Rare autosomal trisomies: comparison of detection through cell-free DNA analysis and direct chromosome preparation of chorionic villus samples.

OBJECTIVE: Direct chromosome preparations of chorionic villus samples (CVS) and cell-free DNA (cfDNA) testing both involve analysis of the trophoblastic cell lineage. The aim of this study was to compare the spectrum of rare autosomal trisomies (RATs) detected by these two approaches and assess the available information on their clinical significance. METHODS: Data from 10 reports on genome-wide cfDNA testing were pooled to determine which chromosomes were most frequently involved in RAT-positive cases, and pregnancy outcome information was reviewed. CVS information was obtained from an updated database of 76 102 consecutive CVS analyses performed over a period of 18 years at TOMA laboratory, in which trophoblastic and mesenchymal layers were analyzed and amniotic fluid cell analysis was recommended for RAT-positive cases. Chromosomes involved and presence of confined placental mosaicism, true fetal mosaicism and uniparental disomy (UPD) for imprinted chromosomes were assessed. Also evaluated were the frequency and types of RATs in products of conception. RESULTS: RATs were present in 634 of 196 662 (0.32%) cfDNA samples and 237 of 57 539 (0.41%) CVS trophoblast samples (P < 0.01). The frequency of RATs varied over 8-fold between the cfDNA reports. Confirmation of abnormality through amniocentesis was more likely when RATs were ascertained through cfDNA (14 of 151; 9.3%) than through CVS trophoblasts (seven of 237; 3.0%) (P < 0.01). In cfDNA-ascertained cases, trisomies 15, 16 and 22, which are associated with fetal loss, were identified proportionately more often. Of 151 cases with RAT identified by cfDNA and outcome information available, 41.1% resulted in normal live birth; 27.2% in fetal loss; 7.3% had phenotypic abnormality detected through ultrasound or other follow-up evaluation; 2.0% had a clinically significant UPD; and 14.6% had fetal growth restriction or low birth weight. All autosomes were involved in trisomies in products of conception; the most common RATs detected were trisomies 16, 22 and 15 with a frequency of > 9% each. CONCLUSIONS: Although there are strong parallels between RATs ascertained through cfDNA analysis and direct chromosome preparation of CVS, caution is needed in applying conclusions from CVS analysis to cfDNA testing, and vice versa. RATs identified through genome-wide cfDNA tests have uncertain risks for fetal loss, growth restriction or fetal abnormality. Copyright © 2019 ISUOG. Published by John Wiley & Sons Ltd.

Genetic analysis of osteogenesis imperfecta in a large Brazilian cohort.

INTRODUCTION: Osteogenesis imperfecta (OI) is a genetically and clinically heterogeneous disorder caused by disruption of type I collagen synthesis. Previous Brazilian molecular OI studies have been restricted to case reports or small cohorts. The Brazilian OI Network (BOIN) is a multicenter study collecting clinical OI treatment data from five reference centers in three regions of Brazil. OBJECTIVE: To describe the molecular analysis of a large cohort of OI registered at BOIN. METHODS: Targeted next-generation sequencing (NGS) was performed at a centralized laboratory with the Ion Torrent platform, covering 99.6 % of the coding regions of 18 OI-associated genes. Clinical information was obtained from a clinical database. RESULTS: We included 156 subjects in the molecular analyses. Variants were detected in 121 subjects: 65 (53.7 %) in COL1A1, 42 (34.7 %) in COL1A2, 2 (1.7 %) in IFITM5, one (0.8 %) in CRTAP, three (2.5 %) in P3H1, two (1.7 %) in PPIB, four (3.3 %) FKBP10, one (0.8 %) in SERPINH1, and one (0.8 %) in TMEM38B. Ninety-one distinct variants were identified, of which 26 were novel. Of the 107 variants identified in COL1A1 and COL1A2, 24.5 % cause mild OI, while the remaining 75.5 % cause moderate, severe, or lethal OI, of which 49.3 % are glycine to serine substitutions. A single variant in FKBP10 (c.179A>C; p.Gln60Pro) was found in three unrelated and non-consanguineous participants living in the same geographic area in Northeast Brazil, suggesting a possible founder effect. CONCLUSION: Consistent with the literature, 88.4 % of the subjects had a variant in the COL1A1 and COL1A2 genes, with 10 % inherited in an autosomal recessive manner. Notably, one variant in FKBP10 with a potential founder effect requires further investigation. Data from this large cohort improves our understanding of genotype-phenotype correlations for OI in Brazil.

