Expression of Mcm-2, Ki-67 and geminin in benign and malignant salivary gland tumours

Background:  Recent studies have proposed that minichromosome maintenance (Mcm) proteins may be sensitive proliferation markers and may serve as novel biomarkers for prognostication and diagnosis of various pre-malignant and malignant lesions. The aims of this study were to determine the expression of Mcm-2, Ki-67 and geminin in salivary gland (SG) tumours, and to evaluate their usefulness for diagnosis or for prediction of tumour behaviour.
Methods:  Tissue from 62 SG tumours was assembled in tissue microarray format. There were 13 adenoid cystic carcinomas (ACC), 10 carcinoma ex pleomorphic adenomas (CEPA), 10 mucoepidermoid carcinomas (MEC), 10 polymorphous low-grade adenocarcinomas (PLGA), 10 pleomorphic adenomas (PA) and nine acinic cell carcinomas (AcCC). Clinicopathological data were collected retrospectively and immunohistochemical analyses of Mcm-2, Ki-67 and geminin were performed on all lesions. Labelling index (LI) for each marker was determined by counting the percentage of positive cells in six random fields from three arrays per case.
Results:  Mcm-2 expression was higher than Ki-67 and geminin in all tumours studied. Mcm-2 LI was higher in ACC (28.2 ± 19.2%) than in CEPA, AcCC, MEC, PA and PLGA (5.3 ± 4.1%, P  = 0.001). Mcm-2 LI was higher in CEPA (20.4 ± 5.0%) than in PA (6.9 ± 5.0%, P  = 0.001). LI did not correlate to tumour grade or outcome for any of the markers or tumour types.
Conclusions:  The findings suggest that Mcm-2 may be a sensitive proliferation marker in SG tumours and may be useful for differential diagnosis between PA and CEPA, and ACC and PLGA. Further studies are warranted to assess the value of Mcm-2 as a predictor of recurrence and survival.     

DNA-Ploidie und proliferative Aktivität bei Speicheldrüsentumoren

DNA ploidy and S-Phase fraction (SPF) of 279 salivary gland tumours were analysed using high-resolution DNA flow cytometry. All 229 benign neoplasms were diploid while 12 of 50 malignant tumours showed cell populations with aneuploid DNA content. The SPF values of diploid malignancies were significantly higher if compared with pleomorphic adenomas but did not differ from that of the zystadenolymphoma (Warthin tumour) group. While aneuploidy represents a distinct indicator of malignancy SPF values are of minor relevance for dignity assessment in salivary gland tumours.     

Flow cytometric S-phase fraction contributes to diagnosis of diploid malignant salivary gland tumours

DNA ploidy studies on salivary gland tumours have shown that the proportion of aneuploid cases, although confined to the malignant entities, is considerably lower than for other solid malignancies. To analyse whether the S-phase fraction (SPF) may contribute to discrimination of diploid malignant from benign tumours, DNA flow cytometric data from 45 different malignant salivary gland tumours was compared with that of 121 pleomorphic adenomas. All benign tumours were diploid. Twelve malignant tumours contained aneuploid cell populations. The SPF values for diploid malignancies ranged between 0.9% and 11.0% (mean 3.9%), and between 0.5% and 7.9% (mean 2.7%) for pleomorphic adenomas. A 4% cut-off value gained statistical significance for discriminating diploid malignant tumours from pleomorphic adenomas (P<0.01). The sensitivity for SPF>4% was 46% and the positive predictive value was 40%. A sensitivity of 60% and a positive predictive value of 54% was achieved by combining aneuploidy and SPF>4%. These results show that DNA flow cytometry may contribute to diagnostic assessment in salivary gland tumours.     

DNA content in malignant salivary gland tumours

Objective:  Our aim was to evaluate the DNA content in malignant salivary gland tumours using image cytometry and its possible relationships with clinical and morphologic findings, disease course and prognosis.
Patients and methods:  The study sample comprised 31 patients diagnosed and treated for primary malignant salivary gland tumours. Formalin-fixed, paraffin-embedded surgical specimens of all patients were Feulgen-stained for DNA content analysis by image cytometry. Statistical analysis was used to investigate possible relationships between DNA content variables and clinical and histological findings, disease course and patient survival.
Results:  Seventeen (55%) cases of our sample were graded as DNA diploid, four (13%) as DNA aneuploid and 10 (32%) as DNA multiploid. In 15 (48%) cases, the 5c exceeding rate (5cER) was higher than 1.7%. DNA ploidy correlated with N stage and tumour size. DNA ploidy and 5cER had a statistically significant prognostic influence on overall and disease-free survival in univariate analysis. However, in multivariate analysis, stage classification was the only parameter with an independent prognosis value.
Conclusion:  Abnormal DNA content is a common finding in salivary gland cancers. Our results suggest an important role of DNA content analysis in the evaluation of these tumours.     

