Exploring the combination of tumor-stroma ratio, tumor-infiltrating lymphocytes, and tumor budding with WHO histopathological grading on early-stage oral squamous cell carcinoma prognosis.

BACKGROUND: While the relevance of the World Health Organization histopathological grading system as a prognostic tool for oral squamous cell carcinoma has received many critics, other histopathological features such as tumor-stroma ratio, tumor-infiltrating lymphocytes, and tumor budding are displaying promising results. Here, we evaluated the prognostic impact of the incorporation of tumor-stroma ratio, tumor-infiltrating lymphocytes, and tumor budding into World Health Organization histopathological grading for patients with oral squamous cell carcinoma. METHODS: A total of 95 patients with early-stage oral squamous cell carcinoma were enrolled in the study, and World Health Organization tumor grading, tumor-stroma ratio, tumor-infiltrating lymphocytes, and tumor budding were evaluated in surgical slides stained with hematoxylin and eosin. Survival analyses for cancer-specific survival and disease-free survival were performed using Cox regression models, and receiver operating characteristic curves were applied for assessment of the performance of the combinations. RESULTS: Tumor-stroma ratio (stroma-rich) was significantly and independently associated with both shortened cancer-specific survival and poor disease-free survival, individually and in combination with World Health Organization histopathological grading. The combination of tumor-stroma ratio with World Health Organization grading did not improve the discriminatory ability compared to tumor-stroma ratio alone. Although low tumor-infiltrating lymphocytes were associated with shortened cancer-specific survival, the association did not withstand multivariate analysis. However, in combination with World Health Organization grading, low tumor-infiltrating lymphocytes were independently associated with poor cancer-specific survival. The combination of tumor-infiltrating lymphocytes and World Health Organization histopathological grading displayed a better discrimination of poor cancer-specific survival than tumor-infiltrating lymphocytes alone, but not at a significant level. CONCLUSION: Our findings support tumor-stroma ratio as a potential prognostic marker for patients with oral squamous cell carcinoma, and the incorporation of tumor-infiltrating lymphocytes into the World Health Organization grading system improves the prognostic ability of the tumor grading alone.

Prognostic significance of the neural invasion in oral squamous cell carcinoma.

BACKGROUND: Although nerve involvement can predict recurrence and prognosis in oral squamous cell carcinomas, there still have controversies and limitations regarding the standardization for its detection. In this study, we explore the impact of neural invasion in oral squamous cell carcinomas prognosis, comparing intraneural invasion (tumor cells inside nerve structure) and perineural invasion (cells involving the nerve, but not invading its sheath). METHODS: Surgical slides stained with hematoxylin and eosin from 235 patients with oral squamous cell carcinomas were carefully verified for the presence of intraneural invasion and perineural invasion. The location in the tumor (intratumoral vs. peritumoral) and number of foci (unifocal or multifocal) were also explored. Survival analyses for cancer-specific survival and disease-free survival were performed with Cox proportional model. RESULTS: Neural invasion was identified in 74 cases, 64.9% displayed intraneural invasion and 35.1% displayed perineural invasion. Univariate analysis revealed a significantly poorer cancer-specific survival, but not disease-free survival, in patients with intraneural invasion, in contrast to cases with perineural invasion that did not achieve significant association with both cancer-specific survival and disease-free survival. Further analyses revealed that the location in the tumor and number of foci had little impact on discriminatory ability of intraneural invasion. Multivariate analysis confirmed that intraneural invasion is significantly and independently associated with poor cancer-specific survival (hazard ratio: 2.50, 95% CI: 1.31-3.79, p = 0.003). CONCLUSION: This study provides evidence that intraneural invasion, but not perineural invasion, is a relevant predictor of survival in patients with oral squamous cell carcinomas, suggesting that its association with other clinical and pathological prognostic factors should be consider in determining the optimal treatment protocol and prognosis of these patients.

Overexpression of heat-shock protein 47 impacts survival of patients with oral squamous cell carcinoma.

