Oral Squamous Cell Research Articles

Identification of Potential Lead Compounds against BCL-2 through In-silico Screening of Phytochemicals of Nigella sativa and Cuscuta reflexa for Oral Squamous Cell Carcinoma Management

Significance of the study: In silico analysis is frequently used in physicochemical characterisation, the discovery and improvement of novel compounds with affinity to a target, and the elucidation of absorption, distribution, metabolism, excretion, and toxicity features. Aim and Objective: The aim of this study is to assess the efficacy of Nigella sativa and Cuscuta reflexa in targeting Bcl2-antiapoptotic protein in Oral Squamous Cell Carcinoma through in silico analysis. Materials and Methods: The present study was designed to formulate a drug against Oral Squamous cell carcinoma against the target protein Bcl-2. Protein Database (PDB) was used to select and analyse the protein. Based on the literature search, the original molecules selected were thymoquinone, tannin pyrogallol, Cuscutin, kaempferol, nigellicine and the ligand structures were obtained ZINC15 database. Target protein structure was recognised through Protein sequence from PB (Protein Data Bank). Swiss ADME was used for assessing the properties of the molecules. Twenty new molecules were identified from zinc pharmer and docking was done for all the 20 using Swiss dock. Binding energy was assessed and compared with the original molecules.ZINC73690300 and ZINC73690304 had better binding energy than that of original molecules. Both molecules can be potential candidates for Bcl2 inhibition to prevent OSCC. Results: ZINC73690300 and ZINC73690304 had better binding energy than that of the original molecules. Both molecules can be potential candidates for Bcl-2 inhibition to prevent OSCC. Conclusion: The present study evaluated the binding energy and bioavailability of the combined extract, thus attempting to predict the target ligand binding between the compounds in OSCC before attempting in vitro studies.-

Effect of 5-Aminolevulinic Acid (5-ALA) in "ALADENT" Gel Formulation and Photodynamic Therapy (PDT) against Human Oral and Pancreatic Cancers.

Oral squamous-cell and pancreatic carcinomas are aggressive cancers with a poor outcome. Photodynamic therapy (PDT) consists of the use of photosensitizer-induced cell and tissue damage that is activated by exposure to visible light. PDT selectively acts on cancer cells, which have an accumulation of photosensitizer superior to that of the normal surrounding tissues. 5-aminolevulinic acid (5-ALA) induces the production of protoporphyrin IX (PpIX), an endogenous photosensitizer activated in PDT. This study aimed to test the effect of a new gel containing 5% v/v 5-ALA (ALAD-PDT) on human oral CAL-27 and pancreatic CAPAN-2 cancer cell lines. The cell lines were incubated in low concentrations of ALAD-PDT (0.05%, 0.10%, 0.20%, 0.40%, 0.75%, 1.0%) for 4 h or 8 h, and then irradiated for 7 min with 630 nm RED light. The cytotoxic effects of ALAD-PDT were measured using the MTS assay. Apoptosis, cell cycle, and ROS assays were performed using flow cytometry. PpIX accumulation was measured using a spectrofluorometer after 10 min and 24 and 48 h of treatment. The viability was extremely reduced at all concentrations, at 4 h for CAPAN-2 and at 8 h for CAL-27. ALAD-PDT induced marked apoptosis rates in both oral and pancreatic cancer cells. Elevated ROS production and appreciable levels of PpIX were detected in both cell lines. The use of ALA-PDT as a topical or intralesional therapy would permit the use of very low doses to achieve effective results and minimize side effects. ALAD-PDT has the potential to play a significant role in complex oral and pancreatic anticancer therapies.

Therapeutic Potential of Dicentrine in Human Disorders: A Review On Medicinal Importance and Pharmacological Activities of Aporphine Alkaloid

Backgrounds: Plants have always been an important source of food and drugs, in addition to their application in medical and cosmetic sectors. The popularity of herbal remedies has increased in many countries in recent decades. Herbal medicines contain numerous active phytocomponents with different biological activities. Alkaloid class phytochemicals have diverse chemical structures and pharmacological activities in nature. Additionally, Aporphinoids are an important class of plant secondary metabolites that have been used for the treatment of numerous human disorders for a long time in traditional medicine. Dicentrine is an aporphine class phytochemical isolated from numerous medicinal plants, including Actinodaphne sesquipedalis and Lindera megaphylla. Methods: The biological potential of dicentrine, an aporphine alkaloid derivative, has been described in the present work. Scientific data on dicentrine were collected here from different scientific databases and presented in this paper in order to know the biological importance of dicentrine in medicine for the treatment of human complications. Further, detailed pharmacological activities and scientific data on dicentrine were also analyzed in order to determine its therapeutic potential in medicine. Moreover, its analytical aspects were also described in this work to understand its separation and isolation methods. Results: The present work described the biological potential of dicentrine in medicine and its analytical aspects. It signified the biological potential of dicentrine in cancer, breast cancer, oral squamous cell carcinoma, lung adenocarcinoma, blood pressure, inflammatory disorders, hyperlipidaemia, arrhythmia, stomach muscle, and glomerulonephropathy. Further, its effectiveness in medicine was found to be mainly because of its antiplatelet, alpha 1-adrenoceptor, epidermal growth factor, antiprotozoal, larvicidal, antimicrobial, topoisomerase II, and acetylcholinesterase inhibitory potential. Moreover, other scientific data also signified its metabolism and pharmacokinetic parameters in terms of its analytical aspects in medicine. Conclusion: The present review gives us an updated summary of the scientific information for pharmacological activities and analytical aspects of dicentrine in medicine. It also signified the potential contribution of dicentrine in medicine for the development of a newer class of drug molecules for human disorders.

