Mapping Conformational Changes in the Saliva Proteome Potentially Associated with Oral Cancer Aggressiveness.

Diverse proteomics-based strategies have been applied to saliva to quantitatively identify diagnostic and prognostic targets for oral cancer. Considering that these targets may be regulated by events that do not imply variation in protein abundance levels, we hypothesized that changes in protein conformation can be associated with diagnosis and prognosis, revealing biological processes and novel targets of clinical relevance. For this, we employed limited proteolysis-mass spectrometry in saliva samples to explore structural alterations, comparing the proteome of healthy control and oral squamous cell carcinoma (OSCC) patients with and without lymph node metastasis. Thirty-six proteins with potential structural rearrangements were associated with clinical patient features including transketolase and its interacting partners. Moreover, N-glycosylated peptides contribute to structural rearrangements of potential diagnostic and prognostic markers. Altogether, this approach utilizes saliva proteins to search for targets for diagnosing and prognosing oral cancer and can guide the discovery of potential regulated sites beyond protein-level abundance.

Adverse post-operative events of salivary gland biopsies: A systematic review and meta-analysis.

OBJECTIVES: The study aimed to perform a systematic review and meta-analysis of the complications following major and minor salivary gland biopsy. MATERIALS AND METHODS: Observational studies assessing postoperative complications of minor salivary gland biopsy and indexed at Medline/PubMed, EMBASE, Cinahl, LILACS, or Scopus were selected. This review was registered under the protocol number: CRD42020211169. The level of significance considered was 0.05, and the R software (The R Foundation) was used for the meta-analysis. RESULTS: Twenty-seven studies reporting 3208 patients were included in this review. The combined prevalence of postsurgical complications was 11% (95% CI, 8 to 13%, p = 0.01). The percentage of the combined prevalence of neurological complications was 3% (95% CI, 1-6%, p = 0.01). The surgical technique did not influence the frequency of overall and neurological complications. CONCLUSION: Minor salivary gland biopsies are a safe and predictable procedure that should be performed on the lower lip. Postoperative complications are more common than previously reported, but permanent complaints are uncommon.

Semaphorins and neuropilins expression in salivary gland tumors.

BACKGROUND: Salivary gland tumors (SGT) account for 3-10% of all head and neck neoplasms, and little is known about their angiogenic properties. Despite semaphorins and neuropilins have been demonstrated to be prognostic determinants in many human cancers, they remain to be investigated in SGT. Therefore, the objective of this study was to analyze the clinical significance of the expression of class 3 semaphorins A (Sema3A) and B (Sema3B) and neuropilins-1 (Np-1) and neuropilins-2 (Np-2), in SGT. METHODS: Two hundred and forty-eight SGT were organized in tissue microarray paraffin blocks and expression of CD34, Sema3A, Sema3B, Np-1, and Np-2 was determined through immunohistochemistry. The immunoreactions were quantified using digital algorithms and the results correlated with clinicopathological parameters. RESULTS: Malignant tumors had an increased vascular density than their benign counterparts and their increased vascular area significantly correlated with recurrences (P < 0.05). Patients older than 40 years and the presence of recurrences determined an inferior survival rate (P = 0.0057 and P = 0.0303, respectively). In normal salivary glands, Np-1 and Np-2 expression was restricted to ductal cells, whereas Sema3A and Sema3B were positive in the serous acinar compartment. Tumors were positive for all markers and the co-expression of Np-1/Np-2 significantly correlated with the presence of paresthesia and advanced stages of the tumors (P = 0.01 and P = 0.04, respectively). CONCLUSION: Sema3A, Sema3B, Np-1, and Np-2 may be involved in the pathogenesis of SGT, but their expression did not present a statistically significant prognostic potential in this study.

High incidence of DNA ploidy abnormalities and increased Mcm2 expression may predict malignant change in oral proliferative verrucous leukoplakia.

AIMS: To assess the DNA content of cases of oral proliferative verrucous leukoplakia (PVL) and correlate the DNA ploidy findings with the expression of Mcm2, geminin, and Ki67, and with clinicopathological data. METHODS AND RESULTS: DNA quantification was performed by image cytometry using the ACIS III Automated Cellular Imaging System. Expression of Ki67, Mcm2 and geminin was determined by immunohistochemistry. There were 21 cases of PVL, the female/male ratio was 6:1, and the average age was 65.5 years. Seventeen patients (81.0%) did not report tobacco and alcohol consumption. Nine patients (42.9%) developed verrucous or squamous cell carcinoma. Levels of Mcm2 expression showed a positive correlation with increasingly severe epithelial changes (P = 0.03). Twenty patients had their DNA examined by ACIS III, and 19 (95%) showed aneuploidy. The frequency and severity of aneuploidy (P < 0.0001), the mean values of the DNA heterogeneity index (P < 0.0001) and the 5n-exceeding fractions (P = 0.0007) increased according to epithelial alterations. Abnormal DNA content was observed even in the more indolent lesions. CONCLUSIONS: Mcm2 expression and DNA ploidy analysis could be used to predict areas of malignant transformation. The clinicopathological findings associated with the immunohistochemical and DNA ploidy results support the distinct and aggressive profile of this entity.

