Oral manifestations of amyloidosis and the diagnostic applicability of oral tissue biopsy.

BACKGROUND: Amyloidosis exhibits a variable spectrum of systemic signs and oral manifestations that can be difficult to diagnose. This study aimed to characterize the clinical, demographic, and microscopic features of amyloidosis in the oral cavity. METHODS: This collaborative study involved three Brazilian oral pathology centers and described cases with a confirmed diagnosis of amyloidosis on available oral tissue biopsies. Clinical data were obtained from medical records. H&E, Congo-red, and immunohistochemically stained slides were analyzed. RESULTS: Twenty-six oral biopsies from 23 individuals (65.2% males; mean age: 59.6 years) were included. Oral involvement was the first sign of the disease in 67.0% of cases. Two patients had no clinical manifestation in the oral mucosa, although the histological analysis confirmed amyloid deposition. Amyloid deposits were distributed in perivascular (88.0%), periacinar and periductal (80.0%), perineurial (80.0%), endoneurial (33.3%), perimuscular (88.2%), intramuscular (94.1%), and subepithelial (35.3%) sites as well as around fat cells (100.0%). Mild/moderate inflammation was found in 65.4% of cases and 23.1% had giant cells. CONCLUSIONS: Amyloid deposits were consistently found in oral tissues, exhibiting distinct deposition patterns. Oral biopsy is less invasive than internal organ biopsy and enables the reliable identification of amyloid deposits even in the absence of oral manifestations. These findings corroborate the relevance of oral biopsy for the diagnosis of amyloidosis.

Central giant cell granulomas of the jaws stromal cells harbour mutations and have osteogenic differentiation capacity, in vivo and in vitro.

BACKGROUND: Central giant cell granulomas (CGCG) of the jaws are osteolytic lesions that may behave aggressively and respond poorly to surgery. Microscopically, in addition to giant cells, there is a mononuclear cell population composed of macrophage/monocytic cells and spindle-shaped cells of mesenchymal origin. Seventy two percent of these tumours harbour mutually exclusive TRPV4, KRAS and FGFR1 mutations. We aimed to assess the mutational status of mononuclear and giant cells and the osteogenic potential of stromal cells in vitro and in vivo. METHODS AND RESULTS: We screened CGCG for signature mutations and used laser-capture microdissection to demonstrate that the mutations are restricted to the mononuclear cells. Additionally, we established CGCG primary cell culture and observed that the cells retained the mutations throughout passages. By flow cytometry, we observed predominance of CD14- CD51- CD61- cells, consistent with the expected profile for stromal cells. Considering the mesenchymal origin of stromal cells, we assessed the osteogenic differentiation potential of CGCG cells in culture by cytochemistry (von Kossa and alizarin red staining), alkaline phosphatase (ALP) activity assay and gene expression of osteogenic markers. CGCG cells presented self-capacity to increase ALP levels in a time-dependent manner and under osteogenic induction presented increasing number of calcium deposits, and overall higher expression of osteocalcin, RUNX2, ALPL and osteopontin than cells without osteogenic induction. A patient-derived xenograft model for CGCG was established, and osteoid material deposition was observed. CONCLUSION: Collectively, the results confirm that the signature mutations are restricted to stromal cells in CGCG, and the in vitro and in vivo results support that these cells have the capacity to differentiate into osteoblasts, in line with the bone formation often observed in the stroma of these lesions.

Adenoid ameloblastoma with dentinoid is molecularly different from ameloblastomas and adenomatoid odontogenic tumors.

BACKGROUND: Adenoid ameloblastoma is a rare epithelial neoplasm, histologically characterized by the presence of ameloblastoma-like features, duct-like structures, epithelial whorls, and cribriform architecture. Dentinoid material is usually present. Some advocate adenoid ameloblastoma is an ameloblastoma variant. However, there are overlapping features not only with ameloblastoma, but also with adenomatoid odontogenic tumor. Most ameloblastomas are characterized by the presence of BRAF p.V600E mutations and adenomatoid odontogenic tumors harbor signature KRAS mutations. The molecular features of adenoid ameloblastoma remain unknown. METHODS: Nine adenoid ameloblastoma cases were screened by TaqMan allele-specific qPCR to assess BRAF p.V600E, ameloblastoma signature mutation, and KRAS p.G12V and p.G12R, adenomatoid odontogenic tumor signature mutations. RESULTS: BRAF and KRAS mutations were not detected in any of the adenoid ameloblastoma cases. CONCLUSION: The molecular results support adenoid ameloblastoma as an entity distinct from adenomatoid odontogenic tumor and ameloblastoma.

