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ABSTRACT: Viral load measurements may predict whether human papillomavirus (HPV) type16 infections may become persistent and eventually lead to cervical lesions. Today, multiple PCR methods exist to estimate viral load. We tested three protocols to investigate viral load as a predictor of HPV clearance. We measured viral load in 418 HPV16-positive cervical smears from 224 women participating in the Ludwig-McGill cohort study by low-stringency PCR (LS-PCR) using consensus L1 primers targeting over 40 known HPV types, and quantitative real-time PCR (qRTPCR) targeting the HPV16-E6 and L1 genes. HPV16 clearance was determined by MY09/11 and PGMY PCR testing on repeated smears collected over five years. Correlation between viral load measurements by qRTPCR (E6 vs. L1) was excellent (r=0.88), but decreased for L1 qRTPCR versus LS-PCR (r=0.61). Viral load by LS-PCR was higher for HPV16 and related types independently of other concurrent HPV infections. Median duration of infection was longer for smears with high copy number by all three PCR protocols (log rank p<0.05). Viral load is inversely related to HPV16 clearance independently of concurrent HPV infections and PCR protocol.
Journal of General Virology 05/2013; · 3.36 Impact Factor
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ABSTRACT: OBJECTIVE: To evaluate reproducibility of oral rinse self-collection for human papillomavirus (HPV) detection and investigate associations between oral HPV, oral lesions, immune and sociodemographic factors, we performed a cross-sectional study of older adults with human immunodeficiency virus (HIV) infection. STUDY DESIGN: We collected oral rinse samples from 52 subjects at 2 different times of day, followed by an oral examination and interview. We identified HPV with the use of polymerase chain reaction platforms optimized for detection of mucosal and cutaneous types. RESULTS: Eighty-seven percent of individuals had oral HPV, of which 23% had oncogenic alpha, 40% had nononcogenic alpha, and 46% had beta or gamma HPV. Paired oral specimens were concordant in all parameters tested. Significant associations observed for oral HPV with increased HIV viral load, hepatitis C seropositivity, history of sexually transmitted diseases, and lifetime number of sexual partners. CONCLUSIONS: Oral cavity may be a reservoir of subclinical HPV in older adults who have HIV infection. Understanding natural history, transmission, and potential implications of oral HPV warrants further investigations.
Oral surgery, oral medicine, oral pathology and oral radiology. 01/2013;
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Yufeng Li,
Shuting Bai,
William Carroll,
Dan Dayan,
Joseph C Dort,
Keith Heller,
George Jour,
Harold Lau,
Carla Penner,
Michael Prystowsky,
Eben Rosenthal, Nicolas F Schlecht,
Richard V Smith,
Mark Urken,
Marilena Vered,
Beverly Wang,
Bruce Wenig,
Abdissa Negassa,
Margaret Brandwein-Gensler
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ABSTRACT: The Risk Model is a validated outcome predictor for patients with head and neck squamous cell carcinoma (Brandwein-Gensler et al. in Am J Surg Pathol 20:167-178, 2005; Am J Surg Pathol 34:676-688, 2010). This model may potentially shift treatment paradigms for patients with low-stage cancers, as current protocols dictate that they might receive only primary surgery. Here we test the hypothesis that the Risk Model has added prognostic value for low-stage oral cavity squamous cell carcinoma (OCSCC) patients. 299 patients with Stage I/II OCSCC were characterized according to the Risk Model (Brandwein-Gensler et al. in Am J Surg Pathol 20:167-178, 2005; Am J Surg Pathol 34:676-688, 2010). Cumulative incidence and competing risk analysis were performed for locoregional recurrence (LRR) and disease-specific survival (DSS). Receiver operating characteristic analyses were performed for worst pattern of invasion (WPOI) and the risk categories. 292 patients were analyzed; 30 T1N0 patients (17 %) and 26 T2N0 patients (23 %) developed LRR. Disease-specific mortality occurred in 9 T1N0 patients (6 %) and 9 T2N0 patients (10 %). On multivariable analysis, the Risk Model was significantly predictive of LRR (p = 0.0012, HR 2.41, 95 % CI 1.42, 4.11) and DSS (p = 0.0005, HR 9.16, 95 % CI 2.65, 31.66) adjusted for potential confounders. WPOI alone was also significantly predictive for LRR adjusted for potential confounders with a cut-point of either WPOI-4 (p = 0.0029, HR 3.63, 95 % CI 1.56, 8.47) or WPOI-5 (p = 0.0008, HR 2.55, 95 % CI 1.48, 4.41) and for DSS (cut point WPOI-5, p = 0.0001, HR 6.34, 95 % CI 2.50, 16.09). Given a WPOI-5, the probability of developing locoregional recurrence is 42 %. Given a high-risk classification for a combination of features other than WPOI-5, the probability of developing locoregional recurrence is 32 %. The Risk Model is the first validated model that is significantly predictive for the important niche group of low-stage OCSCC patients.