BACKGROUND: The expression of heat-shock protein 47 (HSP47) has been linked to collagen synthesis control and implicated in fibrotic disorders, but more recent studies have demonstrated its role in solid tumors. In this study, we explored the prognostic impact of HSP47 in oral squamous cell carcinomas (OSCC) and determined the in vitro effects of its loss-of-function on viability, proliferation, migration, invasion, and resistance to cisplatin of OSCC cells. METHODS: The HSP47 expression in tumor samples was assessed by immunohistochemistry in two independent cohorts totaling 339 patients with OSCC, and protein levels were associated with clinicopathological features and survival outcomes. The OSCC cell lines HSC3 and SCC9 were transduced with lentivirus expressing short hairpin RNA to stably silence HSP47 and used in assays to measure cellular viability, proliferation, migration, and invasion. RESULTS: HSP47 was overexpressed in OSCC samples, and its overexpression was significantly and independently associated with poor disease-specific survival and shortened disease-free survival in both OSCC cohorts. The knockdown of HSP47 showed no effects on cell viability or cisplatin sensitivity, but impaired significantly proliferation, migration, and invasion of OSCC cells, with stronger effects on SCC9 cells. CONCLUSION: Our results show a significant prognostic impact of HSP47 overexpression in OSCC and reveal that HSP47 inhibition impairs the proliferation, migration, and invasion of OSCC cells. HSP47 may represent a potential therapeutic target for OSCC.

Trophoblast cell surface antigen 2 expression predicts outcome in oral squamous cell carcinomas.

BACKGROUND: Trophoblast cell surface antigen 2 (TROP2) has unclear clinical role in oral squamous cell carcinomas (OSCC). Here, we investigated the association of TROP2 immunoexpression with clinicopathological parameters and survival of OSCC patients. SUBJECTS AND METHODS: Cancer-specific survival (CSS) and disease-free survival (DFS) were assessed in a cohort composed of 266 OSCC. An independent cohort with 88 OSCC samples matched with the normal oral tissue, as well as 17 metastatic lymph nodes, was used for validation. RESULTS: Multivariate analysis showed TROP2 as an independent marker of favorable prognosis for both CSS (HR: 0.60, 95% CI: 0.40-0.90, p = .01) and DFS (HR: 0.57, 95% CI: 0.36-0.89, p = .01). Furthermore, TROP2 protein expression was significantly higher in morphologically normal tissues compared to primary tumors (p < .0001) and lymph node metastases (p = .001), and it was significantly associated with CSS (HR: 0.26, 95% CI: 0.09-0.74, p = .008) in the validation cohort. A pooled mRNA analysis performed on the Oncomine™ database confirmed the underexpression in OSCC compared with normal tissues (p = .014). CONCLUSIONS: In summary, our results point to a favorable prognostic significance of TROP2 overexpression in a large cohort of oral cancer patients, suggesting it as an attractive clinical marker.

Association Between Hand Digit Ratio (2D:4D) and Nonsyndromic Orofacial Clefts.

OBJECTIVES: Digit ratio (2D:4D) has been considered to be a marker in studies evaluating an individual's susceptibility to diseases, especially those diseases that show sex differences in their occurrence. We aimed to assess whether 2D:4D ratios are associated with nonsyndromic cleft lip and/or palate (NSCL/P) and verify the existence of a specific pattern of 2D:4D ratio in individuals affected by orofacial clefts. DESIGN: This was a case-control study. METHODS: Digital measurements of index and ring finger lengths of both hands of patients with NSCL/P (n = 54) and age- and gender-matched controls (n = 54) were obtained using a digital vernier caliper. Mean ratios between the second and fourth digits were compared. Data were analyzed by Student's t test and Mann-Whitney test with a significance level of 5%. RESULTS: No significant difference was found between the mean digit ratios of the right and left hands between the groups for any analysis (P > .05), neither for the whole sample nor for the distributions by type of cleft and by gender. CONCLUSIONS: Although the development of the fingers and the occurrence of NSCL/P can be regulated by the actions of similar genes, our results are not consistent with an association between 2D:4D ratio and this craniofacial deformity. This suggests that intrauterine exposure to fetal androgens, assessed using this marker, is similar between patients with NSCL/P and healthy individuals. We highlight the need for further studies in populations with different ancestries.