Monoterpene antifungal activities: evaluating geraniol, citronellal, and linalool on Candida biofilm, host inflammatory responses, and structure-activity relationships.

Introduction: Despite the rising concern with fungal resistance, a myriad of molecules has yet to be explored. Geraniol, linalool, and citronellal are monoterpenes with the same molecular formula (C10H18O), however, neither the effect of these compounds on inflammatory axis induced by Candida spp. nor the antibiofilm Structure-Activity Relationship (SAR) have been well-investigated. Herein we analyzed geraniol, linalool and citronellal antifungal activity, cytotoxicity, and distinctive antibiofilm SAR, also the influence of geraniol on Candida spp induced dysregulated inflammatory axis, and in vivo toxicity. Methods: Minimal inhibitory (MIC) and fungicidal (MFC) concentrations against Candida spp were defined, followed by antibiofilm activity (CFU-colony forming unit/mL/g of dry weight). Cytotoxic activity was assessed using human monocytes (THP-1) and oral squamous cell (TR146). Geraniol was selected for further analysis based on antifungal, antibiofilm and cytotoxic results. Geraniol was tested using a dual-chamber co-culture model with TR146 cells infected with C. albicans, and THP-1 cells, used to mimic oral epithelium upon fungal infection. Expression of Candida enzymes (phospholipase-PLB and aspartyl proteases-SAP) and host inflammatory cytokines (interleukins: IL-1β, IL-6, IL-17, IL-18, IL-10, and Tumor necrosis factor-TNF) were analyzed. Lastly, geraniol in vivo toxicity was assessed using Galleria mellonella. Results: MIC values obtained were 1.25-5mM/mL for geraniol, 25-100mM/mL for linalool, and 100-200mM/mL for citronellal. Geraniol 5 and 50mM/mL reduced yeast viability during biofilm analysis, only 500mM/mL of linalool was effective against a 72h biofilm and no biofilm activity was seen for citronellal. LD50 for TR146 and THP-1 were, respectively: geraniol 5.883 and 8.027mM/mL; linalool 1.432 and 1.709mM/mL; and citronellal 0.3006 and 0.1825mM/mL. Geraniol was able to downregulate expression of fungal enzymes and host pro-inflammatory cytokines IL-1β, IL-6, and IL-18. Finally, safety in vivo parameters were observed up to 20mM/Kg. Discussion: Despite chemical similarities, geraniol presented better antifungal, antibiofilm activity, and lower cytotoxicity when compared to the other monoterpenes. It also showed low in vivo toxicity and capacity to downregulate the expression of fungal enzymes and host pro-inflammatory cytokines. Thus, it can be highlighted as a viable option for oral candidiasis treatment.

TGF-β Signaling in Progression of Oral Cancer.

Oral cancer is a common malignancy worldwide, accounting for 1.9% to 3.5% of all malignant tumors. Transforming growth factor β (TGF-β), as one of the most important cytokines, is found to play complex and crucial roles in oral cancers. It may act in a pro-tumorigenic and tumor-suppressive manner; activities of the former include cell cycle progression inhibition, tumor microenvironment preparation, apoptosis promotion, stimulation of cancer cell invasion and metastasis, and suppression of immune surveillance. However, the triggering mechanisms of these distinct actions remain unclear. This review summarizes the molecular mechanisms of TGF-β signal transduction, focusing on oral squamous cell and salivary adenoid systemic carcinomas as well as keratocystic odontogenic tumors. Both the supporting and contrary evidence of the roles of TGF-β is discussed. Importantly, the TGF-β pathway has been the target of new drugs developed in the past decade, some having demonstrated promising therapeutic effects in clinical trials. Therefore, the achievements of TGF-β pathway-based therapeutics and their challenges are also assessed. The summarization and discussion of the updated knowledge of TGF-β signaling pathways will provide insight into the design of new strategies for oral cancer treatment, leading to an improvement in oral cancer outcomes.