Head and Neck Pathology 12/2012;
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Thomas Harris,
Lizandra Jimenez,
Nicole Kawachi,
Jian-Bing Fan,
Jing Chen,
Tom Belbin,
Andrew Ramnauth,
Olivier Loudig,
Christian E Keller,
Richard Smith,
Michael B Prystowsky, Nicolas F Schlecht,
Jeffrey E Segall,
Geoffrey Childs
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ABSTRACT: Small, noncoding microRNAs (miRNAs) have been shown to be abnormally expressed in every tumor type examined. We used comparisons of global miRNA expression profiles of head and neck squamous cell carcinoma (HNSCC) samples and adjacent normal tissue to rank those miRNAs that were most significantly altered in our patient population. Rank Consistency Score analysis revealed miR-375 to have the most significantly lowered miRNA levels in tumors relative to matched adjacent nonmalignant tissue from the same patient among 736 miRNAs that were evaluated. This result has been previously observed by other groups; however, we extend this finding with the unique observation that low miR-375 expression levels correlate significantly with cancer survival and distant metastasis. In a study of 123 primary HNSCC patients using multivariable Cox proportional hazard ratios (HR) and 95% confidence intervals (CI), both death from disease (HR: 12.8, 95% CI: 3 to 49) and incidence of distant metastasis (HR: 8.7, 95% CI: 2 to 31) correlated with lower expression levels of miR-375 regardless of the site or stage of the tumor. In addition, we found that oral cavity tumor cell lines (eg, UMSCC1 and UMSCC47) overexpressing miR-375 were significantly less invasive in vitro than their matched empty vector controls. We conclude that miR-375 represents a potential prognostic marker of poor outcome and metastasis in HNSCC and that it may function by suppressing the tumor's invasive properties.
American Journal Of Pathology 03/2012; 180(3):917-28. · 4.89 Impact Factor
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Nicolas F Schlecht,
Margaret Brandwein-Gensler,
Richard V Smith,
Nicole Kawachi,
Darcy Broughel,
Juan Lin,
Christian E Keller,
Paul A Reynolds,
Frank J Gunn-Moore,
Thomas Harris,
Geoffrey Childs,
Thomas J Belbin,
Michael B Prystowsky
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ABSTRACT: Members of the 4.1 superfamily of proteins, including ezrin, moesin, merlin, and willin regulate many normal physiologic processes such as cellular shape, motility, and proliferation. In addition, they contribute both to tumor development and tumor progression. We reported previously that strong cytoplasmic ezrin expression was independently associated with poorer patient survival. One hundred and thirty-one histologically confirmed primary head and neck squamous cell carcinomas were examined prospectively for cancer progression and survival at a large health care center in the Bronx, NY, USA. Immunohistochemical analysis of ezrin, moesin, merlin, and willin expression in tissue microarray samples of primary head and neck squamous cell carcinoma revealed a significant association of increased cytoplasmic ezrin with poor cancer survival. Global RNA analyses suggest that cancers with high cytoplasmic ezrin have a more invasive phenotype. This study supports our previous findings associating cytoplasmic ezrin with more aggressive behavior and poorer outcome and indicates the need for a multi-institutional study to validate the use of cytoplasmic ezrin as a biomarker for treatment planning in head and neck squamous cell carcinoma.
Head and Neck Pathology 01/2012; 6(2):232-43.
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Danielle Bottalico,
Zigui Chen,
Anne Dunne,
Janae Ostoloza,
Sharod McKinney,
Chang Sun, Nicolas F Schlecht,
Mahnaz Fatahzadeh,
Rolando Herrero,
Mark Schiffman,
Robert D Burk
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ABSTRACT: Human papillomaviruses (HPVs) primarily sort into 3 genera: Alphapapillomavirus (α-HPV), predominantly isolated from mucosa, and Betapapillomavirus (β-HPV) and Gammapapillomavirus (γ-HPV), predominantly isolated from skin. HPV types might infect body sites that are different from those from which they were originally isolated.
We investigated the spectrum of HPV type distribution in oral rinse samples from 2 populations: 52 human immunodeficiency virus (HIV)-positive men and women and 317 men who provided a sample for genomic DNA for a prostate cancer study. HPV types were detected with the MY09/MY11 and FAP59/64 primer systems and identified by dot blot hybridization and/or direct sequencing.