The significance of DNA flow cytometry in predicting survival and delayed clinical manifestation of occult lymph node metastasis to the untreated neck in patients with oral squamous cell carcinoma

A total number of 116 clinically neck-negative patients with squamous cell carcinoma of the oral cavity who underwent radical primary tumour surgery without simultaneous neck treatment were entered into this prospective study. The 5 year overall survival rate was 87% for patients with flow cytometrically diploid tumours and 58% for the aneuploid group (P < 0.05). By multivariate survival analysis, tumour stage (P < 0.05) and DNA ploidy (P < 0.05) were significantly associated with the outcome. The cumulative 3 year rate of delayed clinical manifestation of lymph node metastasis to the previously untreated neck was 12.6% for patients with flow cytometrically diploid tumours and 41.3% for the aneuploid group (P < 0.01). By multivariate analysis, the DNA ploidy status of the primary tumour was the only factor among tumour stage, localization and degree of histological differentiation predictive of occult metastasis development (P < 0.05). Also, patients with T1 tumours who frequently are not considered to benefit from elective neck dissection were at high risk of subclinical lymph node involvement if the primary tumours were aneuploid (47%), whereas only 10% of the diploid T1 sample showed occult neck disease. Particularly in patients with less extensive oral carcinomas, DNA aneuploidy is therefore an important decisive factor in elective neck dissection.     

增殖细胞核抗原PCNA在涎腺肿瘤中的表达

采用抗增殖细胞核抗原(proliferating cell nueler antigen,PCNA)单克隆抗体PC10,对49例涎腺肿瘤标本进行免疫组化染色.结果显示:涎腺恶性肿瘤与正常涎腺组织和良性肿瘤间,高分化与低分化粘液表皮样癌间及腺样囊性癌实质型与筛孔型之间PCNA标记指数均存在显著差异.提示:PCNA表达对判断涎腺肿瘤的增殖活性有一定意义.     

Ploidy studies of pleomorphic adenomas and minor salivary gland carcinomas of the palate

Ploidy studies of tumors are a diagnostic and prognostic aid. The aim of the present study was to evaluate the DNA content of palate aggressive pleomorphic adenomas (PA) and adenocarcinomas of the salivary glands. Twelve cases of salivary gland tumors of the palate were selected from the archives of the Oral Pathology Department, Faculty of Dentistry, University of Buenos Aires (1966-2001). Six cases corresponded to aggressive pleomorphic adenomas (PA) and the remaining six to adenocarcinomas (AD). Myxoid and epithelial areas of PA were evaluated. The epithelial areas of the most aggressive cases of PA exhibited a high DNA content. The myxoid areas of same cases of PA had a 2C ploidy level. The difference in ploidy values between the myxoid and epithelial areas of PA would suggest the presence of different cell populations. DNA content and the detection of aneuploidy would be prognostic aids in palate salivary gland carcinomas.     

DNA flow cytometry of reclassified subtypes of malignant salivary gland tumors

Malignant salivary gland tumors are rare, constitute a heterogeneous group and are often difficult to diagnose histologically. This is borne out by the fact that in the present study 43.2% of 118 salivary gland tumors originally diagnosed as mucoepidermoid, acinic cell and adenoid cystic carcinomas had their original diagnosis altered upon reclassification. Patients with confirmed adenoid cystic carcinomas had a much worse prognosis than those with mucoepidermoid and acinic cell carcinomas.
DNA flow cytometry showed that very few of the above mentioned three types of malignant neoplasms revealed aneuploid DNA stemlines, indicating that this is not a relevant prognostic tumor marker within the groups. However, several of the tumors that had their diagnosis changed, mostly to undifferentiated adenoor squamous cell carcinomas, showed aneuploid DNA stemlines. The survival time of patients with aneuploid tumors was considerably reduced compared to those with diploid tumors. Among confirmed acinic cell, mucoepidermoid and adenoid cystic carcinomas the S-phase fraction was a significant prognostic factor, as it was among all tumors examined. This indicates that DNA aneuploidy and S-phase fractions are potential prognostic factors for malignant salivary gland tumors, and that DNA flow cytometry may assist the characterization of such tumors.     

