Anti-inflammatory potency of novel ecto-5'-nucleotidase/CD73 inhibitors in astrocyte culture model of neuroinflammation

Three novel cytosine-derived alpha,beta-methylene diphosphonates designated MRS4598, MRS4552, and MRS4602 were tested in the range of 1 x 10(-9) to 1 x 10(-3) M for their efficacy and potency in inhibiting membrane-bound ecto-5'-nucleotidase/CD73 activity in primary astrocytes in vitro. The compounds were also tested for their ability to attenuate the reactive astrocyte phenotype induced by proinflammatory cytokines. The main findings are as follows: A) The tested compounds induced concentration-dependent inhibition of CD73 activity, with maximal inhibition achieved at similar to 1 x 10(-3)M; B) All compounds showed high inhibitory potency, as reflected by IC50 values in the submicromolar range; C) All compounds showed high binding capacity, as reflected by Ki values in the low nanomolar range; D) Among the tested compounds, MRS4598 showed the highest inhibitory efficacy and potency, as reflected by IC50 and Ki values of 0.11 mu M and 18.2 nM; E) Neither compound affected astrocyte proliferation and cell metabolic activity at concentrations near to IC50; E) MRS4598 was able to inhibit CD73 activity in reactive astrocytes stimulated with TNF- alpha and to induce concentration-dependent inhibition of CD73 in reactive astrocytes stimulated with IL-1 beta, with an order of magnitude higher IC50 value; F) MRS4598 was the only compound tested that was able to induce shedding of the CD73 from astrocyte membranes and to enhance astrocyte migration in the scratch wound migration assay, albeit at concentration well above its IC50 value. Given the role of CD73 in neurodegenerative diseases, MRS4598, MRS4552, and MRS4602 are promising pharmacological tools for the treatment of neurodegeneration and neuroinflammation.

Automated summary

In this in vitro study, three novel cytosine-derived alpha,beta-methylene diphosphonates (MRS4598, MRS4552, and MRS4602) were tested for their potency in inhibiting CD73 activity and attenuating reactive astrocyte phenotype. The results showed that all compounds exhibited concentration-dependent inhibition of CD73 activity with high inhibitory potency and binding capacity. Among them, MRS4598 was the most effective in inhibiting CD73 activity and inducing reactive astrocyte phenotype inhibition, making it a promising tool for the treatment of neurodegeneration and neuroinflammation.