Clinical and Psychosocial Impact of Communication about Oral Potentially Malignant Disorders: A Scoping Review.

Delivering bad news has been widely studied in cancer, thus, this scoping review aims to identify the available evidence concerning the communication of oral potentially malignant disorders (OPMDs) and their clinical and psychosocial impacts. A search was performed using electronic databases (Medline/PubMed, Scopus, Embase, and Web of Science) and one grey literature database (Google Scholar). Studies focused on communicating the diagnosis of OPMDs and the patients' perceptions were included. Study selection and data extraction were performed by two authors in a two-phase process. Five publications were included in the qualitative analysis. Differences regarding the study design, population, OPMDs assessed, and outcomes of professional-patient communication were found in each study. Protocols for OPMD communication have not yet been reported and there is a need to standardize strategies as communication skills may provide better clinical outcomes for patients diagnosed with potentially malignant disorders. Although future studies are needed, a brief list recommending the aspects that must be communicated is proposed.

Expression pattern of PLUNC proteins as an auxiliary tool for the diagnosis of high-grade mucoepidermoid carcinoma of the salivary gland.

BACKGROUND: Mucoepidermoid carcinomas are the most frequent malignant neoplasia of the salivary glands and are histologically classified as low, intermediate, and high grade. At present, histochemical stains such as periodic acid-Schiff or mucicarmine are useful tools in making a diagnosis. Recently, expression of the PLUNC proteins has been described in mucin-producing salivary gland tumors, with the suggestion that they could provide a powerful tool for the diagnosis of difficult cases. METHODS: This study evaluates the expression of PLUNC proteins in 30 cases of salivary gland mucoepidermoid carcinomas. Tumors were reviewed and classified according to histological grade. Periodic acid-Schiff, mucicarmine, and immunohistochemical staining for SPLUNC1, LPLUNC1, SPLUNC2, and LPLUNC2 were carried out. Immunostaining was classified as positive or negative. RESULTS: The majority of the tumors (63%) were classified as low grade, 13% were intermediate grade, and 23% were high grade. SPLUNC1 (90%) and LPLUNC1 (93%) were positive in the majority of cases, mainly in mucous cells, mucin plugs, and intermediate cells. SPLUNC2 and LPLUNC2 did not present significative expression within the tumors; however, LPLUNC2 was found to stain positively in mast cells in 83% of the samples. CONCLUSIONS: SPLUNC1 and LPLUNC1 showed a similar pattern of expression and could prove useful in the diagnosis of high-grade cases because of the differential staining in intermediate and epidermoid cells. The expression of LPLUNC2 in mast cells has not previously been reported, but further studies are necessary to validate this finding and to determine its significance.

The Role of Immunohistochemistry for Primary Oral Diagnosis in a Brazilian Oral Pathology Service.

A proper antibody panel selection is one of the most important factors to reach an adequate diagnosis in challenging cases. This retrospective study was designed to determine the contribution of immunohistochemistry (IHC) in the primary diagnosis of oral diseases in one of the main services of oral pathology in the State of São Paulo, Brazil, and to identify the most common antibodies used, and recommend diagnostic algorithms based on our experience with challenging lesions. A total of 1698 IHC stains were performed in 401 cases from a total of 28,804 cases received from public dental clinics and private dental practitioners within a period of 13 years, representing a frequency of 1.4% of IHC solicitations. Among these, 112 (28%) were mandatory to reach a final diagnosis and 255 (63.6%) were confirmative. In 34 (8.4%) cases, it was not possible to reach a conclusive/final diagnosis, even with IHC. Regarding the nature of the lesions, 210 (52.3%) were benign, 163 (40.6%) were malignant tumors, 13 (3.2%) were reactive, 10 (2.5%) were premalignant, and 5 (1.2%) were lesions of uncertain malignancy. Small amount of tissue of some incisional biopsies, overlapping features of spindle cell lesions (epithelial, neural, melanocytic, smooth muscle, endothelial, and fibroblastic/myofibroblastic cell differentiation), and overlapping features of salivary gland lesions were the most frequent challenges in which IHC stains were requested. Spindle cell lesions were the most frequent (22%) among all cases that required IHC to reach a final diagnosis. The implementation of IHC for routine practice requires a wide range of markers, proper antibody selection, and knowledge to interpret the subjectivity of staining. The inherent limitation of incisional biopsies was pointed as a reason to inconclusive diagnosis, despite a wide range of antibodies that our laboratory displays.

