Change of antigenic and neutralizing specificity in substitutional epitope peptides of hemophilia B inhibitor.
We have previously described the epitope mapping and functional neutralization of three factor IX inhibitors in hemophilia B (HB-1, 3, and 7) by synthetic peptides (13). However, A concentration of synthetic peptide of about 1000 times the concentration of factor IX in plasma was essential to neutralize the purified antibodies. We now report that substitutional synthetic peptides of epitope are able to neutralize the factor IX inhibitor with a lower concentration. Using two major epitope peptides, 156Val-Asn-Ser-Thr-Glu-Ala-Glu-Thr-Ile164 and 168Ile-Thr-Gln-Ser-Thr-Gln-Ser-Phe-Asn176, we designed changes of antigenicity using the systematic substitution of different amino acids at each residue of the epitope peptides and neutralization rate of factor IX inhibitors by a lower concentration of substitutional synthetic peptides conjugated with bovine serum albumin (BSA). Twenty-five substitutional peptides for HB-1, twenty substitutional peptides for HB-3 and forty-four substitutional peptides for HB-7 reacted stronger than the native sequences. One of the peptides, 1.0 microM of 156Val-Asn-Ser-Thr-Glu-Tyr-Glu-Thr-Ile164 conjugated with BSA, neutralized 26.5% of inhibitor in HB-1's plasma maximally. Our data show that high antigenicity peptides conjugated with BSA ranging in concentration from 0.1 microM to 1.0 microM are able to neutralize factor IX inhibitors in plasma and there is a possibility of peptide neutralization inhibitor therapy.
Takahashi
I
I
Japanese Red Cross Aichi Blood Center, Seto, Japan. isaotaka@mxa.usen-net.or.jp
Yako
F
F
Saito
H
H
Kamiya
T
T
D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000906:Antibodies / Q000276:immunology*; D001665:Binding Sites; D000939:Epitopes / Q000276:immunology*; D005164:Factor IX / Q000037:antagonists & inhibitors / Q000276:immunology*; D002836:Hemophilia B / Q000276:immunology*; D006801:Humans; D008301:Maleimides / Q000378:metabolism; D009500:Neutralization Tests; D010455:Peptides / Q000138:chemical synthesis* / Q000276:immunology / Q000627:therapeutic use; D011485:Protein Binding; D012710:Serum Albumin, Bovine
D000906:Antibodies; D000939:Epitopes; D008301:Maleimides; D010455:Peptides; C043592:maleimide; D012710:Serum Albumin, Bovine; D005164:Factor IX