Oral rinse specimens from 35 (67%) of 52 HIV-positive individuals and 117 (37%) of 317 older male participants tested positive for HPV DNA. We found 117 type-specific HPV infections from the HIV-positive individuals, including 73 α-HPV, 33 β-HPV, and 11 γ-HPV infections; whereas, the distribution was 46 α-HPV, 108 β-HPV, and 14 γ-HPV infections from 168 type-specific infections from the 317 male participants.
The oral cavity contains a wide spectrum of HPV types predominantly from the β-HPV and γ-HPV genera, which were previously considered to be cutaneous types. These results could have significant implications for understanding the biology of HPV and the epidemiological associations of HPV with oral and skin neoplasia.
The Journal of Infectious Diseases 09/2011; 204(5):787-92. · 6.41 Impact Factor
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ABSTRACT: Detection of human papillomavirus (HPV) in head and neck cancer has therapeutic implications. In situ hybridization and immunohistochemistry for p16 are used by surgical pathologists. We compared the sensitivity and specificity of three popular commercial tests for HPV detection in head and neck squamous cell carcinomas with a 'gold standard' HPV PCR assay. A total of 110 prospectively collected, formalin-fixed tumor specimens were compiled onto tissue microarrays and tested for HPV DNA by in situ hybridization with two probe sets, a biotinylated probe for high-risk (HR) HPV types 16/18 (Dako, CA, USA) and a probe cocktail for 16/18, plus 10 additional HR types (Ventana, AZ, USA). The p16(INK4) expression was also assessed using a Pharmingen immunohistochemistry antibody (BD Biosciences, CA, USA). Tissue microarrays were stained and scored at expert laboratories. HPV DNA was detected by MY09/11-PCR, using Gold AmpliTaq and dot-blot hybridization on matched-fresh frozen specimens in a research laboratory. HPV 16 E6 and E7-RNA expression was also measured using RT-PCR. Test performance was assessed by a receiver operating characteristic analysis. HR-HPV DNA types 16, 18 and 35 were detected by MY-PCR in 28% of tumors, with the majority (97%) testing positive for type 16. Compared with MY-PCR, the sensitivity and specificity for HR-HPV DNA detection with Dako in situ hybridization was 21% (95% confidence interval (CI): 7-42) and 100% (95% CI: 93-100), respectively. Corresponding test results by Ventana in situ hybridization were 59% (95% CI: 39-78) and 58% (95% CI: 45-71), respectively. The p16 immunohistochemistry performed better overall than Dako (P=0.042) and Ventana (P=0.055), with a sensitivity of 52% (95% CI: 32-71) and specificity of 93% (95% CI: 84-98). Compared with a gold standard HPV-PCR assay, HPV detection by in situ hybridization was less accurate for head and neck squamous cell carcinoma on tissue microarrays than p16 immunohistochemistry. Further testing is warranted before these assays should be recommended for clinical HPV detection.
Modern Pathology 05/2011; 24(10):1295-305. · 4.79 Impact Factor
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Roberto A Lleras,
Leslie R Adrien,
Richard V Smith,
Benjamin Brown,
Naheed Jivraj,
Christopher Keller,
Cathy Sarta, Nicolas F Schlecht,
Thomas M Harris,
Geoffrey Childs,
Michael B Prystowsky,
Thomas J Belbin
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ABSTRACT: Identification of epigenetically affected genes has become an important tool for understanding both normal and aberrant gene expression in cancer. Here we report a whole-genome analysis of DNA methylation profiles in fresh-frozen oropharyngeal squamous cell carcinoma (OPSCC) tissues and normal mucosa samples using microarray technology with patient genomic DNA. We initially compared whole-genome patterns of DNA methylation among 24 OPSCC primary tumors and 24 matched normal mucosal samples. From a survey of 27,578 CpG dinucleotide loci spanning more than 14,000 genes, we identified 958 CpG loci in which measurements of DNA methylation were altered in the primary tumors relative to the normal mucosal samples. These alterations were validated in an independent set of 21 OPSCC patients. A survey of these loci by chromosomal location revealed an abnormally high number of differentially methylated loci on chromosome 19. Many of the loci on chromosome 19 are associated with genes belonging to the Krüppel-type zinc finger protein genes. Hypermethylation was accompanied by a significant decrease in expression of these genes in OPSCC primary tumors relative to adjacent mucosa. This study reports the epigenetic silencing of Krüppel-type zinc finger protein genes on chromosome 19q13 in oropharyngeal cancer. The aberrant methylation of these genes represents a new avenue of exploration for pathways affected in this disease.
