Calibration improves the agreement in grading oral epithelial dysplasia-Findings from a National Workshop in Malaysia.

INTRODUCTION: A major pitfall of many of the established oral epithelial dysplasia (OED) grading criteria is their lack of reproducibility and accuracy to predict malignant transformation. The main objective of this study was to determine whether calibration of practicing oral pathologists on OED grading could improve the reproducibility of the WHO 2017 and the binary OED grading systems. METHODS: A nationwide online exercise was carried out to determine the influence of calibration on the reproducibility of the WHO 2017 and the binary OED grading systems. RESULTS: A significant improvement was observed in the inter-observer agreement for the WHO 2017 OED grading system (K 0.196 vs. 0.448; Kw 0.357 vs. 0.562) after the calibration exercise. The significant difference (p = 0.027) in the level of agreement between those with five or more years and less than 5 years of experience was no more observed (p = 0.426) after the calibration exercise. The percent agreement for binary grading was significantly higher (91.8%) for buccal mucosal lesions as compared to lesions on the tongue after the calibration exercise. CONCLUSION: This study validates the significance of calibration in improving the reproducibility of OED grading. The nationwide exercise resulted in a statistically significant improvement in the inter-observer agreement for the WHO 2017 OED grading system among a large number of oral pathologists. It is highly recommended that similar exercises should be organized periodically by professional bodies responsible for continuing education among oral pathologists to improve the reliability of OED grading for optimal treatment of oral potentially malignant disorders.

Salivary biomarkers: Effective diagnostic tool for oral leukoplakia and oral squamous cell carcinoma.

OBJECTIVE: To identify potential salivary biomarkers for the diagnosis and monitoring of disease progression in oral squamous cell carcinoma and oral leukoplakia. MATERIALS AND METHODS: An advance search from PubMed and Hindawi was performed with keywords; oral leukoplakia/oral squamous cell carcinoma, salivary biomarker and diagnosis/prognosis. An additional search of articles was done through a manual search from the Google Scholar database. RESULTS: Twenty studies involving salivary biomarkers as diagnostic tools for oral squamous cell carcinoma and/or oral leukoplakia were identified. A narrative review was carried out. CONCLUSION: Single or multiple salivary biomarkers reported by most studies have shown great potential as diagnostic tools for oral squamous cell carcinoma and oral leukoplakia. However, the validation of sensitivity and specificity should be carried out to ensure the accuracy of the biomarkers. Furthermore, a standardised method for saliva collection should be established to prevent variability in the expression of biomarkers.

Image collection and annotation platforms to establish a multi-source database of oral lesions.

OBJECTIVE: To describe the development of a platform for image collection and annotation that resulted in a multi-sourced international image dataset of oral lesions to facilitate the development of automated lesion classification algorithms. MATERIALS AND METHODS: We developed a web-interface, hosted on a web server to collect oral lesions images from international partners. Further, we developed a customised annotation tool, also a web-interface for systematic annotation of images to build a rich clinically labelled dataset. We evaluated the sensitivities comparing referral decisions through the annotation process with the clinical diagnosis of the lesions. RESULTS: The image repository hosts 2474 images of oral lesions consisting of oral cancer, oral potentially malignant disorders and other oral lesions that were collected through MeMoSA® UPLOAD. Eight-hundred images were annotated by seven oral medicine specialists on MeMoSA® ANNOTATE, to mark the lesion and to collect clinical labels. The sensitivity in referral decision for all lesions that required a referral for cancer management/surveillance was moderate to high depending on the type of lesion (64.3%-100%). CONCLUSION: This is the first description of a database with clinically labelled oral lesions. This database could accelerate the improvement of AI algorithms that can promote the early detection of high-risk oral lesions.

High referral accuracy for oral cancers and oral potentially malignant disorders using telemedicine.

