Antisense Oligonucleotides Internally Labeled with Peptides Show Improved Target Recognition and Stability to Enzymatic Degradation

Maria Taskova, Charlotte S. Madsen, Knud Jørgen Jensen, Lykke Haastrup Hansen, Birte Vester, Kira Astakhova

Research output: Contribution to journalJournal articleResearchpeer-review

Abstract

Specific target binding and stability in diverse biological media is of crucial importance for applications of synthetic oligonucleotides as diagnostic and therapeutic tools. So far, these issues have been addressed by chemical modification of oligonucleotides and by conjugation with a peptide, most often at the terminal position of the oligonucleotide. Herein, we for the first time systematically investigate the influence of internally attached short peptides on the properties of antisense oligonucleotides. We report the synthesis and internal double labeling of 21-mer oligonucleotides that target the BRAF V600E oncogene, with a library of rationally designed peptides employing CuAAC "click" chemistry. The peptide sequence has an influence on the specificity and affinity of target DNA/RNA binding. We also investigated the impact of locked nucleic acids (LNAs) on the latter. Lysine residues improve binding of POCs to target DNA and RNA, whereas the distance to lysine correlates exclusively with a decrease in binding of mismatched RNA targets. Glycine and tyrosine residues affect target binding as well. Importantly, the resistance of POCs to enzymatic degradation is dramatically improved by the internal attachment of peptides but not by LNA alone. Independently of the peptide sequence, the conjugates are stable for up to 24 h in 90% human serum and duplexes of POCs with complementary DNA for up to 160 h in 90% human serum. Such excellent stability has not been previously reported for DNA and makes internally labeled POCs an exciting object of study, i.e., showing high target specificity and simultaneous stability in biological media.
Original languageEnglish
JournalBioconjugate Chemistry
Volume28
Issue number3
Pages (from-to)768-774
Number of pages7
ISSN1043-1802
DOIs
Publication statusPublished - 2017
Externally publishedYes

Cite this

Taskova, Maria ; Madsen, Charlotte S. ; Jensen, Knud Jørgen ; Hansen, Lykke Haastrup ; Vester, Birte ; Astakhova, Kira. / Antisense Oligonucleotides Internally Labeled with Peptides Show Improved Target Recognition and Stability to Enzymatic Degradation. In: Bioconjugate Chemistry. 2017 ; Vol. 28, No. 3. pp. 768-774.
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abstract = "Specific target binding and stability in diverse biological media is of crucial importance for applications of synthetic oligonucleotides as diagnostic and therapeutic tools. So far, these issues have been addressed by chemical modification of oligonucleotides and by conjugation with a peptide, most often at the terminal position of the oligonucleotide. Herein, we for the first time systematically investigate the influence of internally attached short peptides on the properties of antisense oligonucleotides. We report the synthesis and internal double labeling of 21-mer oligonucleotides that target the BRAF V600E oncogene, with a library of rationally designed peptides employing CuAAC {"}click{"} chemistry. The peptide sequence has an influence on the specificity and affinity of target DNA/RNA binding. We also investigated the impact of locked nucleic acids (LNAs) on the latter. Lysine residues improve binding of POCs to target DNA and RNA, whereas the distance to lysine correlates exclusively with a decrease in binding of mismatched RNA targets. Glycine and tyrosine residues affect target binding as well. Importantly, the resistance of POCs to enzymatic degradation is dramatically improved by the internal attachment of peptides but not by LNA alone. Independently of the peptide sequence, the conjugates are stable for up to 24 h in 90{\%} human serum and duplexes of POCs with complementary DNA for up to 160 h in 90{\%} human serum. Such excellent stability has not been previously reported for DNA and makes internally labeled POCs an exciting object of study, i.e., showing high target specificity and simultaneous stability in biological media.",
author = "Maria Taskova and Madsen, {Charlotte S.} and Jensen, {Knud J{\o}rgen} and Hansen, {Lykke Haastrup} and Birte Vester and Kira Astakhova",
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Antisense Oligonucleotides Internally Labeled with Peptides Show Improved Target Recognition and Stability to Enzymatic Degradation. / Taskova, Maria; Madsen, Charlotte S.; Jensen, Knud Jørgen; Hansen, Lykke Haastrup; Vester, Birte; Astakhova, Kira.