Loss of heterozygosity and immunoexpression of PTEN in oral epithelial dysplasia and squamous cell carcinoma.

INTRODUCTION: Oral epithelial dysplasia (OED) is a risk factor for developing subsequent oral squamous cell carcinoma (OSCC). Loss of heterozygosity (LOH) profiles have been validated as risk predictors of malignant transformation of OED. It is still unclear if Phosphatase and tensin homologue deleted on chromosome 10 (PTEN) allelic loss also occurs in initial stage malignant lesions and if the allelic loss is involved as one of the mechanisms of oral carcinogenesis. Thus, this study objective investigate LOH of PTEN gene and the immunohistochemical expression of the protein in OED and OSCC samples. MATERIAL AND METHODS: Formalin-fixed paraffin-embedded samples of 19 OEDs and 16 OSCCs were included to immunohistochemistry and LOH analysis. Two polymorphic microsatellite markers (AFMA086WG9 and D10S1765) located in chromosome 10 were used in this study for LOH analysis. For immunohistochemical analysis, 5 random fields with 400× magnification were evaluated quantitatively and qualitatively in epithelial and neoplastic cells. RESULTS: AFMA086WG9 marker only demonstrated LOH in OEDs cases (10.5%). D10S1765 marker demonstrated LOH in 57.2% of OEDs and 50% of OSCCs. Higher nuclear immunostaining was detected in cases of OSCCs when compared to OEDs (p < .001) and there was strong cytoplasmic immunoexpression in OSCCs (p < .045). CONCLUSIONS: We provide evidence that the allelic loss of PTEN is present in premalignant oral lesions and OSCCs, however the LOH of PTEN does not seems to influence its protein expression.

First insights for targeted therapies in odontogenic myxoma.

OBJECTIVE: Odontogenic myxoma (OM) occasionally responds poorly to surgical treatment. The MAPK pathway is constitutively activated in several neoplasms and we aimed to test if the MAPK pathway is activated in OM, in order to pave the way for an alternative therapy for aggressive and recurrent cases. MATERIALS AND METHODS: The immunoexpression of phosphorylated ERK1/2 (pERK1/2) was assessed in OM. We established a 3D organotypic culture model for the in vitro study and patient-derived xenografts (PDX) in mice for the in vivo study. The MEK inhibitor U0126 was used to inhibit phosphorylation of ERK1/2 in the in vitro and in vivo models. RESULTS: All OM showed strong pERK1/2 immunoexpression, consistent with MAPK pathway activation. Treatment of the 3D culture with U0126 resulted in a reduced pERK1/2/ERK1/2 ratio. Consistent with the in vitro results, all PDX of animals treated with U0126 showed a decreased volume fold change compared with controls. CONCLUSIONS: The MAPK pathway is activated in OM and its inhibition leads to tumor shrinkage in PDX and cell culture models. CLINICAL RELEVANCE: Our results offer a pre-clinical frame for OM-targeted therapy. Further work is needed to determine if this initial finding holds clinical promise.

KRAS mutations drive adenomatoid odontogenic tumor and are independent of clinicopathological features.

