaya, Dmitrii V. Pyshnyi, Elena I. Ryabchikova and Ilya S. Dovydenko
Int. J. Mol. Sci. 2021, 22(18), 9784; https://doi.org/10.3390/ijms22189784 - 10 Sep 2021
Cited by 14 | Viewed by 4518
Abstract
Small interfering RNA (siRNA) is the most important tool for the manipulation of mRNA expression and needs protection from intracellular nucleases when delivered into the cell. In this work, we examined the effects of siRNA modification with the phosphoryl guanidine (PG) group, which, [...] Read more.
Small interfering RNA (siRNA) is the most important tool for the manipulation of mRNA expression and needs protection from intracellular nucleases when delivered into the cell. In this work, we examined the effects of siRNA modification with the phosphoryl guanidine (PG) group, which, as shown earlier, makes oligodeoxynucleotides resistant to snake venom phosphodiesterase. We obtained a set of siRNAs containing combined modifications PG/2′-O-methyl (2′-OMe) or PG/2′-fluoro (2′-F); biophysical and biochemical properties were characterized for each duplex. We used the UV-melting approach to estimate the thermostability of the duplexes and RNAse A degradation assays to determine their stability. The ability to induce silencing was tested in cultured cells stably expressing green fluorescent protein. The introduction of the PG group as a rule decreased the thermodynamic stability of siRNA. At the same time, the siRNAs carrying PG groups showed increased resistance to RNase A. A gene silencing experiment indicated that the PG-modified siRNA retained its activity if the modifications were introduced into the passenger strand. Full article
(This article belongs to the Special Issue Precision Nucleic Acid Therapeutics)
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14 pages, 1156 KB  
Article
Allele-Specific PCR for KRAS Mutation Detection Using Phosphoryl Guanidine Modified Primers
by Alexey S. Chubarov, Igor P. Oscorbin, Maxim L. Filipenko, Alexander A. Lomzov and Dmitrii V. Pyshnyi
Diagnostics 2020, 10(11), 872; https://doi.org/10.3390/diagnostics10110872 - 26 Oct 2020
Cited by 32 | Viewed by 7043
Abstract
Establishing the Kirsten rat sarcoma (KRAS) mutational status is essential in terms of managing patients with various types of cancer. Allele-specific real-time polymerase chain reaction (AS-PCR) is a widely used method for somatic mutations detection. To improve the limited sensitivity and specificity, [...] Read more.
Establishing the Kirsten rat sarcoma (KRAS) mutational status is essential in terms of managing patients with various types of cancer. Allele-specific real-time polymerase chain reaction (AS-PCR) is a widely used method for somatic mutations detection. To improve the limited sensitivity and specificity, several blocking methods have been introduced in AS-PCR to block the amplification of wild-type templates. Herein, we used a novel modified oligonucleotide with internucleotide phosphates reshaped 1,3-dimethyl-2-imino-imidazolidine moieties (phosphoryl guanidine (PG) groups) as primers and blockers in the AS-PCR method. Four common KRAS mutations were chosen as a model to demonstrate the advantages of the PG primers and blockers utilizing a customized PCR protocol. The methods were evaluated on plasmid model systems providing a KRAS mutation detection limit of 20 copies of mutant DNA in a proportion as low as 0.1% of the total DNA, with excellent specificity. PG-modification can serve as the universal additional mismatch-like disturbance to increase the discrimination between wild-type and mutated DNA. Moreover, PG can serve to increase primer specificity by a synergetic effect with additional mismatch and would greatly facilitate medical research. Full article
(This article belongs to the Section Pathology and Molecular Diagnostics)
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