Differential expression of p63 isotypes (DeltaN and TA) in salivary gland neoplasms: biological and diagnostic implications.

To determine the association between the expression of p63 gene isoforms (TA and DeltaN) and salivary gland tumorigenesis, we performed reverse transcription-polymerase chain reaction analysis of these markers in 71 benign and malignant salivary gland neoplasms. The results were correlated with the expression of Notch ligand JAG1 gene and the clinicopathologic features and the full-length p63 protein expression by immunohistochemistry. Both p63 isoforms were either negative or weakly expressed in normal salivary gland tissues. TAp63 was highly expressed in most benign tumors and was either negative or weakly positive in most carcinomas. Conversely, DeltaNp63 was negative or faintly positive in most benign neoplasms and was highly expressed in adenoid cystic, mucoepidermoid, and myoepithelial carcinomas. Immunohistochemical analysis using anti-full-length p63 protein showed ubiquitous nuclear staining in basal and myoepithelial cells in both benign and malignant neoplasms. JAG1 was expressed in most benign and malignant tumors and did not correlate with p63 isoforms expression. We conclude that (1) p63 isoforms are differentially expressed in most benign and malignant tumors and may play distinct biological roles in certain salivary gland neoplasms; (2) p63 immunostaining do not correlate with the isoforms expression; and (3) isoform-specific antibodies are required for better cellular localization and biological correlations.

Gene expression screening of salivary gland neoplasms: molecular markers of potential histogenetic and clinical significance.

Salivary gland neoplasms comprise phenotypically and biologically diverse lesions of uncertain histogenesis. The molecular events associated with their development and clinicopathological heterogeneity remain unknown. To reveal these events, we performed microarray expression analysis using a nylon-filter membrane platform on 18 primary lesions representing the most common benign and malignant types. Our study identified a small set of genes that are differentially altered between normal salivary gland tissues and benign and malignant tumors. Of the 5000 genes arrayed, 136 genes were differentially expressed by normal tissue, benign tumors, and various malignant neoplasms. Hierarchical clustering analysis differentiated between adenoid cystic carcinomas (ACCs) and other malignant subtypes. Non-ACC specimens manifested overlapping patterns of gene expression within and between tumors. Most of the differentially expressed genes share functional similarities with members of the adhesion, proliferation, and signal transduction pathways. Our study identified: 1) a set of genes that differentiate normal tissue from tumor specimens, 2) genes that differentiate pleomorphic adenoma and ACCs from other malignant salivary gland neoplasms, and 3) different patterns of expression between ACCs arising from major and minor salivary gland sites. The differentially expressed genes provide new information on potential genetic events of biological significance in future studies of salivary gland tumorigenesis.

Sarcomatoid carcinoma of the head and neck: molecular evidence for evolution and progression from conventional squamous cell carcinomas.

The underlying events associated with the development of sarcomatoid head and neck squamous carcinoma and the biologic significance remain unknown. To investigate the genetic events involved in the evolution of this entity, comparative analysis of matched microdissected epithelial and sarcoma-like components from 11 primary sarcomatoid carcinomas was performed using microsatellite markers. Nine markers on chromosomes 4p, 9p, and 17p regions (3 per each chromosomal region) were selected based on their informativeness, small product size, and the high alterations in head and neck squamous carcinomas. In this study, loss of heterozygosity (LOH) in at least one marker in either component was noted in all 11 tumors, and instability was found in 10 instances (six in 3 paired specimens and four in the sarcomatoid area only). Concordant results in both components were found in 58 (79.5%) reactions (37 LOH and 21 retention of heterozygosity), and paradoxical findings were noted in 15 instances (20.5%). The latter included LOHs in only two conventional epithelial components and 13 sarcomatoid components. Both keratin-positive and -negative sarcomatoid tumors had a comparable frequency of LOH. The most frequently altered markers in both components were D9S168 and D9S171 (75% each) and D4S1587 (66%). The sarcomatoid components manifested distinctly high alterations at marker D17S520 on chromosome 17p. Our study supports: 1) an evolution of sarcomatoid carcinoma from the conventional epithelial-type, 2) a malignant nature of the sarcomatoid component, and 3) that molecular progression is associated with the sarcomatoid transformation.

Molecular analysis of chromosome 16q regions in dermal analogue tumors of salivary glands: a genetic link to dermal cylindroma?