Adenomatoid odontogenic tumor is a benign encapsulated epithelial odontogenic tumor that shows an indolent clinical behavior. We have reported in a few adenomatoid odontogenic tumors mutations in KRAS, which is a proto-oncogene frequently mutated in cancer such as lung, pancreas, and colorectal adenocarcinomas. We aimed to assess KRAS mutations in the hotspot codons 12, 13, and 61 in a large cohort of adenomatoid odontogenic tumors and to test the association of these mutations with clinical (age, site, tumor size, follicular/extrafollicular subtypes) and histopathological parameters. Thirty eight central cases were studied. KRAS codon 12 mutations were assessed by TaqMan allele-specific qPCR (p.G12V/R) and/or Sanger sequencing, and codon 13 and 61 mutations were screened by Sanger. Histological tumor capsule thickness was evaluated by morphometric analysis. Additionally, the phosphorylated form of the MAPK downstream effector ERK1/2 was investigated. Statistical analysis was carried out to test the association of KRAS mutations with clinicopathological parameters. KRAS c.35 G >T mutation, leading to p.G12V, was detected in 15 cases. A novel mutation in adenomatoid odontogenic tumor, c.34 G >C, leading to p.G12R, was detected in 12 cases and the other 11 were wild-type. Codon 12 mutations were not associated with the clinicopathological parameters tested. RAS mutations are known to activate the MAPK pathway, and we show that adenomatoid odontogenic tumors express phosphorylated ERK1/2. In conclusion, a high proportion of adenomatoid odontogenic tumors (27/38, 71%) have KRAS codon 12 mutations, which occur independently of the clinicopathological features evaluated. Collectively, these findings indicate that KRAS mutations and MAPK pathway activation are the common features of this tumor and some cancer types. Although it is unclear why different codon 12 alleles occur in different disease contexts and the complex interactions between tumor genotype and phenotype need clarification, on the basis of our results the presence of KRAS p.G12V/R favors the adenomatoid odontogenic tumor diagnosis in challenging oral neoplasm cases.

Lack of association between denture trauma and loss of heterozygosity confronts the proposed pathologic role of chronic mucosal trauma in oral carcinogenesis.

Chronic mucosal trauma is suggested as an additional etiologic risk factor for oral squamous cell carcinoma (OSCC), but there is a lack of experimental-molecular data. If chronic trauma of the oral mucosa is carcinogenic, it should be associated with early genetic alterations seen during typical progression of OSCC, like loss of heterozygosity (LOH). We investigated LOH in the key chromosomal arms 3p, 9p and 17p in inflammatory fibrous hyperplasia associated with removable dental prosthesis and also in normal oral mucosa, by using the polymorphic microsatellite markers D3S1300 at 3p14.2, D9S1748 at 9p21, D17S1289 at 17p12 and D17S974 at 17p13 and capillary electrophoresis. LOH was detected in 2/15 (13%) fibrous hyperplasia samples similarly to other reactive and inflammatory lesions. None of the normal mucosa samples presented LOH. Our experimental-molecular results do not support the hypothesis that trauma associated with dental prosthesis has an important role in oral carcinogenesis.

Cohesin subunits, STAG1 and STAG2, and cohesin regulatory factor, PDS5b, in oral squamous cells carcinomas.

BACKGROUND: Cohesin complex is responsible for sister chromatid cohesion. STAG1/STAG2 is part of the complex, which is regulated by PDS5B. Alterations in these genes were described in tumors. PDS5B is a negative regulator of cell proliferation. We aimed to assess molecular alterations in these genes in oral squamous cell carcinoma (OSCC) and predict their expression by the expression of 84 cell cycle genes. In addition, we investigated whether pds5b protein expression impacted ki-67 and p53 immunopositivity. METHODS: We assessed loss of heterozygosity (LOH) at STAG1 and STAG2 loci in 15 OSCC using three polymorphic markers. Associations between the immunoexpression of pds5b and ki-67 and p53 were tested in 62 samples. Differences between transcriptional levels of STAG1, STAG2, and PDS5B between OSCC and normal oral mucosa (NM) were evaluated by qPCR. An 84 cell cycle genes qPCR array was carried with OSCC samples, and STAG1, STAG2, and PDS5B were independently used as response variables in multiple linear regression models. RESULTS: Loss of heterozygosity in at least one marker was observed in three samples. pds5b, p53, and ki-67 were highly expressed, and no association was found between pds5b immunoexpression and ki-67 or p53 (P > 0.05). OSCC and NM showed similar transcriptional levels of STAG1, STAG2, and PDS5B. STAG1 and CUL3 expression seem to be related (P = 0.004). CONCLUSIONS: There is LOH at STAG1 and STAG2 loci in OSCC, but OSCC and NM showed similar transcriptional levels of STAG1, STAG2, and PDS5B. pds5b immunoexpression in OSCC was high, but it was not associated with proliferation cell index.