Microarray application in prenatal diagnosis: a position statement from the cytogenetics working group of the Italian Society of Human Genetics (SIGU), November 2011.

A precise guideline establishing chromosomal microarray analysis (CMA) applications and platforms in the prenatal setting does not exist. The controversial question is whether CMA technologies can or should soon replace standard karyotyping in prenatal diagnostic practice. A review of the recent literature and survey of the knowledge and experience of all members of the Italian Society of Human Genetics (SIGU) Committee were carried out in order to propose recommendations for the use of CMA in prenatal testing. The analysis of datasets reported in the medical literature showed a considerable 6.4% incidence of pathogenic copy number variations (CNVs) in the group of pregnancies with sonographically detected fetal abnormalities and normal karyotype. The reported CNVs are likely to have a relevant role in terms of nosology for the fetus and in the assessment of reproductive risk for the couple. Estimation of the frequency of copy number variations of uncertain significance (VOUS) varied depending on the different CMA platforms used, ranging from 0-4%, obtained using targeted arrays, to 9-12%, obtained using high-resolution whole genome single nucleotide polymorphism (SNP) arrays. CMA analysis can be considered a second-tier diagnostic test to be used after standard karyotyping in selected groups of pregnancies, namely those with single (apparently isolated) or multiple ultrasound fetal abnormalities, those with chromosomal rearrangements, even if apparently balanced, and those with supernumerary marker chromosomes.

Prenatal BACs-on-Beads™ : a new technology for rapid detection of aneuploidies and microdeletions in prenatal diagnosis.

OBJECTIVE: Molecular cytogenetic techniques on uncultured prenatal samples are the sole tests applied in some countries in cases with advanced maternal age (AMA) or increased risk after prenatal screening. Moreover, there is a trend to perform invasive prenatal diagnosis (PD) during the first trimester before ultrasound manifestations, so new rapid and reliable assays are necessary to investigate microdeletions not detectable with the conventional karyotype. We report the validation study of the prenatal bacterial artificial chromosomes-on-Beads™ (BoBs™ ; CE-IVD), a bead-based multiplex assay detecting chromosomes 13, 18, 21, X/Y aneuploidies and nine microdeletion regions having an overall detection rate of 1/1700. METHOD: We retrospectively studied 408 selected samples and prospectively tested 212 consecutive samples ascertained for conventional karyotyping. RESULTS: We did not find false-positive results. Triploidies were not detected. Maternal cell contamination of male samples up to 90% was unmasked inspecting gonosome profiles. Mosaic conditions at 20 to 30% were revealed. Failures were due to low amount of DNA. CONCLUSION: Prenatal BoBs™ is a robust technology for the investigation of fetuses with normal karyotype with or without sonographic abnormalities. Running in parallel with the karyotype analysis, it can be proposed instead of rapid FISH or QF-PCR providing rapid results on common aneuploidies and additional information regarding the microdeletion syndromes.

Effectiveness of standard cervical physiotherapy plus diaphragm manual therapy on pain in patients with chronic neck pain: A randomized controlled trial.

BACKGROUND: Literature concerning the effect of diaphragm treatment to reduce neck pain symptoms is scarce. Aim of this trial was to investigate the effects of diaphragm manual therapy associated with standard physiotherapy treatment on pain in patients with Chronic Neck Pain (CNP). METHODS: In a private practice clinic, subjects with CNP were randomly assigned to receive three 30-min treatment sessions of standard cervical physiotherapy and Diaphragm Manual Therapy (DMT) or Sham Diaphragm Technique (SDT). Participants and assessors were blinded to the assignment. Primary outcome was pain, secondary outcomes were cervical active range of motion, pain pressure threshold, disability and quality of life measured at baseline, before and after each session, at 3 and 6-months. Adverse events were monitored. A non-parametric multivariate approach (combined permutation test) was applied to assess the effect of the treatment on all the outcomes. An intention to treat analysis was performed. RESULTS: Forty patients were randomly allocated to DMT and SDT groups. Combined permutation test showed a significant higher improvement in DMT group compared to SDT group (p-value = 0.0002). The between-group comparisons on single outcomes showed a statistically significant improvement only for pain pressure threshold on upper trapezius (adjusted p-value = 0.029). No adverse events related to the intervention were registered. CONCLUSIONS: In patients with CNP, addition of diaphragm manual techniques to standard cervical treatment seems to give a better global outcome, but this improvement is of unclear clinical relevance; the primary outcome seems not to have a role. Further studies are needed to confirm and clarify these results. TRIAL REGISTRATION: Release Date: July 18, 2017 Registered in ClinicalTrial.gov database ID: NCT03223285A.

