New Cleavable Spacers for Tandem Synthesis of Multiple Oligo­nucleotides

Abstract

In solid-phase oligonucleotide synthesis, a single oligonucleotide is generally acquired from a column loaded with a specific solid support. Herein, we have developed new cleavable spacer (CS) derivatives for tandem synthesis of multiple oligonucleotides on a single column. Four CS analogs were designed, synthesized, and inserted between two oligonucleotide sequences using an automated oligonucleotide synthesizer. The CS derivatives bearing a cyclic cis-1,2-diol exhibited efficient release of the two oligonucleotides under commonly employed basic conditions of aqueous ammonia. Among the CS analogues, it was found that CS with a robust structure can potentially be applied as a spacer molecule in the tandem synthesis of multiple oligonucleotides in a single sequence.

Benzo-Fused 7-Oxabicyclo[2.2.1]heptane-2,3-diol Derivatives as Universal Linkers for Solid-Phase Oligonucleotide Synthesis

“…Dihydroxylation using a catalytic amount of osmium tetroxide (OsO 4 ) in the presence of 4methylmorpholine N-oxide (NMO) afforded the corresponding diols 5b-d in 80-90% yields. Furthermore, the dihydroxylation of 7-oxabicyclo[2.2.1]hept-2-ene stereoselectively provided the exo-cis-glycol as the sole isomer 21 and benzo-fused 7-oxabicyclo[2.2.1]hept-2-ene (4a) was converted into solely the exo-cis-glycol 5a; 23 therefore, diols 5b-d would have the same configuration, as shown. Subsequently, the desired carboxylic acid compounds 2b-d were synthesized by DMTr protection followed by succinylation.…”

“…Compound 2a was synthesized from 4a via 5a according to our previous report. 23 5,8-Dimethyl-1,4-dihydro-1,4-epoxynaphthalene ( 4b ) 26 and 5,8-dimethoxy-1,4-dihydro-1,4-epoxynaphthalene ( 4c ) 27 were prepared according to the literature. 5,8-Bis(methoxymethoxy)-1,4-dihydro-1,4-epoxynaphthalene ( 4d ) was synthesized using a Diels–Alder­ reaction between a benzyne, generated from 2-bromo-1,4-bis(methoxymethoxy)benzene, 28 and furan (see the experimental section).…”

“…The scaffolds of the designed compounds are based on a benzo-fused 7-oxabicyclo[2.2.1]heptane-2,3-diol, which is expected to have reactivity comparable to that of the imide-ring-fused type with a 7-oxabicyclo[2.2.1]heptane-2,3-diol skeleton. 23 Unsubstituted derivative 2a , disubstituted derivatives (dimethyl ( 2b ), dimethoxy ( 2c ), dimethoxymethoxy ( 2d ), diphenyl/terphenyl ( 2e )), and dibenzo-fused (phenanthrene) derivative 3a were designed. Moreover, derivatives 3b – d , which are PS-conjugated phenanthrene-type (PT) linkers with different linking groups, were designed in addition to 3a with a succinic spacer.…”

MicroRNA Pools Synthesized Using Tandem Solid-Phase Oligonucleotide Synthesis

“…The organic phases were collected, reduced under pressure, and purified by column chromatography (10−35% EtOAc in heptane) to obtain the title compound S2 as a white foam (5.08 g, 91%). R f = 0.21 (EtOAc/Hept, 30:70); 1 H NMR (400 MHz, Acetone-d 6 ) δ 7.53− 7.15 (m, 18H, 2xDMTr), 6.90−6.81 (m, 8H, 2xDMTr), 3.77 (s, 12H, OCH3), 3.54−3.43 (m, 1H, H4), 3.07 (t, J = 6.5 Hz, 4H, H1,H7), 1.86−1.36 (m, 8H, H2,H3,H5,H6); 13 1,7-Di-O-dimethoxytrityl-heptane-4-O-(2-cyanoethyl)-N,N-diisopropylphosphoramidite (S3). 16 Compound S3 was prepared by coevaporating S2 (400 mg, 531.25 μmol) with anhydrous DCE (7 mL) and redissolved in anhydrous DCM (4 mL).…”

“…The isomers were not isolated but found in a ratio of 0.4:0.6 (7a:7b) in the UV-trace from LC−MS and confirmed by integration of H1′ in NMR spectra. R f = 0.39 (EtOAc/Hept, 80:20); 1 H NMR (400 MHz, DMSO-d 6 ) δ 11.43 (s, 1H,NH), 7.70 (2 × d, J = 8.1 Hz, 1H, H6,7a + 7b), 7.44−7.20 (m, 9H, aromatic, 7a + 7b), 6.95−6.87 (m, 4H, aromatic, 7a + 7b), 5.92 (d, J = 4.5 Hz, 0.4H, H1′, 7a), 5.82 (d, J = 5.8 Hz, 0.6H, 2′-OH, 7b), 5.76 (d, J = 5.6 Hz, 0.6H, H1′, 7b), 5.58 (d, J = 6.2 Hz, 0.4H, 3′-OH, 7a), 5.46 (d, J = 7.9 Hz, 0.6H, H5, 7b), 5.41 (d, J = 8.0 Hz, 0.4H, H5, 7a), 5.27 (t, J = 5.1 Hz, 0.4H, H2′,7a), 5.14 (t, J = 5.1 Hz, 0.6H, H3′, 7b), 4.45−4.19 (m, 1H, H3′ 7a, H2′ 7b), 4.12 (q, J = 4.2 Hz, 0.6H, H4′, 7b), 4.01−3.97 (m, 0.4H, H4′, 7a), 3.75 (s, 6H, 2 × OCH3, 7a + 7b), 3.32−3.18 (m, 2H, H5′/H5″, 7a + 7b), 2.09 (s, 1.2H, CH3, 7a), 2.07 (s, 1.8H, CH3, 7b); 13 17 Compound S8 was prepared by co-evaporating S7 (204 mg, 340 μmol) in a dry round-bottom flask with dry DCE (7 mL) and redissolved in dry DCM (5 mL) followed by addition of 2-cyanoethyl N,N,N′,N′tetraisopropylphosphoramidite (205 mg, 216 μL, 680 μmol) and diisopropylammonium tetrazolide (116.4 mg, 680 μmol). The reaction was stirred under argon o/n.…”