Transmission and clearance of human papillomavirus infection in the oral cavity and its role in oropharyngeal carcinoma - A review.

The majority of sexually active individuals becomes infected with human papillomavirus (HPV) at least once in their lifetime. Pathways for HPV transmission vary across different mucosal sites per individual. They include autoinoculation within one host, direct transmission between individuals (including perinatal transmission and transmission during sexual activity), and indirect transmission through contact with hands. The authors aim to clarify the prevalence and route of transmission per anatomic site, inter- and intra-individually, using a narrative review of the literature. In conclusion, transmission of HPV to the oral cavity and oropharynx is hypothesised to occur mainly through sexual contact. Transmission of particles through saliva has not been proven and daily living activities are not a documented source of HPV infection. Oropharyngeal HPV related cancer survivors and their partners do not show increased risk of infection during sexual intercourse. Transmission of HPV to the oral cavity (autoinoculation with fingers or transmission through saliva in deep kissing) is probably of limited importance.

Reducing lead uptake by plants as a way to lead-free food.

In this research, an attempt was made to produce safe food from lead-contaminated soil. It was assumed that an increased amount of calcium (Ca) in plants would prevent them from lead (Pb) uptake. A new-generation agricultural product - an activator of Ca transport in plants "InCa" (from Plant Impact) - was used. The study was conducted on several crop species, Cucumis sativus L., Linum usitatissimum L., Medicago sativa L. and Solanum lycopersicum L., cultivated in mineral medium. The leaves were sprayed with InCa activator while the roots received Pb from the substrate in the form of Pb(NO3)2 dissolved in the medium. It was shown that spraying the leaves with InCa reduced Pb concentration in the roots of S. lycopersicum to 73%, in C. sativus to 60%, and in L. usitatissimum to 57%. Finally, it was found that foliar application of InCa reduced the concentration of Pb in plant roots by 53%, and in plant shoots by 57% (on average by about 55%). These observations were confirmed using histochemical and electron microscopy techniques. It was shown that one of the InCa activator components - Ca(NO3)2 - is responsible for such effects. This result was verified by using another experimental method - the Allium epidermis test. Visualization of Pb in epidermal cells of Allium cepa. L. using the Leadmium™Green fluorescent probe (confocal microscopy) showed a reduction in the amount of Pb that entered the epidermal cells after the application of the tested solutions. For the first time, it was shown that it is possible to reduce Pb uptake by plants by up to 55%. In the future, this offers the possibility of developing a foliar calcium preparation aimed at lowering the concentration of Pb in plants and thereby reducing the amount of Pb in the food chain.

Micro-Evolutionary Processes in Armeria maritima at Metalliferous Sites.

Tolerance to heavy metals in plants is a model process used to study adaptations to extremely unfavorable environments. One species capable of colonizing areas with high contents of heavy metals is Armeria maritima (Mill.) Wild. A. maritima plants growing in metalliferous areas differ in their morphological features and tolerance levels to heavy metals compared to individuals of the same species growing in non-metalliferous areas. The A. maritima adaptations to heavy metals occur at the organismal, tissue, and cellular levels (e.g., the retention of metals in roots, enrichment of the oldest leaves with metals, accumulation of metals in trichomes, and excretion of metals by salt glands of leaf epidermis). This species also undergoes physiological and biochemical adaptations (e.g., the accumulation of metals in vacuoles of the root's tannic cells and secretion of such compounds as glutathione, organic acids, or HSP17). This work reviews the current knowledge on A. maritima adaptations to heavy metals occurring in zinc-lead waste heaps and the species' genetic variation from exposure to such habitats. A. maritima is an excellent example of microevolution processes in plants inhabiting anthropogenically changed areas.

The holmium:YAG laser lithotripsy-a non-invasive tool for removal of midsize stones of major salivary glands.

