Potent inhibition of protein-tyrosine phosphatase by phosphotyrosine-mimic containing cyclic peptides

Abstract

In an effort to derive potent and bioavailable protein-tyrosine phosphatase inhibitors, we have previously reported hexameric peptides based on the epidermal growth factor receptor sequence EGFR988–993 (Asp-Ala-Asp-Glu-Xxx-Leu, where Xxx = Tyr), in which the tyrosyl residue has been replaced by the non-hydrolyzable phosphotyrosyl mimics phosphonomethylphenylalanine (Pmp), difluorophosphonomethylphenylalanine (F2Pmp) and O-malonyltyrosine (OMT). Inhibitory potencies (IC50 values) of these peptides against the tyrosine phosphatase PTP 1B were 200, 0.2 and 10 μM, respectively. Since cellular penetration of peptides containing highly charged phosphonate residues is compromised, and good bioreversible protection strategies for the F2Pmp residue have not yet been reported, the OMT residue is of particular interest in that it affords potential new prodrug approaches. In the current study we have prepared cyclized versions of the OMT-containing EGFR988–993 peptide in order to increase its proteolytic stability and restrain conformational flexibility. Three different cyclic analogues were synthesized. Two of these were cyclized through the peptide backbone (‘head to tail’) using in one case a single glycine spacer (heptamer peptide) and in the second instance, two glycines (octamer peptide). In a PTP1-based assay the cyclic heptamer experienced a two-fold loss of potency (Ki= 25.2±3.9 μM) relative to the linear hexamer parent (Ki= 13±0.9 μM), while the cyclic octamer demonstrated a five-fold increase in potency (Ki= 2.60±0.11 μM).The third peptide was cyclized by means of a sulfide bridge between the side chain of a C-terminally added cysteine residue and the β-carbon of a N-terminal acetyl residue. Although the overall size of this ring was identical to that exhibited by the preceding backbone-cyclized octamer, it displayed a three-fold enhancement in potency (Ki= 0.73±0.03 μM). The structural basis for the observed results are discussed. Conformational restrictions induced by cyclization could aid in defining geometries for peptidomimetic design. Finally, it can be speculated that cyclization of other linear PTP-inhibitory peptides, such as the F2Pmp-containing hexamer, may also increase their potency.