FRAGMENTS OF HUMAN CHORIONIC-GONADOTROPIN (HCG) - IMPACT ON PREGNANCYAND TUMOR-DIAGNOSIS

The pregnancy and tumor marker human chrionic gonadotropin (hCG) belon gs to the family of the glyco-protein hormones. Information on epitope forming sequences of hCG and its subunits hCGalpha and hCGbeta has si gnificant impact on the examination of intra- and extracellular metabo lism and the standardization of diagnostic assay systems. Variants of hCG appear in biological fluids with variable modifications on differe nt parts of the molecule. These changes may influence the binding patt erns of monoclonal antibodies (MCA), thereby causing erroneous results in hCG immunoassays. The aim of the present work was to investigate t he influence of peptide bond cleavages and the loss of certain segment s of the molecule, which were induced by proteases on the expression o f the seven hCGalpha(alpha1-alpha7), nine hCGbeta- (beta1-beta9) and f our hCGbeta-core-fragment-epitopes (beta10-beta13), previously identif ied by us [1-10]. To this end, we digested hCGalpha and hCGbeta with c hymotrypsin. Hormone fragments were separated by high performance liqu id chromatography (HPLC) and subsequently immunochemically examined by direct binding radioimmunoassay (DB-RIA), competitive RIA and immunoe nzymometric assays (IEMA). Fractions containing hCG-like immunoreactiv ity were sequenced by Edman and carboxypeptidase-Y degradation. It app eared that: (I) Amino acids (AA) alpha 41-47 and the peptide bonds bet ween AA alpha 40/41, alpha 47/48 and alpha 29/30 do not influence the expression of the 7 alpha-epitopes. (II) The absence of the hCGbeta N- terminus plays a crucial role for the formation of epitopes beta10 and beta13. (III) Neither the presence nor the absence of the C-terminal peptide of hCGbeta (hCGbetaCTP, AA beta 114-145) has any importance fo r the expression of epitopes beta1-beta7 and beta10-beta13. (IV) Since epitopes beta11 and beta12 are not detectable on chymotryptically dig ested hCGbeta, we conclude that AA beta 1-5, beta 115-145 and the inte rruption of the peptide bond between the AA beta 45/46 contribute neit her directly nor indirectly to the formation of those epitopes. Thus, immunometric assay systems based on carefully selected MCA can measure enzymatically altered and metabolic products of hCGalpha and -beta mo lecules and are therefore suitable tools for hCG-diagnostic tests whic h are reliable with respect to both specifity and sensitivity.