Chrysophanol-Induced Autophagy Disrupts Apoptosis via the PI3K/Akt/mTOR Pathway in Oral Squamous Cell Carcinoma Cells.

Background and Objectives: Natural products are necessary sources for drug discovery and have contributed to cancer chemotherapy over the past few decades. Furthermore, substances derived from plants have fewer side effects. Chrysophanol is an anthraquinone derivative that is isolated from rhubarb. Although the anticancer effect of chrysophanol on several cancer cells has been reported, studies on the antitumor effect of chrysophanol on oral squamous-cell carcinoma (OSCC) cells have yet to be elucidated. Therefore, in this study, we investigated the anticancer effect of chrysophanol on OSCC cells (CAL-27 and Ca9-22) via apoptosis and autophagy, among the cell death pathways. Results: It was found that chrysophanol inhibited the growth and viability of CAL-27 and Ca9-22 and induced apoptosis through the intrinsic pathway. It was also found that chrysophanol activates autophagy-related factors (ATG5, beclin-1, and P62/SQSTM1) and LC3B conversion. That is, chrysophanol activated both apoptosis and autophagy. Here, we focused on the roles of chrysophanol-induced apoptosis and the autophagy pathway. When the autophagy inhibitor 3-MA and PI3K/Akt inhibitor were used to inhibit the autophagy induced by chrysophanol, it was confirmed that the rate of apoptosis significantly increased. Therefore, we confirmed that chrysophanol induces apoptosis and autophagy at the same time, and the induced autophagy plays a role in interfering with apoptosis processes. Conclusions: Therefore, the potential of chrysophanol as an excellent anticancer agent in OSCC was confirmed via this study. Furthermore, the combined treatment of drugs that can inhibit chrysophanol-induced autophagy is expected to have a tremendous synergistic effect in overcoming oral cancer.

Novel mucoadhesive celecoxib-loaded cubosomal sponges: Anticancer potential and regulation of myeloid-derived suppressor cells in oral squamous cell carcinoma

Oral squamous-cell carcinoma (OSCC) is a widespread health problem. Myeloid-derived suppressor cells (MDSCs) are major tumor microenvironment (TME) population that govern many carcinogenesis aspects by establishing immunosuppressive milieu favoring tumor aggressiveness and treatment resistance. Cyclooxygenase-2 (COX-2) regulates MDSCs activity, hence, COX-2-selective inhibition by celecoxib (CXB) showed good anticancer effect at relatively high doses with possible subsequent cardiovascular complications. Therefore, targeted CXB delivery to MDSCs may represent a promising OSCC treatment strategy. Novel mucoadhesive-cubosomal buccal sponges were prepared for MDSCs targeting and were evaluated for their in-vitro quality attributes, ex-vivo mucoadhesion using buccal chicken-mucosa. Optimally-selected formulation showed considerable uptake by CD33+/11b+MDSCs in human OSCC cell-line (SCC-4) when quantitatively analyzed by flow-cytometry and examined using confocal-laser microscope. Optimum formulations loaded with low CXB doses (12 mg) were promoted to in-vivo studies via local application, using 4-nitroquinoline-1-oxide-induced OSCC in rats, and compared to their corresponding CXB gels. SP16 revealed the highest ability to decrease MDSC activation, recruitment and TME-immunosuppression in the isolated tumors. Consequently, SP16 exerted the greatest capacity to reduce histologic tumor grade, the OSCC-specific serum tumor markers levels, cancer hallmarks and stemness markers. CXB-loaded cubosomal sponges preferentially target MDSCs with noticeable anticancer potential and may exemplify novel mucoadhesive nanocarriers for OSCC treatment.

Evaluation of Immunohistochemical Expression of BCL-2 in Oral Squamous Cell Carcinoma

Introduction: Immunohistochemical (IHC) expression ofBcl-2 oncogene is helpful in characterization and prognosisof oral cancer. In the present study, we evaluated theimmunoexpression of Bcl-2 oncogene in oral squamouscell carcinoma (OSCC) tissue specimen and correlated itclinicohistopathologically.Material and Method: A total of 42 OSCC tissue specimenwere subjected to evaluation for immunohistochemicalexpression of Bcl-2 oncogene. IHC expression was scored as“-” (<5% positive cells), + (5–24% positive cells), ++ (25–50%positive cells) and +++ (>50% positive cells). Pathological Tstage, nodal status, histopathological grade, lymphovascularinvasion (LVI), perineural invasion (PNI) and mitotic index (MI)were noted. Data was analysed using SPSS 21 software.Chi-square, ANOVA and Kruskall-Wallis tests were used tocompare the data.Results: Mean age of patients was 49.05 ± 13.28 years,majority of them were males (83.3%). There was a dominanceof well/moderately differentiated (81%) cases. Lymphovascularand perineural invasion were seen in 16.7% and 26.2% cases.Majority had mitotic index 2 (66.7%), pT stage 3/4 (73.8%)and no nodal involvement (54.8%). Bcl-2 expression wasseen in 34 (81%) specimen. It was +, ++ and +++ in 6 (14.3%),15 (35.7%) and 13 (31.0%) cases. A significant correlationof Bcl-2 expression was seen with higher histopathologicalgrades, mitotic index and pT stage. There was no correlationof Bcl-2 expression with age, sex, LVI, PNI, MI and pNstatus.Conclusion: There was a high IHC expression of Bcl-2 oncogenewhich was significantly associated with histopathological gradeand pT stage. Bcl-2 expression could have a prognostic value.

