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Plashkevych, O
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Publications (10 of 32) Show all publications
Moriou, C., Da Silva, A. D., Vianelli Prado, M. J., Denhez, C., Plashkevych, O., Chattopadhyaya, J., . . . Clivio, P. (2018). C2 '-F Stereoconfiguration As a Puckering Switch for Base Stacking at the Dinucleotide Level. Journal of Organic Chemistry, 83(4), 2473-2478
Open this publication in new window or tab >>C2 '-F Stereoconfiguration As a Puckering Switch for Base Stacking at the Dinucleotide Level
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2018 (English)In: Journal of Organic Chemistry, ISSN 0022-3263, E-ISSN 1520-6904, Vol. 83, no 4, p. 2473-2478Article in journal (Refereed) Published
Abstract [en]

Fluorine configuration at C2′ of the bis(2′-fluorothymidine) dinucleotide is demonstrated to drive intramolecular base stacking. 2′-β F-Configuration drastically reduces stacking compared to the 2′-α series. Hence, base stacking emerges as being tunable by the C2′-F stereoconfiguration through dramatic puckering variations scrutinized by NMR and natural bond orbital analysis. Accordingly, 2′-β F-isomer photoreactivity is significantly reduced compared to that of the 2′-α F-isomer.

Place, publisher, year, edition, pages
AMER CHEMICAL SOC, 2018
National Category
Organic Chemistry
Identifiers
urn:nbn:se:uu:diva-351016 (URN)10.1021/acs.joc.7b03186 (DOI)000427094200085 ()29364674 (PubMedID)
Funder
Swedish Research CouncilEU, European Research CouncilNational Supercomputer Centre (NSC), Sweden
Available from: 2018-05-24 Created: 2018-05-24 Last updated: 2018-05-25Bibliographically approved
Plashkevych, O., Li, Q. & Chattopadhyaya, J. (2017). How RNase HI (Escherichia coli) promoted site-selective hydrolysis works on RNA in duplex with carba-LNA and LNA substituted antisense strands in an antisense strategy context?. Molecular Biosystems, 13(5), 921-938
Open this publication in new window or tab >>How RNase HI (Escherichia coli) promoted site-selective hydrolysis works on RNA in duplex with carba-LNA and LNA substituted antisense strands in an antisense strategy context?
2017 (English)In: Molecular Biosystems, ISSN 1742-206X, E-ISSN 1742-2051, Vol. 13, no 5, p. 921-938Article in journal (Refereed) Published
Abstract [en]

A detailed kinetic study of 36 single modified AON-RNA heteroduplexes shows that substitution of a single native nucleotide in the antisense strand (AON) by locked nucleic acid (LNA) or by diastereomerically pure carba-LNA results in site-dependent modulation of RNase H promoted cleavage of complementary mRNA strands by 2 to 5 fold at 5'-GpN-3' cleavage sites, giving up to 70% of the RNA cleavage products. The experiments have been performed using RNase HI of Escherichia coli. The 2nd best cleavage site, being the 5'-ApN-3' sites, cleaves up to 23%, depending upon the substitution site in 36 isosequential complementary AONs. A comparison of the modified AON promoted RNA cleavage rates with that of the native AON shows that sequence-specificity is considerably enhanced as a result of modification. Clearly, relatively weaker 5'-purine (Pu)-pyrimidine (Py)-3' stacking in the complementary RNA strand is preferred (giving similar to 90% of total cleavage products), which plays an important role in RNase H promoted RNA cleavage. A plausible mechanism of RNase H mediated cleavage of the RNA has been proposed to be two-fold, dictated by the balancing effect of the aromatic character of the purine aglycone: first, the locally formed 9-guanylate ion (pK(a) 9.3, similar to 18-20% N1 ionized at pH 8) alters the adjoining sugar-phosphate backbone around the scissile phosphate, transforming its sugar N/S conformational equilibrium, to preferential S-type, causing preferential cleavage at 5'-GpN-3' sites around the center of 20 mer complementary mRNA. Second, the weaker nearest-neighbor strength of 50-Pu-p-Py-30 stacking promotes preferential 5'-GpN-3' and 5'-ApN-3' cleavage, providing similar to 90% of the total products, compared to similar to 50% in that of the native one, because of the cLNA/ LNA substituent effect on the neighboring 50-Pu-p-Py-30 sites, providing both local steric flexibility and additional hydration. This facilitates both the water and water/Mg-2+ ion availability at the cleavage site causing sequence-specific hydrolysis of the phosphodiester bond of scissile phosphate. The enhancement of the total rate of cleavage of the complementary mRNA strand by up to 25%, presented in this work, provides opportunities to engineer a single modification site in appropriately substituted AONs to design an effective antisense strategy based on the nucleolytic stability of the AON strand versus RNase H capability to cleave the complementary RNA strand.

