To search for TNF-alpha (tumor necrosis factor alpha) converting enzyme (TACE) inhibitors, we designed a new class of macrocyclic hydroxamic acids by linking the P1 and P2' residues of acyclic anti-succinate-based hydroxamic acids. A variety of residues including amide, carbamate, alkyl, sulfonamido, Boc-amino, and amino were found to be suitable P1-P2' linkers. With an N-methylamide at P3', the 13-16-membered macrocycles prepared exhibited low micromolar activities in the inhibition of TNF-alpha release from LPS-stimulated human whole blood. Further elaboration in the P3'-P4' area using the cyclophane and cyclic carbamate templates led to the identification of a number of potent analogues with IC(50) values of </=0.2 microM in whole blood assay (WBA). Although the P3' area can accommodate a broad array of structurally diversified functional groups including polar residues, hydrophobic residues, and amino and carboxylic acid moieties, in both the cyclophane series and the cyclic carbamate series, a glycine residue at P3' was identified as a critical structural component to achieve both good in vitro potency and good oral activity. With a glycine residue at P3', an N-methylamide at P4' provided the best cyclophane analogue, SL422 (WBA IC(50) = 0.22 microM, LPS-mouse ED(50) = 15 mg/kg, po), whereas a morpholinylamide at P4' afforded the most potent and most orally active cyclic carbamate analogue, SP057 (WBA IC(50) = 0.067 microM, LPS-mouse ED(50) = 2.3 mg/kg, po). Further profiling for SL422 and SP057 showed that these macrocyclic compounds are potent TACE inhibitors, with K(i) values of 12 and 4.2 nM in the porcine TACE assay, and are broad-spectrum MMP inhibitors. Pharmacokinetic studies in beagle dogs revealed that SL422 and SP057 are orally bioavailable, with oral bioavailabilities of 11% and 23%, respectively.