Global prevalence of human papillomavirus-related oral and oropharyngeal squamous cell carcinomas: a systematic review and meta-analysis.

OBJECTIVE: To conduct a systematic review to determine the global prevalence of HPV in oral squamous cell carcinoma (OSCC) and oropharyngeal squamous cell carcinoma (OPSCC). MATERIALS AND METHODS: Literature was searched through October 2022 in main databases to address the question "What is the global prevalence of Human Papillomavirus in oral and oropharyngeal cancer?" Studies had to identify HPV by PCR, ISH, or p16 immunohistochemistry to be eligible. Quality was assessed using the JBI checklist for prevalence studies. Meta-analyses were performed, and reporting followed PRISMA guidelines. RESULTS: Sixty-five studies were included, and most of them had methodological limitations related to sampling and the HPV detection tool. The pooled prevalence of HPV-positivity was 10% (event rate = 0.1; 95% CI: 0.07, 0.13; P < 0.01; I2 = 88%) in the oral cavity and 42% (event rate = 0.42; 95% CI: 0.36, 0.49; P = 0.02; I2 = 97%) in oropharynx. The highest HPV prevalence in OSCC was reached by Japan, meanwhile, in OPSCC, Finland and Sweden were the most prevalent. HPV16 is the genotype most frequent with 69% in OSCC and 89% in OPSCC, being the tonsils the intraoral location more affected by HPV (63%, p < 0.01, I2 76%). CONCLUSION: The evidence points to an apparent burden in HPV-related OPSCC, mostly in North America, Northern Europe, and Oceania, especially due to the HPV16 infection suggesting different trends across continents. CLINICAL RELEVANCE: This updated systematic review and meta-analysis provide sufficient evidence about the global HPV prevalence in OSCC and OPSCC and the most frequent HPV subtype worldwide.

Influence of variants in serotonin modulating genes on the risk, aggressiveness, and prognosis of oropharynx cancer.

BACKGROUND: Serotonin (5-HT) is involved in regulating tumor growth, as well as psychiatric disorders. It is synthesized by tryptophan hydroxylase (TPH) and acts through 5-HT receptors (HTRs). Single-nucleotide variations (SNVs) in TPH1 rs623580 (T>A), TPH2 rs4570625 (G>T), and HTR1D rs674386 (G>A) may affect 5-HT levels. However, the effect of these SNVs on oropharynx carcinoma (OPC) is unknown. METHODS: DNA from 251 patients with OPC and 254 controls was analyzed by RT-PCR. Transcriptional activity of TPH1 rs623580 and HTR1D rs674386 was studied by luciferase assays. Multivariate statistical tests were utilized to evaluate group differences and survival outcomes. RESULTS: TPH1 TT was more frequent in patients than in controls (OR: 1.56, p = 0.03). Patients with HTR1D GG/GA showed invasive tumors (p = 0.01) and shorter survival (HR: 1.66, p = 0.04). TPH1 TT (0.79-fold, p = 0.03) and HTR1D GG (0.64-fold, p = 0.008) presented lower transcriptional activity. CONCLUSION: Our data suggest that SNVs in 5-HT modulating genes can influence OPC.

Association of Inherited Copy Number Variation in ADAM3A and ADAM5 Pseudogenes with Oropharynx Cancer Risk and Outcome.

Inherited copy number variations (CNVs) can provide valuable information for cancer susceptibility and prognosis. However, their association with oropharynx squamous cell carcinoma (OPSCC) is still poorly studied. Using microarrays analysis, we identified three inherited CNVs associated with OPSCC risk, of which one was validated in 152 OPSCC patients and 155 controls and related to pseudogene-microRNA-mRNA interaction. Individuals with three or more copies of ADAM3A and ADAM5 pseudogenes (8p11.22 chromosome region) were under 6.49-fold increased risk of OPSCC. ADAM5 shared a highly homologous sequence with the ADAM9 3'-UTR, predicted to be a binding site for miR-122b-5p. Individuals carrying more than three copies of ADAM3A and ADAM5 presented higher ADAM9 expression levels. Moreover, patients with total deletion or one copy of pseudogenes and with higher expression of miR-122b-5p presented worse prognoses. Our data suggest, for the first time, that ADAM3A and ADAM5 pseudogene-inherited CNV could modulate OPSCC occurrence and prognosis, possibly through the interaction of ADAM5 pseudogene transcript, miR-122b-5p, and ADAM9.

