ABSTRACT
Growth arrest and DNA damage-inducible-ß (Gadd45ß) is a stress-response protein involved in a number of processes, including cell cycle control, DNA repair, survival and death control, and stress signaling, depending on its interactions. Gadd45ß expression is dysregulated in numerous types of cancer, functioning as either a tumor promoter or a tumor suppressor. However, the functions of Gadd45ß in cholangiocarcinoma (CCA), particularly in metastasis, has not been studied. The immunohistochemical analysis of Gadd45ß expression revealed that 75% of histological specimens from patients with CCA expressed high levels of Gadd45ß, and that high Gadd45ß expression was associated with metastasis. The role of Gadd45ß in CCA was examined using siRNA-mediated gene knockdown in HuCCA-1, a human CCA cell line established from a Thai patient. The effects of Gadd45ß downregulation upon cell viability and death, invasion, migration, matrix metalloproteinase (MMP) activity and epithelial-mesenchymal transition (EMT) marker expression were investigated. Gadd45ß knockdown impaired cell viability, which was associated with the induction of apoptosis. In addition, there was a marked reduction in invasion and migration, although MMP activity was unaffected. Impairment of these metastatic properties was accompanied by the decreased expression of EMT markers, including Slug, vimentin, claudin-1 and zona occludens protein 1, whereas E-cadherin expression was increased. The present study suggests that Gadd45ß is involved in regulating the viability and the metastatic potential of CCA cells, which may be mediated by the modulation of the EMT pathway.
ABSTRACT
Cholangiocarcinoma (CCA) is a lethal malignancy of the biliary epithelium. CCA is resistant to currently available chemotherapy; therefore, new drugs as well as new molecular targets must be identified for the development of an effective treatment for CCA. The present study showed that RAD001 (everolimus), a derivative of rapamycin and an orally bioavailable mammalian target of rapamycin (mTOR) inhibitor, exhibits cytotoxic and antimetastatic effects in a CCA cell line, RMCCA-1. Treatment with low concentrations of RAD001 resulted in a significant reduction of in vitro invasion and migration of RMCCA-1, concomitant with a reduction of filopodia and alteration of the actin cytoskeleton. Although, matrix metalloproteinase-9 and -14 activities were unaltered. However, at high concentrations, RAD001 exhibited cytotoxic effects, reducing cell proliferation and inducing apoptotic cell death. Overall, RAD001 exhibits multiple effects mediated by the inhibition of the mTOR, which may serve as a promising agent for the treatment of CCA.
ABSTRACT
BACKGROUND: Genetic BRCA2 insufficiency is associated with breast cancer development; however, in sporadic breast cancer cases, high BRCA2 expression is paradoxically correlated with poor prognosis. Because DSS1, a mammalian component of the transcription/RNA export complex, is known to stabilize BRCA2, we investigated how the expression of DSS1 is associated with clinical parameters in breast cancers. METHODS: DSS1 mRNA and p53 protein were examined by RT-PCR and immunohistochemical staining of breast cancer specimens to classify DSS1(high) and DSS1(low) or p53(high) and p53(low) groups. Patient survival was compared using Kaplan-Meier method. DSS1(high) or DSS1(low) breast cancer cells were prepared by retroviral cDNA transfection or DSS1 siRNA on proliferation, cell cycle progression, and survival by flow cytometric analyses with or without anti-cancer drugs. RESULTS: In comparison to patients with low levels of DSS1, high-DSS1 patients showed a poorer prognosis, with respect to relapse-free survival period. The effect of DSS1 was examined in breast cancer cells in vitro. DSS1 high-expression reduces the susceptibility of MCF7 cells to DNA-damaging drugs, as observed in cell cycle and apoptosis analyses. DSS1 knockdown, however, increased the susceptibility to the DNA-damaging drugs camptothecin and etoposide and caused early apoptosis in p53 wild type MCF7 and p53-insufficient MDA-MB-231 cells. DSS1 knockdown suppresses the proliferation of drug-resistant MDA-MB-231 breast cancer cells, particularly effectively in combination with DNA-damaging agents. CONCLUSION: Breast cancers with high DSS1 expression have worse prognosis and shorter relapse-free survival times. DSS1 is necessary to rescue cells from DNA damage, but high DSS1 expression increases drug resistance. We suggest that DSS1 expression could be a useful marker for drug resistance in breast cancers, and DSS1 knockdown can induce tumor apoptosis when used in combination with DNA-damaging drugs.