Metachronous ameloblastic fibro-odontoma and dentigerous cyst in the posterior mandible.

An ameloblastic fibro-odontoma (AFO) is a rare mixed odontogenic tumor with histologic features of an ameloblastic fibroma in conjunction with the presence of dentin and enamel. It usually appears as a well-circumscribed radiolucency with radiopaque foci and slow growth and is commonly seen in children and young adults. A 13-year-old boy presented with an asymptomatic swelling in the posterior right region of the mandible and the right ascending ramus. The clinical, imaging, and histopathologic findings confirmed the diagnosis of an AFO. After 8 months, a radiolucent lesion involving the unerupted mandibular left third molar was observed; a final diagnosis of a dentigerous cyst (DC) was established for this lesion. Although coincidental events, metachronous odontogenic lesions suggest a possible common genetic origin, since both can be caused by related cellular signaling pathways. Complete enucleation is recommended for both AFOs and DCs; rates of recurrence are low.

Prevalence of Dental Anomalies in Patients With Nonsyndromic Cleft Lip and/or Palate in a Brazilian Population.

Objective : Many studies have demonstrated a high frequency of dental anomalies in patients with cleft lip and/or palate. Because dental anomalies may complicate dental treatment, we investigated the prevalence of dental anomalies in a group of Brazilian patients with nonsyndromic cleft lip and/or palate. Design, Participants, Setting : Retrospective analysis was performed using clinical records of 296 patients aged between 12 and 30 years with repaired nonsyndromic cleft lip and/or palate without history of tooth extraction and orthodontic treatment. Associations between oral clefts and presence of dental anomalies outside the cleft area were investigated. Results : Dental anomalies were identified in 39.9% of the nonsyndromic cleft lip and/or palate patients, and tooth agenesis (47.5%), impacted tooth (13.1%), and microdontia (12.7%) were the most common anomalies. Cleft lip patients were less affected by dental anomalies compared with cleft palate or cleft lip and palate patients (p  =  .057). Specifically, patients with unilateral cleft lip and palate were significantly more affected by dental anomalies than those with bilateral cleft lip and palate (p  =  .00002), and individuals with unilateral complete cleft lip and palate (p  =  .002) and complete cleft palate (p  =  .01) were significantly more affected by tooth agenesis than other cleft types. Agenesis of the premolars (p  =  .043) and maxillary lateral incisors (p  =  .03) were significantly more frequent in patients with unilateral complete cleft lip and palate. Conclusions : The present study revealed a high frequency of dental anomalies in nonsyndromic cleft lip and/or palate patients and further demonstrated that patients with unilateral cleft lip and palate were frequently more affected by dental anomalies than those with bilateral cleft lip and palate. Moreover, our results demonstrate that dental anomalies should be considered during dental treatment planning for individuals affected by nonsyndromic cleft lip and/or palate.

Contribution of polymorphisms in genes associated with craniofacial development to the risk of nonsyndromic cleft lip and/or palate in the Brazilian population.

BACKGROUND AND OBJECTIVE: Nonsyndromic cleft lip and/or palate (NSCL/P) is a complex disease associated with both genetic and environmental factors. One strategy for identifying of possible NSCL/P genetic causes is to evaluate polymorphic variants in genes involved in the craniofacial development. DESIGN: We carried out a case-control analysis of 13 single nucleotide polymorphisms in 9 genes related to craniofacial development, including TBX1, PVRL1, MID1, RUNX2, TP63, TGFß3, MSX1, MYH9 and JAG2, in 367 patients with NSCL/P and 413 unaffected controls from Brazil to determine their association with NSCL/P. RESULTS: Four out of 13 polymorphisms (rs28649236 and rs4819522 of TBX1, rs7940667 of PVRL1 and rs1057744 of JAG2) were presented in our population. Comparisons of allele and genotype frequencies revealed that the G variant allele and the AG/GG genotypes of TBX1 rs28649236 occurred in a frequency significantly higher in controls than in the NSCL/P group (OR: 0.41; 95% CI: 0.25-0.67; p=0.0002). The frequencies of rs4819522, rs7940667 and rs1057744 minor alleles and genotypes were similar between control and NSCL/P group, without significant differences. No significant associations among cleft types and polymorphisms were observed. CONCLUSION: The study suggests for the first time evidences to an association of the G allele of TBX1 rs28649236 polymorphism and NSCL/P.