Role of a genetic variation in the microRNA-4421 binding site of ERP29 regarding risk of oropharynx cancer and prognosis.

We conducted a two-stage association study on patients with oropharynx (OP) squamous cell carcinoma (SCC) and healthy controls to identify single nucleotide variants (SNVs) located at the microRNA (miR)-binding sites of carcinogenesis genes associated with risk and prognosis of the disease. In stage 1, 49 patients and 49 controls were analyzed using Genome-Wide Human SNV Arrays to identify variants in the 3'-untranslated region (3'-UTR) of carcinogenesis-related genes, and one SNV was selected for data validation in stage 2 by TaqMan assays in 250 OPSCC patients and 250 controls. The ERP29 c.*293A > G (rs7114) SNV located at miR-4421 binding site was selected for data validation among 46 SNVs. The ERp29 and miR-4421 levels were evaluated by quantitative-PCR and Western blotting. Interaction between miR-4421 with 3'-UTR of ERP29 was evaluated by luciferase reporter assay. Event-free survival (EFS) was calculated by Kaplan-Meier and Cox methods. ERP29 GG variant genotype was more common in OPSCC patients than in controls (6.4% vs 3.6%, p = 0.02; odds ratio: 5.67; 95% confidence interval (CI) 1.27-25.26). Shorter EFS were seen in the base of tongue (BT) SCC patients with GG genotype (0.0% vs 36.2%, p = 0.01; hazard ratio: 2.31; 95% CI: 1.03-5.15). Individuals with ERP29 AG or GG genotypes featured lower levels of ERP29 mRNA (p = 0.005), ERp29 protein (p < 0.001) and higher levels of miR-4421 (p = 0.02). The miR-4421 showed more efficient binding with 3'-UTR of the variant G allele when compared with wild-type allele A (p = 0.001). Our data suggest that ERP29 rs7114 SNV may alter the risk and prognosis of OPSCC due to variation in the ERp29 production possibly modulated by miR-4421.

Impact of genetic variants in clinical outcome of a cohort of patients with oropharyngeal squamous cell carcinoma.

Tobacco- or human papillomavirus- driven oropharyngeal squamous cell carcinomas (OpSCC) represent distinct clinical, biological and epidemiological entities. The aim of this study was to identify genetic variants based on somatic alterations in OpSCC samples from an admixed population, and to test for association with clinical features. The entire coding region of 15 OpSCC driver genes was sequenced by next-generation sequencing in 51 OpSCC FFPE samples. Thirty-five percent of the patients (18/51) were HPV-positive and current or past tobacco consumption was reported in 86.3% (44/51). The mutation profile identified an average of 2.67 variants per sample. Sixty-three percent of patients (32/51; 62.7%) were mutated for at least one of the genes tested and TP53 was the most frequently mutated gene. The presence of mutation in NOTCH1 and PTEN, significantly decreased patient's recurrence-free survival, but only NOTCH1 mutation remained significant after stepwise selection, with a risk of recurrence of 4.5 (HR 95% CI = 1.11-14.57; Cox Regression p = 0.034). These results show that Brazilian OpSCC patients exhibit a similar clinical and genetic profile in comparison to other populations. Molecular characterization is a promising tool for the definition of clinical subgroups, aiding in a more precise tailoring of treatment and prognostication.

CASP8 (rs3834129) and CASP3 (rs4647601) polymorphisms in oropharynx cancer risk, tumor cell differentiation, and prognosis in a cohort of the Brazilian population.