Incorporeal lithotripsy by using holmium:YAG laser has been recently successfully spread and used in the treatment of urolithiasis. Although this method is not as common in salivary gland surgery, it is also recommended for endoscopic treatment of sialolithiasis. In this study, we decided to analyse effectiveness of holmium:YAG laser method among patients with midsize stones as the treatment in this particular population is still not determined and remains in dispute. Thus, the primary aim of the study is a prospective, complex analysis of incorporeal holmium:YAG laser lithotripsy effectiveness and safety in patients suffering from sialolithiasis. The examined group consisted of 32 consecutive patients, 17 female (53%) and 15 male (47%). The average age was 42 (age range 22-70). In our series, sialoendoscopy with intraductal lithotripsy was performed in patients with wide range of stone's diameter (3 to 10 mm). The calculi were removed with foreign body forceps or retrieval basket, after stone fragmentation with holmium:YAG laser. The power of the laser during surgery was set at 2.5 to 3.5 W. One shot energy equalled 0.5-0.7 J with frequency of 5 Hz. The bivariate Pearson Correlation and a point-biserial correlation were used to analyse the data. Total success rate of holmium:YAG laser lithotripsy in this consecutive group was equal to 90% and efficacy in the group with excluded cases when endoscopic approach difficulties were experienced (stone not visible) reached 93%. The long-term follow-up revealed that in 26 patients (84%), complete recovery was achieved. Two patients developed Wharton's duct stenosis and one Stensen's duct orifice stenosis (9.5%), of whom one required reoperation. Two patients eventually underwent total excision of submandibular gland. Sialoendoscopy combined with incorporeal laser-assisted lithotripsy proved to be effective and safe method for treatment of sialothiasis with midsize stones (4-8 mm) in population of clinic. Even taking into account the prevalence of complications associated with holmium:YAG laser, it is still safer alternative compared to open surgery where there is a significant risk of the bleeding, wound infection or injury of the facial nerve.

"Sandwich technique" enables preservation of hearing and antivertiginous effect in cholesteatomatous labyrinthine fistula.

PURPOSE: The aim of the study was to assess hearing, surgical and clinical results of the treatment in patients with cholesteatomatous labyrinthine fistula (LF) focusing on the different techniques and materials used in the management. METHODS: Study group included 465 patients. Cases with LFs discovered or confirmed during surgical procedure were thoroughly analyzed. RESULTS: LFs were noted in 11.4% of all cases. Thirty-eight patients, with all follow-up data available, were included into the further analysis. Most LFs were located in the lateral semicircular canal (87%). LFs were assessed as small in 2 cases, as medium in 24 patients while 12 were described as large. Based on Dornhoffer and Milewski classification, 50% of LFs were classified as IIa, 24% as IIb, 6 LFs were very deep (type III), while 4-superficial (type I). The size and type of LF did not influence postsurgical complaints (p = 0.1070, p = 0.3187, respectively). Vertigo was less frequent in LFs treated by "sandwich technique", especially those with opened endosteum. In 30 (79%) patients, hearing improved or did not change after surgery. Hearing outcomes were significantly better in the ears operated by means of CWU technique (p = 0.0339), in LFs with intact membranous labyrinth (p = 0.0139) and when "sandwich technique" was performed (p = 0.0159). Postsurgical bone conduction thresholds levels were significantly better in LFs covered by "sandwich method" (p = 0.0440). CONCLUSION: "Sandwich technique" (temporal fascia-bone pate-temporal fascia) enables preservation of hearing as well as antivertiginous effect in patients with cholesteatomatous labyrinthine fistula.

Facial nerve paresis in the course of masked mastoiditis as a revelator of GPA.

PURPOSE: The aim of this study was to present a series of 6 patients with facial nerve palsy and masked mastoiditis which constituted as revelators of localized granulomatosis with polyangiitis (GPA) and to evaluate the utility of the ACR/EULAR 2017 provisional classification criteria for GPA in such cases. METHODS: Study group included 58 patients with GPA. Cases with facial nerve palsy and masked mastoiditis were thoroughly analyzed. RESULTS: The mean age of patients was 37 years. All manifested unilateral facial nerve palsy and hearing loss, while only 2 reported aural complaints suggesting inflammatory cause of the disease. All cases were qualified for surgical intervention. Intraoperative findings were similar: granulation tissue in tympanic cavity and/or pneumatic spaces of the mastoid process. Only 50% of histopathological results suggested vasculitis. In all cases, elevated levels of antineutrophil cytoplasmic antibodies (ANCA) against peroxidase 3 (PR3-ANCA) were determined. Two patients presented rapid progression of the disease and died within 1 week and 2 months, respectively. Four other patients manifested gradual improvement of hearing and facial nerve function after treatment. CONCLUSION: GPA should be included into differential diagnosis in all cases of persistent facial nerve palsy especially when otological symptoms coexist. Even localized GPA could be very aggressive, revelating generalized form of the disease. Rapid systemic treatment of GPA can protect hearing and facial nerve from permanent severe dysfunction. The ACR/EULAR 2017 provisional classification criteria for GPA seem to be valuable tool in diagnosing ENT patients with localized otological form of the disease.

