The discovery of cyclophilin A (CypA) inhibitor is now of special interest in the treatment of immunological
disorders. In this work, using a strategy integrating focused combinatorial library design, virtual screening,
chemical synthesis, and bioassay, a series of novel small molecular CypA inhibitors have been discovered.
First, using the fragments taken from our previously discovered CypA inhibitors (Bioorg. Med. Chem. 2006,
14, 2209-2224) as building blocks, we designed a focused combinatorial library containing 255 molecules
employing the LD1.0 program (J. Comb. Chem. 2005, 7, 398-406) developed by us. Sixteen compounds
(1a-e, 2a-b, 3a-b, and 4a-g) were selected by using virtual screening against the X-ray crystal structure
of CypA as well as druglike analysis for further synthesis and bioassay. All these sixteen molecules are
CypA binders with binding affinities (KD values) ranging from 0.076 to 41.0 M, and five of them (4a, 4c,
and 4e-g) are potent CypA inhibitors with PPIase inhibitory activities (IC50 values) of 0.25-6.43 M. The
hit rates for binders and inhibitors are as high as 100% and 31.25%, respectively. Remarkably, both the
binding affinity and inhibitory activity of the most potent compound increase 10 times than that of the
most active compound discovered previously. The high hit rate and the high potency of the new CypA
inhibitors demonstrated the efficiency of the strategy for focused library design and screening. In addition,
the novel chemical entities reported in this study could be leads for discovering new therapies against the
CypA pathway.