Prognostic Significance of Tumor Immunity in Surgically Resected Pulmonary Pleomorphic Carcinoma.

BACKGROUND: Pulmonary pleomorphic carcinoma (PPC) is a rare aggressive neoplasm, with dismal prognosis. Whether tumor immunity is associated with the progressive biological behavior of PPC remains unclear. The purpose of this study was to examine the prognostic significance of tumor immunity-related markers such as programmed death-1 ligand (PD-L1) and CD4+ or CD8+ tumor-infiltrating lymphocytes (TILs) in patients with surgically resected PPC. PATIENTS AND METHODS: Ninety-nine patients with surgically resected PPC were assessed by immunohistochemistry. The expression of PD-L1, CD4, and CD8 was examined in specimens of the resected tumors. RESULTS: PD-L1 was highly expressed in 61% (60/99) of lesions and high expression of CD4 and CD8 was identified in 42% (42/99) and 51% (51/99) of lesions, respectively. There was no relationship between the expression PD-L1 and the numbers of CD4+ or CD8+ TILs. The expression of PD-L1 was not identified as a significant prognostic marker; however, a low number of CD4+ TILs was identified as an independent marker for predicting a worse outcome after surgical resection of PPC, especially in patients with an epithelial component of adenocarcinoma or early stage of disease. By univariate analysis, a low number of CD8+ TILs was found to be a significant prognostic marker linked to poor overall survival in patients with non-adenocarcinoma components. CONCLUSION: A low number of CD4+ TILs is an independent marker for predicting a favorable prognosis after surgical resection in patients with PPC, especially in patients with adenocarcinoma components or early stage of disease.

Immunoprofile of c-MET/PI3K signaling in human salivary gland tumors.

OBJECTIVE: The aim of this study was to analyze the expression pattern of proteins in the HGF/c-MET/PI3K signaling pathway in salivary gland tumors (SGTs) and to correlate the findings with the proliferative index and clinical parameters. STUDY DESIGN: We assembled tissue microarrays (TMAs) of 108 cases of SGTs, including 69 cases of pleomorphic adenoma (PA), 24 cases of adenoid cystic carcinoma (AdCC), and 15 cases of mucoepidermoid carcinoma (MEC). An immunohistochemical analysis of hepatocyte growth factor (HGF), MET phosphorylation (p-MET), protein kinase B (AKT) phosphorylation (p-AKT), and Ki-67 proteins was performed. RESULTS: Benign and malignant SGTs presented similar scores of HGF-positive cells (P = .36), whereas, malignant SGTs exhibited higher levels of p-MET (P = .001) and p-AKT (P = .001) than benign SGTs. No correlation of HGF, p-MET, or p-AKT expression was observed with clinical parameters. PA had a lower proliferative index than either AdCC (P = .001) or MEC (P = .001). CONCLUSIONS: The salivary gland carcinomas exhibited increased activation of the HGF pathway, as evidenced by the phosphorylation of the MET receptor, and increased activation of the PI3K pathway, as indicated by p-AKT. These data suggest that the HGF/c-MET/PI3K signaling pathway is active in SGTs, especially in malignant neoplasms.

The role of FGF-2/HGF and fibronectin matrix on pleomorphic adenoma myoepithelial cell morphology and immunophenotype: an in vitro study.

Myoepithelial cells play a central role in glandular tumors, regulating the progression of in situ to invasive neoplasias, with the tumor microenvironment being shown to be involved in both initiation and progression. This study aimed to analyze the in vitro effects of fibroblast growth factor-2 (FGF-2) and hepatocyte growth factor (HGF) in myoepithelial cells under the influence of the fibronectin matrix extracellular protein. Benign myoepithelial cells were obtained from pleomorphic adenoma and cultured on a fibronectin substratum. FGF-2 and HGF were supplemented at different concentrations and time intervals, in order to evaluate cell proliferation, morphology and immunophenotype. Individually, FGF-2 and HGF supplementation did not alter myoepithelial cell proliferation, morphology or immunophenotype. The fibronectin substratum provoked an increase in cell proliferation and immunopositivity for α-smooth muscle actin and FGF-2. The myoepithelial cell morphology changed when the fibronectin substratum and FGF-2 acted together, highlighting the importance of the fibronectin extracellular matrix protein on the behavior of these cells.