ESX1 gene expression as a robust marker of residual spermatogenesis in azoospermic men.

BACKGROUND: It would be of value to identify ongoing spermatogenesis molecular markers which can predict successful sperm recovery in patients with non-obstructive azoospermia undergoing conventional or microsurgical testicular sperm extraction (TESE/microTESE). ESX1 is an X-linked homeobox gene expressed in testis, placenta, brain and lung in humans and specifically in pre- and post-meiotic germ cells of the testis in mice. METHODS: We investigated the sequence, expression (by RT-PCR) and epigenetic status (by promoter pyrosequencing) of ESX1 in testicular tissue samples, obtained from 81 azoospermic subjects in the context of surgical sperm extraction, to check a possible association between ESX1 alterations and impaired spermatogenesis, as determined by histological analysis. RESULTS: The ESX1 transcript was detected in 100% of cases diagnosed as obstructive azoospermia (33), hypospermatogenesis (18) and incomplete maturation arrest (MA) (2), and sperm recovery was also successful in 100% of these cases. ESX1 mRNA was also detected in 5 of 6 patients with incomplete Sertoli cell-only syndrome, in 4 of 6 subjects with complete MA but in only 3 of 16 cases of complete Sertoli cell-only syndrome (cSCOS), whereas sperm recovery was successful in 4 of 6, 2 of 6 and 5 of 16 of these patients, respectively. In cases of focal spermatogenesis, ESX1 expression and sperm retrieval were concordant in 14 of 19 (74%) cases subjected to TESE, but in only 3 of 11 (27%) men who underwent microTESE. With TESE, but not with microTESE, both samples originated from adjacent testicular areas. The pyrosequencing of the ESX1 CpG island revealed methylation levels that were significantly lower in ESX1 expressors when compared with non-expressors. CONCLUSIONS: ESX1 emerges as a potentially reliable spermatogenesis molecular marker, whose clinical value as a predictor of successful sperm retrieval warrants further studies.

Arthrodesis of the ankle joint in septic osteoarthritis: six years long term outcomes in authors' personal experience.

OBJECTIVE: Our goal was to evaluate the long-term clinical outcome of ankle arthrodesis, obtained by an extramedullary internal fixation with or without bone-grafting in the treatment of ankle' septic arthritis. PATIENTS AND METHODS: All patients treated with arthrodesis by extramedullary internal fixation for septic arthritis of the ankle joint between January 2011 and December 2016 in the same hospital were included in our retrospective study. Patients were followed-up for a minimum of two years. To evaluate the quality of life, each patient filled in a short form of the physical and mental health summary scale and a visual analogue scale for pain. For the functional evaluation, the American Orthophaedic Foot and Ankle Society Score was used. Demographics and clinical data, including perioperative and postoperative complications, were evaluated. RESULTS: From January 2011 to December 2016, we performed 57 arthrodeses of the ankle joint with cannulated screws in 52 patients. Mean age was 52 years old. 48 patients (92%) had post-traumatic septic arthritis. The most frequently isolated pathogens were Staphylococcus aureus and Pseudomonas aeruginosa. 48% of patients reported a postoperative complication after three months; the most commonly reported complications were weight bearing ankle-foot pain (27%) and surgical wound dehiscence (12.25%). Nonunion was reported only in 8.75% of cases. CONCLUSIONS: Ankle arthrodesis could allow painless gait, improving patients life quality, even after long-term follow-up, and is, therefore, a solution to be considered in patients affected by septic arthritis.

Chromosome 11 segmental paternal isodisomy in amniocytes from two fetuses with omphalocoele: new highlights on phenotype-genotype correlations in Beckwith-Wiedemann syndrome.

