Proliferation Of OSCC Cells Research Articles

Brucea javanica oil inhibited the proliferation, migration, and invasion of oral squamous carcinoma by regulated the MTFR2 pathway

IntroductionOral squamous cell carcinoma (OSCC) is one of the most common malignant tumors in oral and maxillofacial region. The development of new chemotherapy agents and new drug combinations may improve patient survival and quality of life, but both surgery and radiotherapy have significant functional side effects and drug resistance, ultimately resulting in a 5-year survival rate of no more than 60% for OSCC patients. Studies have shown that Brucea javanica oil (BJO) extracts have anti-cancer effects against a variety of cancers, but little research has been reported on OSCC. MethodsCCK8, Colony formation, Scratch test and Transwell invasion assays were applied to determine the effects of BJO on the proliferation, migration, and invasion ability of OSCC cells in vitro. MTFR2 knockdown (shRNA) and overexpression (cDNA) OSCC cells were constructed to evaluate the effect of MTFR2 on the proliferation and invasion of OSCC cells. The nude mouse model of subcutaneous xenograft tumor was used to evaluate the effect of BJO on OSCC cells in vivo. PCR, western blot and immunohistochemistry were used to verify the expression of MTFR2, glycolysis markers and related pathway molecules after BJO treatment.ResultsIn vivo experiments using nude mice with xenografted OSCC cells and in vitro experiments with OSCC cell lines demonstrated that BJO treatment significantly inhibited the proliferation, migration, and invasiveness of OSCC cells. WB and PCR proved that BJO could effectively reduce the expression levels of MTFR2 and SOD2/H2O2 related signal transduction pathways. At the same time, the expression of oxidative phosphorylation markers increased, the expression of glycolytic markers decreased, and glycolysis-mediated decomposition of reactive oxygen species decreased, and H2O2 and oxygen levels decreased.In addition, when MTFR2 expression increased or decreased, SOD2/H2O2 expression also increased or decreased.DiscussionIn this study, we concluded through in vitro and in vivo experiments that BJO may affect the SOD2/H2O2 signaling pathway by down-regulating MTFR2-mediated aerobic glycolysis, thereby inhibiting cell proliferation, Migration, and Invasion. The elucidation of this mechanism helps us to understand the molecular mechanism ofinhibiting OSCC invasion and metastasis by BJO, which has important clinical value or improving the survival rate of OSCC patients.

The mechanism of arsenic trioxide and microwave ablation in the treatment of oral squamous cell carcinoma based on high throughput sequencing

AbstractOral squamous cell carcinoma (OSCC) is a common head and neck malignant tumour with high incidence and poor prognosis. Arsenic trioxide (ATO) has therapeutic effects on solid tumours. Microwave ablation (MWA) has unique advantages in the treatment of solid tumours. However, the therapeutic mechanism of ATO and MWA, as well as their combined effect on OSCC were largely unelucidated. Cal‐27 cell‐bearing nude mice were treated with ATO and/or MWA, respectively. RNA sequencing was used to obtain gene expression profiles in tumour tissues of mice treated by ATO or MWA. RNA sequencing results were verified by real‐time polymerase chain reaction (PCR). The lncRNA‐miRNA‐mRNA co‐expression network was constructed based on the competitive endogenous RNA (ceRNA) theory. Gene ontology and Kyoto Encyclopedia of Genes and Genomes analyses were performed using differentially expressed genes. The combined effect of ATO and MWA on OSCC was evaluated. Finally, CCK‐8 assay, EdU assay and transwell migration assay were performed to detect the effect of HSPA6 on the proliferation and migration of OSCC cells. The reduced volume of tumour tissues was observed in both ATO‐ and MWA‐treated groups. 37.8% decreased in the ATO group and 35.0% in the MWA group. A total of 207 and 539 differentially expressed mRNAs and lncRNAs were identified in the ATO group. And a total of 200 and 522 differentially expressed mRNAs and lncRNAs in the MWA group were identified. The expression levels of 8 genes were verified by real‐time PCR. The differentially expressed genes were closely related to “chemical carcinogenesis”, “herpes simplex infection”, “porphyrin and chlorophyll metabolism”, and “MAPK signalling pathway”. The lncRNA‐miRNA‐mRNA co‐expression networks were constructed. The combined treatment with ATO and MWA showed a better inhibitive effect on OSCC than either of them. The synergistic effect of ATO and MWA was related to the upregulation of HSPA6. The downregulation of HSPA6 could promote the proliferation and migration of OSCC cells. This study detected the long non‐coding RNA and mRNA expression profiles related to the treatment of OSCC and constructed corresponding ceRNA networks. Arsenic trioxide and MWA have a synergistic effect on OSCC, which was related to the upregulation of HSPA6.