Is the 3D exoscope better than the surgical microscope in parotid surgery: a prospective, randomized single-center study.

BACKGROUND: High-definition, three-dimensional (3D) exoscopes are being used to perform a growing number of head and neck surgeries. However, the use of the 3D exoscope in parotid gland surgery has not been previously described. Our initial experience with the VITOM 3D exoscope in the surgical treatment of parotid gland tumors is detailed here. METHODS: We made a prospective study of patients with benign parotid gland tumors indicated for surgical resection. Between January and December 2018, patients were randomly assigned to undergo surgery assisted with the VITOM 3D system (n = 31) or an operating microscope (n = 40). Visualization quality (greater auricular nerve, digastric muscle, tragal pointer), operating time, conversion rates, and surgical outcomes were compared. RESULTS: A total of 71 patients underwent superficial (n = 18) or total parotidectomy (n = 53). No exoscope-related complications were observed. Five patients undergoing exoscope-guided deep lobe surgery required intraoperative conversion to a microscope. No differences were observed in the subjective quality of intraoperative visualization of key anatomical structures. However, a significantly higher percentage of patients in the exoscope group developed transient facial nerve paralysis (n = 9; 29% vs. n = 4, 10%). CONCLUSIONS: These findings suggest that the VITOM 3D is a valid visualization tool for parotid gland surgery, comparable to the operating microscope but with higher resolution 3D visualization, an increased degree of freedom of movement, and better ergonomics. However, the high rate of transient nerve palsy, possibly related to decreased depth perception and the brief learning curve, merits further investigation.

Informed consent for suspension microlaryngoscopy: what should we tell the patient? A consensus statement of the European Laryngological Society.

INTRODUCTION: Informed consent for any surgical intervention is necessary, as only well-informed patients can actively participate in the decision-making process about their care, and better understand the likely or potential outcomes of their treatment. No consensus exists on informed consent for suspension microlaryngoscopy (SML). MATERIALS AND METHODS: Informed consent procedures in nine countries on five continents were studied. RESULTS: Several risks can be discerned: risks of SML as procedure, anesthesiologic risks of SML, specific risks of phonosurgery, risks of inadequate glottic exposure or unexpected findings, risks of not treating. SML has recognized potential complications, that can be divided in temporary (minor) complications, and lasting (major) complications. CONCLUSION: SML is a safe procedure with low morbidity, and virtually no mortality. Eleven recommendations are provided.

Global DNA Methylation Profiling Reveals Differentially Methylated CpGs between Salivary Gland Pleomorphic Adenomas with Distinct Clinical Course.

Pleomorphic adenomas (PAs) are the most frequently diagnosed benign salivary gland tumors. Although the majority of PAs are characterized by slow growth, some develop very fast and are more prone to recur. The reason for such differences remains unidentified. In this study, we performed global DNA methylation profiling using the Infinium Human Methylation EPIC 850k BeadChip Array (Illumina) to search for epigenetic biomarkers that could distinguish both groups of tumors. The analysis was performed in four fast-growing tumors (FGTs) and four slow-growing tumors (SGTs). In all, 85 CpG dinucleotides differentiating both groups were identified. Six CpG tags (cg06748470, cg18413218, cg10121788, cg08249296, cg18455472, and cg19930657) were selected for bisulfite pyrosequencing in the extended group of samples. We confirmed differences in DNA methylation between both groups of samples. To evaluate the potential diagnostic accuracy of the selected markers, ROC curves were constructed. We indicated that CpGs included in two assays showed an area under the curve with an acceptable prognostic value (AUC > 0.7). However, logistic regression analysis allowed us to indicate a more optimal model consisting of five CpGs ((1) cg06748470, (2) cg00600454, (3) CpG located in chr14: 77,371,501−77,371,502 (not annotated in GRCh37/hg19), (4) CpG2 located in chr16: 77,469,589−77,469,590 (not annotated GRCh37/hg19), and (5) cg19930657) with AUC > 0.8. This set of epigenetic biomarkers may be considered as differentiating factors between FGT and SGT during salivary gland tumor diagnosis. However, this data should be confirmed in a larger cohort of samples.

Survival on railway tracks of Geranium robertianum-a glyphosate-tolerant plant.