The objective of this research was to assess the association of genetic polymorphisms related to intrinsic apoptosis pathway CASP8 rs3834129 and CASP3 rs4647601 with the risk, clinical and pathological aspects, and survival of oropharynx squamous cell carcinoma (OPSCC) patients that received cisplatin and radiotherapy. The genotypes were identified in 198 patients with OPSCC and 200 controls using polymerase chain reaction methods. Chi square or Fisher's exact test and logistic regression were applied for the detection of differences between groups. Patients' genotypes were statistically evaluated considering the event-free survival and overall analysis using Kaplan-Meier estimate and Cox regression. CASP3 rs4647601 GG genotype (44.4% vs. 30.0%, p = 0.03) and G allele (63.9% vs. 55.5%, p = 0.04) were more common in patients with OPSCC than in controls. Carriers of GG genotype and G allele were under 1.78-fold and 1.40-fold increased risk of OPSCC than others, respectively. The frequency of CASP8 rs3834129 DD genotype was higher in patients with OPSCC with poorly differentiated or undifferentiated tumors when compared to others (34.5% vs. 16.1%, p = 0.02). No influence of CASP8 and CASP3 polymorphisms on OPSCC patients' survival was seen in this study. Our results indicate that inherited genetic variants in the intrinsic apoptosis pathway related to CASP3 rs4647601 and CASP8 rs3834129 polymorphisms may be an important determinant of OPSCC risk and tumor cell differentiation.

Clinicopathological significance of SNPs in RAD51 and XRCC3 in oral and oropharyngeal carcinomas.

BACKGROUND: This study investigated the influence of single nucleotide polymorphisms (SNP) in RAD51 and XRCC3 on susceptibility to oral and oropharyngeal squamous cell carcinomas (SCC) and determined their clinicopathological significance. SUBJECTS AND METHODS: SNPs rs1801320 and rs1801321 in RAD51 and rs861539 in XRCC3 were genotyped in 81 patients presenting oral SCC, 45 presenting oropharyngeal SCC, and 130 healthy controls, using TaqMan allelic discrimination assays. Multiple logistic regression models were used to explore the association between SNPs and cancer development, as well as gene-gene (GxG) interaction and gene-environmental factor (GxE) interaction. Clinicopathological associations were verified through the chi-square test, and univariate and multivariate methods were applied for survival analyses. RESULTS: Although allelic and genotypic models and the GxG interaction analysis were nonsignificant, the GxE analysis revealed synergistic effects of the risk alleles of rs1801320, rs1801321, and rs861539 with smoking and alcohol consumption on susceptibility to oral and oropharyngeal SCC. Furthermore, oropharyngeal SCC patients carrying the XRCC3 rs861539 GT/TT genotype (T risk allele) presented a shorter overall survival than GG genotype carriers. CONCLUSION: Combined effects of RAD51 (rs1801320 and rs1801321) and XRCC3 (rs861539) SNPs with environmental carcinogens (tobacco and alcohol) are associated with oral and oropharyngeal SCC development.

Oncogenic drivers in 11q13 associated with prognosis and response to therapy in advanced oropharyngeal carcinomas.

