Abstract:[reaction: see text] 2'-Deoxy-3'-deutero pyrimidines have been synthesized in high yields and incorporated into deoxyoligonucleotides using standard phosphoramidite chemistry. A key synthetic step is a stereospecific reduction of 3'-keto nucleosides using sodium triacetoxyborodeuteride to give 3'-deuterated thymidine and 2'-deoxy uridine nucleosides. Conversion of the corresponding phorphoramidites 7a and 7b to 4-triazolo derivatives has, for the first time, enabled incorporation of 2'-deoxy-3'-deutero cytidin… Show more
“…Therefore, 4-triazolyluridine was an attractive option to attach a nucleoside to an amino-functionalized solid phase. The activated nucleoside 2 was synthesized in three steps from uridine according to standard protocols (22,23) (Scheme 1). Scheme 1.( A ) Synthesis of the activated uridine analogue 2 : (i) 1.…”
Section: Resultsmentioning
confidence: 99%
“…2′,3′,5′-protected uridine 1 (22,23) (300 mg, 388 µmol) and 1,2,4-triazole (428 mg, 6.20 mmol) were suspended in anhydrous acetonitrile (3.5 ml) and triethylamine (1.23 ml, 8.91 mmol) under argon, and after stirring for 5 min a clear, colourless solution was obtained. The flask was put at 0°C and POCl 3 (71.0 µl, 776 µmol) was added dropwise, whereupon a white solid precipitated.…”
Nucleic acids possess the unique property of being enzymatically amplifiable, and have therefore been a popular choice for the combinatorial selection of functional sequences, such as aptamers or ribozymes. However, amplification typically requires known sequence segments that serve as primer binding sites, which can be limiting for certain applications, like the screening of on-bead libraries. Here, we report a method to amplify and sequence on-bead RNA libraries that requires not more than five known nucleotides. A key element is the attachment of the starting nucleoside to the synthesis resin via the nucleobase, which leaves the 3′-OH group accessible to subsequent enzymatic manipulations. After split-and-mix synthesis of the oligonucleotide library and deprotection, a poly(A)-tail can be efficiently added to this free 3′-hydroxyl terminus by Escherichia coli poly(A) polymerase that serves as an anchored primer binding site for reverse transcription. The cDNA is joined to a DNA adapter by T4 DNA ligase. PCR amplification yielded single-band products that could be cloned and sequenced starting from individual polystyrene beads. The method described here makes the selection of functional RNAs from on-bead RNA libraries more attractive due to increased flexibility in library design, higher yields of full-length sequence on bead and robust sequence determination.
“…Therefore, 4-triazolyluridine was an attractive option to attach a nucleoside to an amino-functionalized solid phase. The activated nucleoside 2 was synthesized in three steps from uridine according to standard protocols (22,23) (Scheme 1). Scheme 1.( A ) Synthesis of the activated uridine analogue 2 : (i) 1.…”
Section: Resultsmentioning
confidence: 99%
“…2′,3′,5′-protected uridine 1 (22,23) (300 mg, 388 µmol) and 1,2,4-triazole (428 mg, 6.20 mmol) were suspended in anhydrous acetonitrile (3.5 ml) and triethylamine (1.23 ml, 8.91 mmol) under argon, and after stirring for 5 min a clear, colourless solution was obtained. The flask was put at 0°C and POCl 3 (71.0 µl, 776 µmol) was added dropwise, whereupon a white solid precipitated.…”
Nucleic acids possess the unique property of being enzymatically amplifiable, and have therefore been a popular choice for the combinatorial selection of functional sequences, such as aptamers or ribozymes. However, amplification typically requires known sequence segments that serve as primer binding sites, which can be limiting for certain applications, like the screening of on-bead libraries. Here, we report a method to amplify and sequence on-bead RNA libraries that requires not more than five known nucleotides. A key element is the attachment of the starting nucleoside to the synthesis resin via the nucleobase, which leaves the 3′-OH group accessible to subsequent enzymatic manipulations. After split-and-mix synthesis of the oligonucleotide library and deprotection, a poly(A)-tail can be efficiently added to this free 3′-hydroxyl terminus by Escherichia coli poly(A) polymerase that serves as an anchored primer binding site for reverse transcription. The cDNA is joined to a DNA adapter by T4 DNA ligase. PCR amplification yielded single-band products that could be cloned and sequenced starting from individual polystyrene beads. The method described here makes the selection of functional RNAs from on-bead RNA libraries more attractive due to increased flexibility in library design, higher yields of full-length sequence on bead and robust sequence determination.
