Assessment of MicroRNA (miR)-365 Effects on Oral Squamous Carcinoma Cell Line Phenotypes

miR-365 is a microRNA that regulates transcription and has been demonstrated to promote oncogenesis and metastasis in some cancers while suppressing these effects in others. Virtually no information is known about the presence or function of miR-365 in oral cancers. Based upon this information, the primary goal of this project was to evaluate the expression of miR-365 in existing oral cancer cell lines. Five commercially available oral cancer cell lines (SCC4, SCC9, SCC15, SCC25, and CAL27) were obtained and cultured. RNA was then screened by PCR using primers specific for miR-365, as well as matrix metalloproteinase (MMP-2) and a downstream cancer stem cell regulator (NKX2.1), and structural and metabolic standards (beta actin, GAPDH). miR-365 was detected among these oral cancers, and some cells also expressed NKX2.1 and MMP-2, which correlated with miR-365 levels. The relative expression of miR-365, NKX2.1, and MMP-2 RNA was higher than expected. Transfection of miR-365 resulted in a significant increase in proliferation, which was not observed in the negative controls. These data appear to confirm miR-365 expression in oral cancers, which may also be correlated with MMP-2 and NKX2.1 expression. Moreover, the fastest growing oral cancers with the highest viability produced the most miR-365. In addition, miR-365 transfected into cells significantly increased growth, even in normal cells. More research is needed to elucidate the pathways responsible for these observations.

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miR-365 (microRNA): Potential Biomarker in Oral Squamous Cell Carcinoma Exosomes and Extracellular Vesicles

International Journal of Molecular Sciences ◽

10.3390/ijms21155317 ◽

2020 ◽

Vol 21 (15) ◽

pp. 5317

Author(s):

Jeffery Coon ◽

Karl Kingsley ◽

Katherine M. Howard

Keyword(s):

Oral Cancer ◽

Cell Line ◽

Cell Lines ◽

Cancer Cell ◽

Extracellular Vesicles ◽

Squamous Cell ◽

Cancer Cell Lines ◽

Oral Cancers ◽

Oral Cancer Cell ◽

Potential Biomarker

Introduction: miR-365 is a non-coding microRNA that regulates transcription and has been demonstrated to promote oncogenesis and metastasis in some cancers, while suppressing these effects in others. Many microRNAs are produced and then exported extracellularly in exosomes, which are small extracellular vesicles ranging from 30 to 100 nm that are found in eukaryotic fluids and facilitate many cellular functions. Exosomes and extracellular vesicles are produced by many cell types, including oral cancer cells—although no study to date has evaluated miR-365 and oral cancer exosomes or extracellular vesicles. Based on this information, our research question was to evaluate whether oral cancers produce exosomes or extracellular vesicles containing miR-365. Materials and Methods: Two commercially available oral cancer cell lines (SCC25 and CAL27) and a normal oral keratinocyte (OKF4) were grown in serum-free media, supplemented with exosome-depleted fetal bovine serum. Extracellular vesicles and exosomes were then isolated using the Invitrogen total exosome RNA and protein isolation kit for processing using the hsa-miR-365a-5p microRNA qPCR assay kit. Results: RNA was successfully isolated from the exosome-depleted supernatant from each cell line—SCC9, SCC15, SCC25, and CAL27 (oral squamous cell carcinomas) and OKF4 (oral epithelial cell line). Relative concentrations of RNA were similar among each cell line, which were not significantly different, p = 0.233. RNA quality was established by A260:A280 absorbance using a NanoDrop, revealing purity ranging 1.73–1.86. Expression of miR-16 was used to confirm the presence of microRNA from the extracted exosomes and extracellular vesicles. The presence of miR-365 was then confirmed and normalized to miR-16 expression, which demonstrated an increased level of miR-365 in both CAL27 and SCC25. In addition, the normalized relative quantity (RQ) for miR-365 exhibited greater variation among SCC25 (1.382–4.363) than CAL27 cells (1.248–1.536). Conclusions: These results confirm that miR-365 is not only expressed in oral cancer cell lines, but also is subsequently exported into exosomes and extracellular vesicles derived from these cultures. These data may help to contextualize the potential for this microRNA to contribute to the phenotypes and behaviors of oral cancers that express this microRNA. Future research will begin to investigate these potential mechanisms and pathways and to determine if miR-365 may be useful as an oral cancer biomarker for salivary or liquid biopsies.

