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Highlights • Regucalcin expression is reduced in the metastatic tumors of prostate cancer patients. • The progression-free survival rate is prolonged in patients with a higher regucalcin expression. • Overexpressed regucalcin suppresses the growth and bone activity of human prostate cancer cells. • Overexpressed regucalcin regulates many proteins in cell signaling and transcription processes. • Regucalcin is suppressor in human prostate cancer, suggesting a novel therapeutic strategy., Prostate cancer, which is a bone metastatic cancer, is the second leading cause of cancer-related death in men. There is no effective treatment for metastatic prostate cancer. Regucalcin has been shown to contribute as a suppressor in various types of human cancers. In the present study, furthermore, we investigate an involvement of regucalcin in suppression of prostate cancer. Regucalcin expression was compared in 131 primary tumor tissues and 19 metastatic tumor tissues in prostate cancer patients. Regucalcin expression in the metastatic tumor was found to be reduced as compared with that in primary tumor. The progression-free survival rate was prolonged in patients with a higher regucalcin expression. Translationally, overexpression of regucalcin in bone metastatic human prostate cancer PC-3 and DU-145 cells suppressed colony formation and cell growth in vitro. Mechanistically, overexpressed regucalcin enhanced the levels of p53, Rb, and p21, and decreased the levels of Ras, PI3 kinase, Akt, and mitogen-activated protein kinase, leading to suppression of cell growth. Furthermore, higher regucalcin expression suppressed the levels of nuclear factor-κB p65, β-catenin, and signal transducer and activator of transcription 3, which regulate a transcription activity. Cell growth was promoted by culturing with the calcium agonist Bay K 8644. This effect was blocked by overexpression of regucalcin. Notably, overexpressed regucalcin suppressed bone metastatic activity of PC-3 and DU-145 cells when cocultured with preosteoblastic or preosteoclastic cells. Regucalcin may suppress the development of human prostate cancer, suggesting that gene delivery systems in which its expression is forced may be a novel therapeutic strategy.

Ras mutations are present in only a subset of sporadic human cutaneous squamous cell carcinomas (cSCC) even though Ras is activated in most. This suggests that other mechanisms of Ras activation play a role in the disease. The aberrant expression of RasGRP1, a guanyl nucleotide exchange factor for Ras, is critical for mouse cSCC development through its ability to increase Ras activity. However, the role of RasGRP1 in human keratinocyte carcinogenesis remains unknown. Here we report that RasGRP1 is significantly elevated in human cSCC and that high RasGRP1 expression in human primary keratinocytes triggered activation of endogenous Ras and significant morphological changes including cytoplasmic vacuole formation and growth arrest. Moreover, RasGRP1-expressing cells were autophagic as indicated by LC3-II increase and the formation of LC3 punctae. In an in vitro organotypic skin model, wild type keratinocytes generated a well-stratified epithelium, while RasGRP1-expressing cells failed to do so. Finally, RasGRP1 induced transformation-like changes in skin cells from Li-Fraumeni patients with inactivating p53 mutations, demonstrating the oncogenic potential of this protein. These results support a role for RasGRP1 in human epidermal keratinocyte carcinogenesis and might serve as an important new therapeutic target.