OBJECTIVE: To evaluate the accuracy of MeMoSA®, a mobile phone application to review images of oral lesions in identifying oral cancers and oral potentially malignant disorders requiring referral. SUBJECTS AND METHODS: A prospective study of 355 participants, including 280 with oral lesions/variants was conducted. Adults aged ≥18 treated at tertiary referral centres were included. Images of the oral cavity were taken using MeMoSA®. The identification of the presence of lesion/variant and referral decision made using MeMoSA® were compared to clinical oral examination, using kappa statistics for intra-rater agreement. Sensitivity, specificity, concordance and F1 score were computed. Images were reviewed by an off-site specialist and inter-rater agreement was evaluated. Images from sequential clinical visits were compared to evaluate observable changes in the lesions. RESULTS: Kappa values comparing MeMoSA® with clinical oral examination in detecting a lesion and referral decision was 0.604 and 0.892, respectively. Sensitivity and specificity for referral decision were 94.0% and 95.5%. Concordance and F1 score were 94.9% and 93.3%, respectively. Inter-rater agreement for a referral decision was 0.825. Progression or regression of lesions were systematically documented using MeMoSA®. CONCLUSION: Referral decisions made through MeMoSA® is highly comparable to clinical examination demonstrating it is a reliable telemedicine tool to facilitate the identification of high-risk lesions for early management.

A validated predictive model for risk of nodal metastasis in node negative oral squamous cell carcinoma of the buccal mucosa and tongue.

BACKGROUND: Lymph node metastasis in oral squamous cell carcinoma (OSCC) is influenced by clinical and histopathological variables. The aim of this study was to develop a simple model to predict nodal metastasis of OSCC in clinically negative necks (cN0). METHODS: Data from patients who underwent surgery for treatment of OSCC of the tongue or buccal mucosa with neck dissection were used for model development and validation. RESULTS: Nodal metastasis was significantly associated with gender, age, tumor size, site, pattern of invasion and depth of invasion on univariate analysis. All the five variables except age were retained at the variable selection step of the model development and were used in the final model because it was not significant at 0.10 significance level after adjusting for other variables. Regression coefficients of the model were used to estimate risks of nodal metastases for each combination of clinicopathological characteristics. A 10-fold cross-validation was used to assess the model. The average of the resultant 10 AUCs (along with its 95% confidence interval estimated using bootstrap) was used as the overall validated measure of the model. A risk chart was produced using probability of nodal metastasis predicted by the model for each combination of five characteristics. The model's ability to identify patients with nodal metastases as assessed by the area under the ROC curve (AUC) was 0.752. CONCLUSION: The model based on established clinicopathological variables has been internally validated on a large cohort of patients and offers practicability for use in OSCCs of the tongue and buccal mucosa.

Effectiveness of "OralDETECT": a Repetitive Test-enhanced, Corrective Feedback Method Competency Assessment Tool for Early Detection of Oral Cancer.

Early diagnosis of oral cancer results in less aggressive treatment and improves the quality of life and overall 5-year survival rate. Well-trained dental professionals can play a crucial role in the early detection of oral cancers. The present study aims to determine the effectiveness of the training program "OralDETECT", a spaced repetitive, test-enhanced learning tool with a corrective feedback mechanism for early detection of oral cancer. Thirty-two dentists and 259 dental students from three Malaysian dental schools were involved in this study. All participants were trained and calibrated to recognize oral potentially malignant disorders (OPMD) and oral cancer using "OralDETECT", which is comprised of a series of pre-test, lecture, post-tests and review sessions. The percentage of correct answers (scores) for each test given by the participants was calculated and analysed using a paired t test. It was found that the overall percentage of diagnostic accuracy for both dental professionals and student groups increased to above 80% from the pre-tests to the final post-tests. There was a significant improvement in overall scores between the pre-tests and all three post-tests for the dental professional groups and the student groups. The diagnostic accuracy for individual OPMD and lesions suspicious of oral cancer also increased to above 80% for both groups. The results of our study demonstrate that the "OralDETECT" is an efficient and effective competency tool which can be used to train dental professionals and students for the early detection of OPMD and oral cancer.

Survival of Oral Cancer Patients in Different Ethnicities.

Previous studies found that ethnicity influences oral cancer patients' survival; however, most studies were limited to certain ethnic groups particularly from the West, thus of limited relevance to Asians where the disease is most prevalent. We investigated the relationship between ethnicity and patient survival in multi-racial Malaysia. 5-year survival rate was 40.9%. No statistically significant difference was observed in survival between Malays, Chinese, Indians and Indigenous peoples (45.7%, 44.0%, 41.3%, 27.7% respectively). Increased tumor size, lymph node involvement and advanced tumor were predictive of poor survival. We conclude that ethnicity has no effect on survival or its prognostic indicators.

