Constrained peptide analogues of transforming growth factor-alpha residues cysteine 21-32 are mitogenically active. Use of proline mimetics to enhance biological potency.
Two proline mimetics, the enantiomers of 2-aza-bicyclo[2,2,1]heptane-3-carboxylic acid, have been incorporated in place of Pro30 into synthetic peptides based on the B-loop beta-sheet sequence of human transforming growth factor-alpha (TGF-alpha) (residues Cys21-Cys32). The peptides were further modified by inclusion of an N-terminal phenylalanine and constrained by formation of an intramolecular disulfide bond. While no mitogenic response was observed in the parental NR6 cell line, the peptides stimulated DNA synthesis in NR6/HER cells (NR6 fibroblasts transfected with the human epidermal growth factor receptor). Induction of DNA synthesis was dose dependent, with EC50 values in the range 130-330 microM; in the presence of low doses of TGF-alpha, the mitogenic effect of the peptides was additive, up to the plateau response achieved by maximal doses of TGF-alpha alone. These effects are consistent with the peptides acting via the same mechanism as TGF-alpha. Analysis of the structure of the peptides by NMR indicated that the presence of the mimetics significantly increased the propensity of the peptidyl-proline bond to adopt the cis conformation. These data confirm the role of the beta-sheet in receptor activation, and emphasize the importance of presentation of peptides in an appropriate conformation for recognition.
Chamberlin
S G
SG
Cancer Research Campaign Medical Oncology Unit, Southampton General Hospital, United Kingdom.
Sargood
K J
KJ
Richter
A
A
Mellor
J M
JM
Anderson
D W
DW
Richards
N G
NG
Turner
D L
DL
Sharma
R P
RP
Alexander
P
P
Davies
D E
DE
D000595:Amino Acid Sequence; D000818:Animals; D001643:Bridged Bicyclo Compounds / Q000737:chemistry*; D002455:Cell Division / Q000187:drug effects; D002460:Cell Line; D006801:Humans; D009682:Magnetic Resonance Spectroscopy; D051379:Mice; D018716:Molecular Mimicry; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry / Q000494:pharmacology*; D011392:Proline / Q000737:chemistry*; D011487:Protein Conformation; D013237:Stereoisomerism; D016211:Transforming Growth Factor alpha / Q000737:chemistry / Q000494:pharmacology*
C095673:2-azabicyclo(2.2.1)heptane-3-carboxylic acid; D001643:Bridged Bicyclo Compounds; D010455:Peptides; D016211:Transforming Growth Factor alpha; D011392:Proline