Region 8q24 is a susceptibility locus for nonsyndromic oral clefting in Brazil.

BACKGROUND: Nonsyndromic cleft lip with or without cleft palate is a relatively common craniofacial defect with multifactorial inheritance. The association of the rs987525 single nucleotide variant, located in a gene desert at 8q24.21 region, has been consistently replicated in European populations. We performed a structured association approach combined with transcriptional analysis of the MYC gene to dissect the role of rs987525 in oral clefting susceptibility in the ethnically admixed Brazilian population. METHODS: We performed the association study conditioned on the individual ancestry proportions in a sample of 563 patients and 336 controls, and in an independent sample of 221 patients and 261 controls. The correlation between rs987525 genotypes and MYC transcriptional levels in orbicularis oris muscle mesenchymal stem cells was also investigated in 42 patients and 4 controls. RESULTS: We found a significant association in the larger sample (p = 0.0016; OR = 1.80 [95% confidence interval {CI}, 1.21-2.69], for heterozygous genotype, and 2.71 [95% CI, 1.47-4.96] for homozygous genotype). We did not find a significant correlation between rs987525 genotypes and MYC transcriptional levels (p = 0.14; r = -0.22, Spearman Correlation). CONCLUSIONS: We present a positive association of rs987525 in the Brazilian population for the first time, and it is likely that the European contribution to our population is driving this association. We also cannot discard a role of rs987515 in MYC regulation, because this locus behaves as an expression quantitative locus of MYC in another tissue.

Stress induced phosphoprotein 1 overexpression controls proliferation, migration and invasion and is associated with poor survival in oral squamous cell carcinoma.

Objective: Although there have been remarkable achievements in the molecular landscape of oral squamous cell carcinoma (OSCC) in recent years, bringing advances in the understanding of its pathogenesis, development and progression, little has been applied in the prognosis and choosing the optimal treatment. In this study, we explored the influence of the stress induced phosphoprotein 1 (STIP1), which is frequently reported to be highly expressed in many cancers, in OSCCs. Methods: STIP1 expression was assessed in the TCGA database and in two independent cohorts by immunohistochemistry. Knockdown strategy was applied in OSCC cell lines to determine the impact of STIP1 on viability, proliferation, migration and invasion. The zebrafish model was applied for studying tumor formation and metastasis in vivo. The association of STIP1 and miR-218-5p was explored by bioinformatics and mimics transfection. Results: STIP1 was highly expressed in OSCCs and significantly associated with shortened survival and higher risk of recurrence. STIP1 down-regulation decreased proliferation, migration and invasion of tumor cells, and reduced the number of metastases in the Zebrafish model. STIP1 and miR-218-5p were inversely expressed, and the transfection of miR-218-5p mimics into OSCC cells decreased STIP1 levels as well as proliferation, migration and invasion. Conclusion: Our findings show that STIP1 overexpression, which is inversely associated with miR-218-5p levels, contributes to OSCC aggressiveness by controlling proliferation, migration and invasion and is a determinant of poor prognosis.

Ectrodactyly-ectodermal dysplasia-clefting syndrome associated with p63 mutation and an uncommon phenotype.

