1,3-Dialkylxanthine Derivatives Having High Potency as Antagonists at Human A_2B Adenosine Receptors

The structure-activity relationships (SAR) of alkylxanthine derivatives as antagonists at the recombinant human adenosine receptors were explored in order to identify selective antagonists of A_2B receptors. The effects of lengthening alkyl substituents from methyl to butyl at 1- and 3-positions and additional substitution at the 7- and 8-positions were probed. K_i values, determined in competition binding in membranes of HEK-293 cells expressing A_2B receptors using ¹²⁵I-ABOPX (¹²⁵I-3-(4-amino-3-iodobenzyl)-8-(phenyl-4-oxyacetate)-1-propylxanthine), were approximately 10 to 100 nM for 8-phenylxanthine functionalized congeners. Xanthines containing 8-aryl, 8-alkyl, and 8-cycloalkyl substituents, derivatives of XCC (8-[4-[[[carboxy]methyl]oxy]phenyl]-1,3-dipropylxanthine) and XAC (8-[4-[[[[(2-aminoethyl)amino]carbonyl]methyl]-oxy]phenyl]-1,3-dipropylxanthine), containing various ester and amide groups, including L- and D-amino acid conjugates, were included. Enprofylline was 2-fold more potent than theophylline in A_2B receptor binding, and the 2-thio modification was not tolerated. Among the most potent derivatives examined were XCC, its hydrazide and aminoethyl and fluoroethyl amide derivatives, XAC, N-hydroxyethyl-XAC, and the L-citrulline and D-p-aminophenylalanine conjugates of XAC. An N-hydroxysuccinimide ester of XCC (XCC-NHS, MRS 1204) bound to A_2B receptors with a K_i of 9.75 nM and was the most selective (at least 20-fold) in this series. In a functional assay of recombinant human A_2B receptors, four of these potent xanthines were shown to fully antagonize the effects of NECA-induced stimulation of cyclic AMP accumulation.