OBJECTIVES: To identify potential molecular drivers associated with prognosis and response to treatment in advanced oropharyngeal squamous cell carcinomas (OPSCC). MATERIALS AND METHODS: Thirty-three OPSCC biopsies from untreated Brazilian patients were evaluated for human papilloma virus genotyping, genome wide copy number alterations and gene expression profiling. Data were integrated using CONEXIC algorithm. Validation with TCGA dataset and confirmation by RT-qPCR of candidate genes were performed. RESULTS: High-risk HPV positive cases, detected in 55% of advanced OPSCC, were associated with better outcome. Losses of 8p11.23-p11.22, 14q11.1-q11.2 and 15q11.2, and gains of 11q13.2 and 11q13.2-q13.3 were detected as recurrent alterations. Gains of 3q26.31 and 11q13.2 and losses of 9p21.3 were exclusively detected in HPV-negative tumors. Two clusters of expression profiles were observed, being one composed mostly by HPV positive cases (83%). HPV-positive enriched cluster showed predominantly immune response-related pathways. Integrative analysis identified 10 modulators mapped in 11q13, which were frequently cancer-related. These 10 genes showed copy number gains, overexpression and an association with worse survival, further validated by TCGA database analyses. Overexpression of four genes (ORAOV1, CPT1A, SHANK2 and PPFIA1) evaluated by RT-qPCR confirmed their association with poor survival. Multivariate analysis showed that PPFIA1 overexpression and HPV status are independent prognostic markers. Moreover, SHANK2 overexpression was significantly associated with incomplete response to treatment. CONCLUSION: The integrative genomic and transcriptomic data revealed potential driver genes mapped in 11q13 associated with worse prognosis and response to treatment, giving fundamentals for the identification of novel therapeutic targets in OPSCC.

Association between polymorphisms in genes related to DNA base-excision repair with risk and prognosis of oropharyngeal squamous cell carcinoma.

PURPOSE: We examined the influence of OGG1 c.977C>G (rs1052133), APEX1 c.444T>G (rs1130409), XRCC1 c.-77T>C (rs3213245), c.580C>T (rs1799782), c.839G>A (rs25489) and c.1196G>A (rs25487) single-nucleotide polymorphisms (SNPs), involved in base-excision repair (BER) pathway, on oropharyngeal squamous cell carcinoma (OPSCC) risk and prognosis. METHODS: Aiming to identify the genotypes, DNA from 200 consecutive OPSCC patients and 200 controls was analyzed by PCR-RFLP. The prognostic impact of genotypes of SNPs on progression-free survival (PFS) and overall survival of OPSCC patients was examined using the Kaplan-Meier estimates and Cox regression analyses. RESULTS: XRCC1 c.580CT or TT genotypes (19.5 vs. 11.0 %, P = 0.04) and XRCC1 TTGG haplotype from c.-77T>C, c.580C>T, c.839G>A and c.1196G>A SNPs (17.5 vs. 10.0 %, P = 0.04) were more common in patients with OPSCC than in controls. Carriers of combined genotypes of c.580C>T and TTGG haplotype of XRCC1 gene were under 3.35- and 3.22-fold increased risk of OPSCC than others. For survival analysis, we selected only patients with tumor at stage IV. The median follow-up time was 24.5 months. At 24 months of follow-up, PFS was shorter in patients with OGG1 c.977CC genotype when compared with others genotypes (35.5 vs. 52.1 %, log-rank test, P = 0.03). After multivariate Cox analysis, patients with OGG1 c.977CC genotype had more chance to present tumor progression when compared with others (HR 1.68, P = 0.02). CONCLUSIONS: Our data present, for the first time, evidence that inherited OGG1 c.977C>G; XRCC1 c.-77T>C, c.580C>T, c.839G>A and c.1196G>A abnormalities of DNA BER pathway are important determinants of OPSCC and predictors of patient outcomes.

Pathway Analysis using Gene-expression Profiles of HPV-positive and HPV-negative Oropharyngeal Cancer Patients in a Hispanic Population: Methodological Procedures.