American Journal Of Pathology 05/2011; 178(5):1965-74. · 4.89 Impact Factor
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ABSTRACT: Lymphocytic host response (LHR) in the Risk Model is histologically quantified as the density of lymphocytes at the tumor interface (Brandwein-Gensler in Am J Surg Pathol, 34:676-688, 1; in, Am J Surg Pathol 29:167-178, 2). It is classified as strong, intermediate or weak, and is inversely associated with the risk of decreased time to disease progression. In this study, we test the hypothesis that strong LHR corresponds to a greater degree of adaptive cytotoxic T cell response as compared to moderate LHR. We studied resection specimens of primary oral squamous carcinoma classified as having either strong (n = 16), intermediate (n = 20) or weak (n = 4) LHR. CD20+, CD4+, & CD8+ cells were detected by immunohistochemistry and quantified at 40× with a grid; counting the 10 fields with the most lymphocytes at the tumor interface and within tumors. Mean counts/tumor were analyzed by the 2-sided T-test. Statistically significant differences were observed for interface CD8 cells with respect to strong versus moderate LHR, strong versus weak LHR, and moderate versus weak LHR, and tumor infiltrating CD8 cells with respect to strong versus weak LHR. Statistically significant differences were also observed for interface CD4 cells with respect to strong versus weak LHR, and moderate versus weak LHR. Statistically significant differences in interface B cell counts were seen with respect to strong versus weak LHR, and moderate versus weak LHR. Decreased CD8+ T cells were significantly associated with higher stage at presentation (P = 0.005); a direct, but nonsignificant correlation was seen between decreased CD8+ T cells and decreased survival time. Immune response at the tumor interface correlates with an adaptive T cell response; the degree of cytotoxic CD8+ cells infiltrate can distinguish between strong and intermediate LHR at the interface of oral carcinomas.
Head and Neck Pathology 02/2011; 5(2):117-22.
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Margaret Brandwein-Gensler,
Richard V Smith,
Beverly Wang,
Carla Penner,
Andrea Theilken,
Darcy Broughel,
Bradley Schiff,
Randall P Owen,
Jonathan Smith,
Cathy Sarta,
Tiffany Hebert,
Rick Nason,
Marie Ramer,
Mark DeLacure,
David Hirsch,
David Myssiorek,
Keith Heller,
Michael Prystowsky, Nicolas F Schlecht,
Abdissa Negassa
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ABSTRACT: Half of the patients with head and neck squamous cell carcinoma (HNSCC) can be expected to fail therapy, indicating that more aggressive treatment is warranted for this group. We have developed a novel risk model that can become a basis for developing new treatment paradigms. Here we report on the performance of our model in a new multicenter cohort.
Eligible patients from 3 institutions (Montefiore Medical Center, University of Manitoba, and New York University Medical Center) were identified and pathology slides from their resection specimens were reviewed by Margaret Brandwein-Gensler; risk category was assigned as previously published. Kaplan-Meier analysis was performed for disease progression and survival. Cox proportional hazards regression was performed, adjusted for potential confounders. A teaching module was also developed; attending pathologists were asked to score coded slides after a lecture and multiheaded microscope teaching session. Agreement was assessed by calculating Cohen unweighted kappa coefficients.
The validation cohort consisted of 305 patients, from the above institutions, with 311 primary HNSCC of the oral cavity, oropharynx, and larynx. The median follow-up period for all patients was 27 months. Risk category predicts time to disease progression (P=0.0005), locoregional recurrence (P=0.013), and overall survival (P=0.0000) by Kaplan-Meier analysis. High-risk status is significantly associated with decreased time to disease progression, adjusted for clinical confounders (P=0.015, hazard ratio 2.32, 95% confidence interval 1.18-4.58) compared with collapsed intermediate and low-risk groups. We also demonstrate substantial interrater agreement (kappa=0.64), and very good rater agreement when compared with the standard (kappa=0.87).
We demonstrate significant predictive performance of the risk model in a new cohort of patients with primary HNSCC, adjusted for confounders. Our training experience also supports the feasibility of adapting the risk model in clinical practice.
The American journal of surgical pathology 05/2010; 34(5):676-88. · 4.06 Impact Factor
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ABSTRACT: The transporter associated with antigen processing (TAP) is essential in assembling MHC-I proteins. Human papillomavirus (HPV) evades immune recognition by decreasing class I MHC cell surface expression through down-regulation of TAP1 levels. Consistent with heterogeneity in MHC expression is the individual variability in clearing detectable HPV infections. Genetic polymorphisms in TAP genes may affect protein structure, function, and the ability to clear HPV infection.
Case-control study of women with cervical intraepithelial neoplasia (CIN) II or III (n = 114) and women without high-grade CIN (n = 366). Five nonsynonymous single nucleotide polymorphisms (SNP) in TAP1 and TAP2 were genotyped using DNA collected in cervicovaginal lavage samples using microsphere array technology (Luminex xMAP). HPV typing was done using a PCR-based system with MY09/MY11 primers. TAP1 and TAP2 SNPs were validated by direct sequencing.