Hedgehog/Gli2 signaling triggers cell proliferation and metastasis via EMT and wnt/β-catenin pathways in oral squamous cell carcinoma

BackgroundOral squamous cell carcinoma (OSCC) is the most lethal oral malignant tumor, however, clinical outcomes remain unsatisfactory. The Hedgehog/Gli2 pathway plays a pivotal role in tumor progression, yet the regulatory mechanism governing its involvement in the malignant evolution process of OSCC remains elusive. MethodsOSCC animal tissue samples were used to detect the activation of the Hedgehog/Gli2 pathway in OSCC. Based on the clinical information of oral cancer patients in TCGA database, the role of this pathway in patients was analyzed, and the activation status of this pathway was verified in human OSCC cells. After activating or inhibiting the Hedgehog pathway, the effects of this pathway on the biological function of OSCC cells and its regulatory mechanism were examined. Interfering the expression of Gli2, a key transcription factor in this pathway, revealed the role of Hedgehog/Gli2 pathway in the malignant evolution of OSCC cells. ResultsThe Hedgehog pathway exhibits abnormal activation in animal models of OSCC. Clinical data from TCGA demonstrate a significant enrichment of the Hedgehog pathway in patients with OSCC, and Gli2, a key downstream factor of this pathway, is closely associated with the occurrence and progression of OSCC. Cellular studies have revealed aberrant activation of this pathway in human OSCC cells, which exerts its function by modulating the activation of epithelial-mesenchymal transition (EMT) and Wnt/β-catenin pathways. Subsequent investigations further confirm the pivotal involvement of Gli2 in the Hedgehog pathway activation, underscoring its potential as a therapeutic target for inhibiting malignant proliferation and metastasis of OSCC cells through modulation of EMT and Wnt/β-catenin pathways. ConclusionThe Hedgehog/Gli2 pathway induces EMT and activates Wnt/β-catenin pathway to trigger the malignant proliferation and metastasis of OSCC cells, and Gli2 plays a key role in this process, which suggests that targeting Gli2 may be a promising therapeutic strategy for inhibiting the proliferation and metastasis of OSCC.

JARID2, a novel regulatory factor, promotes cell proliferation, migration, and invasion in oral squamous cell carcinoma

BackgroundAccurate regulation of gene expression is crucial for normal development and function of cells. The prognostic significance and potential carcinogenic mechanisms of the related gene JARID2 in OSCC are not yet clear, but existing research has indicated a significant association between the two.Methods and materialsThe relationship between the expression of the JARID2 gene in tumor samples of OSCC patients and clinical pathological factors was analyzed using immunohistochemistry experiments and RT-qPCR analysis. Based on the clinical pathological data of patients, bioinformatics analysis was conducted using public databases to determine the function of JARID2 in OSCC. Knockdown OSCC cell lines were constructed, and the impact of JARID2 on the biological behavior of OSCC cell lines was assessed through CCK-8, wound healing assay, and transwell analysis.ResultsImmunohistochemistry experiments confirmed the correlation between JARID2 and the prognosis of OSCC patients, while RT-qPCR experiments demonstrated its expression levels in tissue and cells. CKK-8 experiments, wound healing assays, and Transwell experiments indicated that knocking down JARID2 had a negative impact on the proliferation, invasion, and migration of OSCC cells. Bioinformatics analysis results showed that the expression of JARID2 in OSCC is closely associated with patient gene co-expression, gene function enrichment, immune infiltration, and drug sensitivity.ConclusionOur study indicates that JARID2 is a novel prognostic biomarker and potential therapeutic target for OSCC.

AKR1B10 accelerates glycolysis through binding HK2 to promote the malignant progression of oral squamous cell carcinoma