Prevalence of oral cancer, oral potentially malignant disorders and other oral mucosal lesions in Cambodia.

OBJECTIVES: To obtain data on the prevalence of oral mucosal lesions (OMLs) among Cambodians, and to assess the relationship between known risk habits of oral diseases with prevalence of oral potentially malignant disorders (OPMDs). DESIGN: This was a population-based, cross-sectional study whereby subjects were adults aged 18 years old and above. A workshop on the identification of OML was held to train and calibrate dental officers prior to data collection in the field. Sociodemographic and risk habits data were collected via face-to-face interview, whilst presence of OML and clinical details of lesions such as type and site were collected following clinical oral examination by the examiners. Data analysis was carried out using the Statistical Package for Social Science (SPSS) version 12.0. The association between risk habits and risk of OPMD was explored using logistic regression analysis. RESULTS: A total of 1634 subjects were recruited. Prevalence of OML for this population was 54.1%. Linea alba was the most common lesion seen (28.7%). This study showed an overall OPMD prevalence of 5.6%. The most common type of OPMD was leukoplakia (64.8%), followed by lichen planus (30.8%). Subjects who only smoked were found to have an increased risk for OPMD of almost four-fold (RR 3.74, 95%CI 1.89-7.41). The highest risk was found for betel quid chewers, where the increased risk observed was more than six times (RR 6.75, 95%CI 3.32-13.72). Alcohol consumption on its own did not seem to confer an increased risk for OPMD, however when practiced concurrently with smoking, a significant risk of more than five times was noted (RR 5.69 95%CI 3.14-10.29). CONCLUSION: The prevalence of OML was 54.1%, with linea alba being the most commonly occurring lesion. Smoking, alcohol consumption and betel quid chewing were found to be associated with the prevalence of OPMD, which was 5.6%.

Prognostic significance of VEGF-C in predicting micrometastasis and isolated tumour cells in N0 oral squamous cell carcinoma.

BACKGROUND: Lymph node metastasis in oral squamous cell carcinoma (OSCC) is a well-known independent prognostic factor. However, the identification of occult tumour cells within the lymph nodes has remained a challenge for the pathologist as well as the clinician. OBJECTIVE: The aim of this study was to determine the prevalence of micrometastasis and isolated tumour cells (ITCs) in pathologically staged N0 OSCC of the tongue and buccal mucosa and to assess its correlation with vascular endothelial growth factor C, (VEGF-C) expression in the primary tumour. METHODS: Thirty-four cases of N0 OSCC comprising of 17 cases each from the tongue and buccal mucosa were evaluated by immunohistochemistry for VEGF-C expression. The corresponding lymph nodes from levels I and II were pathologically examined and cross-detected for micrometastasis and ITCs with desmoglein 3 (DSG3). RESULTS: The prevalence of micrometastasis and ITCs in OSCC of the tongue and buccal mucosa was 23.5% and 17.6%, respectively. A total of 12 out of 151 lymph nodes contained micrometastatic tumour foci and ITCs. A higher expression of VEGF-C in the primary tumour was associated with a greater probability for the occurrence of micrometastasis and ITCs in the lymph nodes. CONCLUSION: High expression of VEGF-C in the primary tumour may be a good determinant for detection of occult tumour cells in the lymph nodes of OSCC cases.

Mobile Phone Imaging in Low Resource Settings for Early Detection of Oral Cancer and Concordance with Clinical Oral Examination.

