Discovery of BRL 50481 [3-(N,N-dimethylsulfonamido)-4-methyl-nitrobenzene], a Selective Inhibitor of Phosphodiesterase 7: In Vitro Studies in Human Monocytes, Lung Macrophages, and CD8⁺T-Lymphocytes

Abstract

The biochemical and pharmacological characteristics in human proinflammatory cells of BRL 50481 [3-(N,N-dimethylsulfonamido)-4-methyl-nitrobenzene], a novel and selective inhibitor of phosphodiesterase (PDE) 7, are described. BRL 50481 inhibited the activity of hrPDE7A1 expressed in baculovirus-infected Spodoptera frugiperda 9 cells in a competitive manner (K i value of 180 nM) and was 416 and 1884 times less potent against PDE3 and 38 and 238 times less potent against PDE4 at a substrate concentration of 1 M and 50 nM cAMP, respectively. Western blotting identified HSPDE7A1 but not HSPDE7A2 in three human cell types that are implicated in the pathogenesis of chronic obstructive lung disease, namely, CD8 ϩ T-lymphocytes, monocytes, and lung macrophages. BRL 50481 had no effect on the proliferation of CD8 ϩ Tlymphocytes and only marginally (ϳ2-11%) reduced the generation of tumor necrosis factor (TNF)␣ from blood monocytes and lung macrophages. However, in the presence of BRL 50481 the inhibitory effect of rolipram was enhanced on all three cell types. The expression of HSPDE7A1 was increased in a time-dependent manner in monocytes that were "aged" in culture medium. Under this condition, BRL 50481 now inhibited TNF␣ generation in a concentration-dependent manner. In aged monocytes, rolipram, Org 9935 (a PDE3 inhibitor), and prostaglandin E 2 inhibited TNF␣ generation in a concentrationdependent manner and interacted additively with BRL 50481. BRL 50481 is the first fully documented PDE7 inhibitor that has acceptable selectivity for in vitro studies. Furthermore, although BRL 50481 had only a modest inhibitory effect per se on the proinflammatory cells studied, it acted at least additively with other cAMP-elevating drugs, especially when HSPDE7A1 was up-regulated.

Identification in Silico and Experimental Validation of Novel Phosphodiesterase 7 Inhibitors with Efficacy in Experimental Autoimmune Encephalomyelitis Mice

“…The data set is characterized by a high structural diversity since it is formed by iminothiadiazoles, 28,29 benzene sulfonamides, 30 quinazolines, 16 thiazoles, 31 and spirotricyclic derivatives, 32 among others (Figure 1). …”

“…Two different compound concentrations (10 and 30 μM) were used, and in both cases we observed an increase of cAMP even more effective than the standard PDE7 inhibitor BRL50481. 30 We have recently reported this class of compounds, the ITDZs, as substrate competitive inhibitors of GSK3 with potential for the treatment of neurodegenerative diseases. 21 As synergistic interactions between PDE4B and GSK3 inhibitors have been suggested due to the influence of increased cAMP levels induced by PDE4B inhibitors on GSK3 function, 43 our ITDZ compounds offer a unique potential to be explored as innovative multifunctional neurological drugs without the emetic effects present in PDE4B inhibitors.…”

PDE7A1 hydrolyzes cCMP

“…The PDE7-specific competitive inhibitor BRL-50481 [23] inhibited PDE7A1/2-mediated cCMP hdrolysis with a K i value of 98 nM or 91 nM for CMP formation or cCMP hydrolysis, respectively. This corresponds well to the K i value of 180 nM that was previously reported for inhibition of cAMP hydrolysis by PDE7A1 expressed in Sf9 cells [23].…”

“…BRL-50481 is a specific competitive inhibitor of PDE7A-mediated cAMP hydrolysis [23]. PDE7A1/2 activity for cCMP (10 lM) was reduced by 50% in the presence of $100 nM of BRL-50481 (Fig.…”

“…Since high PDE7A1 expression was reported for human lymphoblast cell line (HuT 78) lymphoma cells, in contrast to rather low expression levels in human promonocytic cell line (U-937) cells [23], we chose these two cell lines for Western blotting. In fact, U937 cells (Suppl.…”