Diego Antonio Costa Arantes

Immunosuppressive mediators of oral squamous cell carcinoma in tumour samples and saliva

The goal of this study was to compare the salivary concentrations of IL-10, TGF-β1 and soluble HLA-G (sHLA-G) in patients with oral squamous cell carcinoma (OSCC) to those in healthy individuals (control group), and to correlate the expression of these mediators in saliva with that in the tumour microenvironment. Neoplastic tissue and saliva samples from patients with OSCC (n=22) were analysed by immunohistochemistry and enzyme-linked immunosorbent assay (ELISA) respectively. We detected high expression of IL-10 and HLA-G in the tumour microenvironment when compared to healthy oral mucosa samples. Determination of IL-10 salivary concentration enabled us to distinguish patients with OSCC from healthy individuals (P=0.038), which showed correlation with tissue expression of this cytokine. HLA-G salivary release was similar in both groups (P=0.17) and no correlation with tumour expression was observed. TGF-β1 expression was low or absent in tumours, and salivary concentration was similar between groups. Our results suggest that of the three markers analysed, IL-10 is a potential salivary biomarker. Furthermore, the elevated expression of HLA-G and IL-10 in tumour sites could favour the escape of tumour cells from immune defense mechanisms.

Overexpression of immunosuppressive cytokines is associated with poorer clinical stage of oral squamous cell carcinoma

OBJECTIVE The aim of this study was to evaluate the expression of IL-10 and TGF-β2 in oral squamous cell carcinoma (OSCC) and its relationship with prognostic clinical and microscopic parameters. DESIGN Immunohistochemistry was used to assess the expression of IL-10 and TGF-β2 in OSCC samples from 43 patients who had undergone surgical excision and neck dissection. Metastatic lymph nodes were included in the study (n=23). Samples of healthy oral mucosa (n=20) were used as controls. The sections were evaluated using a semi-quantitative method in conjunction with staining intensity. RESULTS Our findings showed that the expression of IL-10 and TGF-β2 by neoplastic and stromal cells was high in most of the OSCC samples (>70% of samples), especially when compared to the controls (≅10% of samples) (P<0.05). OSCC neoplastic cells in cervical lymph nodes were also positive for IL-10 and TGF-β2. An association between high expression of IL-10 by neoplastic cells and advanced clinical stage (T3-T4) was verified (P=0.02). Although not statistically significant, the expression of TGF-β2 was also augmented in advanced stage tumours. CONCLUSIONS These data suggest that the ability of OSCC neoplastic cells to secrete immunosuppressive cytokines could contribute to clinical progression by maintaining a microenvironment conducive to evasion and tumour proliferation.

Characterization of dendritic cells in lip and oral cavity squamous cell carcinoma.

BACKGROUND There may be differences in the antitumor immunity induced by dendritic cells (DCs) during the development of squamous cell carcinoma (SCC) located in the lip rather than in the oral cavity. The aim of this study was to evaluate the number of immature and mature DCs in SCC and potentially malignant disorders of the oral cavity and lip. METHODS Immunohistochemistry was used to identify the number (cells/mm(2) ) of immature (CD1a(+) ) or mature (CD83(+) ) DCs in samples of oral cavity SCC (OCSCC) (n = 39), lip SCC (LSCC) (n = 23), leukoplakia (LK) (n = 21), actinic cheilitis (AC) (n = 13), and normal mucosa of the oral cavity (OC control, n = 12) and the lip (lip control, n = 11). RESULTS The number of CD1a(+) cells tended to be higher in the OC control samples compared with the LK (P = 0.04) and OCSCC (P = 0.21). Unlike, this cell population was lower in the lip control than in AC or LSCC (P < 0.05). The number of CD83(+) cells was increased in the LSCC samples compared with the AC and lip control (P = 0.0001) and in OCSCC compared with both the LK (P = 0.001) and OC control (P = 0.0001) samples. LSCC showed an elevated number of CD1a(+) and CD83(+) cells compared with OCSCC (P = 0.03). The population of mature DCs was lower than the population of immature DCs in all of the tested groups (P < 0.05). CONCLUSION There were a greater number of both mature and immature DC populations in the LSCC samples than in the OCSCC, which could contribute to establishing a more effective immune antitumor response for this neoplasm.

Immune response in cervical lymph nodes from patients with primary oral squamous cell carcinoma.

BACKGROUND Deficient immune response in the cervical lymph nodes of patients with head and neck squamous cell carcinoma may contribute to dissemination of metastatic neoplastic cells. This study evaluates the immune response in cervical lymph nodes from patients with primary oral cavity squamous cell carcinoma (OCSCC). METHODS The density of immature (CD1a(+)) and mature (CD83(+)) dendritic cells (DCs), cytotoxic T lymphocytes CD8(+) /perforin(+) (CTLs), and Foxp3(+) regulatory T (Tregs) cells in the lymph nodes of patients with OCSCC without cervical lymph node metastases (LN1) (negative) (n = 10) were identified through immunohistochemistry. From patients with cervical lymph node metastases, samples were obtained of lymph nodes both with (LM2) (positive) (n = 10) and without (LN2) (negative) (n = 10) metastases. RESULTS The results demonstrated that the number of CD1a(+) and Foxp3(+) cells was significantly higher in the LM2 group than in either the LN1 or the LN2 group. In addition, the number of CD8(+) /perforin(+) and CD83(+) cells was significantly lower in the LM2 group than in the other groups. CONCLUSION The results of this study demonstrate a higher density of immature DCs and Tregs cells and a lower density of mature DCs and activated CTLs in metastatic than in non-metastatic lymph nodes. These findings might indicate an immunosuppressive microenvironment, which could be involved in the spread of neoplastic cells to cervical lymph nodes.