INTRODUCTION: This study examined the concordance in clinical diagnosis of high-risk lesions in the oral cavity and referral decisions between clinical oral examination (COE) and teledentistry. MATERIALS AND METHODS: Sixteen individuals with a range of oral potentially malignant disorders (OPMD) and normal oral mucosa were included. Five areas of the oral cavity were photographed by three dentists using mobile phone cameras with 5 MP-13 MP resolutions. On the same day, the patients were given COE by two oral medicine specialists (OMS) and 3 weeks later, they reviewed the images taken using the phone, and concordance was examined between the two by Kappa statistics. The sensitivity and specificity of clinical diagnosis using the phone images were also measured. Pre- and post-program questionnaires were answered by both the dentists and the OMS to determine the feasibility of integrating teledentistry in their clinical practice. RESULTS: The Kappa values in determining the presence of lesion, category of lesion (OPMD or not), and making referral decision were moderate to strong (0.64-1.00). The overall sensitivity was more than 70% and specificity was 100%. The false negative rate decreased as the camera resolution increased. All dentists agreed that the process could facilitate early detection of oral mucosal lesion, and was easy to use in the clinic. CONCLUSIONS: This study provides evidence that teledentistry can be used for communication between primary care and OMS and could be readily integrated into clinical setting for patient management.

Caveolin 1 (Cav-1) and actin-related protein 2/3 complex, subunit 1B (ARPC1B) expressions as prognostic indicators for oral squamous cell carcinoma (OSCC).

Caveolin-1 (Cav-1) and Actin-Related Protein 2/3 Complex, Subunit 1B (ARPC1B) have been implicated in various human cancers, yet its role in tumorigenesis remains controversial. Therefore, this study aims to determine the protein expression of these two genes in oral squamous cell carcinomas (OSCCs) and to evaluate the clinical and prognostic impact of these genes in OSCC. Protein expressions of these two genes were determined by immunohistochemistry technique. The association between Cav-1 and ARPC1B with clinico-pathological parameters was evaluated by Chi-square test (or Fisher exact test where appropriate). Correlation between the protein expressions of these 2 genes with survival was analyzed using Kaplan-Meier and Cox regression models. Cav-1 and ARPC1B were found to be significantly over-expressed in OSCC compared to normal oral mucosa (p = 0.002 and p = 0.033, respectively). Low level of ARPC1B protein expression showed a significant correlation with lymph node metastasis (LNM) (p = 0.010) and advanced tumor staging (p = 0.003). Kaplan-Meier survival analyses demonstrated that patients with over-expression of Cav-1 protein were associated with poor prognosis (p = 0.030). Adjusted multivariate Cox regression model revealed that over-expression of Cav-1 remained as an independent significant prognostic factor for OSCC (HRR = 2.700, 95 % CI 1.013-7.198, p = 0.047). This study demonstrated that low-expression of ARPC1B is significantly associated with LNM and advanced tumor staging whereas high expression of Cav-1 can be a prognostic indicator for poor prognosis in OSCC patients.

Collagen Triple Helix Repeat Containing-1 (CTHRC1) Expression in Oral Squamous Cell Carcinoma (OSCC): Prognostic Value and Clinico-Pathological Implications.

BACKGROUND: Collagen Triple Helix Repeat Containing 1 (CTHRC1) is a protein often found to be over-expressed in various types of human cancers. However, correlation between CTHRC1 expression level with clinico-pathological characteristics and prognosis in oral cancer remains unclear. Therefore, this study aimed to determine mRNA and protein expression of CTHRC1 in oral squamous cell carcinoma (OSCC) and to evaluate the clinical and prognostic impact of CTHRC1 in OSCC. METHODS: In this study, mRNA and protein expression of CTHRC1 in OSCCs were determined by quantitative PCR and immunohistochemistry, respectively. The association between CTHRC1 and clinico-pathological parameters were evaluated by univariate and multivariate binary logistic regression analyses. Correlation between CTHRC1 protein expressions with survival were analysed using Kaplan-Meier and Cox regression models. RESULTS: Current study demonstrated CTHRC1 was significantly overexpressed at the mRNA level in OSCC. Univariate analyses indicated a high-expression of CTHRC1 that was significantly associated with advanced stage pTNM staging, tumour size ≥ 4 cm and positive lymph node metastasis (LNM). However, only positive LNM remained significant after adjusting with other confounder factors in multivariate logistic regression analyses. Kaplan-Meier survival analyses and Cox model demonstrated that patients with high-expression of CTHRC1 protein were associated with poor prognosis and is an independent prognostic factor in OSCC. CONCLUSION: This study indicated that over-expression of CTHRC1 potentially as an independent predictor for positive LNM and poor prognosis in OSCC.

Downregulation of CRNN gene and genomic instability at 1q21.3 in oral squamous cell carcinoma.