In: Bioconjugate Chemistry, Vol. 28, No. 3, 2017, p. 768-774.

Research output: Contribution to journalJournal articleResearchpeer-review

TY - JOUR

T1 - Antisense Oligonucleotides Internally Labeled with Peptides Show Improved Target Recognition and Stability to Enzymatic Degradation

AU - Taskova, Maria

AU - Madsen, Charlotte S.

AU - Jensen, Knud Jørgen

AU - Hansen, Lykke Haastrup

AU - Vester, Birte

AU - Astakhova, Kira

PY - 2017

Y1 - 2017

N2 - Specific target binding and stability in diverse biological media is of crucial importance for applications of synthetic oligonucleotides as diagnostic and therapeutic tools. So far, these issues have been addressed by chemical modification of oligonucleotides and by conjugation with a peptide, most often at the terminal position of the oligonucleotide. Herein, we for the first time systematically investigate the influence of internally attached short peptides on the properties of antisense oligonucleotides. We report the synthesis and internal double labeling of 21-mer oligonucleotides that target the BRAF V600E oncogene, with a library of rationally designed peptides employing CuAAC "click" chemistry. The peptide sequence has an influence on the specificity and affinity of target DNA/RNA binding. We also investigated the impact of locked nucleic acids (LNAs) on the latter. Lysine residues improve binding of POCs to target DNA and RNA, whereas the distance to lysine correlates exclusively with a decrease in binding of mismatched RNA targets. Glycine and tyrosine residues affect target binding as well. Importantly, the resistance of POCs to enzymatic degradation is dramatically improved by the internal attachment of peptides but not by LNA alone. Independently of the peptide sequence, the conjugates are stable for up to 24 h in 90% human serum and duplexes of POCs with complementary DNA for up to 160 h in 90% human serum. Such excellent stability has not been previously reported for DNA and makes internally labeled POCs an exciting object of study, i.e., showing high target specificity and simultaneous stability in biological media.

AB - Specific target binding and stability in diverse biological media is of crucial importance for applications of synthetic oligonucleotides as diagnostic and therapeutic tools. So far, these issues have been addressed by chemical modification of oligonucleotides and by conjugation with a peptide, most often at the terminal position of the oligonucleotide. Herein, we for the first time systematically investigate the influence of internally attached short peptides on the properties of antisense oligonucleotides. We report the synthesis and internal double labeling of 21-mer oligonucleotides that target the BRAF V600E oncogene, with a library of rationally designed peptides employing CuAAC "click" chemistry. The peptide sequence has an influence on the specificity and affinity of target DNA/RNA binding. We also investigated the impact of locked nucleic acids (LNAs) on the latter. Lysine residues improve binding of POCs to target DNA and RNA, whereas the distance to lysine correlates exclusively with a decrease in binding of mismatched RNA targets. Glycine and tyrosine residues affect target binding as well. Importantly, the resistance of POCs to enzymatic degradation is dramatically improved by the internal attachment of peptides but not by LNA alone. Independently of the peptide sequence, the conjugates are stable for up to 24 h in 90% human serum and duplexes of POCs with complementary DNA for up to 160 h in 90% human serum. Such excellent stability has not been previously reported for DNA and makes internally labeled POCs an exciting object of study, i.e., showing high target specificity and simultaneous stability in biological media.

U2 - 10.1021/acs.bioconjchem.6b00567

DO - 10.1021/acs.bioconjchem.6b00567

M3 - Journal article

VL - 28

SP - 768

EP - 774

JO - Bioconjugate Chemistry

JF - Bioconjugate Chemistry

SN - 1043-1802

IS - 3

ER -