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Characterization of URST1 as a prognostic biomarker and therapeutic target for lung and oral cancers.

Journal of Clinical Oncology ◽

10.1200/jco.2017.35.15_suppl.e14057 ◽

2017 ◽

Vol 35 (15_suppl) ◽

pp. e14057-e14057

Author(s):

Atsushi Takano ◽

Yoshihiro Yoshitake ◽

Masanori Shinohara ◽

Yohei Miyagi ◽

Yataro Daigo ◽

...

Keyword(s):

Oral Cancer ◽

Cell Lines ◽

Cancer Cell ◽

Therapeutic Target ◽

Prognostic Biomarker ◽

Cancer Cell Lines ◽

Normal Lung ◽

Positive Staining ◽

Oral Cancers ◽

Oral Cancer Cell

e14057 Background:Lung cancer and oral cavity cancer belong to aerodigestive cancers whose clinical outcome after multimodal therapy remains poor. Methods:To identify new biomarkers and therapeutic targets for lung and oral cancers, we screened genes using our original gene expression profile database for various solid cancers, and characterized clinical and oncogenic values of up-regulated in solid tumor 1 (URST1) protein as a candidate. Results: URST1 was highly expressed in the most of lung or oral cancer cell lines and tumor tissues, but was hardly detected in normal lung or oral epithelial cells and tissues as detected by real-time PCR and western blot analyses. Immunocytochemical analysis revealed that URST1 was mainly localized in the nucleus and cytoplasm of lung or oral cancer cell lines. Immunohistochemical analysis using tissue microarray showed that positive staining of URST1 was observed in 231 of 358 (64.5%) lung cancers, but not in normal lung, and it was significantly associated with poor prognosis after curative surgery. In addition, URST1 was expressed in 64 of 96 (66.7%) oral cancers, but not in oral mucosa, and it was significantly correlated with poor clinical outocome after surgery. Multivariate analysis confirmed that URST1 expression was an independent prognostic factor for oral cancer. Suppression of URST1 expression by siRNA or treatment with synthesized inhibitor specific for URST1 activity inhibited mitosis and growth of lung or oral cancer cell lines. Conclusions:URST1 is likely to be a prognostic biomarker and therapeutic target for aerodigestive cancers such as lung and oral cancers.

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PRELIMINARY PHYTOCHEMICAL ANALYSIS AND CYTOTOXICITY POTENTIAL OF PINEAPPLE EXTRACT ON ORAL CANCER CELL LINES

Asian Journal of Pharmaceutical and Clinical Research ◽

10.22159/ajpcr.2016.v9s2.13313 ◽

2016 ◽

pp. 140

Author(s):

Anirudh Menon ◽

Vishnu Priya V ◽

Gayathri R

Keyword(s):

Oral Cancer ◽

Cell Lines ◽

Cancer Cell ◽

Anticancer Drug ◽

Cancer Cell Lines ◽

Phytochemical Analysis ◽

Phytochemical Composition ◽

Tetrazolium Bromide ◽

Oral Cancer Cell ◽

Natural Way

ABSTRACT Objective: This study aims at performing a preliminary phytochemical analysis to evaluate the phytochemical composition of pineapple extract and its cytotoxicity potential on oral cancer cell lines. Methods: Preliminary phytochemical analysis of pineapple extract was done, 3-(4, 5-Dimethyl-2-thiazolyl)-2, 5-diphenyl-2H-tetrazolium bromide assay for evaluating the cytotoxicity potential of the extract on oral cancer cell lines was performed. Results: Phytoconstituents such as flavonoids, coumarins, and phenols were present in the pineapple extract. The extract also exhibited increased cytotoxicity with increased concentration. Conclusion: This study is conducted to see if pineapple extract is effective in treating oral cancer in a natural way instead of harmful treatments. Keywords: Cytotoxicity, Pineapple extract, Anticancer drug.

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