Downregulation of AT-rich interaction domain 2 underliesnatural killer cell dysfunction in oral squamouscell carcinoma.

The immune system plays a significant role in controlling oral squamous cell carcinoma (OSCC) initiation and progression. Natural killer (NK) cells actively participate in antitumor immunity but become dysfunctional or exhausted in the tumor microenvironment. To explore the mechanisms of NK cell dysfunction in OSCC, we characterized the expression and function of AT-rich interaction domain 2 (ARID2) in NK cells in a murine OSCC model. ARID2 was downregulated in tongue NK cells compared with splenic NK cells. Notably, ARID2 was significantly decreased in NK cells with an exhausted phenotype and weakened antitumor function. ARID2 knockdown resulted in the upregulation of programmed cell death protein 1 (PD-1) and downregulation of interferon-gamma (IFN-γ), tumor necrosis factor (TNF), granzyme B and perforin in NK cells. As a result, ARID2 knockdown impaired NK cell cytotoxicity. Besides, ARID2 overexpression suppressed the expression of PD-1 and lymphocyte-activation gene 3, and promoted the expression of IFN-γ, TNF, granzyme B and perforin in NK cells which were adoptively transferred into OSCC-bearing mice. Taken together, our study implies that the OSCC microenvironment triggers ARID2 downregulation in intratumoral NK cells. In turn, ARID2 downregulation results in PD-1 upregulation on NK cells and subsequently impairs NK cell cytotoxicity. Therefore, we uncovered a novel mechanism of NK cell dysfunction in OSCC.

The role of altered microRNAexpression in premalignant and malignant head and neck lesions with epithelial origin.

The premalignant lesions of the oral cavity carry a risk of transformation to malignancy. Hence, early diagnosis followed by timely intervention remarkably affects the prognosis of patients. During tumorigenesis, particular microRNAs (miRNAs) show altered expressions and because of their post transcriptionally regulatory role could provide favorable diagnostic, therapeutic, or prognostic values in head and neck cancers. In this review, we have demonstrated diagnostic, prognostic, and potential therapeutic roles of some miRNAs associated with oral premalignant and malignant lesions based on previous validate studies. It is previously documented that dysregulation of miRNAs contributes to cancer development and progression. MiRNAscould be tumor suppressors that normally suppress cell proliferation, differentiation, and apoptosis or play as oncogenes that improved tumorigenesis process. Altered expression of miRNAs has also been reported in premalignant oral epithelial lesions such as leukoplakia, oral submucous fibrosis, oral lichen planus and some malignant carcinoma like oral squamous cell, verrucous, spindle cell, Merkel cell carcinoma and basal cell. Some of miRNAs could be new therapeutic candidates in miRNA-based target gene therapy. Although more investigations are required to identify the most favorable miRNA candidate, altered expression of some miRNAs could be used as biomarkers in premalignant lesions and oral cancers with high sensitivity and specificity.

DNA Karyometry for Automated Detection of Cancer Cells.