Carcinoma ex pleomorphic adenoma of the salivary glands: distinct clinicopathologic features and immunoprofiles between subgroups according to cellular differentiation.

In carcinoma ex pleomorphic adenoma (CXPA), pleomorphic adenoma (PA) and diverse carcinoma components showing luminal (ductal) or non-luminal (myoepithelial) differentiation coexist. To elucidate the clinicopathological implications of cellular differentiation in CXPA and the potential role of p53, vascular endothelial growth factor (VEGF), c-erbB-2, c-kit, and glucose transporter 1 (Glut-1) in carcinogenesis, we analyzed 11 CXPAs with luminal differentiation (CXPAs-LD) and 6 CXPAs with non-luminal differentiation (CXPAs-NLD) and compared protein expressions in residual PAs and carcinomas by immunohistochemistry. Among the CXPAs-LD, 5 were invasive and 8 were histologically high-grade tumors. The 5-year survival rate was 72.7%. P53, c-erbB-2, VEGF, and Glut-1 were more immunoreactive in carcinoma components than in PAs (P = 0.008, 0.004, 0.002, and 0.024, respectively); c-erbB-2 overexpression was associated with high histological grade (P = 0.024). Carcinoma components frequently lacked c-kit expression (P = 0.009). CXPAs-NLD were all low-grade and invasive with a larger mean tumor size (5.2 cm) than CXPAs-LD (3.3 cm) (P = 0.040). The patients remained disease-free without significant immunohistochemical expression. The immunoprofiles and clinical course of CXPA differed according to cellular differentiation. Therefore, it is important to report the histological subtype and to assess potential biomarkers in diagnostic and therapeutic trials.

Extracellular matrix metalloproteinase inducer expression in salivary gland tumors: a correlation with microvessel density.

The purpose of this study was to investigate the expression pattern of extracellular matrix metalloproteinase inducer (EMMPRIN) as well as the correlation between EMMPRIN and microvessel density (MVD) in salivary gland tumors. Extracellular matrix metalloproteinase inducer expression and MVD were examined immunohistochemically on paraffin-embedded tissue specimens from 95 patients with salivary gland tumors, who underwent surgical resection from 1998 to 2006. Reverse transcription-polymerase chain reaction was used to monitor EMMPRIN mRNA expression in frozen samples. Extracellular matrix metalloproteinase inducer expression in mucoepidermoid carcinomas and adenoid cystic carcinomas was significantly higher than in normal salivary gland tissues and pleomorphic adenomas (P < 0.05). The MVD of mucoepidermoid carcinomas and adenoid cystic carcinomas was significantly higher compared with pleomorphic adenomas (P < 0.05). The MVD of the EMMPRIN-positive expression group was significantly higher than the MVD of the EMMPRIN-negative expression group (P < 0.05). Extracellular matrix metalloproteinase inducer mRNA expression in malignant salivary gland tumors was higher than that in pleomorphic adenomas (P < 0.05). This study suggests that EMMPRIN expression is an important feature of malignant salivary gland tumors and can be used as a biologic marker to characterize salivary gland tumors. Extracellular matrix metalloproteinase inducer is also a positive angiogenic factor in salivary gland tumors.

Immunoglobulin mRNA and protein expression in human oral epithelial tumor cells.

In our previous work, we have reported the expression of immunoglobulin (Ig) molecules by numerous epithelial cancer cells and hyperplastic epithelial cells. In the present study, we extended our investigation to study the frequencies of expression of IgG and IgA in some types of oral epithelial tumor cells, and analyzed the oral tumor-derived V regions characteristic of Ig gamma chain gene transcripts by immunohistochemistry, in situ hybridization, laser capture microdissection-correlated reverse-transcription polymerase chain reaction, and sequencing. IgG and IgA immunoreactivity was prominent in the cytoplasmic or plasma membrane or secretion of malignant cells, pleomorphic adenoma tumor cells, and some normal glandular epithelial cells or squamous cells adjacent to tumors. More importantly, rearranged Ig gene transcripts were identified in these tumor cells, and in some normal glandular epithelial cells, the V-D-J region sequences revealed that IgG transcripts in 2 tested oral tumors were oligoclonal. These results support that the phenomenon of Ig could also be expressed in oral cavity epithelial tumor cells.