Lip cancer and pre-cancerous lesions harbor TP53 mutations, exhibit allelic loss at 9p, 9q, and 17p, but no BRAFV600E mutations.

Molecular mechanisms of lip squamous cell carcinoma (LSCC) and actinic cheilitis (AC) are unclear. We aimed at assessing loss of heterozygosity (LOH) and TP53 and BRAF V600E mutations in these lesions. Formalin-fixed paraffin-embedded (FFPE) samples of 17 LSCC and 16 AC were included, with additional 5 fresh LSCC genotyped for TP53 mutations. LOH was assessed by six polymorphic markers located at 9p22, 9q22, and 17p13 and correlated with cell proliferation (Ki-67) and P53 immunostaining. Direct sequencing of TP53 exons 2-11 (fresh samples), and exons 5-9 (FFPE samples) was carried out. BRAF V600E mutation was genotyped in eight LSCC. LOH occurred in at least one marker in 15/17 LSCC and in 9/16 AC. The marker exhibiting the highest frequency of allelic loss (FAL) in LSCC was D9S157 (8/12 informative cases) and D9S287 in AC (4/11 informative cases). Cell proliferation was not correlated with LOH or with the FAL and no correlation between P53 IHC and 17p LOH was observed. We found TP53 missense mutations in both lesions and nonsense in LSCC, including CC>TT transition, which is a marker of UV damage. BRAF V600E mutation was not detected. LOH and TP53 mutations detected in LSCC and AC may be associated with tumorigenesis, whereas BRAF V600E mutation does not seem to significantly contribute to LSCC pathogenesis.

STAG2 expression in oral cancer and potentially malignant lesions.

Oral cancer is a world health problem, and one of the highest incidence rates of oral cancer worldwide occurs in Brazil. STAG2 is part of the cohesin complex which is responsible for sister chromatid cohesion. STAG2 loss of expression was reported in a range of tumors, and STAG2 loss was found to cause chromosomal instability and aneuploidy in cancer cells. On the basis of these findings, we investigated STAG2 expression in oral cancer and potentially malignant lesions. We investigated STAG2 immunoexpression in oral cancer, lip cancer, oral leukoplakia, and actinic cheilitis, including complete clinical information. Normal oral mucosa samples were included as normal controls. STAG2 protein was highly expressed in all samples. We further tested STAG2 expression in gastric adenocarcinomas and glioblastomas, as these tumor types were previously shown to lose STAG2 expression. We found homogenous expression of STAG2 by these tumor cells. Our results suggest that STAG2 loss of expression is not a common event in oral carcinogenesis.

STAG2 loss of expression is rare in aneuploid malignant salivary gland neoplasms.

BACKGROUND: STAG2 depletion leads to loss of centromere cohesion in vitro, and some human neoplasms have been shown to lose expression of this protein. As a result, STAG2 loss has been shown to cause chromosomal instability and aneuploidy in human cancer cell lines. METHODS: We tested the hypothesis that aneuploid salivary gland tumours lose immunoexpression of STAG2 compared with diploid tumours using image cytometry to determine DNA ploidy and immunohistochemistry to assess STAG2 protein expression in 30 malignant salivary gland neoplasms. RESULTS: There was no difference in the immunoexpression of STAG2 between aneuploid (n = 9) and diploid (n = 21) samples. In all but two samples, more than 50% of cells stained for STAG2. CONCLUSION: Aneuploidy in human salivary gland carcinomas is not driven by loss of expression of STAG2.

Effects of tobacco on the DNA of smokers and non-smokers affected by OSCC: systematic review and meta-analysis.