BackgroundOral squamous cell carcinoma (OSCC) remains a rampant oral cavity neoplasm with high degree of aggressiveness. Aldo–keto reductase 1B10 (AKR1B10) that is an oxidoreductase dependent on nicotinamide adenine dinucleotide phosphate (NADPH) has been introduced to possess prognostic potential in OSCC. The present work was focused on specifying the involvement of AKR1B10 in the process of OSCC and its latent functional mechanism.MethodsAKR1B10 expression in OSCC tissues and cells were detected by RT-qPCR and Western blot analysis. CCK-8 method, EdU staining, wound healing and transwell assays respectively assayed cell viability, proliferation, migration and invasion. Immunofluorescence staining and Western blot evaluated epithelial mesenchymal transition (EMT). Adenosine triphosphate (ATP) contents, glucose consumption and extracellular acidification rate (ECAR) were measured by relevant commercially available kits and Seahorse XF96 Glycolysis Analyzer, severally. The expressions of proteins associated with metastasis and glycolysis were examined with Western blot. Co-IP assay confirmed the binding between AKR1B10 and hexokinase 2 (HK2).ResultsIt was observed that AKR1B10 expression was increased in OSCC tissues and cells. After AKR1B10 was knocked down, the proliferation, migration, invasion and EMT of OSCC cells were all hampered. Additionally, AKR1B10 silencing suppressed glycolysis and bound to HK2 in OSCC cells. Up-regulation of HK2 partially abolished the hampered glycolysis, proliferation, migration, invasion and EMT of AKR1B10-silenced OSCC cells.ConclusionTo sum up, AKR1B10 could bind to HK2 to accelerate glycolysis, thereby facilitating the proliferation, migration, invasion and EMT of OSCC cells.

Inhibiting Effect of Cationic Procyanidin Nanoparticles on Drug-Resistant Oral Squamous Cell Carcinoma Cell Lines

Background: Multidrug resistance (MDR) is one of the primary causes of tumor chemotherapy failure. Therefore, it is essential to uncover new drug treatment protocols. In the current study, employing chitosan (CS) and sodium tripolyphosphate (TPP) as carriers and cross-linking agents, the proanthocyanidins (PHL)/poly (lactic-co-glycolic acid) (PLGA) were encapsulated by emulsion solvent evaporation, through which the cationic CS-PLGA-PHL nanosystem was obtained. The effectiveness of CS-PLGAPHL on the invasion and migration of human oral squamous cell carcinoma cells was discussed, as were their potential mechanisms. Materials and Methods: A CS-PLGA-PHL nanosystem was constructed by emulsion-solvent evaporation. The size distribution, dispersion, and morphology were characterised by the laser particle size analyser and transmission electron microscope. Human oral squamous cell carcinoma drug-resistant cell lines SCC131/R and SCC-15/DDP were cultured in vitro. The two cell types were induced at different concentrations by CS-PLGA-PHL, after which a CCK⁃8 experiment was performed to determine the effect of CS-PLGA-PHL on the proliferation of the two cell lines. The 50% inhibitory concentration (IC50) of CSPLGA- PHL was calculated at different time points. The transwell chamber experiment was performed to identify the effects of CS-PLGA-PHL on the migration and invasion of OSCC cells. The expression levels of MMP-2 and MMP-9 were detected by Western blot and q-PCR. Results: CS-PLGA-PHL is well dispersed. The PDI appeared to be lowest when the mass ratio of chitosan to PLGA equaled 1:15. CS-PLGA-PHL exhibited a marked effect in inhibiting the proliferation of SCC- 131/R and SCC-15/DDP as well as the invasion and migration. CS-PLGA-PHL was able to downregulate the expression of MMP-2 and MMP-9 genes and proteins significantly in drug-resistant cell lines. Conclusion: CS-PLGA-PHL for oral squamous cell carcinoma has been successfully prepared to exert significant inhibition on the proliferation, invasion, and migration of OSCC cells. The mechanism involved was possibly related to the down-regulation of MMP-2 and MMP-9 expression by CS-PLGA-PHL.

Molecular mechanisms underlying the epigallocatechin-3-gallate-mediated inhibition of oral squamous cell carcinogenesis

ObjectivesTo reveal the mechanisms underlying the epigallocatechin-3-gallate (EGCG)-mediated inhibition of carcinogenesis and the related regulatory signaling pathways. DesignThe effect of EGCG on the proliferation of OSCC cells was examined. SuperPred, ChEMBL, Swiss TargetPrediction, DisGeNET, GeneCards, and National Center for Biotechnology Information databases were used to predict the EGCG target genes and oral leukoplakia (OL)-related, oral submucosal fibrosis (OSF)-related, and OSCC-related genes. The binding of EGCG to the target proteins was simulated using AutoDock and PyMOL. The Cancer Genome Atlas (TCGA) dataset was subjected to consensus clustering analysis to predict the downstream molecules associated with these targets, as well as their potential functions and pathways. ResultsEGCG significantly inhibited OSCC cell proliferation (p < 0.001). By comparing EGCG target genes with genes linked to oral potentially malignant disorder (OPMD) and OSCC, a total of eleven potential EGCG target genes were identified. Furthermore, EGCG has the capacity to bind to eleven proteins. Based on consensus clustering and enrichment analysis, it is suggested that EGCG may hinder the progression of cancer by altering the cell cycle and invasive properties in precancerous lesions of the oral cavity. Some possible strategies for modifying the cell cycle and invasive properties may include EGCG-mediated suppression of specific genes and proteins, which are associated with cancer development. ConclusionsThis study investigated the molecular mechanisms and signaling pathways associated with the EGCG-induced suppression of OSCC. The identification of specific pharmacological targets of EGCG during carcinogenesis is crucial for the development of innovative combination therapies involving EGCG.