Geranium robertianum is a herbaceous plant that prefers shady and fertile forest habitats. However, it also occurs on railway tracks, where there are difficult conditions for plant growth and regular herbicide spraying (in high concentrations, twice a year). One of the most commonly used herbicides in railway areas is glyphosate. The effect of the glyphosate on the G. robertianum plants found on railway tracks and in nearby forests in north-eastern Poland was checked. The aim of the study was to explain how G. robertianum can survive on railway tracks despite spraying with the glyphosate. Increased tolerance to the glyphosate of the G. robertianum plants from track populations was demonstrated compared to the plants from forest populations that had not previously been in contact with the herbicide. After 35 days after treatment with the herbicide, 75% of the plants from the observed forest populations withered, while only 38% did from the track populations. Ultrastructure of plant leaf cells from forest populations was strongly disturbed, which was not observed in plants from track populations. It was also shown that plants from track populations accumulated more glyphosate and AMPA in their tissues than plants from forest populations. The obtained results indicate that long-term use of herbicides may cause formation of biotypes of plants resistant to a given herbicide. This fact explains the possibility of G. robertianum occurring on railway tracks, despite spraying with the glyphosate. It is also a manifestation of microevolutionary processes.

Head and Neck Paragangliomas-A Genetic Overview.

Pheochromocytomas (PCC) and paragangliomas (PGL) are rare neuroendocrine tumors. Head and neck paragangliomas (HNPGL) can be categorized into carotid body tumors, which are the most common, as well as jugular, tympanic, and vagal paraganglioma. A review of the current literature was conducted to consolidate knowledge concerning PGL mutations, familial occurrence, and the practical application of this information. Available scientific databases were searched using the keywords head and neck paraganglioma and genetics, and 274 articles in PubMed and 1183 in ScienceDirect were found. From these articles, those concerning genetic changes in HNPGLs were selected. The aim of this review is to describe the known genetic changes and their practical applications. We found that the etiology of the tumors in question is based on genetic changes in the form of either germinal or somatic mutations. 40% of PCC and PGL have a predisposing germline mutation (including VHL, SDHB, SDHD, RET, NF1, THEM127, MAX, SDHC, SDHA, SDHAF2, HIF2A, HRAS, KIF1B, PHD2, and FH). Approximately 25-30% of cases are due to somatic mutations, such as RET, VHL, NF1, MAX, and HIF2A. The tumors were divided into three main clusters by the Cancer Genome Atlas (TCGA); namely, the pseudohypoxia group, the Wnt signaling group, and the kinase signaling group. The review also discusses genetic syndromes, epigenetic changes, and new testing technologies such as next-generation sequencing (NGS).

Oral-genital human papillomavirus infection in Polish couples: frequent detection of HPV 42.

BACKGROUND: Human papillomavirus (HPV) contributes to the development of cervical and oropharyngeal tumors. The increased incidence of HPV associated oropharyngeal tumors is lately being observed also in Polish population. The worldwide distribution of HPV varies and the studies rarely combine analysis of virus genotypes in both: genital and oropharyngeal locations. Therefore, in our study, we investigated HPV distribution in both anatomical sites of females with previous history of cervical cancer or with pre-cancerous lesion and their partners to establish the dominant types in Polish couples in genital and oropharyngeal areas, as they can be easily sexually transmitted. METHODS: The study group consisted of 197 females and their partners. Each female had current or previous cervical pathology and HPV detected in gynecological swab with the use of Anyplex™ II HPV28 Detection system. This system is based on multiplexed real time PCR and enables detection of 19 high-risk and 9 low-risk HPVs. RESULTS: Beside women, the virus was found in 114/197 of men in their foreskin swabs. Additionally, HPV was detected in oropharyngeal swabs of 39/197 female and 56/197 male participants. HPV 16/31/42/39/54 dominated in female and HPV 66/42/16/31/53 in male genital locations. The incidence of HPV in oropharynx was lower, top five genotypes included: HPV 6/39/42/35/16 in women compared to HPV 39/6/42/40/33 in men. HPV16 was the most frequently detected virus type, found in 70/197 examined cervical swabs. It was significantly more prevalent as single infection in females, previously treated for the cervical cancer (p = 0.035). Moreover, regular presence of low risk type 42 was noticeable in both sexes, in both kind of swabs. There was a trend observed towards its prevalence as single infectious agent in women with previous history of cervical cancer (p = 0.069). CONCLUSIONS: Our results showed the distribution of HPV genotypes in Polish couples, in which each woman is HPV positive, indicating a common infection of HPV 42, regardless of sex and anatomical site. These findings shed new light on HPV 42 significance, however they should be verified on a larger group of Polish participants, followed regularly in 6 months intervals, in oral as well as in genital areas.