Dermal analogue tumor, an uncommon subtype of basal cell monomorphic adenoma of the parotid gland, has a remarkable clinical and histologic resemblance to dermal cylindroma. Molecular studies of familial and sporadic cylindromas have shown frequent alterations at chromosome 16q12-13 that have recently been found to house the cylindromatosis gene (CYLD). To determine the involvement of the chromosome 16q12-13 region in dermal analogue tumors, we performed loss of heterozygosity analysis using microsatellite markers flanking the cylindromatosis gene locus in 21 sporadic dermal analogue salivary tumors and 12 salivary and dermal lesions from two sisters. Loss of heterozygosity was identified in 17 (80.9%) of the 21 sporadic tumors and in nine of the 12 dermal and salivary gland dermal analogue tumors from the two sisters; a parathyroid adenoma from one sister and two lymphoepithelial lesions from the second sister showed no microsatellite alterations. Microsatellite instability was only identified in three sporadic tumors at marker D16S308. Markers D16S409 (centromeric), D16S541, and D16S308 (telomeric) to the CYLD gene showed the highest incidence of loss of heterozygosity (>65%). The minimally deleted region was flanked proximally by marker D16S389 and distally by marker D16S419 and spanned the 771.5-megabase fragment that included the CYLD locus. We conclude that dermal analogue tumor and cylindroma share similar incidence of alterations at the 16q12-13 region, supporting a common molecular origin.

Molecular and clinicopathologic comparisons of head and neck squamous carcinoma variants: common and distinctive features of biological significance.

To investigate, for the first time, the events associated with the phenotypic and clinical diversities of head and neck squamous carcinomas (HNSC), we performed molecular analyses on 92 primary tumors representing the entire spectrum of the morphologic subtypes using microsatellite markers at chromosome 3p, 4p, 8p, 9p, 11q, 17p, and 18q regions and correlated the results with the clinicopathologic features and patients' survival. Loss of heterozygosity (LOH) at D9S168 and D9S171 markers on chromosome 9p regions was commonly identified in all subtypes. Distinctive alterations in certain subtypes were noted at chromosomes 3p, 4p, 8p, and 11p regions. In general, less aggressive types (verrucous, papillary, and well-differentiated conventional) had a significantly lower LOH incidence than the more aggressive (basaloid, sarcomatoid, and high-grade conventional squamous carcinoma) categories. Significant association between LOH and age, stage, nodal status, and patient outcome was found. Survival analysis revealed that pathologic categorization (less versus more aggressive) and LOH at marker D11S4167 and D3S2432 are independent predictors of patients' survival. Our analysis also defined a set of limited markers that account for most of alterations within and across these tumor subtypes. Our study indicates that 1) certain genetic markers are common to all subtypes of HNSC supporting their early involvement in tumorigenesis, 2) inter- and intratumoral genetic differences evolve subsequently and may underlie their morphologic heterogeneity, 3) high incidence of LOH in certain regions characterizes aggressive tumors, 4) categorical classification and LOH at 11p and 3p regions independently correlated with patient survival, and 5) a limited set of markers identify the majority of genetic alterations in these tumors.

Differential expressions of cyclin-dependent kinase inhibitors (p27 and p21) and their relation to p53 and Ki-67 in oral squamous tumorigenesis.

The cyclin-dependent kinase inhibitors p27Kip1 and p21WAF1/Cip1 play important roles in cell-cycle regulation. Although alterations of these genes have been linked to tumorigenesis of several human carcinomas, their involvement in head and neck squamous tumorigenesis is rarely investigated. To determine the role of these genes in the evolution of squamous carcinoma of the head and neck we evaluated their protein expression by immunohistochemistry in non-dysplastic squamous epithelium, premalignant lesions and oral squamous carcinomas. The p53 gene and Ki-67 expressions were correlated with traditional clinicopathologic variables. Our study shows that in histologically non-dysplastic squamous epithelium, p27 expression was noted mainly in superficial differentiated cells, whereas p21, p53 and Ki-67 staining was observed in basal and suprabasal cells. In dysplasia, divergent expression between p27 and p21 was observed: p27 precipitously decreased and p21, p53, and Ki-67 increased with histologic progression. In squamous carcinomas, p27 was mainly expressed in well differentiated tumor cell nests, while the expressions of p21, p53, and Ki-67 were variable in the poorly differentiated tumor areas. A significant inverse relationship between p27 expression and those of p21, p53, and Ki-67 was observed, but no significant association between any of these markers and clinicopathologic factors was noted in this cohort. Our study indicates that: i) down-regulation of p27 and up-regulation of p21 are associated with early progression of HNSC, ii) p21 expression correlates positively with proliferation while p27 correlates positively with cell differentiation and iii) concurrent p27 and p21 expression analysis may allow for better assessment of HNSC progression.

Sinonasal adenocarcinoma: evidence for histogenetic divergence of the enteric and nonenteric phenotypes.

Adenocarcinomas of nonsalivary origin represent approximately 10% to 20% of all sinonasal malignancies and are characterized by varying histopathologic features and uncertain histogenesis. To better understand the histogenesis and phenotypic heterogeneity of these tumors, we performed immunohistochemical analyses for cytokeratin (CK) 7 and CK20 on 12 primary sinonasal adenocarcinomas (SNACs) representing the histopathologic spectrum of these tumors, adjacent normal mucosa, and 2 metastatic adenocarcinomas from colonic primaries. The demographic and clinicopathologic characteristics of our cohort were similar to those in previously published series. Our results indicate that histologically normal respiratory-type epithelium and submucosal seromucous glands show restricted reactivity to CK7. Epithelial metaplasia of surface epithelium associated with enteric SNACs was accompanied by a conversion from CK7 positivity to CK20 positivity. All primary enteric-type carcinomas and the 2 colonic metastases were reactive to CK20, but all nonenteric-type tumors were negative for CK20 (P=0.003) and positive for CK7. In some of the enteric types, coexpression of CK7 and CK20 was noted. We conclude that (1) nonenteric-type (seromucinous) adenocarcinoma may originate directly from surface respiratory-type epithelium or from seromucous glands, (2) metaplastic transformation of surface respiratory to enteric-type epithelium precedes the development of enteric adenocarcinoma, and (3) coordinate analyses of CK7 and CK20 reactivity may aid the differential diagnosis of adenocarcinoma in the sinonasal tract.