Carcinoma ex pleomorphic adenoma of the salivary gland: an immunohistochemical study.

The proliferative activity of the tumor cells and the expression of tumor-associated genes and sex steroid hormone receptors were investigated immunohistochemically in ten cases of carcinoma ex pleomorphic adenoma (Ca-ex-PA) of the salivary glands. These were analyzed in benign and malignant components separately, and then were compared with ten cases of the other malignant tumors [adenocarcinomas, not otherwise specified (ACN) and salivary duct carcinomas (SDC)] and ten cases of pleomorphic adenomas (PA). The results obtained in this study were as follows: (1) malignant component of Ca-ex-PA showed a higher incidence of PCNA and Ki67 than benign component of Ca-ex-PA. A significant difference between benign component of Ca-ex-PA and PA was not observed. (2) A significant difference in the incidence of p53, c-erbB-2, EGFR overexpression was observed only between malignant component of Ca-ex-PA and benign component of Ca-ex-PA. (3) The incidence of PCNA, Ki67, p53, c-erbB-2 overexpression in malignant component of Ca-ex-PA showed the highest data among the four groups. These results suggest that Ca-ex-PA acquired the particular biological behavior in contrast to the other salivary neoplasms in the long-standing process while PA undergoes malignant transformation.

Increased expression of cyclooxygenase-2 and Ki-67 are associated with malignant transformation of pleomorphic adenoma.

OBJECTIVES: In the present study, we attempted to identify cyclooxygenase-2 (COX-2) expression and Ki-67 index in carcinoma ex-pleomorphic adenoma (Ca ex-PA) using quantitative immunohistochemical analysis and to compare the benign component of the neoplasia. We also aimed to relate the overexpression of COX-2 with the pathways of malignant transformation of Ca ex-PA as evidenced by distinct morphological features. MATERIALS AND METHODS: Forty Ca ex-PA from patients treated at Department of Otolaryngology, Yokohama City University Medical Center, Yokohama, Japan, from 1999 to 2005, were selected. All Ca ex-PA showed only one malignant histological component: adenocarcinoma (23 cases), adenoid-cystic carcinoma (10), epithelial-myoepithelial carcinoma (7). The tissues were stained with monoclonal antibodies to COX-2 and Ki-67. The results were analyzed using quantitative immunohistochemical analysis. We also analyzed the association of the histological classification of the carcinomatous component. RESULTS: In the immunohistochemical analysis of COX-2 and Ki-67 index, significant increase was observed in Ca ex-PA, especially with adenocarcinoma, compared to pleomorphic adenoma and sialadenitis. Quantitative assessment is more sensitive as a measure of cellular protein content as compared to standard optical density measurement. CONCLUSIONS: The data support the hypothesis that increased COX-2 expression is associated with early events in malignant transformation of pleomorphic adenoma.

Expressão das integrinas a2B1, a3B1 e a5B1 em adenoma pleomórfico de glândula salivar menor e carcinoma adenóide cístico / Immunohistochemical expression of a2B1, a3B1 a 5B1 integrins in pleomorphic adenoma from minor major salivary glands and adenoid cystic carcinoma