Voice improvement in patients with recurrent respiratory papillomatosis after combined treatment with cidofovir and CO2 laser surgery.

Mechanical debulking of laryngeal papillomas is associated with voice disorders due to impairment of the vocal fold's mucosa. Intraepithelial injection of cidofovir reduces damage to the laryngeal structures and thus improves vocal parameters. The aim of our study was to compare vocal quality before and after cidofovir and CO2 laser treatment by means of objective phoniatric parameters and to obtain evidence concerning voice outcomes. The vocal parameters of 42 patients with RRP were assessed before and after intralesional cidofovir and CO2 laser debulking treatment (Lumenis AcuPulse 40 CO2 laser, wavelength 10.6 µm; Lumenis Ltd., Yokneam, Israel). The laser was used in SuperPulse™ mode with power tailored on the target structures (average 7 W). The depth of tissue penetration was 1 mm with a single burst of energy lasting 0.3 ms. Most of the patients had previously undergone traditional surgery (1-105 procedures); in 7/42, the video stroboscopic examination revealed extensive scar tissue covering the mucosa of the vocal folds, limiting the mucosal wave. The significance level for all calculations was p < 0.05. Statistical analysis was performed using Statistica 10 by StatSoft Polska. The methodology included subjective voice evaluation (GRBAS), videostroboscopy, analysis of the acoustic laryngeal tone (MDVP), spectrographic analysis, and voice handicap index (VHI). The GRBAS scale during spontaneous conversation revealed better phonation results after cidofovir treatment in all 42 patients. Scars were not observed in the presented group. In the MDVP laryngeal tone analysis before surgery parameters defining the amplitude and frequency of laryngeal tone were significantly elevated compared to the state after surgery. After treatment, both jitter and shimmer were reduced in all subjects. Even more pronounced changes were observed in the spectrographic analysis. In all analyzed patients, there was a significant reduction in VHI scores showing improvement in voice self-assessment. A satisfactory improvement in voice quality was observed in all RRP patients treated with cidofovir and CO2 laser.

Recurrent transcriptional loss of the PCDH17 tumor suppressor in laryngeal squamous cell carcinoma is partially mediated by aberrant promoter DNA methylation.

Protocadherins are cell-cell adhesion molecules encoded by a large family of genes. Recent reports demonstrate recurrent silencing of protocadherin genes in tumors and provide strong arguments for their tumor supresor functionality. Loss of protocadherins may contribute to cancer development not only by altering cell-cell adhesion, that is a hallmark of cancer, but also by enhancing proliferation and epithelial mesenchymal transition of cells via deregulation of the WNT signaling pathway. In this study we have further corroborated our previous findings on the involvement of PCDH17 in laryngeal squamous cell carcinoma (LSCC). We used bisulfite pyrosequencing to analyze a cohort of primary LSCC tumors for alterations in PCDH17 promoter DNA methylation as an alternative gene inactivation mechanism to the homozygous deletions reported earlier. Moreover, we analyzed primary LSCC samples by immunohistochemistry for PCDH17 protein loss. We identified recurrent elevation of PCDH17 promoter DNA methylation in 32/81 (40%) primary tumors (P < 0.001) and therein hypermethylation of 12 (15%) cases in contrast to no tumor controls (n = 24) that were all unmethylated. Importantly, DNA demethylation by decitabine has restored low level PCDH17 expression in LSCC cell lines. In conclusion, we provide a mechanistic explanation of recurrently observed PCDH17 silencing in LSCC by demonstrating the role of promoter methylation in this process. In light of these findings and recent reports showing that PCDH17 methylation is detectable in serum of cancer patients we suggest that testing PCDH17 DNA methylation might serve as a potential biomarker in LSCC.

Primary and salvage laser surgery of 341 glottic cancers-Comparison of treatment outcomes between University Head Neck Tertiary Referral Center and Local Head Neck Department.

