The aim of the present study was to characterize the protein expression profiles of pancreatic islet β-cells affected by cancer cells, and to identify the potential islet molecules related to pancreatic cancer-associated diabetes. The cellular proteins of islet β-cell line INS-1 in response to conditioned medium (CM) prepared from pancreatic cancer cells were analyzed using fluorescence-labeled 2D gel-based proteomics (2D-DIGE). 10 proteins were over-expressed and 5 proteins were down-expressed in cancer CM-stimulated β-cells. Five differently expressed proteins were selected for further validation by Western-blot analysis. HSP60 and Peripherin, which are previously reported to be associated with type 1 diabetes, and Prp19, a DNA repair protein, were up-regulated in INS-1 cells after cancer cell CM stimulation; meanwhile, HMOX1 and GRP78 were down-regulated. Furthermore, our immunohistochemical study demonstrated that the islets adjacent to human pancreatic carcinomas showed increased Peripherin expression than normal pancreatic islets. Our findings provide new information regarding the regulation of protein expression in pancreatic cancer-associated diabetes.