BACKGROUND: The phenotypic variability in Beckwith-Wiedemann syndrome (BWS) reflects the genetic heterogeneity of the mechanism which by default leads to the deregulation of genes located at 11p15.5. Genotype-phenotype correlation studies have demonstrated an association between omphalocoele and CDKN1C/p57 mutations or hypermethylation. Paternal uniparental disomy 11 (pUPD11) has been described only in the mosaic condition with both uniparental and biparental cell lines, and no association with omphalocoele has been pointed out. METHODS: Two cases are presented here, in which a paternal segmental UPD11 was detected by molecular investigation of amniotic fluid cell cultures after the presence of apparently isolated omphalocoele was revealed in the fetuses by ultrasound scan. Further studies were performed on additional autoptic feto-placental tissues to characterise the distribution of the uniparental cell line and to unmask any biparental lineage in order to document in more detail the as yet unreported association between omphalocoele and pUPD11. RESULTS: Results on the UPD distribution profile showed that the abdominal organs have a predominant uniparental constitution. This condition could mimic the effect of CDKN1C/p57 inactivation, causing the omphalocoele. CONCLUSION: New genotype-phenotype correlations emerge from the investigated cases, suggesting that molecular analysis be extended to all cases with fetal omphalocoele in order to establish the incidence of pUPD11 in complete BWS and in monosymptomatic/mild forms.

Fetal and placental chromosomal mosaicism revealed by QF-PCR in severe IUGR pregnancies.

A number of genetic and environmental factors are taken into account as responsible for intrauterine growth restriction (IUGR); nevertheless, the relevance of genetic alteration in IUGR aetiology remains to be determined. The aim of this study was to investigate using a combined cytogenetic-molecular approach, improved by a new application of QF-PCR method, the presence of mosaic chromosomal changes in fetal/placental samples from 12 pregnancies with unexplained severe IUGR. This multiple approach allowed us to reveal and quantify subtle chromosomal mosaicisms with less than 5% of trisomic cells even in cases in which cytogenetic and FISH analyses failed to reveal them. These are three pregnancies with a mosaic trisomy for chromosomes 7, 2 and 14; the former case presented matUPD7 and was previously described in this journal (Placenta 22 (2001) 813) in association with pre- and postnatal growth restriction. It is intriguing that chromosomes 7, 2 and 14 are known or suspected to harbour imprinted genes, so that an unbalanced gene dosage in a subset of cells during embryonic development could lead to an early impairment of placental function. Our findings indicate that extensive molecular and cytogenetic studies of IUGR fetal and placental tissues are necessary to reveal at least part of the heterogeneous genetic lesions implicated in IUGR phenotypes.

HIV-1 proviral DNA polymerase chain reaction detection in chorionic villi after exclusion of maternal contamination by variable number of tandem repeats analysis.

OBJECTIVE: The study of the placental HIV infection in cases of seropositive pregnant women after exclusion of maternal contamination of chorionic villi samples by variable number of tandem repeats (VNTR) analysis. METHODS: We studied 30 HIV-positive women: 17 terminated their pregnancy (11 in the first trimester and six in the second) and 13 delivered at term (one was a twin gestation). We selected chorionic villi and ruled out maternal contamination by VNTR analysis. DNA from chorionic villi and cord and maternal blood were tested for HIV by PCR. All infants underwent a paediatric follow-up. RESULTS: All maternal blood samples tested positive for HIV-1 by polymerase chain reaction. No maternal contamination was revealed and HIV was found in six out of 11 first trimester placentas, in all second trimester samples, and in 10 out of 14 at term. Cord blood tested positive in all second trimester cases and in seven out of 14 liveborns. In no case was HIV found in cord blood without infection of the corresponding placenta; conversely, three placentas tested positive but cord blood was negative. Two infants were HIV-positive, 11 were uninfected (one case was lost to follow-up). CONCLUSION: Our study indicates that HIV-1 can infect the placenta from first trimester onwards. HIV was found in two-thirds of our cord blood samples but it is possible that some viral DNA in cord blood may have come from infected placental cells. Additional studies are needed to assess the source of HIV in cord blood and the possible contribution of placental or maternal cells infected with HIV to vertical transmission of the virus.

Loss of heterozygosity of the NOS3 dinucleotide repeat marker in atherosclerotic plaques of human carotid arteries.