PDZ-binding kinase inhibitor OTS514 suppresses the proliferation of oral squamous carcinoma cells.

PDZ-binding kinase (PBK) has been reported as a poor prognostic factor and is a promising molecular target for anticancer therapeutics. Here, we aimed to investigate the effect of specific PBK inhibitor OTS514 on the survival of OSCC cells. Four OSCC cell lines (HSC-2, HSC-3, SAS, and OSC-19) were used to examine the effect of OTS514 on cell survival and apoptosis. DNA microarray analysis was conducted to investigate the effect of OTS514 on gene expression in OSCC cells. Gene set enrichment analysis was performed to identify molecular signatures related to the antiproliferative effect of OTS514. OTS514 decreased the cell survival of OSCC cells dose-dependently, and administration of OTS514 readily suppressed the HSC-2-derived tumor growth in immunodeficient mice. Treatment with OTS514 significantly increased the number of apoptotic cells and caspase-3/7 activity. Importantly, OTS514 suppressed the expression of E2F target genes with a marked decrease in protein levels of E2F1, a transcriptional factor. Moreover, TP53 knockdown attenuated OTS514-induced apoptosis. OTS514 suppressed the proliferation of OSCC cells by downregulating the expression of E2F target genes and induced apoptosis by mediating the p53 signaling pathway. These results highlight the clinical application of PBK inhibitors in the development of molecular-targeted therapeutics against OSCC.

Butyrate promotes oral squamous cell carcinoma cells migration, invasion and epithelial-mesenchymal transition.

BackgroundOral squamous cell carcinoma (OSCC), the most common type of primary malignant tumor in the oral cavity, is a lethal disease with high recurrence and mortality rates. Butyrate, a metabolite produced by periodontal pathogens, has been linked to oral diseases. The purpose of this study was to evaluate the effect of sodium butyrate (NaB) on the proliferation, migration, and invasion of OSCC cells in vitro and to explore the potential mechanism.MethodsTwo OSCC cell lines (HSC-4 and SCC-9) were treated with NaB at different concentrations. The cell proliferation was assayed by CCK-8, ethylene deoxyuridine (EdU), and flow cytometry. Wound healing and transwell assay were performed to detect cell migration and invasion. Changes in epithelial-mesenchymal transition (EMT) markers, including E-cadherin, Vimentin, and SNAI1, were evaluated by quantitative real-time PCR (qRT-PCR), western blot, and immunofluorescent staining. The expression levels of matrix metalloproteinases (MMPs) were analyzed by qRT-PCR and gelatin zymography.ResultsOur results showed that NaB inhibited the proliferation of OSCC cells and induced cell cycle arrest at G1 phase, but NaB significantly enhanced cell migration and invasion compared with the control group. Further mechanistic investigation demonstrated that NaB induced EMT by increasing the expression of Vimentin and SNAI1, decreasing the expression of membrane-bound E-cadherin, and correspondingly promoting E-cadherin translocation from the membrane to the cytoplasm. In addition, the overexpression of MMP1/2/9/13 was closely related to NaB treatment.ConclusionsOur study conclude that butyrate may promote the migration and invasion of OSCC cells by inducing EMT. These findings indicate that butyrate may contribute to OSCC metastasis.

Cytotoxic and Antioxidant Effects of Phoenix dactylifera L. (Ajwa Date Extract) on Oral Squamous Cell Carcinoma Cell Line.

