Six highly active mu-selective opioid peptides identified from two synthetic combinatorial libraries.

Two synthetic combinatorial libraries (SCLs) were prepared, each composed of 52,128,400 L-amino acid hexapeptides, one with and the other without an N-terminal acetyl moiety. The two libraries were used in conjunction with an iterative selection process to identify individual peptides capable of inhibiting the binding of the mu-selective opioid peptide [3H]-[D-Ala2,MePhe4,Gly-ol5]enkephalin to rat brain homogenates. As reported previously, when using the nonacetylated SCL the first five residues identified corresponded exactly to methionine- and leucine-enkephalin, both of which are endogenous opioid peptides. The iterative identification process has now been completed for two additional mixtures found to have activity in the initial screening of this SCL. Two new series unrelated to the enkephalins have been identified: YPFGFO-NH2 and WWPKHO-NH2 (where O = one of the 20 L-amino acids). Individual peptides from each of these were found to be agonists at the mu receptor and have high affinity (IC50 values of the most active peptides were 10-15 nM) and selectivity for the mu receptor. In addition to the acetalins (described previously), two new series have now been identified from the acetylated library: Ac-FRWWYO-NH2 and Ac-RWIG-WO-NH2 (IC50 values of the most active peptides were 5-30 nM). Ac-FRWWYM-NH2 was determined to be an agonist at the mu receptor, whereas Ac-RWIGWR-NH2 was found to be an antagonist at this receptor.