Scientific evidence about genetic and molecular changes in oral squamous cell carcinoma (OSCC) among smokers and non-smokers is inconclusive. This systematic review and meta-analysis assessed the effects of tobacco on the DNA of individuals with OSCC based on protein mutations. Electronic searches were conducted on PubMed, Ovid, Web of Science, and Scopus to identify observational studies published up to January/2022. The Joanna Briggs Institute tool was used for the critical appraisal of studies. The certainty of the evidence was evaluated. Twenty-three studies assessing 4,060 individuals (2,967 smokers vs. 1,093 non-smokers) were included in this review. Fifteen groups of proteins/genes were investigated. Analysis of the quality of articles revealed low risk of bias in most studies. The certainty of the evidence was very low. The meta-analysis confirmed no significant difference between smokers and non-smokers with respect to damage to GSTM1 (OR: 0.60; 95%CI: 0.30-1.18), GSTT1 (OR: 1.18; 95%CI:0.49-2.83), hydrolase proteins (Ku70 and Ku80) (OR: 0.74; 95%CI: 0.18-3.05), and transferase proteins (GSTM1, GSTT1, GSTM3) (OR: 0.74; 95%CI: 0.47-1.18). Most of the studies included showed that smokers are more likely to exhibit genetic instability. However, the meta-analysis revealed that smokers do not necessarily have more genetic alterations in the DNA than non-smokers.

Anti-apoptotic gene transcription signature of salivary gland neoplasms.

BACKGROUND: Development of accurate therapeutic approaches to salivary gland neoplasms depends on better understanding of their molecular pathogenesis. Tumour growth is regulated by the balance between proliferation and apoptosis. Few studies have investigated apoptosis in salivary tumours relying almost exclusively on immunohistochemistry or TUNEL assay. Furthermore, there is no information regarding the mRNA expression profile of apoptotic genes in salivary tumors. Our objective was to investigate the quantitative expression of BCL-2 (anti-apoptotic), BAX and Caspase3 (pro-apoptotic genes) mRNAs in salivary gland neoplasms and examine the association of these data with tumour size, proliferative activity and p53 staining (parameters associated with a poor prognosis of salivary tumours patients). METHODS: We investigated the apoptotic profile of salivary neoplasms in twenty fresh samples of benign and seven samples of malignant salivary neoplasms, using quantitative real time PCR. We further assessed p53 and ki-67 immunopositivity and obtained clinical tumour size data. RESULTS: We demonstrated that BCL-2 mRNA is overexpressed in salivary neoplasms, leading to an overall anti-apoptotic profile. We also found an association between the anti-apoptotic index (BCL-2/BAX) with p53 immunoexpression. A higher proliferative activity was found in the malignant tumours. In addition, tumour size was associated with cell proliferation but not with the transcription of apoptotic genes. CONCLUSION: In conclusion, we show an anti-apoptotic gene expression profile in salivary neoplasms in association with p53 staining, but independent of cell proliferation and tumour size.

DNA damage-related proteins in smokers and non-smokers with oral cancer.

Tobacco smoking involves a high risk of human malignancies, including oral cancer, because it contains multiple carcinogens that cause genetic instability. Thus, a worse prognosis would be expected for cancer patients who are smokers. The aim of this study was to assess the DNA damage response through the expression of checkpoint kinase 2 (CHK2), H2A histone family member X (H2AX), and P53 among smokers and non-smokers with oral squamous cell carcinoma (OSCC). Associations between immunoexpression of proteins and clinicopathological data and histopathological grading were also analyzed. A total of 35 individuals (18 non-smokers and 17 smokers) with OSCC of the tongue and/or floor of the mouth were included. Immunohistochemistry for H2AX was conducted for the identification of double-strand breaks, CHK2, and P53 to evaluate the expression of this protein in cell cycle regulation. The sample consisted of 22 males and 13 females, with a mean age of 63.9±11.8 years. OSCC of non-smokers were well-differentiated tumors in 50% of the cases, and those of smokers were equally distributed into moderately differentiated and poorly differentiated tumors (35.3% each). Overall, 31 (88.6%) cases were CHK2-positive, 27 (77.1%) were H2AX-positive, and 23 (65.7%) were P53-positive, with no difference between smokers and non-smokers (p > 0.05). No association was found between proteins and clinicopathologic data (p > 0.05). Similarities in CHK2, H2AX, and P53 immunohistochemical staining patterns were observed between smokers and non-smokers, and immunoexpression was not associated with clinicopathological parameters. However, the findings indicated consistent expression of these proteins in OSCC.