OBJECTIVE: The incidence of oral cavity and pharyngeal cancer in Puerto Rican men is higher than it is in the men of any other ethnic/racial group in the United States of America (US). The information regarding the effect of the human papilloma virus (HPV) in the gene-expression profile among patients with this cancer is limited in Hispanic community. We aim to describe the methodology for future studies to identify the molecular networks for determining overrepresented signaling and metabolic canonical pathways, based on the differential gene-expression profiles of HPV+ and HPV- samples from patients with oropharyngeal squamous cell carcinoma in Puerto Rico. METHODS: We analyzed the RNA expression of 5 tissue samples from subjects diagnosed with oropharyngeal squamous cell carcinoma, 2 HPV+ and 3 HPV-, using Affymetrix GeneChips. The relative difference between the average gene expressions of the HPV+ and HPV- samples was assessed, based on the fold change (log2-scale). RESULTS: Our analysis revealed 10 up regulated molecules (Mup1, LRP1, P14KA, ALYREF, and BHMT) and 5 down regulated ones (PSME4, KEAP1, ELK3, FAM186B, and PRELID1), at a cutoff of 1.5-fold change. Ingenuity Pathway Analysis showed the following biological functions to be affected in the HPV+ samples: cancer, hematological disease, and RNA post-transcriptional modification. QRT-PCR analysis confirmed only the differential regulation of ALYREF, KEAP1, and FAM186B genes. CONCLUSION: The relevant methodological procedures described are sufficient to detect the most significant biological functions and pathways according to the HPV status in patients with oropharyngeal cancer in Puerto Rico.

Expression of stem cell-regulating miRNAs in oral cavity and oropharynx squamous cell carcinoma.

BACKGROUND: Head and neck squamous cell carcinoma (HNSCC) is the sixth most common tumor worldwide and is histologically heterogeneous. Studies have demonstrated the presence of stem cell markers in HNSCC, and microRNAs (miRNAs) have emerged as powerful regulators of differentiation, controlling the self-renewal of stem cells. miRNAs are non-coding RNA molecules that regulate gene expression post-transcriptionally. Many miRNAs have been described as regulators of stem cells in different types of cancer. METHODS: We have analyzed the expression of let-7a, miR-34, miR-125b, miR-138, miR-145, miR-183, miR-200b, miR-203, and miR-205 by real-time RT-PCR (qPCR), in 35 oral cavity and oropharynx squamous cell carcinoma (SCC) samples and 10 non-neoplastic oral mucosa controls, to determine possible associations between the expression of these miRNAs and clinical and pathological features of these tumors. RESULTS: We observed downregulation of miR-200b and miR-203 in 60.0% and 71.4% of the samples, respectively. Upregulation of miR-138 and miR-183 was observed in 50.0% of the samples. Downregulation of let-7a was associated with perineural invasion. Upregulation of miR-138, miRNA-145, and miR-205 was associated with advanced tumor stages, vascular invasion, and lymph node metastasis, respectively. CONCLUSIONS: Our study provides evidence of the expression of miRNAs associated with stem cell regulation in oral cavity and oropharynx SCC and the association of these miRNAs with clinical and pathological features of these tumors.

Leptin receptor expression and Gln223Arg polymorphism as prognostic markers in oral and oropharyngeal cancer.

The leptin gene product is released into the blood stream, passes through the blood-brain barrier, and finds the leptin receptor (LEPR) in the central nervous system. This hormone regulates food intake, hematopoiesis, inflammation, immunity, differentiation, and cell proliferation. The LEPR Gln223Arg polymorphism has been reported to alter receptor function and expression, both of which have been related with prognostics in several tumor types. Furthermore, several studies have shown a relationship between the Gln223Arg polymorphism and tumor development, and its role in oral and oropharyngeal squamous cell carcinoma is now well understood. In this study, 315 DNA samples were used for LEPR Gln223Arg genotyping and 87 primary oral and oropharyngeal squamous cell carcinomas were used for immunohistochemical expression analysis, such that a relationship between these and tumor development and prognosis could be established. Homozygous LEPR Arg223 was found to be associated with a 2-fold reduction in oral and oropharyngeal cancer risk. In contrast, the presence of the Arg223 allele in tumors was associated with worse disease-free and disease-specific survival. Low LEPR expression was found to be an independent risk factor, increasing the risk for lymph node metastasis 4-fold. In conclusion, the Gln223Arg polymorphism and LEPR expression might be valuable markers for oral and oropharyngeal cancer, suggesting that LEPR might serve as a potential target for future therapies.