National Category
Cell and Molecular Biology
Identifiers
urn:nbn:se:uu:diva-323445 (URN)10.1039/c6mb00762g (DOI)000400659600011 ()28352859 (PubMedID)
Funder
Swedish Research Council
Available from: 2017-07-04 Created: 2017-07-04 Last updated: 2018-01-13Bibliographically approved
Plashkevych, O. & Chattopadhyaya, J. (2011). Molecular Structure of the Core-Modified siRNA Duplexes Containing Diastereomeric Pair of [C6′(R)-OH]- versus [C6′(S)-OH]-carba-LNAs Suggests a Model for RNAi Action. Nucleosides, Nucleotides & Nucleic Acids, 30(11), 815-825
Open this publication in new window or tab >>Molecular Structure of the Core-Modified siRNA Duplexes Containing Diastereomeric Pair of [C6′(R)-OH]- versus [C6′(S)-OH]-carba-LNAs Suggests a Model for RNAi Action
2011 (English)In: Nucleosides, Nucleotides & Nucleic Acids, ISSN 1525-7770, E-ISSN 1532-2335, Vol. 30, no 11, p. 815-825Article in journal (Refereed) Published
Abstract [en]

Molecular structures of native and a pair of modified small interfering RNA-RNA duplexes containing carbocyclic [6'-(R)-OH/7'-(S)-methyl]- and [6'-(S)-OH/7'-(S)-methyl]-carba-LNA-thymine nucleotides, which are two diastereomeric analogs of the native T nucleotide, incorporated at position 13 in the antisense (AS) strand of siRNA, have been simulated using molecular mechanics/dynamics techniques. The main aim of the project has been to find a plausible structural explanation of why modification of siRNA at T(13) position by the [6'(R)-O-(p-Toluoyl)-7' (S)-methyl]-carba-LNA-Thymine [IC(50) of 3.32 +/- 0.17 nM] is ca 24 times more active as an RNA silencing agent against the target HIV-1 TAR RNA than the [6' (S)-O-(p-Toluoyl)-7' (S)-methyl]-counterpart [IC(50) of 79.8 +/- 17 nM] [1]. The simulations reveal that introduction of both C6' (R)-OH and C6' (S)-OH stereoisomers does not lead even to local perturbation of the siRNA-RNA duplex structures compared to the native, and the only significant difference between 6' (S)- and 6' (R)-diastereomers found is the exposure of the 6'-OH group of the 6' (R)-diastereoisomer toward the edge of the duplex while the 6'-hydroxyl group of the 6' (S)-diastereoisomer is somewhat buried in the minor groove of the duplex. This rules out a hypothesis about any possible local distortion by the nature of chemical modification of the siRNA-target the RNA duplex, which might have influenced the formation of the effective RNA silencing complex (RISC) and puts some weight on the hypothesis about the 6'-hydroxy group being directly involved with most probably Ago protein, since it is known from exhaustive X-ray studies [2, 3] that the core residues are indeed involved with hydrogen bonding with the internucleotidyl phosphates.

Keywords
siRNA, chemical modification, carba-LNA, molecular, structure, RNAi
National Category
Cell Biology
Identifiers
urn:nbn:se:uu:diva-172201 (URN)10.1080/15257770.2011.586956 (DOI)000298738200001 ()
Available from: 2012-04-03 Created: 2012-04-02 Last updated: 2017-12-07Bibliographically approved
Dutta, S., Bhaduri, N., Upadhayaya, R. S., Rastogi, N., Chandel, S. G., Vandavasi, J. K., . . . Chattopadhyaya, J. (2011). The R-diastereomer of 6 '-O-toluoyl-carba-LNA modification in the core region of siRNA leads to 24-times improved RNA silencing potency against the HIV-1 compared to its S-counterpart. MedChemComm, 2(11), 1110-1119
Open this publication in new window or tab >>The R-diastereomer of 6 '-O-toluoyl-carba-LNA modification in the core region of siRNA leads to 24-times improved RNA silencing potency against the HIV-1 compared to its S-counterpart
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2011 (English)In: MedChemComm, ISSN 2040-2503, E-ISSN 2040-2511, Vol. 2, no 11, p. 1110-1119Article in journal (Refereed) Published
Abstract [en]