“…The enormous power of the method is revealed in the list of natural product syntheses (or approaches thereto) that have employed -hydroxy-directed reduction of ketones using NaBH(OAc) 3 (or Me 4 NBH(OAc) 3 (in no special order): cyclophellitol, 158 sanglifehrin A, 159 aurisides A and B aglycon, 160 Several groups have reported the -hydroxyl-directed reduction of various ketonucleosides. [188][189][190][191][192][193][194][195][196] Simple systems also undergo this stereoselective reduction (e.g., 65), 197 and the method has been applied to the synthesis of polypropionates. 198 Although a chairlike transition state (Scheme 10) does not operate in the acylborohydride reduction of R-hydroxy ketones, these substrates nevertheless can be reduced via intramolecular hydride delivery leading to syntheses of allopumiliotoxin A, 199 the bruceantin quassinoids, 200 and the rocaglamides 201 (Scheme 12).…”
Section: Hydroxy-directed Reduction Of Ketonesmentioning
confidence: 99%
“…The enormous power of the method is revealed in the list of natural product syntheses (or approaches thereto) that have employed β-hydroxy-directed reduction of ketones using NaBH(OAc) 3 (or Me 4 NBH(OAc) 3 (in no special order): cyclophellitol, 158 sanglifehrin A, 159 aurisides A and B aglycon, 160 calyculin A, 161 nodulisporic acid A, 162 milbemycin, 163 rutamycin B, 164 parasitic wasp lactones, 165 rhizoxins, 166 omuralide and lactacystin, 167 pyripyropene A, 168 thiazinotrienomycin E, 169 scytophycin C, 170 hemibrevetoxin B, 171 FR-900482 analogues, 172 phorboxazole A, 173 bryostatins, 174 phorbol, 175 polycavernoside A, 176 serofendic acids, 177 spongistatin 1, 178 oleandolide, 179 callipeltoside A, 180 rottnestol and raspailols A and B, 181 mycosamine, 182 macrolactins and macrolactinic acid, 183 kendomycin, 184 spirastrellolide A, 185 Several groups have reported the β-hydroxyl-directed reduction of various ketonucleosides. [188][189][190][191][192][193][194][195][196] Simple systems also undergo this stereoselective reduction (e.g., 65), 197 and the method has been applied to the synthesis of polypropionates. 198 Although a chairlike transition state (Scheme 10) does not operate in the acylborohydride reduction of R-hydroxy ketones, these substrates nevertheless can be reduced via intramolecular hydride delivery leading to syntheses of allopumiliotoxin A, 199 the bruc...…”
Acyloxyborohydrides are unsurpassed in their versatility as reagents in organic synthesis. In addition to their superior ability in effecting reductive amination of aldehydes and ketones, acyloxyborohydrides can reduce nitrogen heterocycles (indoles, quinolines, isoquinolines), imines, enamines, oximes, amides, nitriles, benzylic alcohols, aryl ketones, aldehydes (but not ketones), acetals, -hydroxy ketones, and other substrates. The ability to control chemoselectivity, regioselectivity, and stereoselectivity by adjusting the carboxylic acid, borohydride reagent, stoichiometry, and temperature has no parallel in the repertoire of the organic chemist.
“…However, the instability of the 3 0 -keto nucleosides, due to their susceptibility to b-elimination of the nucleobase 29,30 prompted us to choose the appropriate 3-keto glucoside as the suitable precursor.…”
A novel series of exomethylene- and keto-exomethylene-d-glucopyranonucleosides with thymine, uracil, and 5-fluorouracil as heterocyclic bases have been designed and synthesized. Wittig condensation of the 3-keto glucoside 1 gave the corresponding 1,2:5,6-di-O-isopropylidene-3-deoxy-3-methylene-d-glucofuranose (2), which after hydrolysis and acetylation led to the precursor 1,2,4,6-tetra-O-acetyl-3-deoxy-3-methylene-d-glucopyranose (4). Compound 4 was condensed with silylated thymine, uracil, and 5-fluorouracil, respectively, deacetylated and acetalated to afford 1-(3'-deoxy-4',6'-O-isopropylidene-3'-methylene-β-d-glucopyranosyl)pyrimidines 7a-c. Oxidation of the free hydroxyl group in the 2'-position of the sugar moiety led to the formation of the labile 1-(3'-deoxy-4',6'-O-isopropylidene-3'-methylene-β-d-glucopyranosyl-2'-ulose)pyrimidines 8a-c. Finally, deisopropylidenation of the resulted derivatives 8a-c afforded the diol nucleosides 9a-c. The target keto-exomethylene analogs 9a-c were more cytostatic against a variety of tumor cell lines than the corresponding saturated-hydroxy-exomethylene derivatives 6. In particular, the 5-fluorouracil derivative 9c was highly cytostatic at an IC(50) (50% inhibitory concentration) ranging between 0.56 and 9.4 microg/mL, which was comparable to the free parental 5-fluorouracil base.
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