We have investigated 28 atherosclerotic plaques of human carotid arteries with a panel of 39 microsatellite markers for the presence of LOH. The objective of this research was to verify if LOH, described in association with tumorigenic process, could be involved also in benign fibroproliferative disease. Seventy percent of samples demonstrated allelic imbalance: 50% of cases showed LOH at a minimum of one locus, 3.5% at a minimum of two loci and 14.3% at three or more loci. The percentages of LOH ranged between 3.8 and 14.3% and the highest involved polymorphic marker is the NOS3 internal dinucleotide repeat. Our results indicate that, like tumorigenesis, the atherogenic process could also involve LOH mechanism. Furthermore, the finding regarding the NOS3 internal polymorphism suggests a possible role of the gene as cofactor in formation of the atheromas.

First-trimester human trophoblast is class II major histocompatibility complex mRNA+/antigen.

Lack of expression of the polymorphic class I and class II MHC antigens in the cytotrophoblast is one of the major factors determining the privileged immunologic status of the placenta. In this report, we show that first-trimester human placental cytotrophoblast cells display moderate to strong expression of class II MHC (HLA-DR alpha and -DR beta) and Ii chain transcripts, apparently in absence of detectable class II antigens and Ii chain. In addition, DR alpha, DR beta, and Ii mRNAs, but not antigens, are consistently upregulated by IFN-gamma. Constitutive expression and upregulation of mRNAs are detectable in trophoblast cells kept in short term as well as prolonged (2-3 weeks) culture. These results are reminiscent of an analogous mRNA+/antigen- dissociation occurring, in the case of class I MHC gene products, in a subpopulation of first-trimester cytotrophoblast cells. Thus, analogous mechanisms prevent the expression of potentially hazardous class I and II allodeterminants at early stages of semiallogeneic pregnancy.

Post-zygotic origin of complete maternal chromosome 7 isodisomy and consequent loss of placental PEG1/MEST expression.

Maternal UPD of chromosome 7 is associated with pre- and postnatal growth retardation (IUGR, PNGR) and Silver-Russell syndrome (SRS [MIM 180860]). We report a case of IUGR in a newborn with SRS stigmata. Using combined haplotyping and cytogenetic-FISH studies we characterized the lymphocytes, umbilical cord and four placental cotyledons. The results are consistent with complete maternal isodisomy 7 and trisomy 7 mosaicism of post-zygotic origin. The trisomic cell line was prevalent in trophoblast cells from two placental cotyledons. Trisomy 7 of post-zygotic origin is a frequent finding, but maternal isodisomy 7, due to trisomic rescue has never been reported. PEG1/MEST expression was evaluated on placenta cDNA and a specific transcript was revealed only in the cotyledons with a high percentage of trisomic cells and the presence of the paternal chromosome 7 contribution, but not in the placental biopsies with maternal isodisomy 7. The histological features of the four placental fragments revealed that isodisomy 7 correlates with a pattern of cotyledonary hyper-ramification due to an increase of the branching angiogenesis, which could be the result of a defect of angiogenesis caused by the absence of PEG1 product. The severe hypo-ramification of the two cotyledons, showing trisomy 7 mosaicism, may be due to the triplicate dosage of genes on chromosome 7. The delayed fetal growth could be the phenotypic effect of the imbalance between imprinted and non-imprinted genes on chromosome 7 in the fetus or the result of abnormal placental function during pregnancy.

Direct chromosome preparation and culture using chorionic villi: an evaluation of the two techniques.

We present a comparison between direct chromosome preparation and cell culture for first trimester fetal chromosome study using chorionic villi. The 2 techniques have advantages and disadvantages and are demonstrated to be appropriate for routine diagnostic work. The combined use of both methods may optimize the quality of the chromosome study and minimize the possibility of false-positive and false-negative findings.

Genetic diagnosis before the eighth gestational week.

Transabdominal chorionic villus sampling (CVS) by a freehand, ultrasound-guided technique was offered to 210 high-genetic-risk women at 6-7 weeks' gestation. It was carried out in 201 cases and postponed in nine cases (4.3%). Sampling was successful in 86 and 100% of cases after the first and the second needle insertions, respectively. Chorionic tissue specimens weighed at least 20 mg in 86% of cases, and only 2% were below 10 mg. Early complications were present in 7.9% of cases, apparently without any adverse effect on maternal or fetal outcome. The rate of fetal loss was 3.5%. Genetic diagnosis was concluded in 1-3 days by rapid diagnostic methods. Although more extensive laboratory and clinical experience is necessary to evaluate adequately the safety of early transabdominal CVS, it may be advantageous to offer this technique to certain high-genetic-risk patients. The availability of genetic diagnosis before the eighth week makes clinical abortion by antiprogestins and prostaglandins a viable option in cases of affected embryos.