Meta-omics analysis indicates the saliva microbiome and its proteins associated with the prognosis of oral cancer patients.

Saliva is a biofluid that maintains the health of oral tissues and the homeostasis of oral microbiota. Studies have demonstrated that Oral squamous cell carcinoma (OSCC) patients have different salivary microbiota than healthy individuals. However, the relationship between these microbial differences and clinicopathological outcomes is still far from conclusive. Herein, we investigate the capability of using metagenomic and metaproteomic saliva profiles to distinguish between Control (C), OSCC without active lesion (L0), and OSCC with active lesion (L1) patients. The results show that there are significantly distinct taxonomies and functional changes in L1 patients compared to C and L0 patients, suggesting compositional modulation of the oral microbiome, as the relative abundances of Centipeda, Veillonella, and Gemella suggested by metagenomics are correlated with tumor size, clinical stage, and active lesion. Metagenomics results also demonstrated that poor overall patient survival is associated with a higher relative abundance of Stenophotromonas, Staphylococcus, Centipeda, Selenomonas, Alloscordovia, and Acitenobacter. Finally, compositional and functional differences in the saliva content by metaproteomics analysis can distinguish healthy individuals from OSCC patients. In summary, our study suggests that oral microbiota and their protein abundance have potential diagnosis and prognosis value for oral cancer patients. Further studies are necessary to understand the role of uniquely detected metaproteins in the microbiota of healthy and OSCC patients as well as the crosstalk between saliva host proteins and the oral microbiome present in OSCC.

Oral radiation-induced sarcomas: Systematic review.

BACKGROUND: The aim of this study was to integrate the available data published on radiation-induced sarcoma of the oral cavity into an analysis of its clinical features, treatment modalities and prognostic factors. METHODS: An electronic search was undertaken in September 2019. The eligibility criteria included publications that had enough clinical and histological information to confirm the diagnosis. RESULTS: Forty-two publications with 122 radiation-induced sarcoma of the oral cavities (RISOCs) were included. The mean latency period was 114 months and mean radiation total dose was 62.5 Gy. The tumors were more prevalent in males between 50 and 60 years old and the mandible was the most affected site. Osteosarcoma was the most prevalent histological type and patients were mostly treated with radical surgery. CONCLUSIONS: RISOC showed a poor survival rate of 15.1% in 5-year follow-up. Gender and histological type were independently associated with survival.

Integrative analysis to select cancer candidate biomarkers to targeted validation.

Targeted proteomics has flourished as the method of choice for prospecting for and validating potential candidate biomarkers in many diseases. However, challenges still remain due to the lack of standardized routines that can prioritize a limited number of proteins to be further validated in human samples. To help researchers identify candidate biomarkers that best characterize their samples under study, a well-designed integrative analysis pipeline, comprising MS-based discovery, feature selection methods, clustering techniques, bioinformatic analyses and targeted approaches was performed using discovery-based proteomic data from the secretomes of three classes of human cell lines (carcinoma, melanoma and non-cancerous). Three feature selection algorithms, namely, Beta-binomial, Nearest Shrunken Centroids (NSC), and Support Vector Machine-Recursive Features Elimination (SVM-RFE), indicated a panel of 137 candidate biomarkers for carcinoma and 271 for melanoma, which were differentially abundant between the tumor classes. We further tested the strength of the pipeline in selecting candidate biomarkers by immunoblotting, human tissue microarrays, label-free targeted MS and functional experiments. In conclusion, the proposed integrative analysis was able to pre-qualify and prioritize candidate biomarkers from discovery-based proteomics to targeted MS.

Agrin and perlecan mediate tumorigenic processes in oral squamous cell carcinoma.

Oral squamous cell carcinoma is the most common type of cancer in the oral cavity, representing more than 90% of all oral cancers. The characterization of altered molecules in oral cancer is essential to understand molecular mechanisms underlying tumor progression as well as to contribute to cancer biomarker and therapeutic target discovery. Proteoglycans are key molecular effectors of cell surface and pericellular microenvironments, performing multiple functions in cancer. Two of the major basement membrane proteoglycans, agrin and perlecan, were investigated in this study regarding their role in oral cancer. Using real time quantitative PCR (qRT-PCR), we showed that agrin and perlecan are highly expressed in oral squamous cell carcinoma. Interestingly, cell lines originated from distinct sites showed different expression of agrin and perlecan. Enzymatically targeting chondroitin sulfate modification by chondroitinase, oral squamous carcinoma cell line had a reduced ability to adhere to extracellular matrix proteins and increased sensibility to cisplatin. Additionally, knockdown of agrin and perlecan promoted a decrease on cell migration and adhesion, and on resistance of cells to cisplatin. Our study showed, for the first time, a negative regulation on oral cancer-associated events by either targeting chondroitin sulfate content or agrin and perlecan levels.