Epitopic regions for antibodies against tumor necrosis factor alpha. Analysis by synthetic peptide mapping

The location of biologically relevant epitopes on human tumor necrosis factor alpha (hTNF-alpha) was evaluated by testing the immunoreactivity of anti-TNF-alpha antibodies against 149 sequential, overlapping octamer peptides. A goat polyclonal antibody raised against recombinant hTNF-alpha, which neutralizes hTNF-alpha biological activities, reacted with oligopeptides corresponding to hTNF-alpha residues 7-11, 17-23, 30-39, 42-49, 106-112, and 135-142. A possible assembled epitopic region (residues 25, 27, and 144) for neutralizing murine monoclonal antibodies designated 11D7G4 and 9C4G5 was deduced from the fact that they bound to tripeptides, mimicking a discontinuous epitope. These antigenic regions wer found to include of adjoin poorly conserved amino acids and they were located in the turns between beta-sheets on the surface of the molecule. Three of the sequential epitopic regions and an assembled region were closely related to the receptor binding sites proposed in several other studies. These antibodies appear to neutralize TNF-alpha activities by directly masking receptor binding sites.