Author:

Keywords:

ectonucleotidase inhibitors, enzyme assay, p-nitrophenyl 5 '-thymidine monophosphate, NPP1, NPP1 inhibitors, nucleotide pyrophosphatase 1, Science & Technology, Life Sciences & Biomedicine, Pharmacology & Pharmacy, P-NITROPHENYL, ADENOSINE, POTENT, CELLS, MINERALIZATION, TRIPHOSPHATE, ECTOENZYMES, INSIGHTS, PATHWAY, ANALOGS, p-nitrophenyl 5′-thymidine monophosphate, 1115 Pharmacology and Pharmaceutical Sciences, 3214 Pharmacology and pharmaceutical sciences

Abstract:

Nucleotide pyrophosphatase/phosphodiesterase type 1 (NPP1) is a membrane glycoprotein involved in the hydrolysis of extracellular nucleotides. Its major substrate is ATP which is converted to AMP and diphosphate. NPP1 was proposed as a new therapeutic target in brain cancer and immuno-oncology. Several NPP1 inhibitors have been reported to date, most of which were evaluated vs. the artificial substrate p-nitrophenyl 5′-thymidine monophosphate (p-Nph-5′-TMP). Recently, we observed large discrepancies in inhibitory potencies for a class of competitive NPP1 inhibitors when tested vs. the artificial substrate p-Nph-5′-TMP as compared to the natural substrate ATP. Therefore, the goal of the present study was to investigate whether inhibitors of human NPP1 generally display substrate-dependent inhibitory potency. Systematic evaluation of nucleotidic as well as non-nucleotidic NPP1 inhibitors revealed significant differences in determined Ki values for competitive, but not for non- and un-competitive inhibitors when tested vs. the frequently used artificial substrate p-Nph-5′-TMP as compared to ATP. Allosteric modulation of NPP1 by p-Nph-5′-TMP may explain these discrepancies. Results obtained using the AMP derivative p-nitrophenyl 5′-adenosine monophosphate (p-Nph-5′-AMP) as an alternative artificial substrate correlated much better with those employing the natural substrate ATP.