Simple SummaryCancers have to be microscopically established before they can be treated adequately. This can be performed on cell and/or tissue samples. Smears or sedimentations of cells on glass slides are used to microscopically screen large number of specimens for the presence of cancer cells. Such screenings require highly specialized personnel, which is not available in most countries. We present a microscope-based scanner which is able to identify cancer cells in smears from the oral cavity or in body cavity fluids. In addition, using the same system, the degree of malignancy of prostate cancer can be determined. A combination of image features of stained nuclei and their DNA-content is used to raise a suspicion of malignancy. Nuclear diagnostic classifiers were trained by an expert using supervised machine learning. All device-proposed diagnoses can be verified by a specialist. The overall percentage of correct device-derived diagnoses on oral smears was 91.3% as compared to 75.0% for conventional, subjective investigation.Background: Microscopical screening of cytological samples for the presence of cancer cells at high throughput with sufficient diagnostic accuracy requires highly specialized personnel which is not available in most countries. Methods: Using commercially available automated microscope-based screeners (MotiCyte and EasyScan), software was developed which is able to classify Feulgen-stained nuclei into eight diagnostically relevant types, using supervised machine learning. the nuclei belonging to normal cells were used for internal calibration of the nuclear DNA content while nuclei belonging to those suspicious of being malignant were specifically identified. The percentage of morphologically abnormal nuclei was used to identify samples suspected of malignancy, and the proof of DNA-aneuploidy was used to definitely determine the state malignancy. A blinded study was performed using oral smears from 92 patients with Fanconi anemia, revealing oral leukoplakias or erythroplakias. In an earlier study, we compared diagnostic accuracies on 121 serous effusion specimens. In addition, using a blinded study employing 80 patients with prostate cancer who were under active surveillance, we aimed to identify those whose cancers would not advance within 4 years. Results: Applying a threshold of the presence of >4% of morphologically abnormal nuclei from oral squamous cells and DNA single-cell or stemline aneuploidy to identify samples suspected of malignancy, an overall diagnostic accuracy of 91.3% was found as compared with 75.0% accuracy determined by conventional subjective cytological assessment using the same slides. Accuracy of automated screening effusions was 84.3% as compared to 95.9% of conventional cytology. No prostate cancer patients under active surveillance, revealing DNA-grade 1, showed progress of their disease within 4.1 years. Conclusions: An automated microscope-based screener was developed which is able to identify malignant cells in different types of human specimens with a diagnostic accuracy comparable with subjective cytological assessment. Early prostate cancers which do not progress despite applying any therapy could be identified using this automated approach.

Regulation of dietary polyphenols on cancer cell pyroptosis and the tumor immune microenvironment.

Cancer is a major public health problem that threatens human life worldwide. In recent years, immunotherapy has made great progress in both clinical and laboratory research. But the high heterogeneity and dynamics of tumors makes immunotherapy not suitable for all cancers. Dietary polyphenols have attracted researchers' attention due to their ability to induce cancer cell pyroptosis and to regulate the tumor immune microenvironment (TIME). This review expounds the regulation of dietary polyphenols and their new forms on cancer cell pyroptosis and the TIME. These dietary polyphenols include curcumin (CUR), resveratrol (RES), epigallocatechin gallate (EGCG), apigenin, triptolide (TPL), kaempferol, genistein and moscatilin. New forms of dietary polyphenols refer to their synthetic analogs and nano-delivery, liposomes. Studies in the past decade are included. The result shows that dietary polyphenols induce pyroptosis in breast cancer cells, liver cancer cells, oral squamous cells, carcinoma cells, and other cancer cells through different pathways. Moreover, dietary polyphenols exhibit great potential in the TIME regulation by modulating the programmed cell death protein 1(PD-1)/programmed death-ligand 1 (PD-L1) axis, enhancing antitumor immune cells, weakening the function and activity of immunosuppressive cells, and targeting tumor-associated macrophages (TAMs) to reduce their tumor infiltration and promote their polarization toward the M1 type. Dietary polyphenols are also used with radiotherapy and chemotherapy to improve antitumor immunity and shape a beneficial TIME. In conclusion, dietary polyphenols induce cancer cell pyroptosis and regulate the TIME, providing new ideas for safer cancer cures.

Abstract 203: The role of FOXM1 in senescence suppression and radiation resistance in oral cavity squamous cell carcinoma

