ABSTRACT
Cancer drug resistance and poor selectivity towards cancer cells demand the constant search for new therapeutics. PI3K-Akt-mTOR and RAS-MAPK-ERK signaling pathways are key mechanisms involved in cell survival, proliferation, differentiation, and metabolism and their deregulation in cancer can promote development of therapy resistance. We investigated the effects of targeted inhibitors (wortmannin, GSK690693, AZD2014 and tipifarnib) towards these two pathways on early zebrafish and sea urchin development to assess their toxicity in normal, fast proliferating cells. PI3K inhibitor wortmannin and RAS inhibitor tipifarnib displayed highest toxicity while GSK690693, a pan-Akt kinase inhibitor, exhibited a less significant impact on embryo survival and development. Moreover, inhibition of the upstream part of the PI3K-Akt-mTOR pathway (wortmannin/GSK690693 co-treatment) produced a synergistic effect and impacted zebrafish embryo survival and development at much lower concentrations. Dual mTORC1/mTORC2 inhibitor AZD2014 showed no considerable effects on embryonic cells of zebrafish in concentrations substantially toxic in cancer cells. AZD2014 also caused the least prominent effects on sea urchin embryo development compared to other inhibitors. Significant toxicity of AZD2014 in human cancer cells, its capacity to sensitize resistant cancers, lower antiproliferative activity against human normal cell lines and fast proliferating embryonic cells could make this agent a promising candidate for anticancer therapy.
Subject(s)
Antineoplastic Agents/toxicity , Cell Proliferation/drug effects , Enzyme Inhibitors/toxicity , Molecular Targeted Therapy/adverse effects , Signal Transduction/drug effects , Abnormalities, Drug-Induced/enzymology , Abnormalities, Drug-Induced/etiology , Abnormalities, Drug-Induced/pathology , Animals , Arbacia/embryology , Benzamides , Dose-Response Relationship, Drug , Embryonic Development/drug effects , Morpholines/toxicity , Oxadiazoles/toxicity , Pyrimidines , Quinolones/toxicity , Wortmannin/toxicity , Zebrafish/embryologyABSTRACT
OBJECTIVES: Micro RNAs (miRNAs) have a major role in human cancerogenesis.The current study investigated the prognostic significance of miR-183 and miR-21 expression in tongue carcinoma patients. MATERIAL AND METHOD: For qPCR of miR-183 and miR-21 expression, total RNA isolated from 60 fresh-frozen tissue of tongue carcinomas was converted into cDNA by TaqMan MicroRNA Reverse Transcription Kit and quantified by TaqMan MicroRNAs Expression Assays. Fold changes in the miRNAs expression, normalized to RNU6B, were determined using 2-ΔΔCt method, and dichotomized into high and low according to cut-off values derived from ROC curve analysis. RESULTS: miR-183 emerged as promising discriminatory biomarker of poor outcome. Tissue over-expression of miR-183, observed in 68.3% of tongue carcinomas, was associated with clinical stage (p = 0.037), tumor size (p = 0.036), and high alcohol intake (p = 0.034).The patients with miR-183 over-expression had significantly shorter overall survival (p = 0.006) and a 5.666 times higher risk of poor outcome (p = 0.005), while miR-21 over-expression carried a tendency towards poorer survival (p = 0.073). However, multivariate analysis revealed that the recurrences were independent adverse prognostic factors, while miR-183 over-expression lost its significance. CONCLUSION: Our results suggests that over-expression of miR-183 in tumor tissue could be a potential marker of clinical stage and a poor survival of tongue carcinoma patients and may be associated with high alcohol consumption. CLINICAL RELEVANCE: Oncogenic miRNAs, such as the investigated miR-183 and miR-21, could be novel prognostic biomarkers of tumor progression and adverse clinical outcome in oral cancer, as well as novel therapeutic targets in cancer.
Subject(s)
MicroRNAs/analysis , Tongue Neoplasms/genetics , Adult , Aged , Aged, 80 and over , Biomarkers, Tumor/analysis , Disease Progression , Female , Humans , Male , Middle Aged , Neoplasm Staging , Polymerase Chain Reaction , Prognosis , RNA-Directed DNA Polymerase , Survival Rate , Tongue Neoplasms/pathologyABSTRACT
As a key component of the transforming growth factor beta (TGFB) pathway, which regulates the expression of thyroid-specific genes, tumor suppressor SMAD4 is crucial for thyroid development and function. Aberrant expression of SMAD4 in thyroid tumor tissue was reported and mutations affecting the coding region have been detected, but a potential role of mutations in SMAD4 gene regulatory regions remains unexplored. The aim of this study was to analyze SMAD4 gene promoters in thyroid tumors. A total of 76 thyroidectomy specimens were studied, including 42 malignant and 34 benign tumors. The presence of mutations in four SMAD4 gene promoters was analyzed in thyroid tumor tissue and peripheral blood by PCR and DNA sequencing. The expression and intracellular localization of endogenous SMAD4 protein in selected tumor samples was studied by immunostaining and confocal microscopy. Of three novel variants detected, two were within promoter A (-204T/C and -5C/T) and one in promoter D (-180delA). Unlike somatic mutations previously detected in the nearby region, germline mutation -180delA in promoter D doesn't appear to affect SMAD4 expression in the thyroid tumor tissue. However, all newly detected SMAD4 promoter variants affect predicted binding sites of transcription factors involved in cell cycle regulation and should be further characterized functionally. Although not directly involved in carcinogenesis, detected variants may alter SMAD4 transcriptional regulation to some extent. Considering that dosage dependence is of great importance for the role of SMAD4 protein as a tumor suppressor, potential clinical significance of SMAD4 gene promoter mutations is worth further investigation.
