Metformin is an anti-diabetic drug and has anticancer effects on various cancers. Several studies have suggested that metformin reduces cell proliferation and stimulates cell-cycle arrest and apoptosis. However, the definitive molecular mechanism of metformin in the pathophysiological signaling in endometrial tumorigenesis and metastasis is not clearly understood. In this study, we examined the effects of metformin on the cell viability and apoptosis of human cervical HeLa and endometrial HEC-1-A and KLE cancer cells. Metformin suppressed cell growth in a dose-dependent manner and dramatically evoked apoptosis in HeLa cervical cancer cells, while apoptotic cell death and growth inhibition were not observed in endometrial (HEC-1-A, KLE) cell lines. Accordingly, the p27 and p21 promoter activities were enhanced while Bcl-2 and IL-6 activities were significantly reduced by metformin treatment. Metformin diminished the phosphorylation of mTOR, p70S6K and 4E-BP1 by accelerating adenosine monophosphateactivated kinase (AMPK) in HeLa cancer cells, but it did not affect other cell lines. To determine why the anti-proliferative effects are observed only in HeLa cells, we examined the expression level of liver kinase B1 (LKB1) since metformin and LKB1 share the same signalling system, and we found that the LKB1 gene is not expressed only in HeLa cancer cells. Consistently, the overexpression of LKB1 in HeLa cancer cells prevented metformin-triggered apoptosis while LKB1 knockdown significantly increased apoptosis in HEC-1-A and KLE cancer cells. Taken together, these findings indicate an underlying biological/physiological molecular function specifically for metformin-triggered apoptosis dependent on the presence of the LKB1 gene in tumorigenesis.

HPV16 E6 oncoprotein is a member of the human papillomavirus (HPV) family that contributes to enhanced cellular proliferation and risk of cervical cancer progression via viral infection. In this study, interferon regulatory factor-1 (IRF-1) regulates cell growth inhibition and transcription factors in immune response, and acts as an HPV16 E6-binding cellular molecule. Over-expression of HPV16 E6 elevated cell growth by attenuating IRF-1-induced apoptosis and repressing p21 and p53 expression, but activating cyclin D1 and nuclear factor kappa B (NF-κB) expression. The promoter activities of p21 and p53 were suppressed, whereas NF-κB activities were increased by HPV16 E6. Additionally, the cell viability of HPV16 E6 was diminished by IRF-1 in a dose-dependent manner. We found that HPV16 E6 activated vascular endothelial growth factor (VEGF)-induced endothelial cell migration and proliferation as well as phosphorylation of VEGFR-2 via direct interaction in vitro. HPV16 E6 exhibited potent pro-angiogenic activity and clearly enhanced the levels of hypoxia-inducible factor-1α (HIF-1α). By contrast, the loss of function of HPV16 E6 by siRNA-mediated knockdown inhibited the cellular events. These data provide direct evidence that HPV16 E6 facilitates tumour growth and angiogenesis. HPV16 E6 also activates the PI3K/mTOR signalling cascades, and IRF-1 suppresses HPV16 E6-induced tumourigenesis and angiogenesis. Collectively, these findings suggest a biological mechanism underlying the HPV16 E6-related activity in cervical tumourigenesis.

Snail is implicated in tumour growth and metastasis and is up-regulated in various human tumours. Although the role of Snails in epithelial-mesenchymal transition, which is particularly important in cancer metastasis, is well known, how they regulate tumour growth is poorly described. In this study, the possible molecular mechanisms of Snail in tumour growth were explored. Baculoviral inhibitor of apoptosis protein (IAP) repeat-containing protein 3 (BIRC3), a co-activator of cell proliferation during tumourigenesis, was identified as a Snail-binding protein via a yeast two-hybrid system. Since BIRC3 is important for cell survival, the effect of BIRC3 binding partner Snail on cell survival was investigated in ovarian cancer cell lines. Results revealed that Bax expression was activated, while the expression levels of anti-apoptotic proteins were markedly decreased by small interfering RNA (siRNA) specific for Snail (siSnail). siSnail, the binding partner of siBIRC3, activated the tumour suppressor function of p53 by promoting p53 protein stability. Conversely, BIRC3 could interact with Snail, for this reason, the possibility of BIRC3 involvement in EMT was investigated. BIRC3 overexpression resulted in a decreased expression of the epithelial marker and an increased expression of the mesenchymal markers. siSnail or siBIRC3 reduced the mRNA levels of matrix metalloproteinase (MMP)-2 and MMP-9. These results provide evidence that Snail promotes cell proliferation by interacting with BIRC3 and that BIRC3 might be involved in EMT via binding to Snail in ovarian cancer cells. Therefore, our results suggested the novel relevance of BIRC3, the binding partner of Snail, in ovarian cancer development.

The human papillomavirus (HPV)-18 E7 (E7) oncoprotein is a major transforming protein that is thought to be involved in the development of cervical cancer. It is well-known that E7 stimulates tumour development by inactivating pRb. However, this alone cannot explain the various characteristics acquired by HPV infection. Therefore, we examined other molecules that could help explain the acquired cancer properties during E7-induced cancer development. Using the yeast two-hybrid (Y2H) method, we found that the Elk-1 factor, which is crucial for cell proliferation, invasion, cell survival, anti-apoptotic activity, and cancer development, binds to the E7. By determining which part of E7 binds to which domain of Elk-1 using the Y2H method, it was found that CR2 and CR3 of the E7 and parts 1–206, including the ETS-DNA domain of Elk-1, interact with each other. As a result of their interaction, the transcriptional activity of Elk-1 was increased, thereby increasing the expression of target genes EGR-1, c-fos, and E2F. Additionally, the colony forming assay revealed that overexpression of Elk-1 and E7 promotes C33A cell proliferation. We expect that the discovery of a novel E7 function as an Elk-1 activator could help explain whether the E7 has novel oncogenic activities in addition to p53 inactivation. We also expect that it will offer new methods for developing improved strategies for cervical cancer treatment.

Cervical tumors represent a prevalent form of cancer affecting women worldwide; current treatment options involve surgery, radiotherapy, and chemotherapy. Angiogenesis, the process of new blood vessel formation, is a crucial factor in cervical tumor growth. The molecular mechanisms underlying the effects of the liver kinase B1 (LKB1/STK11) tumor suppressor protein on tumor angiogenesis have not been elucidated. Therefore, we investigated the role of LKB1 in cervical tumor angiogenesis both in vitro and in vivo in this study. Our results demonstrated that LKB1 inhibited cervical tumor angiogenesis by suppressing the expression of angiogenesis-related factors such as vascular endothelial growth factor (VEGF) and hypoxia inducible factor-1α. LKB1 directly affected both carcinoma and vascular endothelial cells, resulting in a significant reduction in tumor growth and angiogenesis. Furthermore, LKB1 was found to bind to VEGF receptor 2 (VEGFR-2) and target the VEGFR-2-mediated protein kinase B/ mechanistic target of rapamycin signaling pathway in endothelial cells, thereby reducing cervical tumor growth and angiogenesis. Our study provides new insights into the molecular mechanisms underlying the anti-tumor and anti-angiogenic effects of LKB1 in cervical cancer. These findings will help develop new therapeutic strategies for cervical cancer.