FGFR4 profile as a prognostic marker in squamous cell carcinoma of the mouth and oropharynx.

BACKGROUND: Fibroblast growth factor receptor 4 (FGFR4) is a member of a receptor tyrosine kinase family of enzymes involved in cell cycle control and proliferation. A common single nucleotide polymorphism (SNP) Gly388Arg variant has been associated with increased tumor cell motility and progression of breast cancer, head and neck cancer and soft tissue sarcomas. The present study evaluated the prognostic significance of FGFR4 in oral and oropharynx carcinomas, finding an association of FGFR4 expression and Gly388Arg genotype with tumor onset and prognosis. PATIENTS AND METHODS: DNA from peripheral blood of 122 patients with oral and oropharyngeal squamous cell carcinomas was used to determine FGFR4 genotype by PCR-RFLP. Protein expression was assessed by immunohistochemistry (IHC) on paraffin-embedded tissue microarrays. RESULTS: Presence of allele Arg388 was associated with lymphatic embolization and with disease related premature death. In addition, FGFR4 low expression was related with lymph node positivity and premature relapse of disease, as well as disease related death. CONCLUSION: Our results propose FGFR4 profile, measured by the Gly388Arg genotype and expression, as a novel marker of prognosis in squamous cell carcinoma of the mouth and oropharynx.

Promoter hypermethylation in primary squamous cell carcinoma of the oral cavity and oropharynx: a study of a Brazilian cohort.

Epigenetic silencing of cancer-related genes plays an important role in oral/oropharyngeal squamous cell carcinoma (OSCC). We evaluated promoter hypermethylation of 4 cancer-related genes in OSCCs of a Brazilian cohort and determined its relationship with exposure to alcohol, tobacco, HPV infection and clinicopathological parameters. CDKN2A (cyclin-dependent kinase inhibitor 2A or p16), SFN (stratifin or 14-3-3 σ), EDNRB (endothelin receptor B) and RUNX3 (runt-related transcript factor-3) had their methylation patterns evaluated by MSP analysis in OSCC tumors (n = 45). HPV detection was carried out by PCR/RFLP. Aberrant methylation was detected in 44/45 (97.8 %) OSCC; 24.4 % at CDKN2A, 77.8 % at EDNRB, 17.8 % at RUNX3 and 97.8 % at SFN gene. There was no significant association between methylation patterns and clinical parameters. HPV (subtype 16) was detected in 3 out of 45 patients (6 %). Our findings indicate that HPV infection is uncommon and methylation is frequent in Brazilian OSCCs, however, EDNRB and SFN gene methylation are not suitable OSCC biomarkers due to indistinct methylation in tumoral and normal samples. In contrast, CDKN2A and RUNX3 genes could be considered differentially methylated genes and potential tumor markers in OSCCs.

Cyclin D1 gene polymorphism as a risk factor for squamous cell carcinoma of the upper aerodigestive system in non-alcoholics.

Squamous cell carcinoma of the upper aerodigestive tract (UADT) is associated with environmental factors, especially tobacco and alcohol consumption. Genetic factors, including cyclin D1 (CCND1) polymorphism have been suggested to play an important role in tumorigenesis and progression of UADT cancer. To investigate the relationship between CCND1 polymorphism on susceptibility for UADT cancers, 147 cancer and 135 non-cancer subjects were included in this study. CCND1 genotype at codon 242(G870A) in exon 4 was undertaken using denaturing high performance liquid chromatography (DHPLC) and DNA sequencing. Significant odds ratio (OR) of the AA+GA genotypes [OR=7.5 (95% CI: 1.4-39.7)] was observed in non-drinkers but for non-smokers a non-significant [OR=5.4 (95% CI: 0.9-31.4)] was found in the adjusted model. These results suggest that allele A may be a risk factor for UADT cancer, especially in non-alcoholics. However, further epidemiological studies are needed to establish the exact role of CCND1 polymorphism and the development of UADT cancers.