Mammary Analogue Secretory Carcinoma (MASC) of salivary gland in four Mexican patients

The Clinco-pathological, immunohistochemical and molecular findings of four cases of Mammary Analogue Secretory Carcinoma (MASC) of salivary glands found in Mexico are described. The cases were extracted from 253 salivary gland tumors from a single institution in Mexico City. The 85 candidates for initial selection were: low grade mucoepidermoid carcinoma (MEC) (N=70 ), acinic cell cancinoma (AciCC) (N=14), papillary cystadenocarcinoma (N=1), and adenocarcinoma NOS (N=0). Tumors with some histological features consistent with MASC (N= 17, 6.7%) were studied by immunohistochemistry for mammaglobin, STAT5, and S-100 protein and four cases were positive (1.5%), thus the diagnosis of MASC was established, and these were submitted for molecular studies for ETV6-NTRK3. Fusion gene was demonstrated in three cases, two had been erroneously diagnosed as poorly granulated AciCC, and one as low grade MEC with microcystic pattern. Female gender predominated (3:1); one occurred in the parotid, two in minor salivary glands and one in the submaxillary gland; infiltrating borders, atypical mitosis and lymph node metastases were seen in the parotideal tumor. Two patients with major salivary gland tumors are alive and well at 10 and 20 months respectively, the two patients with minor salivary gland tumors are lost. It can be concluded that is important to think in MASC in poorly granulated AciCC and low grade MEC with microcystic pattern. Immunohistochemisty studies confirm the diagnosis, preferentially supported by molecular studies. MASC may follow aggressive behavior or transform into a high grade neoplasm. Key words:Acinic cell carcinoma, ETV6-NTRK3, Mammary Analogue Secretory Carcinoma, secretory breast carcinoma.     

Intraoral minor salivary gland neoplasms: review of 213 cases.

PURPOSE: Minor salivary gland tumors (MSGTs) constitute a heterogeneous group of neoplasms with great histomorphologic variation. This study reviews a large series of benign and malignant salivary gland tumors of the oral region and determines the incidence and the correlation of the histopathologic features with the clinical characteristics. MATERIALS AND METHODS: Two hundred thirteen cases of MSGT were retrospectively studied. Hematoxylin-eosin-stained slides were examined in all cases. Special stains and immunohistochemical stains were used in selected cases. Clinical characteristics of the neoplasms were also noted. RESULTS: One hundred nineteen tumors were benign (56%), and 94 tumors were malignant (44%). Pleomorphic adenoma was the most common benign tumor (93 of 119). Canalicular adenoma was the second most common benign MSGT in our series (25 of 119). Of the 94 malignant MSGTs, mucoepidermoid carcinoma (MEC) (45 of 94), adenoid cystic carcinoma (22 of 94), and polymorphous low-grade adenocarcinoma (18 of 94) were the most common. Most MECs (34 of 45) were low-grade lesions. Of 5 central MECs, 3 cases occurred in the maxilla and 2 cases arose in the mandible. CONCLUSIONS: Benign intraoral MSGTs are slightly more common than malignant MSGTs. Pleomorphic adenoma is the most common MSGT, and MEC is the most common malignant variety. The palate is the most common site for minor gland neoplasms. Benign labial salivary gland neoplasms are more common in the upper lip, and malignant labial tumors are more common in the lower lip.     

Quality of life in patients submitted to surgical treatment for minor salivary gland neoplasms

This study was aimed at assessing the quality of life in patients submitted to surgical treatment for minor salivary gland neoplasms (MSGN). Twelve patients (10 women and 2 men, mean age: 49.4 years) with histopathologic diagnosis of pleomorphic adenoma (PA, 3 cases), polymorphous low-grade adenocarcinoma (PLGA, 2 cases), cystic adenoid carcinoma (CAC, 4 cases), and muco-epidermoid carcinoma (MEC, 3 cases) were evaluated. All of them were treated by surgical excision; patients with CAC received radiotherapy as well. The patients quality of life was evaluated through a self-administered questionnaire concerning their physical well-being, emotional status, normal daily activities, and family relationships. The results showed that patients with MEC--the youngest among all patients--reported a significantly greater worsening of their physical well-being and emotional status after treatment as compared with patients treated for PA (P<0.05), and also of their functional activities as compared with those treated for PA and PLGA (P<0.05). In conclusion, age of development of the neoplasm and type of disease produce more impact on patients quality of life than does the therapys degree of aggression.     