Ectrodactyly-ectodermal dysplasia-clefting syndrome is an uncommon disorder that includes a clinical spectrum of limb, facial, ocular, internal ear, and urogenital malformations. The disease is caused by heterozygous mutations in the 3q27-29 located p63 gene. In this paper we describe a 17-year-old girl affected by ectrodactyly-ectodermal dysplasia-clefting syndrome with a de novo p63 mutation that predicts a heterozygous missense substitution (arginine to tryptophan substitution caused by a cytosine to thymine transition) at the amino acid 304 (R304W) of the p63 DNA-binding domain. Scattered freckles on face, legs, and abdominal region, an uncommon feature associated with this syndrome, were recognized in our patient. The clinical features and genotype-phenotype correlation with previous p63 mutations related to the syndrome are discussed and compared with those observed in our patient. This case expands the phenotypic spectrum of ectrodactyly-ectodermal dysplasia-clefting syndrome.

Two rare cases of oral metastasis arising from lung adenocarcinoma and esophageal carcinoma.

Metastasis to the oral cavity are rare, representing only 1% of all oral malignancies, and originate from various sites such as the breast, prostate, lung and kidney. Clinically, they can simulate reactive and inflammatory lesions common in the oral cavity, and the clinical and microscopic diagnosis of these metastasis is a challenge. In this article, we report two new cases of esophageal and lung metastasis to oral tissues, highlighting their clinical characteristics and the process of diagnostic elucidation. We emphasize the importance for clinicians to consider the possibility of metastatic lesions in the oral cavity in patients previously diagnosed with malignant lesions in distant tissues and organs. Key words:Diagnosis, esophageal squamous cell carcinoma, adenocarcinoma of lung, oral cavity, metastasis.

Eukaryotic translation elongation factor 1δ, N-terminal propeptide of type I collagen and cancer-associated fibroblasts are prognostic markers of oral squamous cell carcinoma patients.

OBJECTIVE: Identifying markers that influence oral squamous cell carcinoma (OSCC) prognosis is a fundamental strategy to improve the overall survival of patients. Markers such as eukaryotic translation elongation factor 1δ (EEF1D), fascin, N-terminal propeptide of type I collagen (PINP), and cancer-associated fibroblasts (CAFs) have been noticed in OSCCs and their levels are closely related to the prognosis of tumors. Our aim was to confirm the role of those markers in OSCC prognosis. STUDY DESIGN: Immunohistochemistry was performed in 90 OSCC specimens. The associations between clinicopathologic features and expression of markers were assessed by χ2 test. Kaplan-Meier curves and univariate and multivariate Cox regression models were used for survival analysis. Markers were analyzed individually and in combination. RESULTS: High expression of EEF1D (P = .017) and PINP (P = .02) and abundant density of CAFs in tumor stroma (P = .005) predicted significantly poor survival in OSCC patients. Multivariate analysis revealed that all 3 parameters are individually independent prognostic factors of OSCC patients, and their combination improved the discrimination of patients at high risk for poor survival. CONCLUSIONS: Our results suggested that the expression of EEF1D and PINP and the density of CAFs might influence the survival of patients with OSCC.

Oral Manifestation of Lymphomatoid Granulomatosis.

Lymphomatoid granulomatosis (LYG) is a rare B-cell lymphoproliferative disorder driven by Esptein-Barr virus (EBV) that most commonly affects the lungs, although extra pulmonary sites like the central nervous system, skin, liver and kidney can also be involved. It is microscopically characterized by an angiocentric and angiodestructive growth pattern, predominantly composed by small T-cells, although a smaller population of atypical large B-cells is considered the true neoplastic component. Oral cavity involvement of LYG has rarely been described and the diagnosis of this neoplasm is very difficult. The aim of this report is to present a rare case of LYG affecting an 86-year-old female patient that was diagnosed due to an extensive, ulcerated and painful oral lesion affecting the hard palate. Detailed microscopic evaluation together with a large immunohistochemical study were necessary to achieve the correct diagnosis of LYG.

Variable expressivity and novel PTEN mutations in Cowden syndrome.