Quantitative expression analysis of apoptotic/antiapoptotic genes and association with immunolocalization of BAX and BCL-2 in peripheral and central giant cell lesions of the jaws.

Central giant cell lesion (CGCL) and peripheral giant cell lesion (PGCL) of the jaws are characterized by multinucleated osteoclast-like giant cells in a background of mononuclear cells. While mononuclear cells retain proliferative activity in both lesions, giant cells are Ki-67 negative. This observation raised the theory that giant cells are formed by cytoplasmic fusion of mononuclear cells, and also that these lesions are of reactive nature. As the giant cells are not proliferating in CGCL and PGCL, apoptosis of such cells should be investigated. We investigated the transcription of BAX and BCL-2 mRNAs in six fresh samples of CGCL and six fresh samples of PGCL by qRT-PCR (quantitative reverse transcription PCR) and used immunohistochemistry to demonstrate the localization of these proteins, as well as caspase 3 active in six paraffin-embedded samples of CGCL and nine paraffin-embedded samples of PGCL. While both groups showed increased expression of BAX and BCL-2 mRNA, PGCL showed a higher apoptotic index (ratio BAX/BCL-2) than CGCL. The three proteins investigated were expressed almost exclusively in the cytoplasm of giant cells. To further confirm apoptotic activity, we performed TUNEL analysis in the same samples of the immunohistochemistry and found a higher positivity in the giant cells of PGCL compared to the giant cells of CGCL. Our results show increased expression of apoptotic-related genes in both PGCL and CGCL and that the giant cells are probably the main source of these events. Also, it raises a hypothesis that differences in the apoptotic activity might be associated with the different clinical behavior of CGCL and PGCL.

Toxicity of Carnoy's solution toward human keratinocytes: an in vitro study.

The present study aimed to characterize the chemical elements and cytotoxicity of Carnoy's solution (CS) by comparing two different trademarked products (one Brazilian [NCS] and another imported [ICS]) using inductively coupled plasma mass spectrometry (ICP-MS) and human keratinocyte (HaCaT) cultures. For performing ICP-MS, the solutions were diluted according to calibration curves, and the chemical elements were analyzed with a spectrometer. HaCaT cells were exposed to CS concentrations ranging from 0.10% to 20% for 3 or 5 min. Cell viability was evaluated immediately (T0), 24 h (T1), and 7 days (T2) after exposure to CS using 3-(4,5-dimethylthiazol-2-yl)-2, 5-diphenyl-tetrazolium bromide (MTT) reduction assay. Data were analyzed using a t-test for ICP-MS and analysis of variance followed by Tukey's post-hoc test for MTT assay, both considering statistical significance at p<0.05. ICP-MS results revealed that ICS presented significantly lower concentrations of 12 chemical elements than NCS. The results of MTT assay revealed that at T0, ICS was more cytotoxic than NCS regardless of the time of exposure (p < 0.05). At T1, the only difference between the groups was at a concentration of 0.10% after 5 min of exposure. At T2, at a concentration of 0.5%, ICS resulted in a significant reduction in cell viability compared to NCS (p < 0.05). Thus, the results showed that ICS was more cytotoxic than NCS. Collectively, our findings suggest that the individual compositions of different CS formulations should be investigated.

Assessing pathogenic mutations in dental follicles as an attempt to identify early events in odontogenic tumours tumourigenesis.

