Characterization of soluble CD39 (SolCD39/NTPDase1) from Piggy-Bac nonviral system as a tool to control the nucleotides level.

Extracellular ATP (eATP) and its metabolites have emerged as key modulators of different diseases and comprise a complex pathway called purinergic signaling. An increased number of tools have been developed to study the role of nucleotides and nucleosides in cell proliferation and migration, influence on the immune system and tumor progression. These tools include receptors agonists/antagonists, engineered ectonucleotidases, interference RNAs and ectonucleotidases inhibitors that allow the control and quantification of nucleotide levels. NTPDase1 (also called apyrase, ecto-ATPase, CD39)is one of the main enzymes responsible for the hydrolysis of eATP, and purified enzymes, such as apyrase purified from potato, or engineered as soluble CD39 (SolCD39), have been widely used in in vitro and in vivo experiments. However, the commercial apyrase had its effects recently questioned and SolCD39 exhibit limitations, such as short half-life and need of high doses to reach the expected enzymatic activity. Therefore, this study investigated a nonviral method to improve overexpression of SolCD39 and evaluated its impact over other enzymes of the purinergic system. Our data demonstrated that Piggy-Bac transposon system proved to be a fast and efficient method to generate cells stably expressing SolCD39, producing high amounts of enzyme from a limited number of cells and with high hydrolytic activity. In addition, the soluble form of NTPDase1/CD39 did not alter the expression or catalytic activity of other enzymes from the purinergic system. Altogether, these findings set the groundwork for prospective studies on the function and therapeutic role of eATP and its metabolites in physiological and pathological conditions.