Selection of a histidine-containing inhibitor of gelatinases through deconvolution of combinatorial tetrapeptide libraries.
A fully automated peptide synthesizer was used to generate tetrapeptide sublibraries from 24 natural and nonnatural amino acids, from which new inhibitors of gelatinases (matrix metalloproteinases MMP-2 and MMP-9) were selected as potential anticancer drugs. MMP-2 and MMP-9 from mouse Balbc/3T3 fibroblasts conditioned media were assayed in their linear range response by zymography to quantify inhibition at each step of the tetrapeptide library deconvolution. The histidine-epsilon-amino caproic acid-beta-alanine-histidine (His-epsilon Ahx-beta Ala-His) sequence was found to yield optimal inhibition of both MMP-2 and MMP-9. Inhibition by selected tetrapeptides was also evaluated with two other techniques, a native type IV collagen degradation assay and a fluorogenic enzymatic assay, confirming the tetrapeptide potency. The His-epsilon Ahx-beta Ala-His tetrapeptide also inhibited purified human MMP-2 and MMP-9 and the corresponding enzymes present in conditioned media from human tumour cells. Finally, the length of the spacer between the two terminal histidines was found to be crucial to the inhibitory potential. This approach may thus be considered as a-successful strategy to yield specific peptide or pseudopeptide inhibitors, although their potency remains moderate, since it was measured before any chemical optimization was undertaken.
Ferry
G
G
Department of Tumour Biology, Institut de Recherches Servier, Suresnes, France.
Boutin
J A
JA
Atassi
G
G
Fauchère
J L
JL
Tucker
G C
GC
D016475:3T3 Cells; D000595:Amino Acid Sequence; D000818:Animals; D000970:Antineoplastic Agents / Q000138:chemical synthesis* / Q000737:chemistry / Q000494:pharmacology; D017077:Culture Media, Conditioned; D015195:Drug Design; D018093:Gelatinases / Q000037:antagonists & inhibitors*; D006639:Histidine / Q000737:chemistry; D006801:Humans; D020778:Matrix Metalloproteinase 2; D020780:Matrix Metalloproteinase 9; D061965:Matrix Metalloproteinase Inhibitors; D008666:Metalloendopeptidases / Q000037:antagonists & inhibitors; D051379:Mice; D015394:Molecular Structure; D009842:Oligopeptides / Q000138:chemical synthesis / Q000737:chemistry / Q000494:pharmacology; D019151:Peptide Library; D011480:Protease Inhibitors / Q000138:chemical synthesis* / Q000737:chemistry / Q000494:pharmacology; D013329:Structure-Activity Relationship; D014407:Tumor Cells, Cultured
D000970:Antineoplastic Agents; D017077:Culture Media, Conditioned; D061965:Matrix Metalloproteinase Inhibitors; D009842:Oligopeptides; D019151:Peptide Library; D011480:Protease Inhibitors; D006639:Histidine; D018093:Gelatinases; D008666:Metalloendopeptidases; D020778:Matrix Metalloproteinase 2; D020780:Matrix Metalloproteinase 9