The modified siRNA with pure [6'(S)-O-(p-toluoyl)-7'(S)-methyl]-carba-LNA [6'(S)-O-toluoyl-jcLNA] at position T(13) displayed an IC(50) of 79.8 nM, which has been found to be nearly 24-times less potent as a HIV-1 RNAi silencing agent against TAR RNA than that of the corresponding pure [6'(R)-O-(ptoluoyl)-7'(S)-methyl]jcLNA [6'(R)-O-(p-toluoyl)-jcLNA] counterpart [IC(50) 3.3 nM]. The later [6'(R)-O-(p-toluoyl)-jcLNAl-modified siRNAs have been found to be nearly 2-fold more efficient as a silencing agent than the corresponding 6'-deoxy-jcLNA modified siRNA [IC(50) 8.1 nM], and also nearly 3-fold more effective as a silencing agent than that of LNA-modified siRNA [IC(50) 11.7 nM], thereby showing that the 6'-carbon center in the jcLNA-modified siRNA in the core region is relatively more exposed to the Ago protein in the RISC with a clear chirality preference for the siRNA cleavage reaction. It is noteworthy that the IC(50) of jcLNA-modified siRNAs are very comparable to that of the native siRNA [1.8 nM]. The jcLNA derivatized siRNAs, however, have a clear advantage of being, in general, considerably more stable in human serum. The main structural difference in duplexes of the antisense strand of the 6'(R or S)-O-(p-toluoyl)-jcLNA modified siRNA and target RNA duplex is found to be the spatial orientation of the 6'(R)-O-toluoyl group, which is exposed towards the edge of the duplex backbone, while the 6'(S) makes the minor groove relatively inaccessible for the Ago protein in the RISC. Clearly, any further C6'-modification in jcLNA-modified siRNAs with any hydrophobic group for tighter binding and cleavage or for cross-linking in the core region should preferably be done in the 6'(R)-stereochemistry.

National Category
Natural Sciences
Identifiers
urn:nbn:se:uu:diva-166735 (URN)10.1039/c1md00167a (DOI)000297960100013 ()
Available from: 2012-01-13 Created: 2012-01-13 Last updated: 2017-12-08Bibliographically approved
Li, Q., Yuan, F., Zhou, C., Plashkevych, O. & Chattopadhyaya, J. (2010). Free-Radical Ring Closure to Conformationally Locked alpha-L-Carba-LNAs and Synthesis of Their Oligos: Nuclease Stability, Target RNA Specificity, and Elicitation of RNase H. Journal of Organic Chemistry, 75(18), 6122-6140
Open this publication in new window or tab >>Free-Radical Ring Closure to Conformationally Locked alpha-L-Carba-LNAs and Synthesis of Their Oligos: Nuclease Stability, Target RNA Specificity, and Elicitation of RNase H
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2010 (English)In: Journal of Organic Chemistry, ISSN 0022-3263, E-ISSN 1520-6904, Vol. 75, no 18, p. 6122-6140Article in journal (Refereed) Published
Abstract [en]

A new class of conformationally constrained nucleosides, alpha-L-ribo-carbocyclic LNA thymidine (alpha-L-carba-LNA-T, LNA is an abbreviation of locked nucleic acid) analogues and a novel "double-locked" alpha-L-ribo-configured tetracyche thymidine (6,7'-methylene-bridged-alpha-L-carba-LNA-T) in which both the sugar puckering and glyeosidic torsion are simultaneously constrained, have been synthesized through a key step involving 5-exo free-radical intramolecular cyclization. These alpha-L-carba-LNA analogues have been subsequently transformed to corresponding phosphoramidites and incorporated into isosequential antisense oligonucleotides (AONs), which have then been examined for the thermal denaturation of their duplexes, nuclease stability, and RNase H recruitment capabilities. Introduction of a single 6',7'-substituted alpha-L-carba-LNA-T modification in the AON strand of AON/RNA heteroduplex led to T-m reduction by 2-3 degrees C as compared to the native heteroduplex, whereas the parent 2'-oxa-alpha-L-LNA-T modification at the identical position in the AON strand has been found to lead to an increase in the T-m by 3-5 degrees C. This suggests that the 6' and 7' substitutions lead to much reduced thermal stability for the modified heteroduplex, especially the hydrophobic 7'-methyl on alpha-L-carba-LNA, which is located in the major groove of the duplex. All of the AONs incorporating 6',7'-substituted alpha-L-earba-LNA-T have, however, showed considerably improved nuclease stability toward 3'-exonuclease (SVPDE) and in human blood serum compared to the 2'-oxa-alpha-L-LNA-T incorporated AONs. The hybrid duplexes that are formed by 6',7'-substituted alpha-L-carba-LNA-T-modified AONs with complementary RNA have been found to recruit RNase H with higher efficiency than those of the beta-D-LNA-T or beta-D-carba-LNA-T-modified counterparts. These greatly improved nuclease resistances and efficient RNasc H recruitment capabilities elevate the alpha-L-carba-LNA-modified nucleotides into a new class of locked nucleic acids for potential RNA targeting therapeutics.