Differences in allele distribution between women with high-grade cervical neoplasia and women without was seen for TAP1 I333V (P = 0.02) and TAP1 D637G (P = 0.01). The odds ratios (OR) for CIN III were significantly lower among carriers of the TAP1 I333V polymorphism (OR, 0.28; 95% confidence interval, 0.1-0.8), and TAP1 D637G polymorphism (OR, 0.27; 95% confidence interval, 0.1-0.7). These associations remained significant even after restricting the evaluation to women who were positive for high-risk HPV types.
In addition to the down-regulation of MHC-1 by oncogenic HPV, HPV pathogenesis might be facilitated by polymorphisms in the TAP proteins. Identifying TAP polymorphisms may potentially be used to identify women less susceptible to progression to high-grade CIN and cervical cancer.
Clinical Cancer Research 03/2009; 15(3):1019-23. · 7.74 Impact Factor
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Michael B Prystowsky,
Alfred Adomako,
Richard V Smith,
Nicole Kawachi,
Wendy McKimpson,
Peter Atadja,
Quan Chen, Nicolas F Schlecht,
Joanna L Parish,
Geoffrey Childs,
Thomas J Belbin
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ABSTRACT: Head and neck squamous cell carcinoma represents a complex set of neoplasms arising in diverse anatomical locations. The site and stage of the cancer determine whether patients will be treated with single or multi-modality therapy. The HDAC inhibitor LBH589 is effective in treating some haematological neoplasms and shows promise for certain epithelial neoplasms. As with other human cancer cell lines, LBH589 causes up-regulation of p21, G2/M cell cycle arrest, and cell death of human HNSCC cell lines, as measured using flow cytometry and cDNA microarrays. Global RNA expression studies following treatment of the HNSCC cell line FaDu with LBH589 reveal down-regulation of genes required for chromosome congression and segregation (SMC2L1), sister chromatid cohesion (DDX11), and kinetochore structure (CENP-A, CENP-F, and CENP-M); these LBH589-induced changes in gene expression coupled with the down-regulation of MYC and BIRC5 (survivin) provide a plausible explanation for the early mitotic arrest and cell death observed. When LBH589-induced changes in gene expression were compared with gene expression profiles of 41 primary HNSCC samples, many of the genes that were down-regulated by LBH589 showed increased expression in primary HNSCC, suggesting that some patients with HNSCC may respond to treatment with LBH589.
The Journal of Pathology 03/2009; 218(4):467-77. · 6.32 Impact Factor
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Geoffrey Childs,
Melissa Fazzari,
Gloria Kung,
Nicole Kawachi,
Margaret Brandwein-Gensler,
Michael McLemore,
Quan Chen,
Robert D Burk,
Richard V Smith,
Michael B Prystowsky,
Thomas J Belbin, Nicolas F Schlecht
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ABSTRACT: Small noncoding microRNAs (miRNAs) have been shown to be abnormally expressed in every tumor type examined. The importance of miRNAs as potential cancer prognostic indicators is underscored by their involvement in the regulation of basic cellular processes such as cell proliferation, differentiation, and apoptosis. In this study, miRNA expression profiles of head and neck squamous cell carcinoma (HNSCC) tumor and adjacent normal tissue were examined by microarray analysis and validated by quantitative TaqMan real-time polymerase chain reaction. Using TaqMan real-time polymerase chain reaction we measured the quantitative associations between a subset of miRNAs identified on microarrays in primary tumors at diagnosis and cancer survival in a cohort of 104 HNSCC patients undergoing treatment with curative intent. The majority of miRNAs exhibiting altered expression in primary human HNSCC tumors (including miR-1, miR-133a, miR-205, and let-7d) show lower expression levels relative to normal adjacent tissue. In contrast, hsa-miR-21 is frequently overexpressed in human HNSCC tumors. Using univariate and multivariable statistical models we show that low levels of hsa-miR205 are significantly associated with loco-regional recurrence independent of disease severity at diagnosis and treatment. In addition, combined low levels of hsa-miR-205 and hsa-let-7d expression in HNSCC tumors are significantly associated with poor head and neck cancer survival Our results show that miRNA expression levels can be used as prognostic markers of head and neck cancer.