Aim The aim of the current study is to investigate the antioxidant and apoptotic potential of Ajwa date flesh (ADF) and Ajwa date pit (ADP) extract on human squamous cell carcinoma cell line (HSC-2). Method ADF and ADP were extracted with a solvent extraction method using hexane, acetone, and ethanol, which were then subjected to antioxidant assay by 2,2-diphenyl-1-picrylhydrazyl (DPPH). HSC-2 cells were then treated with different concentrations of ADF and ADP extract for 24, 48, and 72 hours. MTT assay was performed to assess the antiproliferative effect, and Annexin V-FITC was used for the detection of cellular apoptosis. Results Acetone extracts of ADF and ADP had the highest radical scavenging and antioxidant activities followed by the ethanolic extracts, whereas ADP appeared to have significantly higher antioxidant effects than ADF. MTT assay demonstrated that acetone extracts of ADF and ADP were significantly cytotoxic against HSC-2 cells in a dose- and time-dependent manner. The half inhibitory concentration (IC50) of ADF was found to be 8.69 mg/ml at 24 h, and the maximum cell growth inhibition was observed at 50 mg/ml. The IC50 for the ADP was found to be 0.97 mg/ml at 24 h, and the maximum cell growth inhibition was observed at 5 mg/ml. Statistical analysis of the flow cytometry assay showed that the treatment with ADF and ADP extracts had a significant apoptotic effect which occurred in a dose-dependent manner. HSC-2 cells were seen in the late apoptotic stage with higher doses of ADF and ADP extract. ADP extract demonstrated higher apoptotic activity than ADF extract. In addition, combined treatment of ADF and ADP was also performed on HSC-2 cells which demonstrated higher apoptotic activity when compared to the single extract. Conclusion Ajwa date fruit has a promising cytotoxic effect by inhibiting the growth and proliferation of OSCC cells and inducing cell death by apoptosis.

Adipose-derived stem cells promote the proliferation, migration, and invasion of oral squamous cell carcinoma cells by activating the Wnt/planar cell polarity signaling pathway.

BackgroundOral squamous cell carcinoma (OSCC) is the most common malignancy of the oral and maxillofacial region. Adipose-derived stem cells (ADSCs) interact with a variety of malignant tumors to promote their proliferation and metastasis. Abnormalities in Wnt/planar cell polarity (PCP) signaling and overactivation of the signaling pathway are considered to be related to the occurrence and development of various malignant tumors. In order to determine whether ADSC can promote tumorigenesis in OSCC and its molecular mechanism, we conducted a series of studies.MethodsThe effect of ADSCs on the occurrence and development of OSCC was studied in vivo and in vitro, and the molecular mechanism was investigated using Western blot and immunofluorescence (IHC) assays.ResultsThe results revealed that ADSCs could promote the proliferation, invasion, and migration of OSCC cells in a dose- and time-dependent manner. With regard to the mechanism, the expression of collagen triple helix repeat-containing protein 1 (CTHRC1) and phospho-c-Jun (p-c-Jun) increased significantly with enhancement of the interaction between ADSCs and OSCC cells, indicating that the Wnt/PCP signaling pathway was overactivated.ConclusionsADSCs promote the pathogenesis of OSCC by activating the Wnt/PCP signaling pathway, suggesting that proteins related to this pathway may be potential therapeutic targets for OSCC.

Upregulated circ_0002141 facilitates oral squamous cell carcinoma progression via the miR-1231/EGFR axis.

The dysregulation of circular RNAs (circRNAs) is implicated in the progression of various cancers. This study was aimed at expounding the role and mechanism of hsa_circ_0002141 in the OSCC progression. Circ_0002141 expression was examined in 52 pairs of OSCC tissues and matched adjacent tissue samples by quantitative real-time polymerase chain reaction (qRT-PCR) assay. After circ_0002141 was overexpressed or knocked down in OSCC cell lines, cell counting kit-8 (CCK-8) assay, Transwell assay, flow cytometry, and Western blotting were conducted to detect the changes in the growth, migration, invasion and apoptosis of OSCC cells. Western blot assay, qRT-PCR and dual-luciferase reporter assay were performed to clarify the interplay among circ_0002141, miR-1231, and epidermal growth factor receptor (EGFR). Circ_0002141 expression was significantly upregulated in OSCC tissues and cell lines. Circ_0002141 overexpression markedly promoted the proliferation, migration, and invasion of OSCC cells whereas reduced the apoptotic of OSCC cells. Also, circ_0002141knockdown suppressed the malignant characteristics of OSCC cells. EGFR was validated as the target of miR-1231. Besides, circ_0002141 could sponge miR-1231 and upregulate EGFR expression in OSCC cells. Circ_0002141/miR-1231/EGFR axis is involved in the progression of OSCC.

LncRNA HOTAIR promotes proliferation and migration of OSCC cells via targeting miR-126.

LncRNA HOTAIR promotes proliferation and migration of OSCC cells via targeting miR-126.

