We used immunohistochemical staining to assess protein over-expression of glial cell-derived neurotrophic factor (GDNF) and its RET receptor tyrosine kinase in patients with pancreatic cancer and benign pancreatic neoplasm, and assessed correlations with clinicopathological features and prognosis. Surgically resected pancreatic cancer patients (40/58, 68.9%) showed positive GDNF immunostaining, a significantly higher frequency than in patients with benign pancreatic tumour (3/11, 27.3%). Intrapancreatic neural invasion by cancer cells was significantly related to over-expression of GDNF. Strongly positive expression of GDNF was significantly more frequent than lesser grades of expression in patients with severe back pain before and 12 months after surgery. Expression of RET was significantly related to lymphatic invasion, survival rate after tumour resection and degree of tumour cell differentiation. We conclude that GDNF may be important in pancreatic cancer proliferation and metastasis, especially in patients with perineural invasion. Strongly positive expression of GDNF may be an indication for early intensified radiotherapy. RET expression in pancreatic cancer tissues may be a useful prognostic marker.