Structures of disease-specific serum alpha-fetoprotein isoforms

Alpha-fetoprotein (AFP) is widely used as a serological marker in the diagn osis of hepatocellular carcinoma (HCC) and non-seminomatous germ cell tumou rs (NSGCT). By application of isoelectric focusing (IEF) disease-specific A FP isoforms can be identified Three major bands are apparent: + I (associat ed with 'benign' liver disease), + II (associated with HCC) and +[I( (assoc iated with NSGCT). Recently we have characterized the predominant glycans o f human serum AFP and now report the application of these findings and elec trospray ionization-mass spectrometry (ESI-MS) to the determination of the glycan composition of the isoforms present in the sera of 12 patients with MCC and of one patient with NSGCT. ESI-MS allowed simultaneous identificati on of various AFP glycoforms in purified serum AFP. Seven glycoforms were i dentified, but with different abundance in the sera of the HCC patients, wh ereas six glycoforms were identified in the serum from the NSGCT patient. T he glycan structures of these glycoforms were deduced from their observed m asses. AFP glycoforms carrying a single biantennary complex-type N-glycan a ppeared as the predominant glycoforms, whereas those carrying both N-glycan and O-glycan appeared as minor glycoforms. Correlation between the abundan ce of the AFP glycoforms and the IEF band intensity suggested that differen t degrees in sialylation cause the formation of isoforms. This contention w as subsequently supported by the ESI-MS and kinetic in vitro desialylation studies on purified Bands + I and + II AFPs. Our findings indicate that HCC -associated isoforms (Band + [I) represent a group of glycoproteins whose c arbohydrate structures are ail characterized by being mono-sialylated, wher eas those associated with benign liver disease and NSGCT are di- and a-sial o species, respectively. Knowledge of the structure of the tumour-specific isoforms should form an important basis for clinically useful assays. (C) 2 000 Cancer Research Campaign.