O adenoma pleomórfico e o carcinoma adenóide (CAC) representam neoplasias de glândula salivar e maligna, respectivamente, as quais compartilham algumas características com a mesma origem celular e uma marcante presença de matriz extracelular, apresentando, porém, comportamento biológicos distintos. O propósito desta pesquisa consistiu em comparar a expressão das integrinas a2B1, a3B1, e a5B1 em adenomas pleomórficos de glândula salivar menor e maior e CACs. Além disso, procurou investigar se havia diferenças na expressão destas integrinas entre os subtipos histológicos do CAC. Foram selecionados 14 casos de adenomas pleomórfico de glândula salivar maior, 14 casos de glândula salivar menor e 10 casos de CACs. Analisou-se a presença ou ausência, localização e intensidade de marcação das integrinas. Os dois grupos de adenomas pleomórfico foram reunidos em um só para fazer a comparação entre os dois tumores. Verificou-se que houve diferença estatística altamente significativa (p<0,0001) para a integrina a2B1 entre os dois tumores, apresentando o adenoma pleomórfico, uma marcação mais intensa para esta integrina. Em relação à integrina a5B1 não foi possível a realização de testes estatísticos, ficando patente, porém, que houve uma tendência da referida integrina ser mais intensamente expressa no adenoma pleomórfico. Para a ánalise comparativa, os CACs foram subdivididos em 2 grupos: sólido e tubular/cribriforme. Para a integrina a2B1 observou-se que não houve diferença estatisticamente significativa e em relação à a3B1 e a5B1 não foi possível a realização do teste estatístico; no entanto, também foi verificada uma clara tendência para os casos do subtipo sólido apresentarem expressão ausente ou reduzida das integrinas avaliadas

Cyclooxygenase-2 regulates the degree of apoptosis by modulating bcl-2 protein in pleomorphic adenoma and mucoepidermoid carcinoma of the parotid gland.

CONCLUSION: These results suggest that COX-2 and bcl-2 protein were overexpressed and that apoptosis was reduced in MEC compared to PMA, and that COX-2 may regulate the degree of apoptosis by modulating bcl-2 protein in PMA and MEC. OBJECTIVE: Cyclooxygenase (COX)-2 plays a crucial role in tumorigenesis and overexpression of COX-2 in vitro accompanied by overexpression of bcl-2 protein has been shown to reduce apoptosis. The purpose of this study was to verify that COX-2 regulates the degree of apoptosis by modulating bcl-2 protein in benign and malignant parotid gland tumors. : We examined archival formalin-fixed, paraffin-embedded tissue sections of 10 pleomorphic adenomas (PMAs) and 10 mucoepidermoid carcinomas (MECs) by immunostaining with anti-COX-2, anti-bcl-2 and anti-single-stranded DNA (ssDNA) antibodies. Labeling indices of the three antibodies were calculated using computer-assisted image analysis. RESULTS: Labeling indices (mean+/-SD) of anti-COX-2 antibody in PMA and MEC were 2.05+/-1.30 and 11.2+/-2.95, respectively (p < 0.001), those of anti-bcl-2 antibody were 2.00+/-1.28 and 9.68+/-4.05, respectively (p < 0.001) and those of anti-ssDNA antibody were 8.06+/-2.54 and 2.08+/-1.47; respectively (p <0.001). Correlation coefficients between the labeling indices of anti-COX-2 antibody and anti-bcl-2 antibody, anti-bcl-2 antibody and anti-ssDNA antibody and anti-COX-2 antibody and anti-ssDNA antibody were 0.88, -0.75 and -0.76, respectively (p <0.001).

Adenoma pleomorfo / Pleomorphic adenoma

El adenoma pleomorfo es una neoplasia benigna y es el tumor más frecuente de aquellos que derivan de los tejidos que forman las glándulas salivales. En años recientes se han publicado casos de transformación maligna de esta neoplasia. En este trabajo se exponen los pincipales resultados de diferentes investigaciones sobre este tema. Asimismo, se revisarán las características clínicas, histogénesis, hallazgos microscópicos, inmunología, transformación maligna, tratamiento y pronóstico

Expression of CD44 standard and variant isoforms in parotid gland and parotid gland tumours.

