Primary Epithelioid Angiosarcoma of the Submandibular Gland-A Case Report with Histology-Cytology Correlation and Comprehensive Molecular Analysis.

BACKGROUND: Angiosarcoma is a sarcoma that occurs in a range of tissue types, and only rarely in the salivary glands, showing a predilection for the parotid glands of older patients. Preoperative diagnosis may be challenging, especially on cytology, with significant morphological overlap with high-grade primary salivary gland carcinomas. The molecular alterations of this rare salivary gland neoplasm are also not well-characterized. METHODS AND RESULTS: We present a case of right submandibular gland swelling in a 73-year-old male. On fine needle aspiration, including immunohistochemical stains on cell block, the tumor was initially diagnosed as poorly differentiated carcinoma. Resection of the submandibular gland revealed epithelioid angiosarcoma. We performed molecular work-up of the tumor, utilizing targeted next-generation sequencing, DNA methylation profiling and fluorescence in-situ hybridization. Histopathologic assessment revealed an infiltrative tumor comprising solid sheets of epithelioid cells. The tumor cells formed haphazardly anastomosing vascular channels with intracytoplasmic lumina containing red blood cells. On immunohistochemistry, the tumor cells were positive for CD31, CD34 and ERG. Approximately 40% of the tumor cells showed nuclear expression of GATA3. A pathogenic TP53 R267W mutation was detected on next-generation sequencing. DNA methylation analysis did not cluster the tumor with any known sarcoma type. Copy number analysis showed possible MYC amplification and CDKN2A losses, although only the latter was confirmed on fluorescence in-situ hybridization. CONCLUSION: Epithelioid angiosarcoma is an important differential diagnosis to high-grade salivary gland carcinoma. In particular, GATA3 expression may be encountered in both angiosarcoma and high-grade salivary gland carcinomas and cause diagnostic confusion. Identification of TP53 mutations and CDKN2A losses suggest shared oncogenic pathways with soft tissue angiosarcomas, and should be further investigated.

Primary desmoplastic small round cell tumor of the submandibular gland: a case report and literature review.

BACKGROUND: Desmoplastic small round cell tumor (DSRCT) is a sporadic, highly malignant tumor with a poor prognosis. The abdomen and pelvis have been reported as the primary localization sites. However, to the best of our knowledge, there are few reports on primary DSRCT in the submandibular gland. CASE PRESENTATION: We report a case of a 26-year-old Chinese man with a mass in the right submandibular gland. Imaging studies showed a hypoechoic mass in the right submandibular region. Intraoperative pathology revealed that the tumor tissue was composed of small round tumor cells and a dense desmoplastic stroma. On immunostaining, the tumor cells showed markers of epithelial, mesenchymal, myogenic, and neural differentiation. The EWSR1 gene rearrangement was detected by fluorescence in situ hybridization. Based on the overall morphological features and immunohistochemical findings, a final diagnosis of DSRCT was made. The patient was treated with comprehensive anti-tumor therapy mainly based on radiotherapy and chemotherapy. CONCLUSIONS: DSRCT is an uncommon malignant neoplasm with rare submandibular gland involvement. In this report, we have described a case of DSRCT in the submandibular gland and reviewed the literature on DSRCT over the past 5 years. Considering the importance of differential diagnosis between DSRCT, especially with rare extra-peritoneal involvement, and small round blue cell tumors, a full recognition of the clinicopathological features will help to better diagnose this neoplasm.

Genomic Alterations in Undifferentiated Malignant Tumors with Rhabdoid Phenotype and Loss of BRG1 Immunoexpression Identified by Fine Needle Aspirates.