Cowden syndrome (CS) is a phosphatase and tensin homolog gene (PTEN)-associated condition characterized by multiple mucocutaneous hamartomas and an increased risk of malignancies. We reported an isolated case and another of several individuals in one family affected by CS. The isolated case showed typical features, including fibrocystic breast disease, benign thyroid nodules, and multiple papillomatous lesions in the face and oral cavity, and the cause was a novel nonsense mutation-guanine (G) to thymine (T) transition at position 940 (c.940 G>T)-in PTEN. In the family, the proband showed erythema nodosum, duodenal ulcer, intestinal polyps, cervical lipoma, renal cysts, and glaucoma, whereas multiple members of her family were found to have intestinal polyps, and a sister had been diagnosed with breast cancer at early age. An intronic mutation-T>G transition at the +32 position of intron 8 (c.1026+32 T>G)-was found in this family, with in silico analysis revealing the creation of a new donor splice site. This study confirmed the involvement of PTEN in CS and the variable clinical expressivity of disease.

Novel mutations in the IRF6 gene in Brazilian families with Van der Woude syndrome.

Van der Woude Syndrome (VWS) is an autosomal craniofacial disorder characterized by lower lip pits and cleft lip and/or palate. Mutations in the interferon regulatory factor 6 (IRF6) gene have been identified in patients with VWS. To identify novel IRF6 mutations in patients affected by VWS, we screened 2 Brazilian families, sequencing the entire IRF6-coding region and flanking intronic boundaries. Two novel heterozygous mutations were identified: a frame shift mutation with deletion of G at the nucleotide position 520 in the exon 6 (520delG), and a missense single nucleotide substitution from T to A at nucleotide position 1135 in exon 8 (T1135A). By using restriction enzyme analysis, we were able to demonstrate the lack of similar mutations in unrelated healthy individuals and non-syndromic cleft lip and palate patients. Our results further confirmed that haploinsufficiency of the IRF6 gene results in VWS.

Apert syndrome: report of a case with emphasis on craniofacial and genetic features.

Apert syndrome is 1 of the 5 craniosynostosis syndromes that shore clinical features and are caused by allelic mutations in the fibroblast growth factor receptor 2 (FGFR2) gene. The purpose of this paper was to report a case of Apert syndrome, with particular emphasis on craniofacial and genetic features, in a 5-year-old female patient. The patient presented with several craniofacial deformities, including severe brachycephaly, midface hypoplasio, flat forehead, proptosis, hypertelorism, and short nose with a bulbous tip. Syndactylies of the hands and feet were also observed. Intraoral findings included arched palate with pseudocleft in the midline, upper lip with symmetric depression resembling pseudoclefts, severe malocclusion, and several decoyed teeth. DNA sequence and restriction enzyme analysis showed a G to C transversion, resulting in a serine to tryptophan amino acid substitution at position 252 (S252W). Identification of the clinical features associated with mutation analysis is important to correctly diagnose Apert syndrome and distinguish it from other clinically similar craniosynostosis syndromes.

A challenging diagnosis: Case report of oral metastasis from gastric adenocarcinoma mimicking pyogenic granuloma.

RATIONALE: Oral metastases occur more commonly in bone, but can also manifest in soft tissues and eventually resemble a reactive lesion. Few cases of oral metastases mimicking reactive lesions in soft tissues have been reported to date. PATIENT CONCERNS: We report a metastasis of gastric carcinoma (GC) to the oral mucosa without bone involvement in a 43 yom clinically and microscopically mimicking a reactive lesion. The patient related that the lesion had 1 month of evolution, and the ulcerated area suggested the lesion was related to trauma. DIAGNOSES: The histopathological examination of the lesion revealed an exuberant granulation tissue with few neoplastic cells, and the initial diagnosis of pyogenic granuloma was considered. In a second analysis, clusters of clear cells morphologically similar to degenerating mucous cells or macrophages, positive for Cytokeratin (CK)-20, and CDX2 were found. At the moment, it was confirmed the presence of a primary GC in the patient. INTERVENTIONS: A palliative radiotherapy/chemotherapy was started. OUTCOMES: However, the patient died 3 months after the diagnosis of oral metastasis. LESSONS: This report highlights the importance of careful clinical and microscopic examinations in cases of oral metastasis that may mimic a reactive lesion.