Properties of nicked and circular dumbbell RNA/DNA chimeric oligonucleotides containing antisense phosphodiester oligodeoxynucleotides

Yamakawa, Hidefumi; Abe, Takayuki; Saito, Takeshi; Takai, Kazuyuki; Yamamoto, Naoki; Takaku, Hiroshi

doi:10.1016/s0968-0896(98)00060-1

Cited by 11 publications

(3 citation statements)

References 27 publications

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“…RNA hairpins have functions in initiating folding and forming tertiary structures and protein binding sites [2,3]; DNA hairpins are involved in regulating replication and transcription [4,5]. Hairpin loops are attractive candidates for the design of antisense therapeutics [6][7][8]. Understanding the factors underlying the stability of this conformation has received a great deal of attention [9][10][11].…”

Section: Introductionmentioning

confidence: 99%

The effect of hydrostatic pressure on the thermal stability of DNA hairpins

Amiri

Macgregor

2011

Biophysical Chemistry

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Section: Introductionmentioning

confidence: 99%

The effect of hydrostatic pressure on the thermal stability of DNA hairpins

Amiri

Macgregor

2011

Biophysical Chemistry

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“…Biochemical syntheses have been described based on enzymatic ligation of either a pair of hairpinlike oligonucleotides with short sticky ends or a single self-complementary, circularizable oligonucleotide (Wemmer and Benight, 1985;Erie et al, 1989;Chu and Orgel, 1992;Clusel et al, 1993;Hunt, 1994, 1997;Yamakawa et al, 1998;Hosoya et al, 1999). Both these approaches are quite effective for assembly of DNA dumbbells with duplex stems measuring 8-25 bp.…”

Section: Introduction Dmentioning

confidence: 99%

High-Purity Preparation of a Large DNA Dumbbell

Kuhn

Frank‐Kamenetskii²,

Demidov³

2001

Antisense and Nucleic Acid Drug Development

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We report on the efficient biochemical synthesis of a large DNA dumbbell starting from a pair of short DNA hairpins with long single-stranded tails of arbitrary sequence. The DNA dumbbell is obtained by enzymatic ligation yielding a 94-bp duplex stem closed at both termini by single-stranded loops of 5 nt. Following ligation, all unligated precursors and monoligated by-products were multiply biotinylated via nick-translation or primer-extension or both. Thus, they could readily be removed from the DNA dumbbell preparation by a mild biomagnetic separation procedure. The closed conformation of the purified DNA dumbbell was verified by its altered gel mobility as compared with unligated or monoligated samples and by an exonuclease assay. Considering the promising therapeutic potential of DNA dumbbells, the developed biosynthetic approach could be used for high-purity preparation of longer, covalently closed DNA decoys.

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“…Another approach to the problem involved circularization of oligonucleotides by formation of covalent linkage between 5¢-and 3¢-ends of the molecule [13,14]. Moreover, there were described the so called "self stabilising" structures in which the resistance to exonucleases was achieved by formation of hairpin or hairpin loop structures using self-complementary ends of oligonucleotide [15][16][17][18][19][20].…”

mentioning

confidence: 99%

Antisense hairpin loop oligonucleotides as inhibitors of expression of multidrug resistance-associated protein 1: their stability in fetal calf serum and human plasma.

Rębowski¹,

Wójcik²,

Boczkowska³

et al. 2001

Acta Biochim Pol

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Multidrug resistance-associated protein (MRP1) is a transmembrane pump protein responsible for the efflux of chemotherapeutic drugs, an important cause of anticancer treatment failure. Trying to circumvent MRP-mediated resistance we designed and synthesized hairpin loops forming antisense oligodeoxyribonucleotides (ODNs), both phosphodiesters (PO-ODNs) and their phosphorothioate analogues (PS-ODNs), to reduce the protein expression by targeting its mRNA in a sequence specific manner. Melting temperature measurements as well as polyacrylamide gel electrophoresis supported the preferential formation of a secondary structure, which was expected to protect ODNs against 3'-exonuclease degradation. ODNs and PS-ODNs designed in this work were successfully tested as antisense inhibitors of the expression of MRP1 in the leukaemia HL60/ADR cell line. Foreseeing the necessity to perform clinical studies with such ODNs we investigated their stability against the 3'-exonuclease activity of fetal calf serum and human plasma. Under the conditions, corresponding to physiological ones, we observed high stability of hairpin loop forming ODNs, especially those containing longer (e.g. 7 base pair) stems. Comparative studies on the stability of chemically unmodified hairpin loop forming ODNs and their PS-counterparts indicated that endonuclease activity did not play any important role in the process of their nucleolytic degradation. Our studies provide strong evidence for high stability of chemically unmodified hairpin loop ODNs, making them an attractive alternative to phosphorothioate analogues commonly used in antisense strategy.

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Properties of nicked and circular dumbbell RNA/DNA chimeric oligonucleotides containing antisense phosphodiester oligodeoxynucleotides

Cited by 11 publications

References 27 publications

The effect of hydrostatic pressure on the thermal stability of DNA hairpins

The effect of hydrostatic pressure on the thermal stability of DNA hairpins

High-Purity Preparation of a Large DNA Dumbbell

Antisense hairpin loop oligonucleotides as inhibitors of expression of multidrug resistance-associated protein 1: their stability in fetal calf serum and human plasma.

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