OBJECTIVE: The main goal was to compare the clinical data of patients with T1 and T2 glottic cancer treated with CO2 transoral laser microlaryngoscopy (TLM) in the Tertiary Referral University Department of Otolaryngology, Head Neck Surgery, Poznan and the local Department of Otolaryngology, Oncology and Maxillofacial Surgery, Bydgoszcz. MATERIAL AND METHODS: Unified databases for the 7-year period January 2005-December 2011 were created to compare these two cohorts. The database contained 341 patients: 231 from the tertiary center and 110 from the local department, of which 298 (87%) were men, and 43 (13%) were women. RESULTS: Cordectomy type I-IV was performed in 250 (73%) patients, cordectomy type V-VI in 84 (25%) patients, and cordectomy enlarged to epiglottic petiole in 7 (2%) patients. Local recurrence was observed in 96 (28%) cases. Among these cases, 81 (87.5%) patients had salvage surgery: 43 re-cordectomy, 10 open partial laryngectomy, and 28 total laryngectomy. The outcomes for the whole cohort, Tertiary Referral Center and Local Department respectively were as follows: larynx preservation rate was 91.8%, 93.6%, and 88%; 3-year disease specific survival was 97.4%, 97.9%, and 93.3%; 3-year overall survival was 93.5%, 96.6%, and 85.5%; 5-year disease specific survival was 95.2%, 95.2%, and 96.3%; and 5-year overall survival was 84.5%, 88.7%, and 76%, respectively. CONCLUSION: Comparison of the cohorts showed that outcomes of primary treatment were similar but there were noticeable differences in salvage treatment efficacy, favoring patients from the Tertiary Referral Center. Lasers Surg. Med. 50:311-318, 2018. © 2017 Wiley Periodicals, Inc.

Outcomes of CO2 laser-assisted posterior cordectomy in bilateral vocal cord paralysis in 132 cases.

The purpose of the study was to assess the role of laser-assisted posterior cordectomy in the management of patients with bilateral vocal cord paralysis. We aimed an analysis of 132 consecutive patients treated by CO2 laser posterior cordectomy, aged 38-91, 31% tracheotomized on admission. Cordectomy was performed under microlaryngoscopy using CO2 laser (Lumenis AcuPulse 40 CO2 laser, wavelength 10.6 µm, Lumenis Ltd., Yokneam, Israel). We looked at the number of laser glottic procedures necessary to achieve decannulation in tracheotomized patients and to achieve respiratory comfort in non-tracheotomized subjects and we evaluated the two groups for differences in patient characteristics. In tracheotomized patients, we also assessed factors affecting the success of decannulation and we evaluated the impact of tracheotomy on patients' lives. Decannulation was performed in 63% of tracheotomized patients. In terms of the number of procedures, 54% (14), 19% (5), and 27% (7) tracheotomized vs. 74% (61), 24% (20), and 2% (2) non-tracheotomized subjects underwent one, two, or three procedures, respectively. In the group of tracheotomized patients who were successfully decannulated, the number of multiple laser-assisted procedures was significantly higher than in the group of non-tracheotomized subjects with respiratory comfort after treatment (p = 0.04). Advanced age (> 66 years), comorbidities (diabetes, gastroesophageal reflux disease (GERD)), multiple thyroid surgeries, and tracheotomy below the cricoid cartilage were found to decrease the likelihood of successful decannulation. Posterior cordectomy is a simple method allowing for airway improvement and decannulation in patients with bilateral vocal cord paralysis. It is less effective in tracheotomized subjects with diabetes or GERD, older than 66 years old, after two or more thyroidectomies.

Recurrent epigenetic silencing of the PTPRD tumor suppressor in laryngeal squamous cell carcinoma.

Cellular processes like differentiation, mitotic cycle, and cell growth are regulated by tyrosine kinases with known oncogenic potential and tyrosine phosphatases that downmodulate the first. Therefore, tyrosine phosphatases are recurrent targets of gene alterations in human carcinomas. We and others suggested recently a tumor suppressor function of the PTPRD tyrosine phosphatase and reported homozygous deletions of the PTPRD locus in laryngeal squamous cell carcinoma. In this study, we investigated other gene-inactivating mechanisms potentially targeting PTPRD, including loss-of-function mutations and also epigenetic alterations like promoter DNA hypermethylation. We sequenced the PTPRD gene in eight laryngeal squamous cell carcinoma cell lines but did not identify any inactivating mutations. In contrast, by bisulfite pyrosequencing of the gene promoter region, we identified significantly higher levels of methylation (p = 0.001 and p = 0.0002, respectively) in 9/14 (64%) laryngeal squamous cell carcinoma cell lines and 37/79 (47%) of primary laryngeal squamous cell carcinoma tumors as compared to normal epithelium of the upper aerodigestive tract. There was also a strong correlation (p = 0.0001) between methylation and transcriptional silencing for the PTPRD gene observed in a cohort of 497 head and neck tumors from The Cancer Genome Atlas dataset suggesting that DNA methylation is the main mechanism of PTPRD silencing in these tumors. In summary, our data provide further evidence of the high incidence of PTPRD inactivation in laryngeal squamous cell carcinoma. We suggest that deletions and loss-of-function mutations are responsible for PTPRD loss only in a fraction of cases, whereas DNA methylation is the dominating mechanism of PTPRD inactivation.