Calcifying epithelial odontogenic tumours: Collaborative study of 32 cases and review of literature

OBJECTIVES To analyse the occurrence of calcifying epithelial odontogenic tumours (CEOT) based on biopsy records from different Brazilian geographic regions and to contrast the data with a review of the literature. MATERIALS AND METHODS A 2-step study was conducted. Step 1 consisted of a collaborative study of biopsies obtained from 1953 to 2017 at six Brazilian oral and maxillofacial pathology centres. Evaluation of 86,268 biopsy records was performed. Demographic and histopathological diagnosis data were assessed. In Step 2, a review of the literature of case reports and cases series of CEOT identified across five electronic databases was conducted. RESULTS In the collaborative study, 32 cases of CEOT were evaluated. This figure represented 0.03% of the oral and maxillofacial lesions and 1.7% of all odontogenic tumours across the centres. Women in the fourth decade of life were more affected. CEOT occurred more in the mandible than in the maxilla (ratio 1.9:1). The review of the literature showed that Asian individuals were more affected by this neoplasm. CONCLUSIONS Useful knowledge on the epidemiology, treatment and follow-up of CEOT has been provided. Demographic data and clinical features of the cases presented in this collaborative study were quite similar to those of studies reported worldwide.

Randomized clinical trial of a mucoadhesive formulation containing curcuminoids (Zingiberaceae) and Bidens pilosa Linn (Asteraceae) extract (FITOPROT) for prevention and treatment of oral mucositis - phase I study

Preclinical repeated-dose toxicity and efficiency studies developed by our group suggest the potential of FITOPROT in treating mucositis. This serious limiting side effect is observed at a rate of 40-100% in patients under antineoplastic therapy and despite different palliative measures and therapeutic agents have been investigated, still no therapy was completely successful. Therefore, this study aimed to establish the safety and recommended phase II dose of FITOPROT for the prevention and treatment of chemoradiotherapy-induced oral mucositis (OM) in patients with head and neck cancer. Twenty healthy adult participants were randomized into two groups that received pre-established concentrations of the collutory: group 1 (FITOPROT A - mucoadhesive formulation containing 10 mg/mL of curcuminoids extract plus 20% v/v of Bidens pilosa L. extract) and group 2 (FITOPROT B - mucoadhesive formulation containing 20 mg/mL of curcuminoids extract, plus 40% v/v of Bidens pilosa L. extract). Participants rinsed their mouths with FITOPROT, three times daily, for ten consecutive days. No participant experienced toxicity or unacceptable discomfort and/or adverse reactions (CTCAE v5.0), with laboratory and clinical parameters under normal conditions. Side effects observed were low intensity and temporary mucosa/dental surface pigmentation (n = 7) and tooth sensitivity (n = 4), which disappeared after formulation use ceased. No significant cellular genotoxic effects were observed (p > 0.05), and micronuclei frequencies were not changed (p > 0.05). Biochemical assays reveled no altered levels of myeloperoxidase (p = 0.2268), malondialdehyde (p = 0.1188) nor nitric oxide (p = 0.5709) concentration, and no significant difference were found in the levels of pro-inflammatory cytokines (p > 0.05). Thus, FITOPROT demonstrated to be safe and tolerable in both tested doses and is suitable for evaluation in a phase II trial as treatment against OM.

Immunohistochemical investigations on the expression of programmed cell death ligand 1, human leukocyte antigens G and E, and granzyme B in intraoral mucoepidermoid carcinoma

OBJECTIVE To identify the expression of nonclassical human leukocyte antigen G and E (HLA-G and -E), programmed cell death ligand-1 (PD-L1) and granzyme B (GB) in intraoral mucoepidermoid carcinomas (MECs), and to assess whether such expressions are related to metastasis, survival, staging, tumor grade and number of GB-positive cells. DESIGN For this cross-sectional study, samples of MEC (n=30) were selected and classified as low-grade (LG), intermediate-grade (IG) or high-grade (HG), according to the WHO grading system. HLA-G, -E and PD-L1 were identified by immunohistochemistry and quantified as the proportion of positive neoplastic cells. The density of GB+ cells was also evaluated. The Kruskal-Wallis test was used with a 5% significance level. RESULTS Expressions of HLA-G, -E and PD-L1 were identified in the majority of epidermoid, intermediate and clear cells, but not in the mucous cells of the MECs. The quantitative analysis of the total percentage of positive neoplastic cells showed overexpression of this set of proteins in all MEC samples. The expression of these proteins and histological grading were positively correlated [HLA-G (LG=79% positive cells, IG=96%, HG=99%; p=0.0004), HLA-E (LG=70%, IG=96%, HG=99%; p<0.0001) and PD-L1 (LG=34%, IG=79%, HG=80%; p=0.01)]. No relationship was observed between the immunosuppressive proteins and other clinicopathological parameters. Low GB density was found in all MEC samples. CONCLUSIONS The augmented expression of HLA-G, -E and PD-L1 in the intraoral MEC might suggest a role of these molecules in the scape of neoplastic cells from immunosurveillance.