Nucleotide Pyrophosphatase/Phosphodiesterase 1 Is Responsible for Degradation of Antisense Phosphorothioate Oligonucleotides

Wójcik, Marzena; Cieślak, Marcin; Stec, Wojciech J.; Goding, James W.; Koziołkiewicz, Maria

doi:10.1089/oli.2007.0021

Cited by 36 publications

(26 citation statements)

References 37 publications

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“…11). NPP1 has been reported to even degrade antisense phosphorothionate oligonucleotides, resulting in the formation of a ladder of shorter products and the release of the mononucleoside 5′-phosphorothionate [368].…”

Section: General Properties and Functional Rolementioning

confidence: 99%

Cellular function and molecular structure of ecto-nucleotidases

Zimmermann

Zebisch

Sträter

2012

Purinergic Signalling

871

922

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Ecto-nucleotidases play a pivotal role in purinergic signal transmission. They hydrolyze extracellular nucleotides and thus can control their availability at purinergic P2 receptors. They generate extracellular nucleosides for cellular reuptake and salvage via nucleoside transporters of the plasma membrane. The extracellular adenosine formed acts as an agonist of purinergic P1 receptors. They also can produce and hydrolyze extracellular inorganic pyrophosphate that is of major relevance in the control of bone mineralization. This review discusses and compares four major groups of ectonucleotidases: the ecto-nucleoside triphosphate diphosphohydrolases, ecto-5′-nucleotidase, ecto-nucleotide pyrophosphatase/phosphodiesterases, and alkaline phosphatases. Only recently and based on crystal structures, detailed information regarding the spatial structures and catalytic mechanisms has become available for members of these four ecto-nucleotidase families. This permits detailed predictions of their catalytic mechanisms and a comparison between the individual enzyme groups. The review focuses on the principal biochemical, cell biological, catalytic, and structural properties of the enzymes and provides brief reference to tissue distribution, and physiological and pathophysiological functions.

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Section: General Properties and Functional Rolementioning

confidence: 99%

Cellular function and molecular structure of ecto-nucleotidases

Zimmermann

Zebisch

Sträter

2012

Purinergic Signalling

871

922

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“…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)(4)(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).…”

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confidence: 99%

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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“…It is now accepted that the human plasma 3 0 -exonuclease possesses the [R P ]-selective nature and its digestion mechanism proceeds with retention of configuration at a phosphorus atom, that is, the reaction occurs via a two-step mechanism with participation of a covalent enzymesubstrate intermediate (Gijsbers et al 2001;Koziolkiewicz et al 2002). This fact, combined with the result obtained from immunoprecipitation assay with mouse monoclonal antibodies 3E8 binding the native form of NPP1 (also known as plasma cell membrane glycoprotein PC-1, EC 3.1.4.1/3.6.1.9) confirmed that the human plasma 3 0 -exonuclease is a soluble form of PC-1 belonging to the nucleotide pyrophosphatases/ phosphodiesterases (NPPs) family (Wojcik et al 2007), widely distributed and phylogenetically conserved membrane-associated or secreted proteins involved in regulation of various physiological processes, including bone mineralization, calcification of ligaments and joint capsules, signaling by nucleotides and insulin, and the differentiation and motility of cells (Bollen et al 2000;Goding et al 2003;Stefan et al 2005). Multiple roles of NPPs are consequence, at least in part, of their action on distinct substrates like nucleotides and nucleotide derivatives as well as lysophospholipids and choline phosphate esters.…”

Section: Introductionmentioning

confidence: 52%

The effect of divalent cations on the catalytic activity of the human plasma 3′-exonuclease

Wójcik

Stec

2010

Biometals

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The 3'-exonuclease from human plasma is a soluble form of nucleotide pyrophosphatase/phosphodiesterase 1 (NPP1) (EC 3.1.4.1/EC 3.6.1.9). Here, the possibility of divalent cation influence for the 3'-exonuclease activity was investigated using the phosphorothioate congener of oligonucleotide containing all phosphorothioate internucleotide linkages of the [R(P)]-configuration ([R(P)-PS]-d[T(12)]) as the substrate for this enzyme. It was found that the 3'-exonuclease is a metalloenzyme, i.e. its phosphodiesterase activity was completely abolished at 0.8 mM concentration EDTA and, in turn, it was restored in the presence of Mg(2+) or Mn(2+) ions. In addition, Mg(2+) can be replaced effectively by Ca(2+), Mn(2+), or Co(2+), but not by Ni(2+) and Cd(2+) during the hydrolysis of the phosphorothioate substrate in human plasma. In addition, the mechanism is postulated, by which a single internucleotide phosphorothioate bond of the S(P)-configuration at the 3'-end of unmodified phosphodiesters (PO-oligos), or their phosporothioate analogs (PS-oligos) protects these compounds against degradation in blood.

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Nucleotide Pyrophosphatase/Phosphodiesterase 1 Is Responsible for Degradation of Antisense Phosphorothioate Oligonucleotides

Cited by 36 publications

References 37 publications

Cellular function and molecular structure of ecto-nucleotidases

Cellular function and molecular structure of ecto-nucleotidases

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

The effect of divalent cations on the catalytic activity of the human plasma 3′-exonuclease

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