Deoxyribonucleoside Phosphoramidites

Pedras-Vasconcelos²,

Wang³

et al. 2005

Nucleic Acids Research

A CpG-containing DNA oligonucleotide functionalized with the 2-(N-formyl-N-methyl)aminoethyl thiophosphate protecting group (CpG ODN fma1555) was prepared from phosphoramidites 1a–d using solid-phase techniques. The oligonucleotide behaved as a prodrug by virtue of its conversion to the well-studied immunomodulatory CpG ODN 1555 through thermolytic cleavage of the 2-(N-formyl-N-methyl)aminoethyl thiophosphate protecting group. Such a conversion occurred at 37°C with a half-time of 73 h. The immunostimulatory properties of CpG ODN fma1555 were evaluated in two in vivo assays, one of which consisted of mice challenged in the ear with live Leishmania major metacyclic promastigotes. Local intradermal administration of CpG ODN fma1555 was as effective as that of CpG ODN 1555 in reducing the size of Leishmania lesions over time. In a different infectious model, CpG ODN 1555 prevented the death of Tacaribe-infected mice (43% survival) when administered between day 0 and 3 post infection. Administration of CpG ODN fma1555 three days before infection resulted in improved immunoprotection (60–70% survival). Moreover, co-administration of CpG ODN fma1555 and CpG ODN 1555 in this model increased the window for therapeutic treatment against Tacaribe virus infection, and thus supports the use of thermolytic oligonucleotides as prodrugs in the effective treatment of infectious diseases.

Section: Introductionsupporting

confidence: 88%

Section: Methodsmentioning

confidence: 99%

Thermolytic CpG-containing DNA oligonucleotides as potential immunotherapeutic prodrugs

Pedras-Vasconcelos²,

Wang³

et al. 2005

Nucleic Acids Research

“…Only oligonucleotides prepared from phosphoramidites 1 exhibited adequate thermolytic phosphate/thiophosphate deprotection kinetics 13 to assess their potential as prodrugs. The removal of 2-(N-formyl-N-methyl)aminoethyl phosphate/thiophosphate protection from DNA oligonucleotides is consistent with an intramolecular cyclodeesterification process [19][20][21] as tentatively proposed in FIG-URE 2. Oligonucleotides prepared through phosphoramidites 2-6 displayed phosphate/thiophosphate deprotection kinetics that were too rapid for thermolytic prodrug applications.…”

Section: Introductionsupporting

confidence: 80%

Design and Development of Thermolytic DNA Oligonucleotide Prodrugs

Annals of the New York Academy of Sciences

Pedras-Vasconcelos

Ausín

et al. 2005

Deoxyribonucleoside phosphoramidites functionalized with the thermolytic 2-(N-formyl-N-methyl)aminoethyl group for phosphorus protection (1a-d) have been prepared and employed in the solid-phase synthesis of CpG ODN fma1555. Given that this modified oligonucleotide can be converted to the immunomodulatory CpG ODN 1555 under neutral conditions at 37 degrees C, its biologic activity was demonstrated in vivo by studies showing that intraperitoneal administration of CpG ODN fma1555 in mice resulted in the activation of cytokine-secreting splenocytes. Furthermore, administration of CpG ODN fma1555 to mice that were challenged intradermally in the ear with live L. major metacyclic promastigotes, reduced the severity of Leishmania skin lesions over time to an extent similar to that obtained with CpG ODN 1555. In another infectious model experiment, CpG ODN fma1555 protected newborn mice from death (65% survival) when administered 3 days before infection with the aggressive Tacaribe (TCRV) virus. A comparable immunoprotection was obtained by treatment of TCRV-infected mice with CpG ODN 1555 administered on the same day of infection (45% survival). However, when TCRV-infected mice were treated with CpG ODN fma1555 on the day of infection, they died as a consequence of the relatively slow conversion of the oligonucleotide prodrug to the bioactive CpG ODN 1555. Co-administration of both CpG ODN 1555 and CpG ODN fma1555 to mice 3 days prior to TCRV infection or on the day of infection provided protection from death (45-65% survival) and thus widened the immunoprotection window against TCRV-infection.

“…Readers are also referred to the Critical Parameters and Troubleshooting sections of Current Protocols articles Grajkowski et al. () and Wilk, Grajkowski, Chmielewski, Beaucage, and Phillips (), which address the critical issues associated with the preparation and use of nucleoside phosphoramidites.…”

Section: Commentarymentioning

confidence: 99%

“…Each reactant (1, 4, 6, and 8) including solid 1H-tetrazole should be dried under high vacuum overnight at ambient temperature prior to being dissolved in commercial anhydrous acetonitrile, which is stored in an amber glass bottle over activated 4-Å (8 to 12 mesh) molecular sieves. Readers are also referred to the Critical Parameters and Troubleshooting sections of Current Protocols articles Grajkowski et al (2008) and Wilk, Grajkowski, Chmielewski, Beaucage, and Phillips (2001), which address the critical issues associated with the preparation and use of nucleoside phosphoramidites.…”

Section: Critical Parameters and Troubleshootingmentioning

confidence: 99%

An Improved Strategy for the Chemical Synthesis of 3′,5′‐Cyclic Diguanylic Acid

CP Nucleic Acid Chemistry

Takáhashi

Kaczyński

et al. 2019

Self Cite

The physiological functions of c‐di‐GMP and its involvement in many key processes led to its recognition as a major and ubiquitous bacterial second messenger. Aside from being a bacterial signaling molecule, c‐di‐GMP is also an immunostimulatory molecule capable of inducing innate and adaptive immune responses through maturation of immune mammalian cells. Given the broad biological functions of c‐di‐GMP and its potential applications as a nucleic‐acid‐based drug, the chemical synthesis of c‐di‐GMP has drawn considerable interest. An improved phosphoramidite approach to the synthesis of c‐di‐GMP is reported herein. The synthetic approach is based on the use of a 5′‐O‐formyl protecting group, which can be rapidly and chemoselectively cleaved from a key dinucleotide phosphoramidite intermediate to enable a cyclocondensation reaction leading to a fully protected c‐di‐GMP product in a yield ∼80%. The native c‐di‐GMP is isolated, after complete deprotection, in an overall yield of 36% based on the commercial ribonucleoside used as starting material. © 2019 by John Wiley & Sons, Inc.