Constitutive activation of glycogen synthase kinase-3beta correlates with better prognosis and cyclin-dependent kinase inhibitors in human gastric cancer

Abstract

Background: Aberrant regulation of glycogen synthase kinase-3beta (GSK-3beta) has been implicated in several human cancers; however, it has not been reported in the gastric cancer tissues to date. The present study was performed to determine the expression status of active form of GSK-3beta phosphorylated at Tyr216 (pGSK-3beta) and its relationship with other tumor-associated proteins in human gastric cancers.

Methods: Immunohistochemistry was performed on tissue array slides containing 281 human gastric carcinoma specimens. In addition, gastric cancer cells were cultured and treated with a GSK-3beta inhibitor lithium chloride (LiCl) for immunoblot analysis.

Results: We found that pGSK-3beta was expressed in 129 (46%) of 281 cases examined, and was higher in the early-stages of pathologic tumor-node-metastasis (P < 0.001). The expression of pGSK-3beta inversely correlated with lymphatic invasion (P < 0.001) and lymph node metastasis (P < 0.001) and correlated with a longer patient survival (P < 0.001). In addition, pGSK-3beta expression positively correlated with that of p16, p21, p27, p53, APC, PTEN, MGMT, SMAD4, or KAI1 (P < 0.05), but not with that of cyclin D1. This was confirmed by immunoblot analysis using SNU-668 gastric cancer cells treated with LiCl.

Conclusions: GSK-3beta activation was frequently observed in early-stage gastric carcinoma and was significantly correlated with better prognosis. Thus, these findings suggest that GSK-3beta activation is a useful prognostic marker for the early-stage gastric cancer.

Figure 1

Representative immunohistochemial features in gastric cancer tissues and normal gastric tissues. Positive versus negative examples of gastric cancer for pGSK-3β (A and B) and cyclin D1 (D and E) (× 400). (C) A normal gastric tissue stained for pGSK-3β (× 200). (F) Negative control treated without the primary antibodies (× 400).

Figure 2

Kaplan-Meier curves for patient survival. pGSK-3β-positive carcinomas showed a more favorable prognosis than pGSK-3β-negative carcinomas (P < 0.001, A). Survival rate was dependent on pGSK-3β expression in early-stage tumors (P = 0.034, B), but not in late-stage tumors (P = 0.918, C).

Figure 3

Protein expressions of pGSK-3β and cyclin D1 in SNU-668 gastric cancer cells. Cells were treated with or without a GSK-3β inhibitor LiCl (30 mM) for 48 hours and immunoblot analysis was performed for pGSK-3β and cyclin D1. β-actin was used as internal control.