National Category
Biological Sciences
Identifiers
urn:nbn:se:uu:diva-134911 (URN)10.1021/jo100900v (DOI)000281585600008 ()
Available from: 2010-12-02 Created: 2010-12-02 Last updated: 2017-12-12Bibliographically approved
Li, Q., Yuan, F., Zhou, C., Plashkevych, O. & Chattopadhyaya, J. (2010). Free-Radical Ring Closure to Conformationally Locked α-l-Carba-LNAs and Synthesis of Their Oligos:: Nuclease Stability, Target RNA Specificity, and Elicitation of RNase H. Journal of Organic Chemistry, 75(18), 6122-6140
Open this publication in new window or tab >>Free-Radical Ring Closure to Conformationally Locked α-l-Carba-LNAs and Synthesis of Their Oligos:: Nuclease Stability, Target RNA Specificity, and Elicitation of RNase H
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2010 (English)In: Journal of Organic Chemistry, ISSN 0022-3263, E-ISSN 1520-6904, Vol. 75, no 18, p. 6122-6140Article in journal (Refereed) Published
Abstract [en]

A new class of conformationally constrained nucleosides, α-L-ribo-carbocyclic LNA thymidine (α-L-carba-LNA-T, LNA is an abbreviation of locked nucleic acid) analogues and a novel "double-locked" α-L-ribo-configured tetracyclic thymidine (6,7'-methylene-bridged-α-L-carba-LNA-T) in which both the sugar puckering and glycosidic torsion are simultaneously constrained, have been synthesized through a key step involving 5-exo free-radical intramolecular cyclization. These α-L-carba-LNA analogues have been subsequently transformed to corresponding phosphoramidites and incorporated into isosequential antisense oligonucleotides (AONs), which have then been examined for the thermal denaturation of their duplexes, nuclease stability, and RNase H recruitment capabilities. Introduction of a single 6',7'-substituted α-L-carba-LNA-T modification in the AON strand of AON/RNA heteroduplex led to T(m) reduction by 2-3 °C as compared to the native heteroduplex, whereas the parent 2'-oxa-α-L-LNA-T modification at the identical position in the AON strand has been found to lead to an increase in the T(m) by 3-5 °C. This suggests that the 6' and 7' substitutions lead to much reduced thermal stability for the modified heteroduplex, especially the hydrophobic 7'-methyl on α-L-carba-LNA, which is located in the major groove of the duplex. All of the AONs incorporating 6',7'-substituted α-L-carba-LNA-T have, however, showed considerably improved nuclease stability toward 3'-exonuclease (SVPDE) and in human blood serum compared to the 2'-oxa-α-L-LNA-T incorporated AONs. The hybrid duplexes that are formed by 6',7'-substituted α-L-carba-LNA-T-modified AONs with complementary RNA have been found to recruit RNase H with higher efficiency than those of the β-D-LNA-T or β-D-carba-LNA-T-modified counterparts. These greatly improved nuclease resistances and efficient RNase H recruitment capabilities elevate the α-L-carba-LNA-modified nucleotides into a new class of locked nucleic acids for potential RNA targeting therapeutics.