OBJECTIVE: Driver oncogenic mutations have been reported in several benign neoplasms. While ameloblastomas show BRAF p.V600E mutations, adenomatoid odontogenic tumours harbour either KRAS p.G12R or p.G12 V. The lack of understanding of the core molecular changes involved in tumour initiation and progression represents a critical barrier to developing new strategies for cancer detection and prevention. Considering the fact that ameloblastoma and adenomatoid odontogenic tumours can originate from dental follicles, we hypothesized that the BRAF and KRAS mutations might be early events in odontogenic tumours tumourigenesis. We aimed to assess BRAF and KRAS mutations in dental follicles associated with asymptomatic impacted teeth. DESIGN: Forty-eight dental follicles containing odontogenic epithelial remnants were included in the study. As ameloblastomas most often occur in the posterior mandible and adenomatoid odontogenic tumours have a predilection for the anterior jaws, we assessed by allele-specific qPCR the presence of BRAF p.V600E in 32 dental follicles associated with impacted 3rd mandibular molar teeth and KRAS p.G12 V and KRAS p.G12R mutations in 16 dental follicle specimens obtained from around impacted anterior teeth. Sanger sequencing was used as an additional method. RESULTS: None of the dental follicle cases tested positive for the mutations. CONCLUSION: In conclusion, we tried to detect the early genetic events associated with odontogenic tumours development in dental follicles, but we were unable to showcase that BRAF p.V600E and KRAS p.G12R or p.G12 V mutations are the early genetic events associated with odontogenic tumours development.

Targeted Next-Generation Sequencing and Allele-Specific Quantitative PCR of Laser Capture Microdissected Samples Uncover Molecular Differences in Mixed Odontogenic Tumors.

The molecular pathogenesis of mixed odontogenic tumors has not been established, and understanding their genetic basis could refine their classification and help define molecular markers for diagnostic purposes. Potentially pathogenic mutations in the component tissues of 28 cases of mixed odontogenic tumors were assessed. Laser capture microdissected tissue from 10 ameloblastic fibromas (AF), 4 ameloblastic fibrodentinomas (AFD), 6 ameloblastic fibro-odontomas (AFO), 3 ameloblastic fibrosarcomas (AFS), and 5 odontomas (OD) were screened by next-generation sequencing and results confirmed by TaqMan allele-specific quantitative PCR. BRAF p.V600E mutation in the mesenchymal component was shown in 4 of 10 AF (40%), 2 of 4 AFD (50%), 2 of 6 AFO (33%), and 2 of 3 AFS (67%), whereas all 5 OD were wild type for BRAF p.V600E. Mutation in the epithelial component was only observed in one AF and one AFO. One AFS contained an area of benign AF, and the mesenchymal component of both (AFS and AF) contained BRAF p.V600E, supporting the concept of malignant progression from a benign AF precursor. KDR, TP53, KIT, and PIK3CA single-nucleotide polymorphisms are reported. In conclusion, AF, AFD, AFO, and AFS show BRAF p.V600E in their mesenchymal component, unlike OD, which are BRAF wild type, suggesting that at least a subset of AF, AFD, and AFO are molecularly distinct from OD, and may represent distinct entities and be neoplastic.

Effects of tobacco on the DNA of smokers and non-smokers affected by OSCC: systematic review and meta-analysis

Abstract Scientific evidence about genetic and molecular changes in oral squamous cell carcinoma (OSCC) among smokers and non-smokers is inconclusive. This systematic review and meta-analysis assessed the effects of tobacco on the DNA of individuals with OSCC based on protein mutations. Electronic searches were conducted on PubMed, Ovid, Web of Science, and Scopus to identify observational studies published up to January/2022. The Joanna Briggs Institute tool was used for the critical appraisal of studies. The certainty of the evidence was evaluated. Twenty-three studies assessing 4,060 individuals (2,967 smokers vs. 1,093 non-smokers) were included in this review. Fifteen groups of proteins/genes were investigated. Analysis of the quality of articles revealed low risk of bias in most studies. The certainty of the evidence was very low. The meta-analysis confirmed no significant difference between smokers and non-smokers with respect to damage to GSTM1 (OR: 0.60; 95%CI: 0.30-1.18), GSTT1 (OR: 1.18; 95%CI:0.49-2.83), hydrolase proteins (Ku70 and Ku80) (OR: 0.74; 95%CI: 0.18-3.05), and transferase proteins (GSTM1, GSTT1, GSTM3) (OR: 0.74; 95%CI: 0.47-1.18). Most of the studies included showed that smokers are more likely to exhibit genetic instability. However, the meta-analysis revealed that smokers do not necessarily have more genetic alterations in the DNA than non-smokers.