Circulating
autoantibodies against the
glucose-regulated
protein of 78 kDa (
GRP78) are present at high levels in
prostate cancer patients and are a
biomarker of aggressive
tumor behavior. We purified the anti-GRP78 IgGs and examined their effect on 1-LN, PC-3, DU145, and LnCap human
prostate cancer cells. We also evaluated its effects on the
breast cancer MDA-MB231 and
melanoma DM413 cell lines. The anti-GRP78 antibody binds only to cells expressing
GRP78 on the surface, to a site also recognized by its physiologic agonist, activated alpha(2)-macroglobulin (alpha(2)M*). This antibody is completely specific for a
peptide, including the primary amino acid sequence CNVKSDKSC, which contains a tertiary structural motif mimicking an
epitope in
GRP78. Tertiary structural analysis suggested the linear
GRP78 primary amino acid sequence LIGRTWNDPSVQQDIKFL (Leu(98)-Leu(115)) as the putative binding site, containing the tertiary structual arrangement described above, which was confirmed experimentally. The anti-GRP78
antibodies from
prostate cancer patients recognize almost exclusively this
epitope. We produced animal
antibodies against both these
peptides, and they are able to mimic the effects of the human antibody. Our experiments also suggest this
epitope as highly immunogenic, thereby explaining the specificity of the immune response against this
epitope in
GRP78, observed in humans. Using 1-LN cells as a model, we show that anti-GRP78
IgG purified from the sera of these patients mimics the proproliferative effects induced by alpha(2)M* via the common receptor,
GRP78. Furthermore, increasing concentrations of human anti-GRP78
IgG show a dose-dependent protective effect on apoptosis induced by
tumor necrosis factor alpha.