The Origin of Stroma Influences the Biological Characteristics of Oral Squamous Cell Carcinoma.

Simple SummaryNormal stromal cells play a significant role in the progression of cancers but are poorly investigated in oral squamous cell carcinoma (OSCC). In this study, we found that stromal cells derived from the gingival and periodontal ligament tissues could inhibit differentiation and promote the proliferation, invasion, and migration of OSCC both in vitro and in vivo. Furthermore, microarray data suggested that genes, such as CDK1, BUB1B, TOP2A, DLGAP5, BUB1, and CCNB2, probably play a role in influencing the different effects of gingival stromal tissue cells (G-SCs) and periodontal ligament stromal cells (P-SCs) on the progression of OSCC. Therefore, both G-SCs and P-SCs could promote the progression of OSCC, which could be a potential regulatory mechanism in the progression of OSCC.Normal stromal cells surrounding the tumor parenchyma, such as the extracellular matrix (ECM), normal fibroblasts, mesenchymal stromal cells, and osteoblasts, play a significant role in the progression of cancers. However, the role of gingival and periodontal ligament tissue-derived stromal cells in OSCC progression is unclear. In this study, the effect of G-SCs and P-SCs on the differentiation, proliferation, invasion, and migration of OSCC cells in vitro was examined by Giemsa staining, Immunofluorescence (IF), (3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium) (MTS), invasion, and migration assays. Furthermore, the effect of G-SCs and P-SCs on the differentiation, proliferation, and bone invasion by OSCC cells in vivo was examined by hematoxylin-eosin (HE) staining, immunohistochemistry (IHC), and tartrate-resistant acid phosphatase (TRAP) staining, respectively. Finally, microarray data and bioinformatics analyses identified potential genes that caused the different effects of G-SCs and P-SCs on OSCC progression. The results showed that both G-SCs and P-SCs inhibited the differentiation and promoted the proliferation, invasion, and migration of OSCC in vitro and in vivo. In addition, genes, including CDK1, BUB1B, TOP2A, DLGAP5, BUB1, and CCNB2, are probably involved in causing the different effects of G-SCs and P-SCs on OSCC progression. Therefore, as a potential regulatory mechanism, both G-SCs and P-SCs can promote OSCC progression.

SOX2-dependent expression of dihydroorotate dehydrogenase regulates oral squamous cell carcinoma cell proliferation

SummaryOral squamous cell carcinoma (OSCC) become a heavy burden of public health, with approximately 300 000 newly diagnosed cases and 145 000 deaths worldwide per year. Nucleotide metabolism fuel DNA replication and RNA synthesis, which is indispensable for cell proliferation. But how tumor cells orchestrate nucleotide metabolic enzymes to support their rapid growth is largely unknown. Here we show that expression of pyrimidine metabolic enzyme dihydroorotate dehydrogenase (DHODH) is upregulated in OSCC tissues, compared to non-cancerous adjacent tissues. Enhanced expression of DHODH is correlated with a shortened patient survival time. Inhibition of DHODH by either shRNA or selective inhibitors impairs proliferation of OSCC cells and growth of tumor xenograft. Further, loss of functional DHODH imped de novo pyrimidine synthesis, and disrupt mitochondrial respiration probably through destabilizing the MICOS complex. Mechanistic study shows that transcriptional factor SOX2 plays an important role in the upregulation of DHODH in OSCC. Our findings add to the knowledge of how cancer cells co-opt nucleotide metabolism to support their rapid growth, and thereby highlight DHODH as a potential prognostic and therapeutic target for OSCC treatment.

Abstract A29: Targeted disruption of PI3K/Akt/mTOR signaling pathway induces cell cycle arrest, apoptosis, autophagy, and inhibits inflammation, invasion, and angiogenesis of OSCC cells