DNA analysis of oral leukoplakia by flow cytometry.

DNA ploidy of 19 oral leukoplakias with and without epithelial dysplasia was investigated and the results were compared with 11 normal gingival biopsies, 14 oral benign tumours and 50 oral squamous cell carcinomas. The results suggest a possible relationship between DNA aneuploidy and oral leukoplakias or squamous cell carcinomas, as 32% of the oral leukoplakias and 48% of the oral squamous cell carcinomas were aneuploid although all the normal gingival biopsies and the benign oral tumours examined were diploid. No significant relationship was observed, however, between DNA ploidy and epithelial dysplasia in the leukoplakias.     

Paediatric mucoepidermoid carcinoma of the palate

The purpose of this paper is to review our experience with mucoepidermoid carcinoma (MEC), a rare tumour in minor salivary glands, in a small series of paediatric patients. A retrospective analysis of minor salivary gland tumours seen by one surgeon from March 1991 to December 1999 was undertaken. A total of 58 cases were identified and of these, five (9%) occurred in children. There were 23 cases of MEC, four (17%) of which occurred in patients under the age of 18 who presented with T1 or T2N0M0 low- to intermediate-grade MEC of the palate and adjacent structures. These patients form the basis of this study. All patients were treated with wide local excision, obtaining tumour-free margins, and followed for a mean number of 58 months. None of these tumours invaded bone and resection of bone was not performed in any case as the periosteum was intact and the tumours were low to intermediate grade. To date, all patients remain free of disease. One patient who went elsewhere for treatment, was treated with local resection only, and has also experienced no recurrence. Wide local excision is the treatment of choice for low to intermediate grade MEC of the minor salivary glands in paediatric patients.     

PCNA, Ki-67 and p53 expressions in submandibular salivary gland tumours

Salivary gland tumours are uncommon with a broad heterogeneity. The most common benign tumour is the pleomorphic adenoma, whereas mucoepidermoid carcinoma and adenoid cystic carcinoma predominate among the malignancies. Most salivary gland tumours occur in the parotid, and consequently clinical and biological data are normally derived from this site. This work describes the expressions of PCNA, Ki-67 and p53 in 15 pleomorphic adenomas, 15 mucoepidermoid carcinomas and 15 adenoid cystic carcinomas of the submandibular gland. Our results showed that all pleomorphic adenomas were negative for p53 and Ki-67 with 66.6% being positive for PCNA. Conversely, p53 was positive in 53% of the mucoepidermoid carcinomas and in 20% of the adenoid cystic carcinomas. Ki-67 was expressed in 47.7% of the mucoepidermoid carcinomas and 40% of the adenoid cystic carcinomas. All malignant tumours were positive for PCNA. These results indicate that the proliferative rate analysed with PCNA and Ki-67 and the expression of p53 in pleomorphic adenoma and adenoid cystic carcinoma of the submandibular gland were similar to those described in the parotid and minor salivary glands. However, mucoepidermoid carcinomas showed higher expression of these markers than those of other salivary glands. This work is the first describing the expression of these immunohistochemical markers exclusively in submandibular salivary gland tumours.     

Maspin expression in normal and neoplastic salivary gland

BACKGROUND: Maspin inhibits cell motility, invasion and metastasis. Loss or reduction in maspin expression has been associated with tumoral progression. METHODS: The presence of maspin was studied immunohistochemically in salivary gland tumours presenting cells with myoepithelial differentiation in their composition, and in normal salivary gland. RESULTS: Pleomorphic adenoma (PA) presented high expression of maspin, except in the spindle cells and occasional luminal cells. Epithelial-myoepithelial carcinoma and tubular adenoid cystic carcinoma (ACC) showed intense expression in all cells. Cribriform ACC evidenced only few positive cells of the luminal type, while solid subtype showed rare positive cells. Normal salivary gland tissue has shown low levels of maspin positivity. CONCLUSIONS: Maspin has small participation in normal salivary gland, is increased in PA, and decreases as the histological malignancy raises. Hence, in salivary gland, its expression is not exclusive of myoepithelial cells; thus, it should not be used as a marker for this cell. Nevertheless, we believe it is an important marker of biological behaviour in these tumours.     