Abstract INTRODUCTION: Advanced oral cavity squamous cell carcinoma (OCSCC) relies on surgery and radiation for cure; with ~30% experiencing locoregional failure. Earlier studies suggest OCSCCs resist radiation response by suppressing senescence. CDKN2A, coding for p16INK4a (p16), a CDK4/6 inhibitory protein, is among the most commonly lost tumor suppressors in OCSCC. Forkhead Box M1 (FOXM1) is an understudied key regulatory protein in OCSCC whose activation, similar to Rb, is controlled by CDK4/6. FOXM1 activity not only impacts cell proliferation, but also promotes DNA damage repair and suppresses senescence. The current study examines how FOXM1 signaling plays a critical role in resistance to radiation in OCSCC. We demonstrate that CDK4/6 inhibition improves radiosensitivity in OCSCC via senescence as a result of FOXM1 suppression. METHODS: Microarray data were analyzed to investigate FOXM1 levels in OSCC tumors. Basal FOXM1 levels in immortalized OCSCC cell lines (HN5 and Cal27) were evaluated using western blot. Cells were treated with palbociclib, radiation (RT), or combination (1µM palbociclib and 4 Gy (P+RT)) to examine the effects on senescence and FOXM1 signaling. β galactosidase colorimetric (β-gal) staining was used to evaluate senescence. Western blot was used to measure protein levels and QPCR to measure FOXM1 expression. To demonstrate senescence as a function of FOXM1 inhibition, knockdown assays (KD) were performed with siRNA targeting FOXM1. To further eliminate off target drug effects knockdown was also performed with siRNA targeting CDK4/6. RESULTS: FOXM1 was upregulated ~2.2-fold in OCSCC tumors vs. matched normal mucosa samples (N=43, p=2.85008E-06). FOXM1 protein was upregulated 7-fold (HN5) and 5-fold (Cal27) compared to normal oral keratinocytes (HOK16B). β-gal staining revealed a dose dependent senescence induction in OCSCC, where P+RT vs. RT showed a robust 5-fold (HN5) and 3-folds (Cal27) increase. Interestingly, FOXM1 protein and transcript expression doubled in samples treated with RT. P+RT vs. RT demonstrated a decrease in FOXM1 protein levels in HN5 (10-fold) and Cal27 (3-fold), with corresponding decreases in mRNA levels. Knockdown of FOXM1 induced senescence, which further increased when combined with radiation: siFOXM1+4 Gy resulted in 60% (HN5) and 50% (CAL27) β-gal positive cells. Compared to untreated controls, CDK4/6 KD demonstrated a 2.5 and 10-fold decrease in FOXM1 protein levels in HN5 and CAL27, respectively. Furthermore, a concurrent inhibition of CDK4/6 with RT not only resulted in a decrease in FOXM1 expression but also resulted in a near-complete senescence in treated cells (95% β-Gal staining). CONCLUSION: Suppressing FOXM1 directly or via CDK4/6 inhibition appears to make OCSCC more susceptible to RT via potentiating senescence. We propose inhibition of FOXM1 as a rational and potentially effective strategy to sensitize OCSCC to radiation. Citation Format: Nitisha Shrivastava, Yisrael Wallach, Daniel Li, Carlos Thomas, Michael B. Prystowsky, Indranil Basu, Chandan Guha, Thomas J. Ow. The role of FOXM1 in senescence suppression and radiation resistance in oral cavity squamous cell carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 203.

Abstract 2142: Prognostic and predictive applications from mesenchymal gene expression subtype analysis for early-stage, HPV(-) head and neck squamous cell carcinoma

Abstract Introduction: Although often presented as one disease for genomic studies, head and neck squamous cell carcinoma (HNSCC) is treated according to clinical factors such as anatomic subsite, and tumor stage. Oral cavity squamous cell carcinoma (OCSCC) comprises 1/3 of all HNSCC and is predominantly HPV(-). Depending upon clinical stage, OCSCC treatment involves surgical resection +/- neck dissection, followed by radiation +/- chemotherapy. Our group and others previously described four mRNA expression patterns (classical, atypical, basal, and mesenchymal), each with unique genomic features and prognosis. Here, we further examine the clinical utility of gene expression subtyping in HNSCC and introduce the potential for predictive applications in HPV(-) HNSCC according to clinically relevant subgroups including oral cavity cancer. Experimental Procedures: Multiple independent HNSCC datasets were obtained from public repositories, totaling 562 patients, including from MD Andersen (MDA) (GSE41117) and the Cancer Genome Atlas (TCGA) sourced from the Genome Data Commons. Cases were included for further analysis if they had N stage and overall survival values. Samples were assigned molecular subtypes (basal, mesenchymal, atypical or classical) using a reduced gene set version of the classifier reported earlier. HPV status was determined by HPV gene expression. The clinical endpoint was overall survival at censured at 36 months. Kaplan-Meier (KM) plots and logrank tests were used to investigate associations between clinical variables and survival. Cox models were used to adjust for potential confounders. All statistics were performed using the survival package in R. Results: Of the 418 training patients from the TCGA dataset that met analysis criteria, nearly 20% presented with stage I and II tumors. In the clinically relevant subgroup of node(-) OCSCC patients, mesenchymal subtype was associated with worse survival (HR 2.4, p=0.021), offering a novel and potentially actionable biomarker in otherwise early-stage and low risk disease. Associations in the MDA validation cohort confirmed the association. Node(-) non-mesenchymal OCSCC patients had far better survival (no deaths observed) compared to node(-) mesenchymal and all node(+) patients which had similarly poor survival. Differential responses as a function of radiation will be presented. Summary and Conclusions: This study confirms that the mesenchymal subtype is associated with worse outcomes across all cases of HNSCC. However, we demonstrate that mesenchymal subtype is associated with poor survival, even in the clinically relevant setting of early-stage, node(-) OCSCC treated with surgical resection. These findings highlight the potential value of gene expression subtyping as an adjunct to pathology for deciding which treatment option is best suited for patients with HNSCC. Citation Format: D. Neil Neil Hayes, MD, MS, MPH, Greg Mayhew, Josh Uronis, Jose Zevallos. Prognostic and predictive applications from mesenchymal gene expression subtype analysis for early-stage, HPV(-) head and neck squamous cell carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 2142.