Place, publisher, year, edition, pages
U. S. A: American Chemical Society (ACS), 2010
Keywords
conformationally constrained nucleoside, free-radical intramolecular cyclization, antisense oligonucleotide, RNA target therapeutic
National Category
Organic Chemistry
Research subject
Chemistry
Identifiers
urn:nbn:se:uu:diva-176562 (URN)10.1021/jo100900v (DOI)
Available from: 2012-06-20 Created: 2012-06-20 Last updated: 2017-12-07Bibliographically approved
Upadhayaya, R. S., Shinde, P. D., Sayyed, A. Y., Kadam, S. A., Bawane, A. N., Poddar, A., . . . Chattopadhyaya, J. (2010). Synthesis and structure of azole-fused indeno[2,1-c]quinolines and their anti-mycobacterial properties. Organic and biomolecular chemistry, 8(24), 5661-5673
Open this publication in new window or tab >>Synthesis and structure of azole-fused indeno[2,1-c]quinolines and their anti-mycobacterial properties
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2010 (English)In: Organic and biomolecular chemistry, ISSN 1477-0520, E-ISSN 1477-0539, Vol. 8, no 24, p. 5661-5673Article in journal (Refereed) Published
Abstract [en]

Prompted by our discovery of a new class of conformationally-locked indeno[2,1-c]quinolines as anti-mycobacterials, compounds 2a and 3a (Fig. 1; MIC < 0.39 mu g mL(-1) and 0.78 mu g mL(-1), respectively)(14) with a freely rotating C2-imidazolo substituent, we herein describe the synthesis of pentacyclic azole-fused quinoline derivatives 4 and 5, in which we have restricted the rotation of the C2-imidazolo moiety by fusing it to the adjacent quinoline-nitrogen to give a five-membered fused azole heterocycle. The idea of locking the flexibility of the system by conformational constraint was simply to reduce its entropy, thereby reducing the overall free-energy of its binding to the target receptor. Out of 22 different azole-fused indeno[2,1-c] quinoline derivatives, seven structurally distinct compounds, 9, 15, 17, 25, 27, 28 and 29, have shown 79-99% growth inhibition of Mycobacterium tuberculosis H37Rv at a fixed dose of 6.25 mu g mL(-1). The efficacies of these compounds were evaluated in vitro for 8/9 consecutive days using the BACTEC radiometric assay upon administration of single dose on day one. Of these, two compounds, 9 and 28, inhibited growth of M. tuberculosis very effectively at MIC < 0.39 mu g mL(-1) (0.89 mu M and 1 mu M, respectively). These active compounds 9, 15, 17, 25, 27, 28 and 29 were screened for their cytotoxic effect on mammalian cells (human monocytic cell line U937), which showed that the human cell survival is almost unperturbed (100% survival), except for compound 25, hence these new compounds with new scaffolds have been identified as potent anti-mycobacterials, virtually with no toxicity. Thus these "hit" molecules constitute our important "leads" for further optimization by structure-activity relationship against TB.

National Category
Biological Sciences
Identifiers
urn:nbn:se:uu:diva-139386 (URN)10.1039/c0ob00445f (DOI)000284610600023 ()20927480 (PubMedID)
Available from: 2010-12-27 Created: 2010-12-23 Last updated: 2017-12-11Bibliographically approved
Zhou, C., Plashkevych, O. & Chattopadhyaya, J. (2009). Double Sugar and Phosphate Backbone-constrained Nucleotides: Synthesis, Structure, Stability and Their Incorporation into Oligodeoxynucleotides. Journal of Organic Chemistry, 74(9), 3248-3265
Open this publication in new window or tab >>Double Sugar and Phosphate Backbone-constrained Nucleotides: Synthesis, Structure, Stability and Their Incorporation into Oligodeoxynucleotides
2009 (English)In: Journal of Organic Chemistry, ISSN 0022-3263, E-ISSN 1520-6904, Vol. 74, no 9, p. 3248-3265Article in journal (Refereed) Published
Abstract [en]