American Journal Of Pathology 02/2009; 174(3):736-45. · 4.89 Impact Factor
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ABSTRACT: It is known that head and neck squamous cell carcinomas (HNSCC) originating from different anatomic locations can exhibit varying behavior that is not predictable by histopathology of the primary tumor. Using a microarray containing 27,323 cDNA clones, we generated sets of gene expression profiles for 36 HNSCC primary tumors (12 oral cavity, 12 oropharynx, and 12 larynx/hypopharynx). From these datasets, we ranked genes according to their ability to differentiate between patients whose disease progressed within a 24 month period (aggressive phenotype) and those that did not (non-aggressive phenotype) based on levels of gene expression. A merging of datasets from the three sites revealed that only a fraction of identified genes were shared between any two sites. This contrasted greatly with the significant overlap (approximately 50%) in down-regulated genes identified in tumor/normal comparisons using cases both from oropharynx and larynx/hypopharynx. From these data, we conclude that HNSCC tumors originating from different anatomic sites share consistent changes in gene expression when comparing primary tumors to normal adjacent mucosa; these common changes most likely reflect alterations required for tumor development. In contrast, once a tumor has developed, tumor-host interactions at the different anatomic sites are likely responsible for the site-specific signatures associated with aggressive versus non-aggressive disease. Predictions of outcome based on gene expression profiling are therefore heavily influenced by the anatomic site of the primary tumor.
Head and Neck Pathology 12/2008; 2(4):243-56.
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ABSTRACT: Topoisomerase IIα and minichromosome maintenance protein 2 are proteins associated with aberrant S-phase induction. The current study evaluated the performance of these biomarkers (ProEx™ C; TriPath Oncology, Burlington, NC) compared with p16INK4A and MiB-1 in distinguishing high-grade squamous intraepithelial lesions (HSILs) from HSIL mimics. We collected archival cervical biopsy, cone, and curettage specimens from 96 cases in which the differential diagnosis of HSIL vs reactive epithelial changes was considered. Hematoxylin- and eosin-stained slides were reviewed independently by three pathologists and scored for the presence or absence of SIL. Immunostains for ProEx C, p16, and MiB-1 were available for 95, 96, and 59 samples, respectively, and classified blinded to histological interpretation. Strong nuclear and cytoplasmic staining for p16 and staining for MiB-1 and ProEx C that extended beyond the lower one-third of the epithelium were scored as positive. χ2-tests and receiver operating characteristic analysis were conducted to statistically compare biomarker immunostaining performance against majority histological interpretation of SIL. Agreement between pathologists was also assessed by the κ-statistic. Inter-observer agreement ranged from fair to moderate (κ=0.37–0.57). All three biomarkers correlated strongly with the majority diagnosis of SIL (P<0.001). Positive staining for ProEx C, p16, and MiB-1 was observed in 87% (N=52/60), 84% (N=51/61), and 94% (34/36), respectively, of SIL and negative in 71% (N=25/35), 63% (N=22/35), and 52% (N=12/23), respectively, of majority diagnoses of NoSIL. The combination of p16/ProEx C predicted more SIL (92%, N=33/36) and NoSIL (61%, N=14/23) than p16 plus MiB-1 (94%, N=34/36 and 43%, N=10/23), although this difference was not statistically significant. ProEx C appears to provide an equivalent level of sensitivity and a higher level of specificity for HSIL alone or in conjunction with p16. Its principal value may be in providing a lower false positive rate for NoSIL relative to MiB-1.Keywords: cervical biopsy, cervical intraepithelial neoplasia, high-grade squamous intraepithelial lesion, MiB-1, p16INK4A, ProEx™ C
Modern Pathology 06/2008; 21(9):1067-1074. · 4.79 Impact Factor
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ABSTRACT: Topoisomerase IIalpha and minichromosome maintenance protein 2 are proteins associated with aberrant S-phase induction. The current study evaluated the performance of these biomarkers (ProEx C; TriPath Oncology, Burlington, NC) compared with p16(INK4A) and MiB-1 in distinguishing high-grade squamous intraepithelial lesions (HSILs) from HSIL mimics. We collected archival cervical biopsy, cone, and curettage specimens from 96 cases in which the differential diagnosis of HSIL vs reactive epithelial changes was considered. Hematoxylin- and eosin-stained slides were reviewed independently by three pathologists and scored for the presence or absence of SIL. Immunostains for ProEx C, p16, and MiB-1 were available for 95, 96, and 59 samples, respectively, and classified blinded to histological interpretation. Strong nuclear and cytoplasmic staining for p16 and staining for MiB-1 and ProEx C that extended beyond the lower one-third of the epithelium were scored as positive. Chi(2)-tests and receiver operating characteristic analysis were conducted to statistically compare biomarker immunostaining performance against majority histological interpretation of SIL. Agreement between pathologists was also assessed by the kappa-statistic. Inter-observer agreement ranged from fair to moderate (kappa=0.37-0.57). All three biomarkers correlated strongly with the majority diagnosis of SIL (P<0.001). Positive staining for ProEx C, p16, and MiB-1 was observed in 87% (N=52/60), 84% (N=51/61), and 94% (34/36), respectively, of SIL and negative in 71% (N=25/35), 63% (N=22/35), and 52% (N=12/23), respectively, of majority diagnoses of NoSIL. The combination of p16/ProEx C predicted more SIL (92%, N=33/36) and NoSIL (61%, N=14/23) than p16 plus MiB-1 (94%, N=34/36 and 43%, N=10/23), although this difference was not statistically significant. ProEx C appears to provide an equivalent level of sensitivity and a higher level of specificity for HSIL alone or in conjunction with p16. Its principal value may be in providing a lower false positive rate for NoSIL relative to MiB-1.