Novel computer-aided reconstruction of soft tissue defects following resection of oral and oropharyngeal squamous cell carcinoma

BackgroundReconstruction of soft tissue defects following surgical tumor resection is important for quality of life in cancer patients with oral and oropharyngeal squamous cell carcinoma (SCC). This study presents a novel computer-aided reconstruction of soft tissue (CARST) technology employed with these patients.MethodsWe first described the CARST technology in detail in a report of a 34-year-old male patient with locally invasive right-sided tongue SCC following a nearly total glossectomy and reported the postoperative outcomes. This digital technology was applied to construct a 3D model from CT images, which was used to delineate surgical resection boundaries and design a personalized reconstruction of the soft tissue defect. A nonuniform rational B-spline (NURBS) was generated and applied to transform the 3D model into a 2D flap-cutting guide printed out using a 3D printer. We then reported a case-series study on oral and oropharyngeal SCC patients who were randomly assigned to receive the CARST (n = 15) or a traditional soft tissue reconstruction (n = 15). Clinicopathological features and short- and long-term postoperative outcomes between the two groups were compared.ResultsThe patient with the tongue SCC had a successful CARST following surgical tumor resection without any complications. His speech and swallowing functions recovered well after surgery and he experienced no significant changes to his appearance following recovery. There was no recurrence within a 3-year follow-up period. Results of the case-series study showed that the CARST group had significantly shorter operative and post-operation hospital-stay time, a higher flap utilization rate, and a trend of less and milder postoperative complications, and they experienced no significant difference in intraoperative blood loss and long-term outcomes compared to the traditional group.ConclusionCARST is a safer and more efficient personalized technology of soft tissue reconstruction following surgical tumor resection in patients with oral and oropharyngeal SCC.

Signal peptidase complex catalytic subunit SEC11A upregulation is a biomarker of poor prognosis in patients with head and neck squamous cell carcinoma.

In the current study, we aimed to investigate the expression of the five microsomal signal peptidase complex (SPC) subunit genes (SEC11A, SEC11C, SPCS1, SPCS2, and SPCS3) in head and neck squamous cell carcinoma (HNSC) and to explore their prognostic value. Data from the HNSC subset of The Cancer Genome Atlas (TCGA) and one previous single-cell RNA-seq dataset was used. Subgroup analysis was conducted in tumors from different anatomic sites. Gene set enrichment analysis (GSEA), and immune cell infiltration analysis were performed to check the influence of SEC11A on the tumor microenvironment. Among the genes significantly upregulated in the tumor group, only SEC11A expression (as a continuous variable) is independently associated with poorer progression-free survival (PFS) (HR: 2.075, 95%CI: 1.447–2.977, p<0.001) and disease-specific survival (DSS) (HR: 2.023, 95%CI: 1.284–3.187, p = 0.002). Subgroup analysis confirmed the prognostic value in tumors from three anatomic origins, including laryngeal squamous cell carcinoma, oral cavity-related squamous cell carcinoma, and oropharynx-related squamous cell carcinoma. SEC11A is expressed in all subtypes of cells in the tumor microenvironment. Its expression showed a moderate positive correlation with its gene-level copy number (Pearson’s r = 0.53, p<0.001). SEC11A expression was negatively correlated with CD8+ T cells and B cells, but was positively correlated with cancer-associated fibroblast and myeloid-derived suppressor cells (MDSCs) in the tumor microenvironment. In summary, SEC11A upregulation is a result of gene amplification in head and neck squamous cell carcinoma. Its upregulation might serve as an independent prognostic biomarker and a predictor of the infiltration of certain types of immune cells.

MiR-214 Modulates the Growth and Migration of Oral Cancer before and after Chemotherapy through Mediating ULK1.

The role of miRNAs as crucial components in carcinogenesis has been well documented. However, whether and how miR-214 influences oral cancer cells' drug resistance remains to be elucidated, and its downstream targets are still under investigation. Hence, this research is aimed at determining miR-214 and ULK1 expression in oral cancer before and after chemotherapy and their correlations with cancer cell growth. Human oral normal epithelial cells and human tongue squamous cell carcinoma CAL-27 cells were cultured to detect miR-214 and ULK1 levels. It was found that before chemotherapy, miR-214 was higher, while ULK1 was underexpressed in CAL-27 cells, versus normal epithelial cells. After chemotherapy, miR-214 decreased obviously in CAL-27 cells, while ULK1 level increased significantly. In addition, autophagy-related genes (Beclin 1, mTOR, and P53) in CAL-27 cells were found to be significantly inhibited before chemotherapy and were obviously increased after chemotherapy. Moreover, to further determine the impacts of miR-214 and ULK1 on oral cancer cell growth after chemotherapy, the two were overexpressed or silenced in CAL-27 cells after transfection. We found that ULK1 could effectively decrease the activity and invasion of CAL-27 cells and increase their apoptosis level, while miR-214 could antagonize its antitumor effect. Therefore, miR-214 can be used as an early prognostic biomarker for oral cancer, and ULK1 is a new candidate therapeutic target.