In this study, we investigated the distribution of the standard form of the CD44 (CD44s) cell adhesion molecule and of its v3 and v6 isoforms in samples of foetal and adult parotid gland tissue, in comparison with samples of parotid gland adenomas and carcinoma ex pleomorphic adenoma. Foetal parotid gland showed CD44s and CD44v3 expression in the peripheral small primordial ducts and acini, while CD44v6 was only focally expressed. Adult parotid gland tissue showed a similar distribution of CD44s and variants, with a predominant expression in acinar structures and a weaker expression at duct level. In parotid gland adenomas, a diffuse and intense expression of CD44s and variants 3 and 6 was observed only in pleomorphic adenomas, while expression of CD44s was prevalent in Warthin's tumour, myoepithelioma and oncocytoma. The malignant areas of carcinoma ex pleomorphic adenoma showed a markedly decreased expression of CD44v3 and CD44v6 in comparison with the adjacent pleomorphic adenoma component. In conclusion, the prevalent expression of CD44s and variants in pleomorphic adenoma in comparison with other adenomas may be related to the abundant extracellular matrix production present in these tumours, while loss of CD44v3 and CD44v6 associated with the onset of carcinoma ex pleomorphic adenoma could promote stromal invasion, eventually contributing to the development of distant metastases.

P53 and Ki-67 antigen expression in small oral biopsy specimens of salivary gland tumors.

OBJECTIVE: To investigate the possibility that various salivary gland tumors that look histologically similar could express p53 oncoprotein and Ki-67 proliferation antigen differentially and possibly aid in distinguishing benign from malignant lesions. STUDY DESIGN: Intraoral paraffin-embedded biopsy specimens of salivary gland tumors were used. Thirty-eight pleomorphic adenomas, 17 adenoid cystic carcinomas, 23 monomorphic adenomas, and 17 polymorphous low-grade adenocarcinomas were stained with p53 and Ki-67 antibodies by using an immunoperoxidase detection system. Each case was evaluated in terms of staining intensity and percentage of cells staining. RESULTS: Ki-67 and p53 antigens are expressed in generally low levels in the histologically well-differentiated salivary tumors that were studied here, both benign and malignant. Only 1 solid-type adenoid cystic carcinoma showed a high percentage of cells expressing p53. CONCLUSIONS: The histologically well-differentiated salivary tumors studied do not show differential expression of p53 and Ki-67, in spite of their differing courses of biologic behavior. These antibodies should not be relied on to distinguish benign from malignant lesions of the salivary glands; however, they might be markers for those lesions that are dedifferentiating histologically and, therefore, might be displaying more aggressive behavior.

Immunohistochemical detection of DNA topoisomerase type II alpha and Ki-67 in adenoid cystic carcinoma and pleomorphic adenoma of the salivary gland.

Immunohistochemical detection of cell proliferation-associated antigens was investigated in 28 cases of adenoid cystic carcinoma (ACC) and 20 cases of pleomorphic adenoma (PA), using antibodies against DNA topoisomerase type II alpha (topo-II) (Ki-S1) and Ki-67 (MIB-1). The correlation of staining indices with clinicopathological data, histological features and prognosis was also studied. The topo-II value was significantly higher in ACC than in PA (P<0.0001), and highest in the solid growth pattern of ACC. In addition, significant relationships were found between topo-II values and clinical features such as local recurrence, surgical margins, and distant metastases. By log-rank test, the topo-II index was also correlated significantly with patient survival (P<0.01). The values of topo-II index paralleled those of Ki-67 index in ACC, and a correlation coefficient of 0.97 was obtained. Topo-II may be considered an additional marker for estimating the proliferating fraction of cells and for predicting the short-term prognosis for patients with salivary gland tumors.

Comparison of integrin alpha chain expression in benign and malignant salivary gland tumors.

OBJECTIVE: This study investigates the distribution of the alpha chain of the integrin family of extracellular matrix receptors in a series of adenomas and carcinomas of salivary gland origin to determine if the malignant phenotype is associated with modification of the expression of these receptors. STUDY DESIGN: Cryostat sections of 36 tumor specimens were stained by a standard streptavidin-biotin-peroxidase technique using primary monoclonal antibodies against alpha 1-6 and alpha v integrin chains. The immunohistochemical reaction was scored using a three-point scale and the results were analyzed using Fisher's exact test. RESULTS: In salivary adenomas, alpha 2, alpha 3, alpha 4, alpha 6, and alpha v chains were widely expressed in most of the cases studied. The alpha 1 subunit was prominently expressed by the epithelial cells of Warthin's tumor, whereas a minority of pleomorphic adenomas showed immunoreactivity for this antigen. We observed alpha 5 subunit expression only in the mesenchymal-like component of pleomorphic adenomas. In salivary carcinomas, integrin alpha chain expression was heterogeneous, varying greatly between different histotypes and within the same histotype. The distribution of the antigens was similar to that of adenomas, except for the alpha 6 chain, which localized not only at the interface between cell and matrix, but also at sites of cell-cell contact. When the immunohistochemical levels of integrin alpha chain expression were compared in adenomas and carcinomas, expression significantly decreased for the alpha 6 and alpha v chains (p = 0.0007; p = 0.002, respectively). CONCLUSIONS: Loss of alpha 6 and alpha v integrin subunits occurring in salivary gland carcinomas could modify the adhesive properties of malignant cells, contributing to the invasive potential of these tumors.

