AbstractHeparin‐binding epidermal growth factor‐like growth factor (HB‐EGF) is a member of the EGF family. Juxtacrine activity of proHB‐EGF (the membrane‐anchored form of HB‐EGF) has been shown to be significantly potentiated when it is coexpressed with CD9 in vitro. The purpose of our study was to investigate the issue of whether proHB‐EGF and CD9 are coexpressed in gastric cancer. HB‐EGF gene expression and protein production in human gastric cancers was investigated, and EGF receptor and CD9 expressions were also evaluated. HB‐EGF mRNA levels in gastric cancers were elevated, compared with normal gastric tissues, especially in the intestinal type. ProHB‐EGF immunoreactivity was detected primarily in the cytoplasm and plasma membrane of gastric cancer cells. Of 66 patients, 40 (60.6%) exhibited proHB‐EGF immunoreactivity and the level of its expression was significantly associated with tumor status (p < 0.01) and histological differentiation (p < 0.001). In addition, proHB‐EGF mRNA was detected at high levels in the intestinal type by in situ hybridization. CD9 immunoreactivity was found to be preserved in 26 of 36 patients (72.2%) and CD9 protein expression was inversely associated with lymph node status (p < 0.05). A significant correlation between its expression and histological differentiation (p < 0.01) was found, and the association of CD9 with proHB‐EGF was increased in the intestinal type, as evidenced by an immunoprecipitation method. These results indicate that the coexpression of proHB‐EGF and CD9 may be involved in the tumorigenesis and/or proliferation of gastric cancers in a juxtacrine manner. © 2002 Wiley‐Liss, Inc.