Abstract. Constitutive activation of the signal transducer and activator of transcription 3 (STAT3) signaling pathway possesses confirmed oncogenic potential in oral squamous cell carcinoma (OSCC). Crosstalk with other molecular pathways contributes to STAT3 regulation in cancer. The effects of mitogen-activated protein kinases (MAPKs) and particularly extracellular signal-regulated kinase 1/2 (Erk1/2) on STAT3 signaling in OSCC have not been thoroughly investigated. The present study examined the effects of Erk1/2 modulation on STAT3 signaling and cell growth in OSCC cells. Constitutive expression levels of phosphorylated (tyrosine and serine) and total STAT3, Erk1/2 and cyclin D1 were assessed in OSCC cell lines. Erk1/2 modulation was achieved by pharmacological agents; siRNA silencing against Erk1/2 was also performed. Cell proliferation and viability were assessed. Erk1/2 inhibition with either U0126 treatment or specific siRNA silencing resulted in decreases in p-ser STAT3 and cyclin D1 levels and increases in p-tyr STAT3 in OSCC cells. Moreover, Erk1/2 inhibition resulted in a dose-dependent reduction in OSCC cell growth and viability. Erk1/2 induction had the opposite effects. Taken together, these results are supportive of an active crosstalk between the oncogenic Erk1/2 and STAT3 pathways in OSCC, the significance of which requires further investigation.

Abstract. Mitogen-activated protein kinases (MAPKs) are a family of protein kinases that link extracellular stimuli with intracellular responses and participate in numerous cellular processes such as growth, proliferation, differentiation, inflammation and apoptosis. Persistent activation of signal transducer and activator of transcription 3 (STAT3), which is accompanied by increases in STAT3 tyrosine phosphorylation, is associated with cell proliferation, differentiation and apoptosis in oral squamous cell carcinoma (OSCC). The role and significance of the activation of MAPKs, particularly of c-Jun N-terminal kinase (JNK), on STAT3 signaling in OSCC have not been thoroughly investigated. The present study examines the effects of JNK1/2 modulation on STAT3 signaling and cellular activities in OSCC cells. The expression levels of STAT3 [total, tyrosine phosphorylated (p-Tyr) and serine phosphorylated (p-Ser)], JNK, c-Jun and cyclin D1 were assessed in the OSCC cell lines SCC25 and SCC9. Inhibition of JNK1/2 was achieved by pharmacological agents (SP600125) and by small interfering RNA (siRNA) silencing, while JNK1/2 was induced by active MAPK kinase 7. Cell proliferation and viability rates were also evaluated. Inhibition of JNK1/2 with either SP600125 treatment or specific siRNA silencing resulted in decreased levels of p-Ser STAT3 and increased levels of p-Tyr STAT3 and cyclin D1 in both cell lines. Furthermore, JNK1/2 inhibition resulted in a dose-dependent increase in cell growth and viability in both cell lines. Opposite results were observed with JNK1/2 induction in both cell lines. The present results are supportive of a potential tumor suppressive role of JNK1/2 signaling in OSCC, which may be mediated through negative crosstalk with the oncogenic STAT3 signaling pathway. The possible therapeutic implications of JNK1/2 inhibition for patients with OSCC require to be investigated.

The present study aims to investigate the expression levels of two critical mammalian target of rapamycin (mTOR) downstream effectors, 4E binding protein 1 (4EBP1) and eukaryotic initiation factor 4E (eIF4E) proteins, in precancerous squamous intraepithelial lesions and cancer of the uterine cervix, and their association with human papilloma virus (HPV) infection status. Uterine cervical biopsies from 73 patients were obtained, including 40 fresh-frozen samples and 42 archival formalin-fixed, paraffin-embedded tissue specimens. Whole protein extracts were analyzed for the expression of 4EBP1 and eIF4E proteins using western blotting. In addition, distribution of 4EBP1 and eIF4E protein expression and 4EBP1 phosphorylation (P-4EBP1) were analyzed by immunohistochemistry in archival tissues and correlated with the degree of dysplasia. The presence of high-risk HPV (HR-HPV) types was assessed by polymerase chain reaction. Using western blot analysis, high expression levels of 4EBP1 and eIF4E were observed in all uterine cervical carcinomas, which significantly correlated with the degree of dysplasia. By immunohistochemistry, overexpression of 4EBP1 and eIF4E was detected in 20 of 21 (95%) and 17 of 21 (81%) samples, respectively, in patients with high-grade dysplasia and carcinomas, compared with 1 of 20 (5%) and 2 of 20 (10%) samples, respectively, in patients with low-grade lesions or normal histology. All 4EBP1-positive cases tested were also positive for P-4EBP1. Furthermore, overexpression of 4EBP1 and eIF4E significantly correlated with the presence of HR-HPV oncogenic types. The present study demonstrated that critical effectors of mTOR signaling, which control protein synthesis initiation, are overexpressed in cervical high-grade dysplasia and cancer, and their levels correlate with oncogenic HPV types. These findings may provide novel targets for investigational therapeutic approaches in patients with cancer of the uterine cervix.

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