Assessment of BRAF V600E (VE1) protein expression and BRAF gene mutation status in codon 600 in benign and malignant salivary gland neoplasms.

BACKGROUND: The role of BRAF mutations in cancerogenesis has been demonstrated in several solid tumor types. However, in salivary gland tumors, this genetic alteration is very uncommon, and its role still remains unclear. Thus, the aim of this study was to analyze BRAF V600E (VE1) protein expression with BRAF mutation status in codon 600, in malignant and benign salivary gland tumors. METHODS: Studies were performed on archived formalin-fixed paraffin-embedded tissue sections derived from 95 patients who underwent surgery for tumors of the salivary gland. Immunohistochemical staining (IHC) on tissue microarray slides was performed for evaluation of BRAF V600E (VE1) protein expression, and the automatic molecular diagnostics platform was used for the evaluation of mutations in codon 600 of BRAF gene. RESULTS: IHC cytoplasmic expression of BRAF V600E (VE1) protein was found in two of 95 cases: one case of adenocarcinoma NOS (one of three; 33%) and one case of carcinoma ex pleomorphic adenoma (one of five; 20%). Although, in IHC studies, nuclear BRAF V600E (VE1) protein expression was found in 14 (15%) of the analyzed cases: nine of 28 (32%) cases of pleomorphic adenoma, three of five (60%) cases of ductal carcinoma, one of nine (11%) case of mucoepidermoid carcinoma, and in one of five (20%) case of carcinoma ex pleomorphic adenoma. All cases were negative for polymerase chain reaction PCR-based analyses of BRAF mutations in codon 600. CONCLUSIONS: In studied salivary gland cancers, no PCR-based prove mutations of BRAF V600 were detected. Further molecular analyses are necessary to rapid molecular arrays for the identification of specific mutations, optimal for individualized targeted therapies.

The role of high-risk human papillomavirus infections in laryngeal squamous cell carcinoma.

The contribution of human papillomavirus (HPV) to the development and clinical outcome of oropharyngeal cancers has been well documented. The association of HPV in laryngeal squamous cell carcinoma (LSCC) has been examined in several studies, but controversy exists regarding its role in carcinogenesis, the outcome of the patients and thus, clinical significance of HPV testing in LSCC. In this review, we give an update of known associations between HPV-positive testing and carcinogenesis in laryngeal cancer. In an early study, the HPV-DNA detection rate in LSCC was documented being 24.0% with significant regional differences. Non-HPV-16 types were more often detected in LSCC when compared to the oropharynx. Later, single institution case series revealed markedly fewer amounts (<10%) of HPV DNA in LSCC and the results suggested that high-risk HPV infections seem to be biologically irrelevant in most LSCC. The significance of p16INK4a (p16) expression as a surrogate marker towards high-risk HPV infection and the outcome in LSCC is doubtful, since only few p16-positive LSCC samples are HPV RNA positive and accordingly there was poor correlation of p16-test results towards the outcome in LSCC. Recent meta-analysis (n = 2739) and large case series (n = 1042) of LSCC revealed the true rate of HPV-driven LSCC being 8.6%, respectively, <5%. In the latter the rate of DNA-, DNA/RNA-, DNA/p16, and DNA/RNA/p16 positivity was 5.7, 3.1, 1.9, and 1.5%, respectively. These results indicate relevant amounts of insignificant/transient HPV infection in LSCC specimen. However, in the same study the rate of transforming HPV infections increased since 2000, and younger patients had higher amounts of HPV-driven LSCC. Serologic testing of E6/E7 antibodies additionally revealed odds ratios between 2 and 5 as a hint for a weak contribution of high-risk HPV infection and the development of LSCC. The contribution of HPV for the development of LSCC needs future investigations, to date, routine HPV testing of LSCC specimen is not warranted.