Developmnet of an efficient oligonucleotide derivatization protocol

Gmeiner, William H.; Luo, Weide; Pon, Richard T.; Lown, J. William

doi:10.1016/s0960-894x(01)81111-2

Cited by 12 publications

(3 citation statements)

References 7 publications

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“…Gels were photographed under UV illumination. a rigid oxycarbonyl hydroxymethyl pyrrolidinol framework [ll], (4) as a carbamate at the N(6)-position of adenosine linked to CPG [12], or ( 5 ) via an aminobutyl 1,3-propane diol backbone [13]. It has been reported that the presence of such a lipophilic group may play a large part in improving the traversal of an oligonucleotide through the cell membrane.…”

Section: Resultsmentioning

confidence: 99%

“…Among the family of steroids only cholesterol has been attached onto oligomers [6][7][8][9][10][11][12][13][14][15][16][17][18]20,21]. This is surprising given the availability of other steroids for possible antisense applications and the presence of specific receptors for steroids in certain cells, particularly neuronal cells, that would be excellent targets for antisense inhibition [22, 231. We report here the synthesis and characterization of other steroids conjugated at the 5'and 3'-positions of mononucleosides and the 5'-position of oligonucleotides, and having a steroid as a bridge between two oligomers of different sequence [24].…”

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confidence: 99%

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Targeting of Antisense: Synthesis of Steroid-Linked and Steroid-Bridged Oligodeoxynucleotides

1996

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Steroid-Linked oligonucleotides could be used as antisense drugs targeted to specific steroid receptors on selected cells. Phosphoramidites of desoxycorticosterone and 21-acetoxypregnenolone have been synthesized and characterized. They have been attached by solid-phase synthesis to the 5'-terminus of thymidine and of oligodeoxynucleotides. The 3-hydroxy group of pregnenolone has also been used to attach a second oligomer at its 3'4erminus. The steroid acts as a rigid bridge between the two noncomplementary oligomers, compared to a flexible triethylene glycol linker that was also prepared. If the two oligomers are complementary to different sites on the same mRNA, a bridged noncomplementary dimer could act as a bifunctional antisense molecule. MATERIALS A N D METHODSTetrahydrofuran, pyridine, diisopropylamine, and triethylamine were dried and distilled following established procedures. Anhydrous acetonitrile and oxidizing reagent solution were purchased from Applied Biosystems. Acetonitrile (HPLC grade) was purchased from Baxter. 2-Cyanoethyl N,Ndiisopropylchlorophosphoramidite, dimethoxytrityl chloride, 4-dimethylaminopyridine, hexane, and ethylacetate were purchased from Aldrich Chemical Co. All the steroids used in this study were from Sigma and Aldrich. Thin-layer chromatography (TLC) was performed on silica gel 60 F254 glass plates and silica gel (70-to 100-mesh or 230-to 400-mesh, Merck). Products were visualized by ultraviolet absorption at 254 nm and by exposing the TLC plates to anisaldehyde spray reagent and then heating the plate to -450°C. Snake venom phosphodiesterase (Crotallus durissus) and alkaline phosphatase were purchased from Sigma.'H-NMR spectra were run in CDC13 or D 2 0 using a Varian XL400 or XL300 instrument; ppm are relative to TMS. 31P-NMR spectra (1 21 MHz) were recorded on a Varian XL300 instrument; ppm are relative to external 85% phosphoric acid. FAB mass spectrometry was carried out using a Hewlett Packard 5988. Samples were dissolved in 9: 1 3-nitrobenzyl alcohol (NBA)/glycerol and recorded in both negative and positive ion modes. 27Drug Delivery Downloaded from informahealthcare.com by University of Western Ontario on 10/25/12For personal use only.

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Section: Resultsmentioning

confidence: 99%

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confidence: 99%

Targeting of Antisense: Synthesis of Steroid-Linked and Steroid-Bridged Oligodeoxynucleotides

1996

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“…It has been shown that cellular uptake of native oligonucleotides and polyanionic analogs may be improved by the attachment to these molecules of terminal lipophilic groups, such as cholesterol, that are known to interact specifically with cell membranes [8][9][10][11][12][13][14][15][16]. But this approach may have a substantial drawback, in that the lipophilic groups of these conjugated oligomers may remain attached to cell membranes, and this anchoring may make the antisense molecules unavailable for binding to their mRNA targets.…”

Section: Introductionmentioning

confidence: 99%

Antisense pro-drugs: 5'-ester oligodeoxynucleotides

Polushin¹,

Cohen²

1994

Nucl Acids Res

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Oligonucleotides bearing a terminal lipophilic group attached through a biodegradable ester bond should be useful as antisense pro-drugs with improved cellular uptake. The synthesis of 5'-ester oligonucleotides is, however, problematic due to lability of the ester bond during aqueous ammonia treatment that is commonly used for the deprotection of synthetic oligonucleotides. The synthesis of 5'-palmitoyl oligodeoxynucleotides was accomplished in good yield by the use of a combination of base-labile tert-butylphenoxyacetyl amino protecting groups (t-BPA), the oxalyl-CPG anchor group, and ethanolamine (EA) as a deprotecting reagent.

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