Alcohol dehydrogenase 3 genotype as a risk factor for upper aerodigestive tract cancers.

OBJECTIVE: To assess alcohol dehydrogenase 3 (ADH3) polymorphism at position Ile349Val as indicator of risk factor for upper aerodigestive tract (UADT) cancer to verify its association with UADT cancer in nonalcoholic or nonsmoking individuals. DESIGN: Cross-sectional study. SETTING: Primary care or referral center. PATIENTS: The study group consisted of 141 consecutive patients with newly diagnosed squamous cell carcinoma of the oral cavity, oropharynx, hypopharynx, or larynx admitted for surgical treatment. The comparison group consisted of 94 inpatients without cancer from the A. C. Camargo or other São Paulo (Brazil) hospital and 40 healthy individuals. INTERVENTION: All participants were interviewed and data were collected using a structured questionnaire. After written informed consent was obtained, 20 mL of blood was collected in heparinized tubes. MAIN OUTCOME MEASURES: Odds ratio for ADH3 genotypes using logistic regression models. RESULTS: After adjustment for sex, age, tobacco use, and history of cancer in first-degree family relatives, a significantly higher odds ratio for UADT cancer was observed among individuals with AA genotype and low cumulative alcohol consumption (< or =100 kg of ethanol) (odds ratio = 3.8 [95% confidence interval, 1.5-9.7]). A 4-fold increase in odds ratio for UADT cancer among individuals with AA genotype and low tobacco consumption (< or =25 pack-years) was also found in the adjusted model. CONCLUSIONS: These results suggest that genotype AA may be a risk factor for UADT cancer, especially in individuals with low alcohol or tobacco consumption. However, further epidemiological case-control or cohort studies, preferably prospective, are needed to establish the exact role of ADH3 polymorphism and its association with the development of UADT cancers.

Micronucleus investigation of alcoholic patients with oral carcinomas

The micronucleus test (MN) is used as an indicator of genotoxic exposition, since it is associated with chromosome aberrations. An increased mutation rate in oral squamous cells, indicated by an increased MN frequency, is also related to the development of oral carcinomas. We evaluated the frequencies of MN and other metanucleated anomalies in the buccal squamous cells of 30 alcoholics with oral or oropharyngeal carcinomas, and compared them to a control group of abstinent health individuals. Microscopic examination was made of 2000 cells per individual from each of three distinct areas of the mouth: around the lesion (A), opposite to the lesion (B) and in the upper gingival-labial gutter (C); C was used as a control region because of low tumor frequency. There was a seven-fold increase in MN frequency in region B, a three-fold increase in region A and a two-fold, though nonsignificant, increase in C; indicating a gradient of frequencies towards carcinogenesis: C ® A ® B. Comparisons of frequencies of various types of metanucleated cells: binucleated, karyorrhexis (KR), karyolysis (KL) and broken egg (BE) in patients and controls showed, with few exceptions, highly significant differences. This gave us a better understanding of the dynamics of this squamous epithelium, supporting a more efficient biomonitor based on these various metanucleated anomalies: the repair index . Also, the apparently contradictory results from regression analysis revealed that the MN frequency decreased with age and alcohol consumption, probably because of slow cell proliferation, and consequently led to a loss of homeostasis due to aging. In addition, in the analysis of nonparametric variables only one CAGE question was significant, confirming the effect of alcohol. In conclusion, the MN test and the repair index could be used for monitoring clinical evolution, by means of intra- and inter-individual cellular comparisons, in subjects with healed or surgically removed tumors or leukoplastic lesions, after chemo- or radiotherapeutic treatments.

Circulating 92-kilodalton matrix metalloproteinase (MMP-9) activity is enhanced in the euglobulin plasma fraction of head and neck squamous cell carcinoma.