OBJECTIVES: This study includes the direct sequencing of cornulin (CRNN) gene to elucidate the possible mechanism of CRNN downregulation and explore the genetic imbalances at 1q21.3 across oral squamous cell carcinoma (OSCC) samples. MATERIALS AND METHODS: In mutation screening of CRNN gene, gDNA from OSCC tissues were extracted, amplified, and followed by direct sequencing. OSCC samples were also subjected to fragment analysis on CRNN gene to investigate its microsatellite instability (MSI) and loss of heterozygosity (LOH). Immunohistochemistry was performed to validate CRNN downregulation in OSCC samples. RESULTS: No pathogenic mutation was found in CRNN gene, while high frequency of allelic imbalances was found at 1q21.3 region. MSI was found more frequent (25.3 %) than LOH (9.3 %). Approximately 22.6 % of cases had high MSI which reflects higher probability of inactivation of DNA mismatch repair genes. MSI showed significant association with no betel quid chewing (p = 0.003) and tongue subsite (p = 0.026). LOH was associated with ethnicity (p = 0.008) and advanced staging (p = 0.039). The LOH at 1q21.3 was identified to be as an independent prognostic marker in OSCC (HRR = 7.15 (95 % CI, 1.41-36.25), p = 0.018). Downregulation of CRNN was found among MSI-positive OSCCs and was associated with poor prognosis (p = 0.044). CONCLUSION: This study showed a significant correlation between LOH/MSI at 1q21.3 with clinical outcomes and that downregulation of CRNN gene could be considered as a prognostic marker of OSCC. CLINICAL RELEVANCE: Insights of the downregulation mode of CRNN gene lays the basis of drug development on this gene as well as revealing its prognostic value.

Overexpression of MMP13 is associated with clinical outcomes and poor prognosis in oral squamous cell carcinoma.

Matrix metalloproteinase 13 (MMP13) plays a central role in the MMP activation cascade that enables degradation of the extracellular matrix and basement membranes, and it is identified as a potential driver in oral carcinogenesis. Therefore, this study aims to determine the copy number, mRNA, and protein expression of MMP13 in oral squamous cell carcinoma (OSCC) and to associate these expressions with clinicopathological parameters. Copy number, mRNA, and protein expression analysis of MMP13 were determined using real-time quantitative PCR and immunohistochemistry methods in OSCC samples. The correlations between MMP13 expressions and clinicopathological parameters were evaluated, and the significance of MMP13 as a prognostic factor was determined. Despite discrepancies between gene amplification and mRNA and protein overexpression rates, OSCC cases showed high amplification of MMP13 and overexpression of MMP13 at both mRNA and protein levels. High level of MMP13 protein expression showed a significant correlation with lymph node metastasis (P = 0.011) and tumor staging (P = 0.002). Multivariate Cox regression model analysis revealed that high level of mRNA and protein expression of MMP13 were significantly associated with poor prognosis (P < 0.050). Taken together, these observations indicate that the MMP13 protein overexpression could be considered as a prognostic marker of OSCC.

First isolation and identification of Cystobasidium calyptogenae from the oral samples of an elderly patient presenting with angular cheilitis.

BACKGROUND: Angular cheilitis, an infection mainly caused by Candida yeasts, is featured by the appearance of inflammatory lesions at the bilateral corners of the mouth, particularly in patients with poor oral hygiene, ill-fitting dentures and old age. The first isolation of an atypical yeast, Cystobasidium calyptogenae, from oral samples of a patient presenting with angular cheilitis is discussed in this study. CASE PRESENTATION: Angular cheilitis was diagnosed in a 60-year-old denture-wearing woman who presented with an irritation fibroma on her right lower buccal sulcus over the premolar region. Primary cultures of her oral swab and oral rinse samples grew a pure culture of an uncommon yeast strain resembling Rhodotorula sp. Sequence analysis of the yeast internal transcribed spacer (ITS) gene region and D1D2 domain showed highest similarity (99.6% and 100%, respectively) to C. calyptogenae CBS 9125 type strain. Following 2 weeks of treatment with miconazole/fusidic acid and mouthwash, the oral lesion showed improvement with less erythema. C. calyptogenae was not isolated from the patient's oral samples upon repeat sampling. CONCLUSION: This is the first report on the isolation of C. calyptogenae from human oral samples. The ability of C. calyptogenae to grow at 37 °C and the fact that it was the only yeast species isolated from the patient's oral samples suggests its pathogenic potential and possible involvement in angular cheilitis. The ubiquitous nature of the Cystobasidium yeast is believed to increase the likelihood of opportunistic infections among immunocompromised individuals. As Cystobasidium is phenotypically indistinguishable from Rhodotorula, an emerging opportunistic pathogen, surveillance using molecular identification in clinical settings is essential in providing accurate diagnosis and treatment of uncommon yeast infections.

