Multivalent dendrimeric conjugates of GPCR ligands may have increased potency or selectivity in comparison to monomeric ligands, a phenomenon that was tested in a model of cytoprotection in mouse HL-1 cardiomyocytes. Quantitative RT-PCR indicated high expression levels of endogenous A(1) and A(2A) adenosine receptors (ARs), but not of A(2B) and A(3)ARs. Activation of the heterologously expressed human A(3)AR in HL-1 cells by AR agonists significantly attenuated cell damage following 4h exposure to H(2)O(2) (750 microM) but not in untransfected cells. The A(3) agonist IB-MECA (EC(50) 3.8 microM) and the non-selective agonist NECA (EC(50) 3.9 microM) protected A(3) AR-transfected cells against H(2)O(2) in a concentration-dependent manner, as determined by lactate dehydrogenase release. A generation 5.5 PAMAM (polyamidoamine) dendrimeric conjugate of a N(6)-chain-functionalized adenosine agonist was synthesized and its mass indicated an average of 60 amide-linked nucleoside moieties out of 256 theoretical attachment sites. It non-selectively activated the A(3)AR to inhibit forskolin-stimulated cAMP formation (IC(50) 66nM) and, similarly, protected A(3)-transfected HL-1 cells from apoptosis-inducing H(2)O(2) with greater potency (IC(50) 35nM) than monomeric nucleosides. Thus, a PAMAM conjugate retained AR binding affinity and displayed greatly enhanced cardioprotective potency.
Published by Elsevier Inc.