Factors associated with recurrence in oral cavity squamous cell carcinoma: A real-world experience from an Indian comprehensive cancer center.

e18018 Background: Oral cavity squamous cell carcinoma (OCSCC) is the third most common malignancies in India with an estimated 72,000 deaths annually. The incidence is increasing owing to rising alcohol and tobacco consumption in last 2 decades. Methods: We performed a retrospective cohort study in patients with OCSCC to identify risk factors for local and metastatic recurrences following definitive treatment including surgery and/or radiation therapy or chemotherapy or both at our institute. Patient records were screened from 2016 till 2021. Eligible patients were followed up for disease status and outcome. Data included sociodemographic details, substance use, tumor differentiation, stage at diagnosis, LVI, PNI, margin status, treatment details and site of recurrence. All statistical analysis was performed on Microsoft excel 2016 and Jamovi version 1.6.23.0. Results: Out of 195 patients files screened, 90 were available for analysis. Demographic details, substance use pattern and tumor profile are highlighted in table. Tongue was the most frequent site involved and most of the tumors were well to moderately differentiated. Majority of the patients presented as locally advanced disease (Stage IVa). LVI and PNI were identified in 5.75% and 29.89% patients respectively while one of the patients had a positive margin after resection. Surgery followed by adjuvant chemo-radiation was the most frequently prescribed treatment (n = 46; 52.8%) whereas 27.59% patients (n = 24) underwent surgery alone. Fifty six (63.3%) patients experienced recurrence out of which 50% were local and 35% were metastatic recurrences. On multivariate analysis, the predictors of disease recurrence were male gender (OR- 3.2; 95% CI- 1.02-10.05; p-0.05), tobacco and alcohol use (OR- 1.93; 95% CI- 0.8-4.6; p- 0.14 and OR- 2.16; 95% CI- 0.82-5.65; p- 0.12 respectively), Stage IVb disease (OR- &gt; 10; 95% CI- 0-10; p-1.0), presence of PNI (OR- 1.39 ; 95% CI- 0.55-3.70; p-0.497) and presence of LVI (OR- 2.54; 95% CI- 0.36-50.8; p- 0.414). Median disease free survival and overall survival of the cohort was 11.3 months and 15 months respectively. Advanced stage, tobacco consumption, presence of LVI and PNI negatively affected overall survival in our cohort. Conclusions: Oral cavity squamous cell carcinoma is among the most preventable malignancies and they contribute to significant morbidity and mortality. This study highlights the role of early detection and management of such patients. Patients may still experience disease recurrence after initial management which may be difficult to treat. Such patients should be encouraged to participate in a clinical trial as treatment options for recurrent disease are limited.[Table: see text]

Palmatine and berberine chloride synergistically Inhibit NanH sialidase of Tannerella forsythia

The periodontal pathogen Tannerella forsythia is associated with severe periodontitis, and expresses NanH sialidases that cleave sialic acids by hydrolyzing the glycosidic bonds to underlying sugars. Palmatine and berberine chloride are plant-derived alkaloids with pharmacological effects, including anti-inflammatory and anti-bacterial properties. Recombinant NanH sialidase was purified using HisTag affinity chromatography while sialidase activity was determined using 4-methylumbelliferyl N-acetyl-α-D-neuraminic acid (MUNANA) as a substrate. The individual and synergistic effects of palmatine and berberine chloride on NanH sialidase inhibition was determined as well as their antimicrobial effects. The IC50 values of palmatine and berberine chloride were found to be 0.143 and 0.474 mM respectively. A significant synergistic effect was observed when a 0.20 mM:0.50 mM Palmatine:Berberine chloride mixture was used, inhibiting NanH sialidase by almost 100%, as compared to 0.2 mM palmatine and 0.5 mM berberine chloride invidually, which inhibit sialidase activityby 60.33 and 55.94%, respectively. Additionally, an antimicrobial viability assay was conducted and, 0.5 mM palmatine and 0.45 mM berberine showed a significant antimicrobial activity against Tannerella forsythia. Lastly, to examine potential toxicity to host cells, thecytotoxic effects of palmatine (0.15 mM) on H357 oral squamous carcinoma cells was investigated using a trypan blue assay and palmatine was found not to be toxic. In summary, a combination of palmatine and berberine display significant synergistic inhibitory effects on NanH with minimal cytotoxic effects as well as potential antimicrobialeffects on the oral pathogen T. forsythia. Suggesting thatthese compounds may have potential for future development.

OC-0432 Patterns of failure in oral cavity squamous cell carcinoma treated with adjuvant radiotherapy

OC-0432 Patterns of failure in oral cavity squamous cell carcinoma treated with adjuvant radiotherapy