Immunogenic regions of the GA733-2 tumour-associated antigen recognised by autoantibodies of patients with colorectal carcinoma. 
The tumour-associated antigen (TAA) GA733-2 is overexpressed by >90% of human colorectal carcinomas (CRC).  The antigen has previously been shown to be recognised by B and T cells.  The aim of the present study was to define B cell epitopes of GA733-2.  Fifteen percent of CRC patients with no previous immunotherapy have recently been shown to elicit an anti-GA733-2 IgG antibody response.  Sera of these patients ( n=136) were analysed by enzyme-linked immunosorbent assay (ELISA) for immunoglobulin G (IgG) antibodies against 23 partly overlapping synthetic peptides (18 amino acids: aa) derived from the extracellular domain of GA733-2.  An 18-aa long sequence at the N-terminal region of the antigen (peptide 2) was found to be an immunodominant B cell epitope.  Fifty percent of the patients had antibodies against peptide 2, while 8% to 9% had antibodies against peptides 1, 4, 7, 8 or 20.  In healthy donors ( n=30) antibodies against peptides 2 and 8 were also detected in 13% and 3% of cases respectively, while no antibodies were found against the other peptides and the complete protein.  Thirteen percent of CRC patients ( n=30) with no IgG antibodies against the GA733-2 antigen elicited antibodies against peptide 2.  The specificity of peptide-reactive sera was verified by inhibition ELISA.  The binding of sera to GA733-2 was significantly inhibited by peptides to which CRC sera bound, but not by control peptides.  Binding to peptide 2 of sera showing both peptide 2 and GA733-2 reactivity was specifically inhibited by the complete GA733-2 antigen, while binding of peptide 2-reactive sera showing no GA733-2 reactivity was not inhibited.  CRC sera interfered with the binding of monoclonal antibody (mAb) 17-1A and mAb C215 that recognise distinct epitopes of GA733-2.  No significant correlation was found between the presence of anti-peptide antibodies in CRC patients and clinical stage or overall survival.  The results provide additional evidence for immune recognition of CRC by the host.