Cytogenetic abnormalities detected by direct analysis in a case of choriocarcinoma.

Malignant trophoblastic cells from a case of choriocarcinoma were cytogenetically investigated by direct analysis of fresh tissue from the tumor. To our knowledge, previous cytogenetic studies have been performed only on established cell lines. In this study, 54 metaphases were observed, of which 41 were fully karyotyped. Three chromosomally abnormal lines were identified. In all of them, trisomy 3 and 10, a supernumerary isochromosome 1q,i(1)(q10), an i(8)(q10) replacing one chromosome 8, and a marker chromosome were observed. In addition, involvement of chromosome 12 was observed in two of the three lines, trisomic in one and an i(12)(q10) in the other.

Multiple clonal chromosome abnormalities in Peyronie's disease.

Numerical and structural chromosome aberrations were found in cell plaque metaphases from 9 of 14 patients with Peyronie's disease. In two cases there was evidence of clonal evolution for some of the chromosomal aberrations observed. The Y chromosome was the most frequently involved in numerical changes. Four of 9 cases with abnormal karyotype showed more than one abnormal, cytogenetically unrelated clone. Our findings suggest the possibility of a multiclonal origin for this benign tumor, and confirm the presence of chromosome instability in this cell growth disorder.

Cytogenetic abnormalities and microsatellite instability in endometrial adenocarcinoma.

Recently various authors described a new mechanism involved in the genesis of some tumors, which is characterized by a tendency for replication mistakes and by genomic instability of microsatellite repeats. This instability can be revealed through the shift in the electrophoretic mobility of the analyzed fragments, which is due to a different number of repeat units. This phenomenon is widely documented in colorectal tumors of patients affected by hereditary nonpolyposis colorectal carcinoma (HNPCC). We performed a cytogenetic and molecular study of 23 endometrial adenocarcinomas to investigate the presence of genomic instability and to evaluate the possibility of a positive correlation with specific chromosomal changes. The study of genomic instability was performed using 23 microsatellites localized over 8 chromosomes. Genomic instability of microsatellites was observed in 3 cases over all 8 analyzed chromosomes. The tumoral stage of cases with microsatellite instability does not differ significantly from the remaining tumors. As a matter of fact several cases showing no evidence of instability were more advanced (II B, III A) than tumors with instability. In ten cases we observed trisomy of chromosome 10, in some as a sole anomaly. The 3 cases with genomic instability revealed a near-diploid karyotype and all showed the presence of a supernumerary marker derived from chromosome 1 rearrangements. A derivative chromosome 1 was revealed in 4 cases without evidence of microsatellite instability. It should be noted that the presence of many unidentified markers and the small number of tumors with instability do not allow us to give a definitive significance to this observation. Our results indicate that there is not an apparent correlation between microsatellite instability and specific chromosomal abnormalities. Moreover, we did not find any correlation between pathological characteristics of the tumor and genomic instability. Microsatellite instability appears to be a relatively rare event in endometrial carcinoma.

Losses of heterozygosity in oral and oropharyngeal epithelial carcinomas.

We analyzed 25 oral and oropharyngeal epithelial carcinomas for loss of heterozygosity (LOH) and microsatellite instability by using 55 oligonucleotide repeat markers located in 45 chromosomal regions. The aim was to identify which chromosomal regions and tumor-suppressor genes (TSGs) are preferentially lost in these tumors and to relate LOH at specific loci to clinicopathologic data. The analysis was performed on tumor tissue and on a corresponding normal tissue (blood lymphocytes) with the use of the polymerase chain reaction technique followed by microsatellite allele separation with denaturing gel electrophoresis. Thirty-two of 45 chromosomal regions demonstrated a significant (>/=20%) incidence of LOH. An allelic loss of >/=50% was found in 9p21 (77.8%), 8p22-23 (70%), 3p12 (61.5%), 1p36.1 and 12q22 (60%), 3q28 (57.1%), 5q23.3 (54.5%), 3p25-26, 3p24, and 7q35 (50%). We did not find any microsatellite instability. Our results suggest that in addition to a group of TSGs, pleiotropic for several tumor types, other suppressor genes are specifically involved in oral and oropharyngeal carcinogenesis.