Overexpression of gamma glutamyl hydrolase predicts extranodal extension in squamous cell carcinoma of the oral cavity.

OBJECTIVE: Extranodal extension (ENE) is an important prognostic factor in oral squamous cell carcinoma (OSCC). However, ENE is only confirmed postoperatively by histologic assessment of the lymph nodes after neck dissection. Accurate identification of ENE preoperatively would help in management of OSCC. STUDY DESIGN: We determined the expression of molecular markers gamma glutamyl hydrolase (GGH), cyclin-dependent kinase inhibitor-3 (CDKN3), and chromobox homolog-7 (CBX7) using immunohistochemistry in OSCC clinical samples (n = 35). The intensity of staining was scored using a semiquantitative index (HSCORE). The association between clinicopathologic parameters and expression of molecular markers with ENE status was analyzed using chi-square test. RESULTS: The number of positive nodes and the highest anatomic level of nodal involvement significantly correlated with ENE (P < .05). High GGH expression was significantly associated with ENE (P < .05), with an increased risk for ENE (odds ratio [OR] 9.9, 95% CI 1.08-91.47, P = .04), whereas no significant association was seen for CDKN3 and CBX7 expression with ENE. However, a trend toward significance was observed with a high level of CDKN3 and a low level of CBX7 expression with ENE. CONCLUSIONS: Gamma glutamyl hydrolase offers potential as a predictor for ENE in OSCC, whereas the role of CDKN3 and CBX7 need to be validated in a larger sample.

Transcriptional analysis highlights three distinct immune profiles of high-risk oral epithelial dysplasia.

Oral potentially malignant disorders (OPMD) are precursors of oral squamous cell carcinoma (OSCC), and the presence of oral epithelial dysplasia (OED) in OPMD confers an increased risk of malignant transformation. Emerging evidence has indicated a role for the immune system in OPMD disease progression; however, the underlying immune mechanisms remain elusive. In this study, we used immune signatures established from cancer to delineate the immune profiles of moderate and severe OED, which are considered high-risk OPMD. We demonstrated that moderate and severe OEDs exhibit high lymphocyte infiltration and upregulation of genes involved in both immune surveillance (major histocompatibility complex-I, T cells, B cells and cytolytic activity) and immune suppression (immune checkpoints, T regulatory cells, and tumor-associated macrophages). Notably, we identified three distinct subtypes of moderate and severe OED: immune cytotoxic, non-cytotoxic and non-immune reactive. Active immune surveillance is present in the immune cytotoxic subtype, whereas the non-cytotoxic subtype lacks CD8 immune cytotoxic response. The non-immune reactive subtype showed upregulation of genes involved in the stromal microenvironment and cell cycle. The lack of T cell infiltration and activation in the non-immune reactive subtype is due to the dysregulation of CTNNB1, PTEN and JAK2. This work suggests that moderate and severe OED that harbor the non-cytotoxic or non-immune reactive subtype are likely to progress to cancer. Overall, we showed that distinct immune responses are present in high-risk OPMD, and revealed targetable pathways that could lead to potential new approaches for non-surgical management of OED.

Barriers to early detection and management of oral cancer in the Asia Pacific region.