Chondroid syringoma: an immunohistochemical study using antibodies to Ca 15-3, KA-93, Ca 19-9, CD44 and BM-1.

The expression of Ca 15-3, KA-93, Ca 19-9, CD44 and BM-1 in normal skin and chondroid syringoma was investigated immunohistochemically using antibodies to these antigens. In the normal skin, the eccrine ducts and/or secretory elements were positive for all these antigens. On the other hand, the apocrine ducts and/or secretory elements were positive for the antibodies to Ca 15-3, KA-93 and CD44. In chondroid syringoma, the luminal cells of tubuloglandular structures were positive for the antibodies to Ca 15-3 and BM-1, and partly positive for those to KA-93 and Ca 19-9. The basal cells of the tubular structures and solid nests were positive only for the antibody to CD44. Stromal cells in the myxoid area were positive for the antibodies to KA-93 and CD44, and the chondroid matrix was positive for the antibody to BM-1. It is suggested that chondroid syringoma might originate from, or differentiate into the ducts and/or secretory elements of the eccrine sweat glands. In addition, the significance of the expression of BM-1 in the chondroid matrix is discussed.

Salivary gland anlage tumor of the nasopharynx: a clinicopathologic and immunohistochemical study of three cases.

The histologic and immunohistochemical features of three congenital pedunculated nasopharyngeal midline masses are reported. The follow-up in all cases was uneventful. The tumors were characterized by solid and cystic squamous nests and ductlike structures focally continuous with the surface squamous mucosa of the tumor. Most of epithelial structures coalesced with densely cellular stroma-like nodules. Immunoperoxidase staining showed the presence of epithelial markers in both spindle cells and epithelial structures. Spindle cells were also reactive to vimentin and smooth muscle actin, revealing their myoepithelial phenotype. Based on these observations, a diagnosis of salivary gland anlage tumor, also called congenital pleomorphic adenoma of the nasopharynx, was made. The similarity of these tumors' architecture and cellular composition to the normally developing salivary gland has led to the hypothesis of a tumorlike, hamartomatous lesion developing in a site in which minor salivary gland tissue occurs. This report reviews 12 identified cases of this tumor, of which all but one (in which the patient died of sepsis) had a favorable outcome. In an infant with respiratory distress and/or feeding difficulties, these tumors must be differentiated from other midline masses such as encephaloceles and teratomas. They appear curable by surgical exeresis only.

Immunohistochemical study of c-erbB-2 expression in carcinoma ex-pleomorphic adenoma.

A series of 19 cases of carcinoma ex-pleomorphic adenoma was studied for the immuno-expression of c-erbB-2 oncoprotein. Twelve tumours showed a malignant component with only one histological type; in the remaining seven there was co-existence of areas of various carcinoma types, adenocarcinoma NOS being the most frequent. Membranous c-erbB-2 reactivity was found in 21.1% of the cases, all corresponding to high-grade adenocarcinomatous areas. The low-grade carcinoma types that formed the malignant mixed tumours components were negative. Benign pleomorphic adenoma areas, either adjacent or intermingled with carcinomatous areas, were also consistently negative, proving that c-erbB-2 accumulation is associated with the acquisition of the malignant phenotype. The finding of a preferential association between c-erbB-2 overexpression and high-grade malignant mixed tumour may indicate prognostic implications for the oncogene protein and may also be indicative of its specific relationship with the putative pathway of malignant transformation in pleomorphic adenomas.