Differential expression of p53 gene family members p63 and p73 in head and neck squamous tumorigenesis.

p73 and p63 are recently cloned genes that share considerable structural and functional homologies with the p53 tumor suppressor gene. These genes, unlike p53, express multiple mRNA isoforms with variable biologic functions, and their suppressor nature has yet to be confirmed. To determine the interrelationship between these genes in the tumorigenesis of head and neck squamous carcinoma (HNSC), we performed immunohistochemical analyses of their protein products and compared the data with clinicopathologic parameters in 38 patients. In histologically normal epithelium, p53 and p73 showed similar basal and/or parabasal expression, but that of p53 was weaker and discontinuous. p63 staining was noted in more suprabasal cellular layers and was stronger. In dysplasias, all three markers manifested variable but gradual increase in extent and intensity of cellular expression with histologic progression. In carcinomas, p63 was the most frequently expressed (94.7%), followed by p73 (68.4%) and p53 (52.6%). Significant statistical correlation was noted only between p63 and p73 expressions (P =.04). Although no statistical correlation was found between p53 and p63 or p73, p53-negative tumors overexpressed either p63 or p73. p73 expression was associated with distant metastasis and perineural/vascular invasion. Our study indicates that (1) p63 and p73 expression may represent an early event in HNSC tumorigenesis, (2) the lack of correlation between p73 or p63 and p53 expression suggests an independent and/or compensatory functional role, (3) p73 expression may play a part in HNSC progression, and (4) p73 and p63 may function as oncogenes in the development of these tumors.

CRTC1/MAML2 fusion transcript in high grade mucoepidermoid carcinomas of salivary and thyroid glands and Warthin's tumors: implications for histogenesis and biologic behavior.

We analyzed 55 primary salivary gland tumors including 22 mucoepidermoid carcinomas (MECs) to determine the association of MECT1/TORC1/CRTC1-MAML2 fusion transcript to tumor types, level of MEC differentiation and clinicopathologic parameters. Our primary salivary gland tumors were composed of 22 MECs, 11 Warthin's tumors, 10 adenoid cystic carcinomas, two basaloid carcinomas, five salivary duct carcinomas, and five adenocarcinomas, not otherwise specified. We also included, for the first time, three primary MECs of the thyroid gland. We used nested RT-PCR and subsequent sequencing techniques for detection and verification of the fusion transcript in fresh and archival specimens. Eighteen (81%) of the 22 primary salivary and one of the three thyroid glands with MEC were positive for the fusion transcript. The transcript was detected equally in low, intermediate and high grade as well as low and high stage MECs. Significant correlation between fusion negative tumors and distant metastasis was noted (P = 0.005). Four (36%) of the 11 Warthin's tumors were also positive for the transcript. None of the 22 primary non-MEC gland salivary carcinomas were positive for the transcript. We conclude that the CRTC1/MAML2 transcript may be detected in both low and high grade MEC, that fusion negative tumors may define a subset of biologically aggressive MEC's tumors, that the fusion is present in primary MECs of the thyroid gland and is also detectable in Warthin's tumor, and that a subset of MECs can be targeted for therapeutic intervention.

Myofibroblastoma of the larynx: a study of two cases.

BACKGROUND: We describe, for the first time, the occurrence of two pure examples of spindle cell neoplasm with fibrous and smooth muscle differentiation in the larynx. MATERIALS AND METHOD: Tissue sections from both tumors stained with hematoxylin and eosin were reviewed. Immunophenotypic analysis using keratin, S-100 protein, SMA, and h-Caldesmon markers was performed. RESULTS: Tumors were seen as raised submucosal nodules in the glottic region of two elderly male patients. Histopathologically, the lesions were unencapsulated and composed of spindle cells that manifested variable cellular anaplasia and expressed SMA and were negative for keratin, S-100, and h-Caldesmon. CONCLUSIONS: We document histologically low-grade malignant myofibroblastic tumors of the larynx. Complete excision is advised.

Plasma cell dyscrasias and the head and neck.

Structures in the head and neck (bones, soft tissues, lymph nodes, mucosa) are variably affected by plasma cell dyscrasias. Involvement can be manifested by localized lesions (extramedullary plasmacytoma or solitary plasmacytoma of bone) or by more diffuse disease (multiple myeloma). We present a contemporary review of these disorders with emphasis on patient outcomes.