Modern Pathology 06/2008; 21(9):1067-74. · 4.79 Impact Factor
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ABSTRACT: Evidence of an etiological role for human papillomavirus (HPV) in Schneiderian inverted papillomas IP arose in the late 1980's; yet almost three decades later, the association between HPV and IP has yet to be universally accepted. This is probably due to the disparate HPV detection rates in IP reported in the literature. We analyzed the weight of published data in order to address the following questions: why do the HPV detection rates in IP vary so greatly? What is the relationship between low-risk (LR) and high-risk (HR) HPV types and HPV detection rates in IP? Is there a relationship between the presence and type of HPV in IP and recurrence and malignant progression?
A search using the Pubmed search engine was performed to identify studies published in English from 01/87 through 12/06 using the MeSH terms ''HPV'' and ''Inverted", "Exophytic", "Oncocytic Schneiderian" or "Fungiform papilloma''. Data was abstracted from publications including histology, HPV target, HPV type, method of detection, etc. HPV results were stratified by histology and other variables. Tests for heterogeneity (between-study variability) were conducted, and weighted prevalence (WP) estimates and 95% confidence intervals (CI) were calculated using a random-effects inverse-variance model stratified on study. The association between HPV IP recurrence was estimated by random-effects inverse-variance weighted odds ratio (OR).
Weighted estimates revealed similar detection rates across detection methods, 26.8% (95%CI 16.4-37.2%) by ISH, 25.2% (95%CI 14.7-35.6%) by consensus PCR, and 23.6% (95%CI 12.2-35.0%) by type-specific PCR. A preponderance of HPV 6/11 is found in IP as compared to HPV 16/18; the overall unadjusted ratio of LR to high-risk HR HPV types is 2.8:1 The HPV detection rates significantly increase (Wald t-test P < 0.02) in IPs with high-grade dysplasia (WP 55.8%, 95%CI 30.5-81.0%) and carcinoma (WP 55.1%, 95%CI 37.0-73.2%) as compared to IPs with no dysplasia or mild dysplasia (WP 22.3%, 95%CI 15.9-28.6%). Furthermore, the preponderance of LR HPV in benign IP (ratio LR/HR = 4.8:1) shifts in dysplastic and malignant IP. The LR/HR ratio is 1.1:1 for IPs with high-grade dysplasias, this ratio is inverted to favor HR HPV (1:2.4) for malignant IP. Recurrences developed in 44 of 236 patients; HPV was detected in 27 of 44 IPs (WP 57.9%, 95%CI 31.6-84.2%) that developed recurrences and in 24 of 192 IPs (WP 9.7%, 95%CI 4.4-15.0%) that did not develop recurrence. The presence of HPV was significantly associated with the likelihood of developing recurrence (weighted OR of 10.2, 95%CI 3.2-32.8).
We hypothesize that LR HPV may induce IP formation, and then are lost as infected cells are shed, as a "hit and run" phenomenon. HPV detection rates increase in dysplastic IP and SCC-ex-IP with increasing ratio of HR to LR HPV types, compared to nondysplastic IP. We believe that one explanation for the variation in HPV detection rates between different studies may be the actual histologic composition of the cohort. That is, if one series contains a higher frequency of dysplastic and malignant IP, it may have a higher detection rate than another series which contains only nondysplastic IP. We hypothesize that the higher rates of HPV detection in dysplastic and malignant IP may be related to HPV integration. The implication of this is that HPV sub-type testing may identify patients at risk for recurrence, or progression to dysplasia and malignancy, and thus may impact surveillance protocols.
Head and Neck Pathology 06/2008; 2(2):49-59.
