N-PHENYLAMIDINES AS SELECTIVE INHIBITORS OF HUMAN NEURONAL NITRIC-OXIDE SYNTHASE - STRUCTURE-ACTIVITY STUDIES AND DEMONSTRATION OF IN-VIVO ACTIVITY

Selective inhibition of the neuronal isoform of nitric oxide synthase (NOS) compared to the endothelial and inducible isoforms may be requir ed for treatment of neurological disorders caused by excessive product ion of nitric oxide. Recently, we described N-(3-aminomethyl)-benzyl)a cetamidine (13) as a slow, tight-binding inhibitor, highly selective f or human inducible nitric oxide synthase (iNOS). Removal of a single m ethylene bridge between the amidine nitrogen and phenyl ring to give N -(3-(aminomethyl)phenyl)acetamidine (14) dramatically altered the sele ctivity to give a neuronal selective nitric oxide synthase (nNOS) inhi bitor. Part of this large shift in selectivity was due to 14 being a r apidly reversible inhibitor of iNOS in contrast to the essentially irr eversible inhibition of iNOS observed with 13. Structure-activity stud ies revealed that a basic amine functionality tethered to an aromatic ring and a sterically compact amidine are key pharmacophores for this class of NOS inhibitors. Maximal nNOS inhibition potency was achieved with N-(3-(aminomethyl)phenyl)-2-furanylamidine (77) (Ki-nNOS = 0.006 mu M; Ki-eNOS = 0.35,mu M; Ki-iNOS = 0.16 mu M). Finally, alpha-fluoro -N-(3-(aminomethyl)phenyl)acetamidine (74) (Ki-nNOS = 0.011 mu M; Ki-e NOS = 1.1 mu M; Ki-iNOS = 0.48 mu M) had excellent brain penetration a nd inhibited nNOS in a rat brain slice assay as well as in the rat bra in (cerebellum) in vivo. Thus, N-phenylamidines should be useful in va lidating the role of nNOS in neurological disorders.