Oral Abstract Presentations #1- 5/1/2017 - 8:00 Am in Vivo Antinuclear Antibody Positivity of Oral Epithelium May Indicate Systemic Connective Tissue Disease

Objective:To determine and compare cytomorphometric changes in buccal mucosal cells of cigarette smokers, naswar users and non-users/smokers. Materials and methods:Cellular diameter CD, nuclear diameter ND and nuclear to cytoplasmic ratio N/C ratio were assessed in buccal smears taken with wooden spatula from clinically normal mucosa of smokers,naswar users and control group. The sample size was 99 subjects of ages 15 yrs-60yrs, divided into three groups (33 each group) as M, S and N i.e control, smokers and naswar users respectively. Slides were stained with three stains Hematoxylin and Eosin Stain, Giemsa Stain and Papanicolaou Stain. Results:The cytomorphometric variables were measured by using stage and ocular micrometers. The mean cellular diameter of group M, S and N was 43.8μm, 54.3μm and 42.7μm respectively. The mean nuclear diameter of M, S and N was 9.97μm, 12.6μm and 11.8μm respectively. And the mean N/C ratio of group M, S and N was 1:4.4, 1:4.3and 1:3.5 respectively. The mean differences between CD, ND and N/C ratio in all three stains, among the three groups, S, N and M was found to be statistically highly significant i.e p = 0.001 on ONE WAY ANOVA. While, on Post hoc tukey test between S and N, CD and N/C ratio were highly significant p=0.000 while ND was not significant. While between S and M, CD and ND both were significant and showed p=0.000 while N/C ratio was not significant.In comparison between N and M, ND and N/C ratio were found to be significant while CD was not significant.Conclusion:The cytomorphometric changes assessed by this study depicts only cause effect relationship with smoking and naswar use. Association of these changes with dysplasia or pre-malignancy needs further verification with the help of specific immune-markers. #135/1/2017 10:24 AM LOSS OF CALPROTECTIN (S100A8/A9) ASSOCIATES WITH POOR EPITHELIAL DIFFERENTIATION AND INCREASED EGFR EXPRESSION IN HEAD AND NECK SQUAMOUS CELL CARCINOMA (HNSCC). P. Argyris, I. Koutlas, D. Kademani, Z. Slama, K. Patel, B. Pakzad, M. C. Herzberg, U of Minnesota and North Memorial Hospital, Minnesota Background: Calprotectin, a heterodimeric complex of the calcium-binding proteins S100A8 and S100A9 (S100A8/A9) encoded on chromosome 1q21, activates the G2/M cell cycle checkpoint, inhibits carcinoma cell migration and invasion, and suppresses tumorigenesis in vitro and in vivo. Calprotectin mRNA and protein expression decreases in poorly differentiated HNSCC and is associated with lower survival rates. Signaling through EGFR is also considered a negative prognosticator for HNSCC. Aims: 1) To investigate changes in S100A8/A9 expression during the continuum of oral carcinogenesis and 2) address whether EGFR expression is affected by S100A8/A9 status. Materials and methods: FFPE sections of 34 HNSCCs including 16 well-differentiated (WD), 8 moderatelydifferentiated (MD), 4 poorly-differentiated (PD) and 6 basaloid HNSCCs, and 15 premalignant epithelial dysplasias were immunohistochemically stained for S100A8 and S100A9. EGFR immunoexpression was studied in 17 HNSCCs. Results: Strong cytoplasmic and nuclear S100A8/A9 staining was seen throughout normal oral mucosal epithelium (NE) except for the basal cell layer which was consistently S100A8/A9 negative. Dysplastic lesions showed significantly less calprotectin expression than NE. WD HNSCCs strongly expressed calprotectin, while S100A8/A9 was progressively lost in MD, PD and basaloid tumors. EGFR expression was limited in S100A8/A9-high HNSCCs. Conversely, S100A8/A9low carcinomas appeared to upregulate EGFR. Conclusions: In HNSCC, S100A8/A9 expression positively associates with the level of squamous differentiation of malignant cells and inversely correlates with EGFR expression. S100A8/A9-associated down-regulation of EGFR could explain better survival of patients with S100A8/A9-high tumors. #145/1/2017 10:36 AM COMPOSITION OF IMMUNE CELL AGGREGATES AT THE INVASION FRONTS OF ORAL CANCER ASSOCIATED WITH CLINICAL OUTCOMES C. Poh, M. Lopes, K. Liu, Y. Liu, C. Nelson, U. of Victoria, Victoria, U. of British Columbia, Vancouver, Federal U. of Rio Grande do Norte, Natal and U. of Electronic Science and Technology, China, OBJECTIVES: Evading immune destruction from tumor microenvironment has been proposed as one of the hallmarks of cancer. However, little is known in oral cancer (OC). The objective is to investigate the immune cellular composition of invasion fronts (IFs) of OC and associated outcome. METHODS: As an exploratory study, two 5-μm serial sections from the cross-section of 12 highly annotated OC samples were used for 2 sets of multi-color immunostaining with antibodies against: CD8, CD163, FoxP3, CD25; and PD-L1, PD-1, CD3, CD20 (Deeley Research Lab, Victoria, BC). RESULTS: PD-L1 tumor-cell expression was heterogeneous in 9 OCs, especially at the IFs. PD-L1tumors had a better outcome with no nodal disease. In addition, PD-L1+ immune cells with dendritic morphology were seen in all cases. In areas with interconnecting clusters of PDL1+ immune cells we observed different proportions of CD25+ immune cells and CD8+-cells co-localizing within T-cell clusters. We did not observe a relationship between the number of CD8+-cells and the outcome; however, clusters of PD-1+ immune cells were commonly seen in patients with poor outcomes. Aggregates containing both expanded B-(CD20) and T-(CD3) cells at the IFs seemed to have a better outcome. CD163 was not exclusively expressed in macrophages. Various patterns of CD25 expression were also found at the IFs of the tumor cells of 11 OCs, including membranous, cytoplasmic, and dot-like. The latter was more evident in poorly differentiated tumors, while well-differentiated tumor nests expressed little to none CD25. CONCLSION: Immune cellular composition at TME, either pro-inflammatory or immune suppressive, are associated with clinical consequences. Further investigation is warranted to shed light of the underlying biology. #155/1/2017 10:48 AM DOWNREGULATION OF DMBT1 PROMOTES INVASION IN HEAD AND NECK CANCER. S. Piao, S. Piao, R. Banerjee, M. Liu, R.C. Inglehart, N. D'Silva, U. of Michigan, Ann Arbor Deleted in malignant brain tumors 1 (DMBT1) is a tumor suppressor that is downregulated in multiple cancers. Objective: In the present study, we investigated the expression, function and mechanism of regulation of DMBT1 in human head and neck cancer. Methods: Using laser capture microdissection we isolated epithelium from head and neck cancer and normal tissue and used quantitative RTPCR to quantify DMBT1 transcripts. The regulation of DMBT1 was investigated by genetic and biochemical approaches in human head and neck cancer cell lines. The function of DMBT1 was investigated using in vitro and in vivo approaches. Results: DMBT1 is downregulated in head and neck cancer compared to normal epithelium. DMBT1 expression is inversely correlated with EZH2, an oncogene that promotes invasion in head and neck cancer. Furthermore, EZH2 silences DMBT1 via histone and promoter methylation. In head and neck cancer cell lines with stable downregulation of EZH2, suppression of DMBT1 rescues the invasive phenotype. Conclusions: DMBT1 is a tumor suppressor that inhibits invasion and is silenced in HNC. (This work was supported by NIDCR grants DE022567 and