BACKGROUND: Cancer lethality is usually the result of local invasion and metastasis of neoplastic cells from the primary tumor. Because of their ability to degrade extracellular matrix components (EMC), matrix metalloproteinases (MMPs) have been implicated in the breakdown of basement membranes and underlying stroma, thereby facilitating tumor growth and invasion. METHODS: The authors quantitated, by gelatin zymography and densitometric analysis, MMP activity in the euglobulin plasma fraction of 50 healthy controls and 91 head and neck squamous cell carcinoma (HNSCC) patients (51 from the larynx and 40 from the oropharynx). RESULTS: The median value for 92-kilodalton (kD) MMP (MMP-9) activity was increased significantly in laryngeal (Md 2.1 arbitrary units (AU)/mL plasma; range, 0.2-6.4) and oropharyngeal patients (Md 2.08 AU/mL; range, 0.0-5.0) with respect to the controls (Md 0.48 AU/mL; range, 0.0-1.8). Both groups of cancer patients showed a similar behavior. Multivariate analysis indicated that circulating 92-kD MMP activity was not predicted by the clinical-pathologic parameters such as tumor stage, histologic grade, and metastatic lymph nodes. There was no association between high levels of MMP-9 activity and either cigarette smoking or alcohol consumption, major risk factors for developing HNSCC. CONCLUSIONS: The authors found a significant increase of MMP-9 plasma activity both in laryngeal and oropharyngeal squamous cell carcinoma patients as compared with healthy controls. Further studies are necessary to establish its usefulness in the clinical management of these patients.

Micronucleus investigation of alcoholic patients with oral carcinomas.

The micronucleus test (MN) is used as an indicator of genotoxic exposition, since it is associated with chromosome aberrations. An increased mutation rate in oral squamous cells, indicated by an increased MN frequency, is also related to the development of oral carcinomas. We evaluated the frequencies of MN and other metanucleated anomalies in the buccal squamous cells of 30 alcoholics with oral or oropharyngeal carcinomas, and compared them to a control group of abstinent health individuals. Microscopic examination was made of 2000 cells per individual from each of three distinct areas of the mouth: around the lesion (A), opposite to the lesion (B) and in the upper gingival-labial gutter (C); C was used as a control region because of low tumor frequency. There was a seven-fold increase in MN frequency in region B, a three-fold increase in region A and a two-fold, though nonsignificant, increase in C; indicating a gradient of frequencies towards carcinogenesis: C --> A --> B. Comparisons of frequencies of various types of metanucleated cells: binucleated, karyorrhexis (KR), karyolysis (KL) and broken egg (BE) in patients and controls showed, with few exceptions, highly significant differences. This gave us a better understanding of the dynamics of this squamous epithelium, supporting a more efficient biomonitor based on these various metanucleated anomalies: the repair index RI=(KL+KR)/(MN+BE). Also, the apparently contradictory results from regression analysis revealed that the MN frequency decreased with age and alcohol consumption, probably because of slow cell proliferation, and consequently led to a loss of homeostasis due to aging. In addition, in the analysis of nonparametric variables only one CAGE question was significant, confirming the effect of alcohol. In conclusion, the MN test and the repair index could be used for monitoring clinical evolution, by means of intra- and inter-individual cellular comparisons, in subjects with healed or surgically removed tumors or leukoplastic lesions, after chemo- or radiotherapeutic treatments.

Detection of oral and oropharyngeal cancer by microsatellite analysis in mouth washes and lesion brushings.

Microsatellite allele losses are characteristic features of head and neck squamous cell carcinoma and can be used as molecular markers for malignancy. We have investigated the detection of microsatellite allele loss in mouth washes and lesions brushings from 19 patients with squamous cell carcinoma of the oral cavity and oropharynx as a means of tumour detection. In 84% of the analysed cases, allele loss previously identified in the tumour of these patients, was detected in these easily obtained specimens. No alterations were found in material derived from 10 healthy individuals. Success of detection was independent of tumour stage, suggesting that this approach may be useful for early diagnosis as well as for follow-up.