OBJECTIVE Hyperglycosylated hCG (hCG-H) is a glycosylation variant of hCG produced by cytotrophoblast cells at implantation of pregnancy and in choriocarcinoma. We investigated the biological function of hCG-H in invasion in vitro and in vivo and the use of hCG-H antibodies in blocking tumorigenesis and cancer growth in vivo. RESULTS hCG-H accounts for 43% to 100% of total hCG immunoreactivity in the culture fluid of choriocarcinoma cell lines and 100% in primary cultures of pregnancy cytotrophoblast cells. We investigated the action of hCG and hCG-H on isolated cytotrophoblast cell primary cultures and on 3 different lines of choriocarcinoma cells cultured on Matrigel basement membrane inserts (culture models for assessing tumor invasion). The addition of hCG-H to medium significantly promoted invasion of membranes with both pregnancy and cancer cell line sources, while regular hCG had no significant effect. JEG-3 human choriocarcinoma cells were transplanted subcutaneously into athymic nude mice. Tumors rapidly formed. B152, mouse monoclonal antibody against hCG-H, and non-specific mouse IgG (control) were administered twice weekly once tumors were clearly visible. While a correlation between time and growth was observed with the control group (r(2)=0.97), no correlation was observed with the B152-treated mice (r(2)=0.15). B152 blocked tumor growth (t test, IgG vs. B152, P=0.003). In a second experiment, antibody B152 or IgG was administered to mice at the time of choriocarcinoma transplantation. B152 significantly inhibited tumorigenesis (t test P=0.0071). CONCLUSIONS hCG-H is a critical promoter in human cytotrophoblast and human choriocarcinoma cell invasion in vivo and in vitro, promoting tumor growth and invasion through an autocrine mechanism. hCG-H is a signal for choriocarcinoma cell invasion, making it a biological tumor marker. Antibodies against hCG-H block tumor formation and growth. Human or humanized antibodies against hCG-H may be useful in treating and managing choriocarcinoma and other gestational trophoblastic malignancies.