The mononucleotide-dependent, nonantisense mechanism of action of phosphodiester and phosphorothioate oligonucleotides depends upon the activity of an ecto-5′-nucleotidase

Koziołkiewicz, Maria; Gendaszewska, E; Maszewska, Maria; Stein, C. A.; Stec, Wojciech J.

doi:10.1182/blood.v98.4.995

Cited by 34 publications

(29 citation statements)

References 40 publications

(43 reference statements)

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“…For instance, products of enzymatic degradation of ODNs can influence cell proliferation. 59 Moreover, G3139 contains two unmethylated CpG motifs within the ODN sequence which are known to activate NK cells and macrophages, 60 and to stimulate B cells. 61 Phosphorothioate ODNs have also been shown to competitively inhibit a variety of nucleases and polymerases.…”

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confidence: 99%

Chemosensitization of myeloma plasma cells by an antisense-mediated downregulation of Bcl-2 protein

et al. 2003

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Chemosensitization of myeloma plasma cells by an antisense-mediated downregulation of Bcl-2 protein

et al. 2003

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“…It has been suggested that extracellular dNMPS and dNMP can be converted to the corresponding nucleoside by 5Ј-nucleotidase (ecto-5Ј-NT) (9). This membrane-bound enzyme preferably releases adenosine from extracellular AMP, but other purine and pyrimidine 5Ј-nucleotides are also dephosphorylated by this protein (11).…”

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confidence: 99%

“…Their hydrolysis in plasma, kidney, and liver proceeds mainly from the 3Ј end, resulting in the appearance of the mononucleoside 5Ј-phosphorothioates identified in urine from PS-oligo-injected animals (6,7). (d)NMPS may exert cytotoxic effects affecting cell proliferation, DNA or RNA synthesis, and other unknown processes (8,9). Recently, the phosphorothioate DNA segments have been identified in bacterial DNA (10), which makes investigations into PS-oligo metabolism even more important.…”

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Histidine Triad Nucleotide-binding Protein 1 (HINT-1) Phosphoramidase Transforms Nucleoside 5′-O-Phosphorothioates to Nucleoside 5′-O-Phosphates

Ozga

Dolot

Janicka

et al. 2010

Journal of Biological Chemistry

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Nucleoside 5-O-phosphorothioates are formed in vivo as primary products of hydrolysis of oligo(nucleoside phosphorothioate)s (PS-oligos) that are applied as antisense therapeutic molecules. The biodistribution of PS-oligos and their pharmacokinetics have been widely reported, but little is known about their subsequent decay inside the organism. We suggest that the enzyme responsible for nucleoside 5-O-monophosphorothioate ((d)NMPS) metabolism could be histidine triad nucleotide-binding protein 1 (Hint-1), a phosphoramidase belonging to the histidine triad (HIT) superfamily that is present in all forms of life. An additional, but usually ignored, activity of Hint-1 is its ability to catalyze the conversion of adenosine 5-O-monophosphorothioate (AMPS) to 5-O-monophosphate (AMP). By mutagenetic and biochemical studies, we defined the active site of Hint-1 and the kinetic parameters of the desulfuration reaction (P-S bond cleavage). Additionally, crystallographic analysis (resolution from 1.08 to 1.37 Å ) of three engineered cysteine mutants showed the high similarity of their structures, which were not very different from the structure of WT Hint-1. Moreover, we found that not only AMPS but also other ribonucleoside and 2-deoxyribonucleoside phosphorothioates are desulfurated by Hint-1 at the following relative rates: GMPS > AMPS > dGMPS > CMPS > UMPS > dAMPS Ͼ Ͼ dCMPS > TMPS, and during the reaction, hydrogen sulfide, which is thought to be the third gaseous mediator, was released. (Fig. 1). 5Ј-O-Phosphorothioates of ribonucleosides (NMPSNMPS and dNMPS (together denoted (d)NMPS)) are formed during the enzymatic hydrolysis of oligo(nucleoside phosphorothioate) (PS-oligos) that contain a sulfur atom attached in non-bridging positions to the phosphorus atom at each or selected internucleotide bond(s). Synthetic PS-oligos have been developed as antisense probes for genomic research and medicinal applications (1, 2). These oligonucleotides are promising therapeutic molecules because they are much more stable against nucleolytic degradation in blood and various cellular systems than their natural, unmodified counterparts (3-5). Their hydrolysis in plasma, kidney, and liver proceeds mainly from the 3Ј end, resulting in the appearance of the mononucleoside 5Ј-phosphorothioates identified in urine from PS-oligo-injected animals (6, 7). (d)NMPS may exert cytotoxic effects affecting cell proliferation, DNA or RNA synthesis, and other unknown processes (8, 9). Recently, the phosphorothioate DNA segments have been identified in bacterial DNA (10), which makes investigations into PS-oligo metabolism even more important.Although several reports have been published on the biodistribution of PS-oligos, little is known about the metabolism of the products of their degradation in vivo. It has been suggested that extracellular dNMPS and dNMP can be converted to the corresponding nucleoside by 5Ј-nucleotidase (ecto-5Ј-NT) (9). This membrane-bound enzyme preferably releases adenosine from extracellular AMP, but other purine and pyrimid...

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“…According to our findings, the influence of extracellular nucleotides and their phosphorothioate analogs depended on the activity of the ecto-5′-nucleotidase present at the cell surface. However, ectonucleotidase activity did not seem to be the only mechanism responsible for the biological activity of these compounds [16].…”

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confidence: 99%

Thymidine 5'-O-monophosphorothioate induces HeLa cell migration by activation of the P2Y6 receptor

Gendaszewska‐Darmach

Szustak

2016

Purinergic Signalling

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ATP, ADP, UTP, and UDP acting as ligands of specific P2Y receptors activate intracellular signaling cascades to regulate a variety of cellular processes, including proliferation, migration, differentiation, and cell death. Contrary to a widely held opinion, we show here that nucleoside 5′-Omonophosphorothioate analogs, containing a sulfur atom in a place of one nonbridging oxygen atom in a phosphate group, act as ligands for selected P2Y subtypes. We pay particular attention to the unique activity of thymidine 5′-Omonophosphorothioate (TMPS) which acts as a specific partial agonist of the P2Y6 receptor (P2Y6R). We also collected evidence for the involvement of the P2Y6 receptor in human epithelial adenocarcinoma cell line (HeLa) cell migration induced by thymidine 5′-O-monophosphorothioate analog. The stimulatory effect of TMPS was abolished by siRNAmediated P2Y6 knockdown and diisothiocyanate derivative MRS 2578, a selective antagonist of the P2Y6R. Our results indicate for the first time that increased stability of thymidine 5′-O-monophosphorothioate as well as its affinity toward the P2Y6R may be responsible for some long-term effects mediated by this receptor.

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The mononucleotide-dependent, nonantisense mechanism of action of phosphodiester and phosphorothioate oligonucleotides depends upon the activity of an ecto-5′-nucleotidase

Cited by 34 publications

References 40 publications

Chemosensitization of myeloma plasma cells by an antisense-mediated downregulation of Bcl-2 protein

Chemosensitization of myeloma plasma cells by an antisense-mediated downregulation of Bcl-2 protein

Histidine Triad Nucleotide-binding Protein 1 (HINT-1) Phosphoramidase Transforms Nucleoside 5′-O-Phosphorothioates to Nucleoside 5′-O-Phosphates

Thymidine 5'-O-monophosphorothioate induces HeLa cell migration by activation of the P2Y6 receptor

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