Two diastereomerically pure carba-LNA dioxaphosphorinane nucleotides [(S-p)- or (R-p)-D-2-CNA], simultaneously conformationally locked at the sugar and the phosphate backbone, have been designed and synthesized. Structural studies by NMR as well as by ab initio   calculations showed that in (S-p)- and (R-p)-D-2-CNA the Mowing occur: (i) the sugar is locked in extreme North-type conformation with P = 11 degrees and Phi(m) (ii) the six-membered 1,3,2-dioxaphosphorinane ring adopts a half-chair conformation; (iii) the fixed phosphate backbone delta, epsilon, and zeta torsions were found to be delta [gauch(+)],   epsilon (cis), zeta[anticlinal(+)] for (S-p)-D-2-CNA, and delta [gaitche(+)], epsilon(cis), zeta[anticlittal(-)] for (R-p)-D-2-CNA. It   has been found that F- ion can catalyze the isomerization of pure (S-p)-D-2-CNA or (R-p)-D-2-CNA to give an equilibrium mixture (K =   1.94). It turned out that at equilibrium concentration the (S-p)-D-2-CNA isomer is preferred over the (R-p)-D-2-CNA isomer by 0.39 kcal/mol. The chemical reactivity of the six-membered   dioxaphosphorinane ring in D-2-CNA was found to be dependent on the   internucleotidic phosphate stereochemistry. Thus, both (Sp)- and  (Rp)-D2-CNA dimers (17a and 17b) were very labile toward nucleophile attack in concentrated aqueous ammonia [t(1/2) = 12 and 6 min, respectively] to give carba-LNA-6',5'-phosphodiester (21) approximate   to 70-90%, carba-LNA-3',5'-phosphodiester (22) approximate to 10%, and   carba-LNA-6',3'-phosphodiester (23) < 10%. In contrasts the (S-p)-D-2-CNA was about 2 times more stable than (Rp)-D2-CNA under hydrazine hydrate/pyficfine/AcOH (pH = 5.6) [t(1/2) = 178 and 99 h, respectively], which was exploited in the deprotection of pure (S-p)-D-2-CNA incorporated antisense oligodeoxynucleotides (AON). Thus, after removal of the solid supports from the (S-p)-D-2-CNA-modified AON by BDU/MeCN, they were treated with hydrazine hydrate in pyridine/AcOH to give pure AONs in 35-40% yield, which was unequivocally   characterized by MALDI-TOF to show that they have an intact six-membered dioxaphosphorinane ring. The effect of pure (S-p)-D-2-CNA   niodification in the AONs was estimated by complexing to the complementary RNA and DNA strands by the thermal denaturation studies. This showed that this cyclic phosphotriester modification destabilizes   the AON/DNA and AON/RNA duplex by about -6 to -9 degrees C/modification. Treatment of (Sp)-D-2-CNA-modified AON with concentrated aqueous ammonia gave cwba-LNA-6',5'-phosphodiester modified AON (similar to 80%) plus a small amount of carba-LNA-3',5'-Phosphodiester-modified AON (similar to 20%). It is noteworthy that Carba-LNA-3',5'-phosphodiester modification stabilized  the AON/RNA duplex by +4 degrees C/modificafion (J. Org. Chem. 2009, 74, 118), whereas carba-LNA-6', 5'-phosphodiester modification   destabilizes both AON/RNA and AON/DNA significantly (by -10 to -19 degrees C/modification), which, as shown in our comparative CD studies, that the cyclic phosphotriester modified AONs as well as carba-LNA-6'.5'-phosphodiester modified AONs are much more weakly   stacked than carba-LNA-3',5'-phosphodiester-modified AONs.

National Category
Biological Sciences
Identifiers
urn:nbn:se:uu:diva-106721 (URN)10.1021/jo900391n (DOI)000265554900002 ()
Available from: 2009-06-29 Created: 2009-06-29 Last updated: 2017-12-13
Zhou, C., Liu, Y., Andaloussi, M., Badgujar, N., Plashkevych, O. & Chattopadhyaya, J. (2009). Fine Tuning of Electrostatics Around the Internucleotidic Phosphate through Incorporations of Functionalized 2', 4'-Carbocyclic-LNAs and –ENAs Lead to Significant Modulation of Antisense Properties. Journal of Organic Chemistry, 74(1), 118-134
Open this publication in new window or tab >>Fine Tuning of Electrostatics Around the Internucleotidic Phosphate through Incorporations of Functionalized 2', 4'-Carbocyclic-LNAs and –ENAs Lead to Significant Modulation of Antisense Properties
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2009 (English)In: Journal of Organic Chemistry, ISSN 0022-3263, E-ISSN 1520-6904, Vol. 74, no 1, p. 118-134Article in journal (Refereed) Published
Abstract [en]