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Miriam S Teixeira,
Olga Camacho-Vanegas,
Yolanda Fernandez,
Goutham Narla,
Analisa DiFeo,
Bryant Lee,
Tamara Kalir,
Scott L Friedman, Nicolas F Schlecht,
Eric M Genden,
Mark Urken,
Margaret Brandwein-Gensler,
John A Martignetti
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ABSTRACT: The Krüppel-like transcription factor (KLF6) gene is a tumor suppressor gene (TSG) reported to be dysregulated and inactivated through loss of heterozygosity (LOH) and/or somatic mutation in a number of major human cancers. The aim of the present study was to examine KLF6 gene status and expression in head and neck squamous cell carcinomas (HNSCC). A collection of 81 well-characterized oral and oropharyngeal HNSCC samples were analyzed for evidence of KLF6 LOH and mutation and differences in expression patterns between normal and cancerous tissues and these findings were correlated with clinicopathological variables. We also tested the effect of KLF6 inhibition in HNSCC cell lines on proliferation and p21 expression. LOH was found in approximately 30% of cases and was strongly correlated with cancer progression, tumor recurrence and decreased patient survival. Overall, median survival of patients with LOH was less than half (19 vs. 41 months, p=0.036, stratified on stage) than those without loss. Risk of death for patients with LOH was 8 times greater independent of tumor size, nodal status, tobacco smoking or treatment modality (HR 7.89, 95% CI: 1.9-32.4). Subsequent analyses revealed KLF6 mutations in only 2 of 20 samples, but altered subcellular protein localization in 64% of tumors. Targeted stable reduction of KLF6 in HNSCC cell lines increased cellular proliferation while decreasing p21 expression. Taken together, these findings suggest that KLF6 LOH represents a clinically-relevant biomarker predicting patient survival and tumor recurrence and that dysregulation of KLF6 function plays an important role in HNSCC progression.
International Journal of Cancer 12/2007; 121(9):1976-83. · 5.44 Impact Factor
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Rashna Madan,
Margaret Brandwein-Gensler, Nicolas F Schlecht,
Kristin Elias,
Eleanor Gorbovitsky,
Thomas J Belbin,
Radma Mahmood,
Dwayne Breining,
Hong Qian,
Geoffrey Childs,
Joseph Locker,
Richard Smith,
Missak Haigentz,
Frank Gunn-Moore,
Michael B Prystowsky
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ABSTRACT: Members of the ezrin-radixin-moesin (ERM) protein family regulate cellular shape, motility, and proliferation and potentially influence ability to metastasize. We investigated the correlation between ERM subcellular localization and survival in patients with squamous cell carcinoma (SCC) METHODS: Tissue microarrays (TMAs) were constructed from paraffin-embedded tissue. TMA sections were evaluated for ERM protein expression immunohistochemically. The results were compared across clinical and histopathologic variables
ERM staining results for 47 patients showed that cytoplasmic ERM expression was prevalent in tumors (>92%). Whereas ezrin and moesin also localized to the membrane, only willin was found in the nucleus of tumors. Multivariable Cox regression analysis demonstrated that strong cytoplasmic ezrin expression was independently associated with poorer survival (p = .04, hazard ratio 1.82)
Both level of expression and subcellular localization of ERM proteins may be important indicators of clinical outcome in SCC. This pilot study justifies the need for an expanded validation study of ERM proteins and clinical outcome.
Head & Neck 11/2006; 28(11):1018-27. · 2.40 Impact Factor
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ABSTRACT: Cohort studies of the natural history of human papillomavirus (HPV) infection and cervical squamous intraepithelial lesions with repeated screening allow the comparison of different macroscopic and microscopic diagnostic methods.
Concurrent visual inspection using cervicography and conventional Pap cytology tests were performed during multiple visits in a cohort of women attending a maternal and child health clinic in São Paulo, Brazil. HPV infection status at the same visits was determined by polymerase chain reaction followed by typing with specific oligonucleotide probing and viral load quantification. Information on reproductive health and hygiene habits was also collected at each visit.
Overall agreement between cervicography and cytology was low (kappa = 0.046), which increased only slightly when high oncogenic-risk HPV types (kappa = 0.120) or high viral burden (>100 copies/cell) (kappa = 0.170) was present. Analysis of reproductive health and hygiene habits revealed somewhat different risk factors for cervical lesions detected by these tests. However, presence of oncogenic HPV DNA (odds ratio = 36.0, 95% CI = 16.6-77.8) and high viral burden (odds ratio = 67.34; 95% CI = 27.1-167.0) were strongly associated with lesions detected by cytology but not by cervicography.
Although changes in the cervix (because of age, gravidity, or hormonal effects) may influence the performance of morphology-based screening tests, the lack of agreement and the different degrees of association with HPV infection measures indicate that a visual inspection method such as cervicography may detect different cervical abnormalities relative to cytology.
Journal of Lower Genital Tract Disease 10/2006; 10(4):229-37. · 1.07 Impact Factor