Abstract Purpose: The phosphoinositide-3-kinase (PI3K) pathway is frequently altered in human cancer. This has led to the development and study of novel PI3K inhibitors for targeted therapy and also to overcome resistance to radiotherapy. Method: The antitumor effects of PI3K inhibitors (PI-828, PI-103 and PX-866) in terms of cell proliferation, colony formation, induction of apoptosis, cell cycle arrest, invasion, autophagy, and pNF- κB/p65 translocation in SCC-4, SCC-9 and SCC-25 cells were studied by performing MTT, clonogenic, DAPI staining, propidium iodide staining, annexin–V binding, Matrigel invasion, acridine orange staining and immunofluorescence assay. Western blot assay was performed to assess the alteration in the expression of various proteins. Result: PI-828 and PI-103 treatment exhibited dose-dependent inhibition of growth and proliferation of OSCC cells with a concomitant induction of apoptosis, altered cell cycle regulation and decreased invasiveness (p &amp;lt;0.01). PX-866 induced apoptosis, cell cycle arrest, autophagy and a significant decrease in the invasiveness of oral cancer cells as compared to untreated cells (p &amp;lt;0.01). These compounds significantly reduced expression of COX-2, Cyclin-D and VEGF in the treated cells besides cytoplasmic accumulation of pNF-κB/p65 protein. In addition to PI3Kα, inactivation of downstream components, i.e., Akt and mTOR was seen. Conclusion: PI3K inhibitors such as PI-103, PI-828 and PX-866 may be developed as potential chemotherapeutic agents for effective treatment of OSCC and other malignancies associated with activated PI3K/Akt pathway. Note: This abstract was not presented at the conference. Citation Format: Sadhna Aggarwal, Sarah John, Satya N. Das, Suresh C. Sharma. Targeted disruption of PI3K/Akt/mTOR signaling pathway induces cell cycle arrest, apoptosis, autophagy, and inhibits inflammation, invasion, and angiogenesis of OSCC cells [abstract]. In: Proceedings of the AACR Special Conference on Targeting PI3K/mTOR Signaling; 2018 Nov 30-Dec 8; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Res 2020;18(10_Suppl):Abstract nr A29.

Abstract 2634: Oral bacteria effects on oral cancer initiation and progression

Abstract Oral squamous cell carcinoma (OSCC) is the most common site of head and neck squamous cell carcinoma. It is estimated to be diagnosed in 50,000 patients this year with an overall 5-year survival rate of 64%. The tumor microenvironment has been shown to play a role in the progression of many human cancers, including OSCC. Given the complexity of the oral microbiome, we hypothesize that OSCC and oral bacteria interact to affect the tumorigenic properties of OSCC. Methods: OSCC cells were cultured in the presence of oral pathogenic (Aggregatibacter actinomycetemcomitans (Aa), Enterococcus faecalis (Ef), Fusobacterium nucleatum (Fn)) and non-pathogenic (Streptococcus parasanguinis (Sp)) bacteria. Human oral keratinocytes (HOK) and OSCC cells were treated with oral bacteria and assessed for changes in cell proliferation and gene expression. Results: HOK treated with Ef demonstrated significantly increased proliferation and increased expression of HES1, MMP11, and CDKN1A(p21). Of the five OSCC cell lines, Ef increased proliferation in the two cell lines that were human papilloma virus positive (HPV+) (2A3, SCC152) and in one of three cell lines that were HPV negative (HPV-, Cal27). Ef increased the expression of HES1, MMP2, CDKN1A, and GLI1 in SCC152 cell and HES1, MMP11, and CDKN1A in 2A3 cells. Ef also increased the invasion of SCC152 cells in a transwell experiment. In HPV- Cal27 cells, Ef increased MMP2 and GLI1 expression. Aa also increased proliferation in 2A3 HPV+ cells. Fn did not increase proliferation of OSCC cells, but it did increase the expression of HES1, CDKN1A, and GLI1 in SCC152 HPV+ cells. Conclusions: Oral bacteria have direct interactions with human oral keratinocytes and cancer cells. In particular, Ef may be involved in both the initiation and progression stages of oral cancer. Citation Format: Stefan Kovac, Tiara S. Napier, Jessica Scoffield, Hope M. Amm. Oral bacteria effects on oral cancer initiation and progression [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 2634.

Isoform specific FBXW7 mediates NOTCH1 Abruptex mutation C1133Y deregulation in oral squamous cell carcinoma

Our group previously identified that the NOTCH1 Abruptex domain contains the most mutations in Chinese OSCC patients, including a hotspot mutation (C1133Y). FBXW7 is an E3 ubiquitin ligase that regulates a network of proteins, including NOTCH1, via degradation. In this study, we first described the co-localization of isoform specific FBXW7-FBXW7β and NOTCH1C1133Y mutation in the same cytoplasmic sites. Gain- and loss-of-function assays were performed to examine the tumor suppressor role of FBXW7β in the proliferation and invasion of OSCC cells. The co-expression of NOTCH1C1133Y and FBXW7β significantly attenuated tumor growth. Meanwhile, FBXW7β reversed the oncogenic phenotype and the activation of the AKT/ERK/NFκB pathway induced by NOTCH1C1133Y mutation. FBXW7β downregulated the stability of NOTCH1C1133Y protein and promoted protein ubiquitination. This was the first time that we selected a NOTCH1 hotspot mutation detected in clinical samples and identified the function of FBXW7β that mediated NOTCH1 mutation degradation in OSCC. The newly identified interaction between FBXW7β and NOTCH1C1133Y protein provides new insights into the progression of OSCC, especially regarding Abruptex domain mutations, and represents a valuable target for OSCC therapy.

