Solid phase combinatorial library of 1,3-azole containing peptides for the discovery of matrix metallo proteinase inhibitors

A solid phase combinatorial library of approximately 240,000 putative inhibitors were prepared and screened for activity against MMP-14. The compounds of the library had the common structure H-XX-azole-XX-NH2 in which X is an arbitrary amino acid and azole is one of four oxazole or thiazole dipeptidomimetics (8a - d). The library was generated by the split and mix strategy resulting in a one-bead-two-compounds library format in such a way that each bead contained a common quenched fluorogenic substrate and a different putative inhibitor. Application of ladder synthesis [22] during library construction enabled full sequence determination by MALDI-TOF mass spectrometry. The library was screened against MMP-14 activity, by performing multiple, sequential incubations in which beads containing active inhibitors were selected by a novel automated bead-sorting apparatus, in order to isolate only the best inhibitors. Statistical analysis of the sequences obtained from overall 184 beads gave rise to a series of 20 sequences; 4 truncated and 16 full sequences were resynthesized. The inhibitors were investigated in an enzyme kinetic assay with MMP-14 revealing compounds with K-i values in the low micromolar area, whereas the truncated sequences derived from the library showed poor inhibitor activity.