细胞外基质金属蛋白酶诱导因子表达与唾液腺肿瘤侵袭性的关系

目的:检测唾液腺肿瘤组织中细胞外基质金属蛋白酶诱导因子(EMMPRIN)的表达和定位,探讨其与唾液腺肿瘤侵袭性生物学行为的关系。方法:采用免疫组织化学方法,检测9例唾液腺正常组织、28例多形性腺瘤(PA)、25例黏液表皮样癌(MC)、33例腺样囊性癌(ACC)中EMMPRIN的表达和定位。采用SPSS10.0软件包进行多个样本率间的χ2检验或确切概率分析。结果:EMMPRIN在正常唾液腺组织、多形性腺瘤、黏液表皮样癌、腺样囊性癌的表达阳性率分别为11.11%、53.71%、84.00%和90.91%(P<0.05)。在正常唾液腺组织的表达主要见于导管上皮细胞的细胞膜;EMMPRIN蛋白在多形性腺瘤、黏液表皮样癌、腺样囊性癌的表达见于肿瘤细胞的胞膜或胞质。腺样囊性癌嗜神经现象组中,EMMPRIN表达的阳性率高于无嗜神经现象组,但差异无统计学意义。结论:EMMPRIN的表达与唾液腺肿瘤侵袭性生物学行为密切相关。     

Salivary duct carcinoma in major and minor salivary glands. A clinicopathological analysis of four cases

Salivary duct carcinomas are an uncommon but distinct group of highly malignant salivary gland tumours. We report the clinical course, pathological findings and surgical treatment of this tumour in 4 patients. In one patient an intraductal tumour developed in a minor salivary gland, while in the other three patients, a major salivary gland was involved by an infiltrating salivary duct carcinoma. We point out the highly aggressive biological behaviour of the tumour when occurring in the major salivary glands, in contrast to the benign course of the intraductal carcinoma in the minor salivary gland.     

Genetic analysis of DNA microsatellite loci in salivary gland tumours: comparison with immunohistochemical detection of hMSH2 and p53 proteins

To investigate genetic alterations in salivary gland tumours, microsatellite instability at eight representative loci and loss of heterozygosity (LOH) on chromosome 17 were analysed by polymerase chain reaction amplification. The results were compared with immunohistochemical expression of the hMSH2 and p53 proteins. Microsatellite instability and expression loss of hMSH2 protein were not recognized in the salivary gland tumours, suggesting a low frequency of abnormalities of the mismatch repair system. LOH associated with the p53 gene was detected in approximately one-half of pleomorphic adenomas and salivary carcinomas, which often showed strong p53 immunoreactivity. These features suggest that the p53 gene plays an important role in malignant transformation of salivary gland tumours. The genetic characteristics of pleomorphic adenomas might reflect a low-grade potential for malignant progression.     

Lack of correlation between DNA ploidy, Langerhans cell population and grading in oral squamous cell carcinoma

This study was undertaken to determine the inter-observer reproducibility of the invasive cell grading method on oral squamous cell carcinomas and to correlate this with the DNA ploidy status and Langerhans cell (LC) population. Fifty formalin-fixed paraffin-embedded blocks that contained both tumor and adjacent normal epithelium were randomly selected. DNA ploidy analysis was performed on a flow cytometer and the LC population was determined using an immunohistochemical technique with anti-S100 and anti-HLADR primary antibodies. The inter-observer correlation of the total malignancy score and individual morphologic features was highly significant. Thirty-two of the 50 tumors were graded as poorly differentiated and 18/50 as moderately differentiated. Thirteen of 25 poorly differentiated tumors had an aneuploid DNA content compared with 9/18 of the moderately differentiated group. No statistical differences in the LC counts between the poorly and moderately differentiated and aneuploid and diploid carcinomas were found. This study showed that the invasive cell grading method is reproducible but no correlation was found between the grading results and the ploidy status or LC count.     

涎腺癌DNA含量的流式细胞分析及其临床病理学意义

本实验应用流式细胞术(FLOW CYTOMETRY)对67例涎腺癌石蜡包埋样本的细胞核DNA含量进行定量检测,结果DNA异倍体的发生率为70％,不同组织学类型及不同分化程度的涎腺癌DNA倍体水平差别有显著意义。这表明DNA倍体异常是涎腺癌的一个重要生物学特征,对于指示涎腺癌的生物学行为、指导治疗和判断预后具有一定的临床意义.