MicroRNA-345 functions as a tumor suppressor via targeting ZEB2 in oral squamous cell carcinoma

ObjectiveThe objective of this study was to explore the role of miRNAs in OSCC and to identify potential novel biomarkers or therapeutic agents in OSCC treatment. DesignMicroarray analysis and quantitative reverse transcription polymerase chain reaction (qRT-PCR) were performed to identify and verify differentially expressed miRNAs in OSCC tissues. The migration, invasion, proliferation and cell cycle of OSCC cells were analyzed to determine the function of miR-345 in OSCC development. Bioinformatics analysis and Dual-luciferase reporter assays were performed to identify and verify the target of miR-345. ResultsThe results showed a total of 17 miRNAs with significantly different expression in OSCC tissues (5 upregulated miRNAs and 12 downregulated miRNAs), including miR-345. The microarray results were also validated by qRT-PCR using 22 pairs of cancerous tissues and matched non-cancerous healthy samples. In particular, miR-345 expression was significantly lower in OSCC tissues. In addition, overexpression of miR-345 mimics in OSCC cells significantly inhibited their migration, invasion and proliferation while inducing cell cycle arrest in the G1 phase. Bioinformatics analysis predicted ZEB2 (zinc finger E-box-binding homeobox 2) as a potential target of miR-345, and luciferase reporter assays confirmed that miR-345 targeted ZEB2 through direct binding the 3′ untranslated region of ZEB2. Furthermore, miR-345 overexpression in OSCC reduced both mRNA and protein expression of ZEB2. ConclusionsThe results of this study indicated that miR-345 functions as a tumor suppressor to target ZEB2 in OSCC. These findings suggest that the miR-345/ZEB2 axis may be used as a potential therapeutic target in OSCC treatment.

Abstract A60: Targeted disruption of PI3K/Akt/mTOR signaling pathway induces cell cycle arrest, apoptosis, and autophagy, and inhibits inflammation, invasion and angiogenesis of OSCC cells

Abstract Purpose: The phosphoinositide-3-kinase (PI3K) pathway is frequently altered in human cancer. This has led to the development and study of novel PI3K inhibitors for targeted therapy and also to overcome resistance to radiotherapy. Method: The antitumor effects of PI3K inhibitors (PI-828, PI-103 and PX-866) in terms of cell proliferation, colony formation, induction of apoptosis, cell cycle arrest, invasion, autophagy, and pNF-κB/p65 translocation in SCC-4, SCC-9 and SCC-25 cells was studied by performing MTT, clonogenic, DAPI staining, propidium iodide staining, annexin-V binding, Matrigel invasion, acridine orange staining and immunofluorescence assay. Western blot assay was performed to assess the alteration in the expression of various proteins. Results: PI-828 and PI-103 treatment exhibited dose-dependent inhibition of growth and proliferation of OSCC cells with a concomitant induction of apoptosis, altered cell cycle regulation and decreased invasiveness (p &amp;lt;0.01). PX-866 induced apoptosis, cell cycle arrest, autophagy and a significant decrease in the invasiveness of oral cancer cells as compared to untreated cells (p &amp;lt;0.01). These compounds significantly reduced expression of COX-2, Cyclin-D and VEGF in the treated cells besides cytoplasmic accumulation of pNF-κB/p65 protein. In addition to PI3Kα, inactivation of downstream components, i.e., Akt and mTOR, was seen. Conclusion: PI3K inhibitors such as PI-103, PI-828 and PX-866 may be developed as potential chemotherapeutic agents for effective treatment of OSCC and other malignancies associated with activated PI3K/Akt pathway. Note: This abstract was not presented at the conference. Citation Format: Sadhna Aggarwal, Sarah John, Suresh C. Sharma, Satya N. Das. Targeted disruption of PI3K/Akt/mTOR signaling pathway induces cell cycle arrest, apoptosis, and autophagy, and inhibits inflammation, invasion and angiogenesis of OSCC cells [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology and Immunotherapy; 2018 Nov 27-30; Miami Beach, FL. Philadelphia (PA): AACR; Cancer Immunol Res 2020;8(4 Suppl):Abstract nr A60.