OBJECTIVE: Oral cancer is amenable to early detection but remains a prominent cause of mortality in the Asia Pacific region. This study aimed to identify barriers to early detection and management of oral cancer in the Asia Pacific region. METHODS: A mixed-methods approach was employed triangulating findings from a survey and focus groups. The survey was conducted among seven representative members of the Asia Pacific Oral Cancer Network (APOCNET) across six countries. Focus groups were conducted to gain deeper insights into the findings of the survey. RESULTS: The identified barriers were a lack of national cancer control strategies and cancer registries and the limited availability of trained health care professionals. Overcoming these challenges in the Asia Pacific region where resources are scarce will require collaborative partnerships in data collection and novel approaches for continuous professional training including eLearning. Further, to overcome the lack of trained health care professionals, innovative approaches to the management of oral potentially malignant lesions and oral cancer including telemedicine were suggested. CONCLUSION: The findings of this study should be taken into account when charting national cancer control plans for oral cancer and will form the basis for future collaborative studies in evaluating effective measures to improve oral cancer detection and management in low- and middle-income countries.

Expression of Ki-67, Cornulin and ISG15 in non-involved mucosal surgical margins as predictive markers for relapse in oral squamous cell carcinoma (OSCC).

BACKGROUND: Local relapse of oral squamous cell carcinoma in non-involved mucosal surgical margins indicated possibility of field alteration in the margins, which could be predicted with certain biomarkers. The objectives were to evaluate the expression of Ki-67, Cornulin and ISG15 in non-involved mucosal surgical margins and the association of clinicopathological prognosticators with local relapse in oral squamous cell carcinoma. METHODS: Surgical margins from the study (relapse) group (n = 23), control (non-relapse) group (n = 32) and normal oral mucosa (n = 5) were immunohistochemically stained using Ki-67, Cornulin and ISG15 antibodies. Association between expression of markers and clinicopathological prognosticators with local relapse in oral squamous cell carcinoma was analyzed statistically. RESULTS: The study group surgical margins demonstrated significantly decreased Cornulin expression (p = 0.032). Low Cornulin expression was significantly associated with local relapse (p = 0.004) and non-tongue primary tumor (p = 0.013). Although not significantly associated with local relapse, expression of Ki-67 was significantly reduced in female patients (p = 0.041). Age above 57.5 years, Chinese & Indian ethnicity, alcohol consumption, epithelial dysplasia in surgical margins, and type III and IV patterns of invasion of tumor were also significantly related to local relapse. Regression analysis showed low expression of Cornulin (p = 0.018), and increased patient's age (p = 0.008) were predictors of local relapse in oral squamous cell carcinoma, with 34-fold risk and 18-fold risk, respectively. Expression of Ki-67 and ISG15 did not show significant association with local relapse in oral squamous cell carcinoma. CONCLUSION: Low expression of Cornulin is an independent predictor of relapse in oral squamous cell carcinoma.

DNA Vaccines Targeting Novel Cancer-Associated Antigens Frequently Expressed in Head and Neck Cancer Enhance the Efficacy of Checkpoint Inhibitor.

HPV-independent head and neck squamous cell carcinoma (HNSCC) is a common cancer globally. The overall response rate to anti-PD1 checkpoint inhibitors (CPIs) in HNSCC is ~16%. One major factor influencing the effectiveness of CPI is the level of tumor infiltrating T cells (TILs). Converting TILlow tumors to TILhigh tumors is thus critical to improve clinical outcome. Here we describe a novel DNA vaccines to facilitate the T-cell infiltration and control tumor growth. We evaluated the expression of target antigens and their respective immunogenicity in HNSCC patients. The efficacy of DNA vaccines targeting two novel antigens were evaluated with or without CPI using a syngeneic model. Most HNSCC patients (43/44) co-expressed MAGED4B and FJX1 and their respective tetramer-specific T cells were in the range of 0.06-0.12%. In a preclinical model, antigen-specific T cells were induced by DNA vaccines and increased T cell infiltration into the tumor, but not MDSC or regulatory T cells. The vaccines inhibited tumor growth and improved the outcome alone and upon combination with anti-PD1 and resulted in tumor clearance in approximately 75% of mice. Pre-existence of MAGED4B and FJX1-reactive T cells in HNSCC patients suggests that these widely expressed antigens are highly immunogenic and could be further expanded by vaccination. The DNA vaccines targeting these antigens induced robust T cell responses and with the anti-PD1 antibody conferring excellent tumor control. This opens up an opportunity for combination immunotherapy that might benefit a wider population of HNSCC patients in an antigen-specific manner.