In the antisense (AS) and RNA interference (RNAi) technologies, the native single-stranded 2'-deoxyoligonucleotides (for AS) or double-stranded RNA (for RNAi) are chemically modified to bind to the target RNA in order to give improved downregulation of gene expression   through inhibition of RNA translation. It is shown here how the fine adjustment of the electrostatic interaction by alteration of the substituents as well as their stereochemical environment around the intemucleotidic phosphodiester moiety near the edge of the minor grove of the antisense oligonucleotides (AON)-RNA heteroduplex can lead to the modulation of the antisense properties. This was demonstrated through the synthesis of various modified carbocyclic-locked nucleic acids (LNAs) and -ethylene-bridged nucleic acids (ENAs) with hydroxyl and/or methyl substituents attached at the carbocyclic pan and their   integration into AONs by solid-phase DNA synthesis. The target affinity toward the complementary RNA and DNA, nuclease resistance, and RNase H elicitation by these modified AONs showed that both the nature of the modification (-OH versus -CH3) and their respective stereochemical orientations vis-a-vis vicinal phosphate play a very important role in modulating the AON properties. Whereas the affinity to the target RNA and the enzymatic stability of AONs were not favored by the hydrophobic and sterically bulky modifications in the center of the minor groove, their positioning at the edge of the minor groove near the phosphate linkage resulted in significantly improved nuclease resisitance without of target affinity. On the other hand, hydrophilic modification, such as a hydroxyl group, close to the phosphate linkage made the internucleotidic phosphodiester especially nucleolytically unstable, and hence was not recommended. The substitutions on the carbocyclic moiety of the carba-LNA and -ENA did not affect significantly the choice of the cleavage sites of RNase H mediated RNA cleavage in the AON/RNA hybrid duplex, but the cleavage rate depended on the modification site in the AON sequence. If the original preferred cleavage site by RNase H was included in the 4-5nt stretch from the 3'-end of the modification site in the AON, decreassed cleavage rate was observed. Upon screening of 52 modified AONs, containing 13 differently modified derivatives at C6' and C7' (or CS') of the carba-LNAs and -ENAs, two excellent modifications in the carba-LNA series were identified, which synergistically gave outstanding antisense properties such as the target RNA affinity, nuclease resistance, and RNase H activity and were deemed to be ideal candidates as potential antisense or siRNA therapeutic agents.

National Category
Biological Sciences Natural Sciences
Identifiers
urn:nbn:se:uu:diva-106719 (URN)10.1021/jo8016742 (DOI)000262004000015 ()
Available from: 2009-06-29 Created: 2009-06-29 Last updated: 2017-12-13
Zhou, C., Plashkevych, O., Liu, Y., Badgujar, N. & Chattopadhyaya, J. (2009). Synthesis and Structure of New Methylene-bridged hexopyranosyl nucleoside (BHNA). Heterocycles, 78(7), 1715-1728
Open this publication in new window or tab >>Synthesis and Structure of New Methylene-bridged hexopyranosyl nucleoside (BHNA)
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2009 (English)In: Heterocycles, ISSN 0385-5414, E-ISSN 1881-0942, Vol. 78, no 7, p. 1715-1728Article in journal (Refereed) Published
Abstract [en]

A new member of hexopyranosyl nucleoside family, methylene-bridged   hexopyranosyl nucleoside (BHNA), has been synthesized through   generation of carbon radical at C6' in [6'S-Me, 7'S-Me]-carba-LNA T  nucleoside, followed by rearrangement to C4' radical which was quenched   by hydrogen atom to give BHNA. The stereoelectronic requirement for   this unusual radical rearrangement has been elucidated by chemical   model building and ab intio calculations to show that the coplanarity of the single electron occupied p-orbital at C6' with sigma*(O4'-C4')   plays an important role for the rearrangement reaction to take place.   The solution structure of BHNA has also been studied using NMR as well   as by ab initio calculations. The new six-membered pyranosyl ring in  BHNA, unlike other known hexopyranosyl nucleosides, adopts a twist   conformation, with base moiety occupying the axial position while  3'-hydroxymethyl and 4'-hydroxyl occupying the equatorial position.

Keywords
Conformationally Constrained Nucleoside, Hexopyranosyl Nucleoside, Radical Rearrangement, Conformational Analysis, ab initio Calculation
National Category
Biological Sciences
Identifiers
urn:nbn:se:uu:diva-106720 (URN)10.3987/COM-09-11660 (DOI)000268441500003 ()
Available from: 2009-06-29 Created: 2009-06-29 Last updated: 2017-12-13
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