OBJECTIVE: Evidence shows that the switch/sucrose nonfermenting chromatin remodeling complex plays a critical role in DNA repair, cancer progression and dedifferentiation. BRG1 is one of its key catalytic subunits. While the loss of BRG1 expression by immunocytochemistry has been identified in a subset of malignancies arising in various sites with undifferentiated/rhabdoid morphology and poor prognosis, the underlying basis for its loss is unclear. METHODS: A retrospective search was conducted in our cytopathology archive for undifferentiated malignant tumors with rhabdoid phenotype and BRG1 loss. Clinical information was obtained from electronic medical records. Next-generation sequencing was performed following macro-dissection of paraffin-embedded cellblock tissue. RESULTS: Three cases were identified; all presented with widely metastatic disease with no previously diagnosed primary malignancy, and subsequently died within 6 months of initial presentation. Cytologically, the aspirates showed dyshesive and undifferentiated cells with rhabdoid features. Extensive immunocytochemical workup demonstrated immunoreactivity with vimentin only and could not establish a specific lineage. BRG1 expression was absent, while INI1 expression was retained. Two cases harbored deleterious mutations in BRG1/SMARCA4. Pathogenic mutations in TP53 were identified in all tumors. CONCLUSIONS: BRG1 deficiency reflects underlying mutation in SMARCA4 gene in some but not all cases, suggesting that additional mechanisms may be causing BRG1 silencing. Pathogenic mutations in TP53 in all tumors are consistent with their highly aggressive nature. Recognizing the cytomorphology of this group of neoplasms and confirming their BRG1-deficient status by immunocytochemistry not only has prognostic implications, but may also impart potentially therapeutic value in the near future.

The repertoire of genetic alterations in salivary duct carcinoma including a novel HNRNPH3-ALK rearrangement.

Salivary duct carcinoma (SDC) is a rare, aggressive malignancy with limited treatment options and poor outcome. Twenty-nine primary resected SDC, including 15 SDC de novo (SDCDN), and 14 SDC ex pleomorphic adenoma (SDCXPA) were subjected to the massive parallel sequencing assay (MSK-IMPACT) targeting 287 to 468 cancer-related genes. TP53 was the most frequently altered gene (69%). TP53 mutations and ERBB2 amplification were more frequent in SDCXPA than in SDCDN (P = .0007 and P = .01, respectively). Potentially targetable mutations were detected in 79% (23/29) of SDC involving ERBB2 (31%), PIK3CA (28%), HRAS (21%), ALK (7%) and BRAF (3%), and 22% (5/23) of those cases harbored possible primary resistance mutations involving CCNE1, NF1 and PTEN. A novel HNRNPH3-ALK rearrangement was found in one SDCDN. In another case, EML4-ALK fusion detected in the primary tumor was associated with ALK G1202R secondary resistance mutation in the post-treatment metastasis. A germline analysis of the DNA repair genes revealed a case with a pathogenic BRCA1 E23fs germline variant. SDCDN and SDCXPA are genetically distinct. Although the majority of SDC may be amenable to molecular targeted therapy, concurrent possible resistance mutations may be found in a significant minority of cases. A broad genomic profiling is necessary to ensure detection of rare but clinically actionable somatic alterations in SDC.

Genomic profiling of a combined large cell neuroendocrine carcinoma of the submandibular gland.

A 69-year-old female with no previous medical history presented with a rapidly growing submandibular mass. Fine needle aspiration cytology suggested a small-cell carcinoma and PET-CT showed increased 18-FDG uptake in the submandibular mass as well as in a lung mass. Submandibular resection and selective neck dissection was performed and histopathologic examination revealed a combined large-cell neuroendocrine carcinoma (LCNEC) with a squamous component and without lymph node metastases. Resection of the lung tumor revealed a papillary adenocarcinoma that was morphologically distinctly different from the LCNEC. The patient died of her lung cancer after 19 months without evidence of recurrence of the LCNEC. Genomic profiling of the salivary gland LCNEC revealed a hypodiploid genome predominated by losses of whole chromosomes or chromosome arms involving chromosomes 3p, 4, 7q, 10, 11, 13, 16q and gains of 3q and 16p. In addition, there was a segmental gain of 9p23-p22.3 including the NFIB oncogene. Continued studies of salivary gland LCNEC may provide new knowledge concerning potential diagnostic biomarkers and may ultimately also lead to the identification of new treatment targets for patients with these aggressive carcinomas.

Extra-prostatic transgene-associated neoplastic lesions in transgenic adenocarcinoma of the mouse prostate (TRAMP) mice.

Male transgenic adenocarcinoma of the mouse prostate (TRAMP) mice are frequently used in prostate cancer research because their prostates consistently develop a series of preneoplastic and neoplastic lesions. Disease progression in TRAMP mouse prostates culminates in metastatic, poorly differentiated carcinomas with neuroendocrine features. The androgen dependence of the rat probasin promoter largely limits transgene expression to the prostatic epithelium. However, extra-prostatic transgene-positive lesions have been described in TRAMP mice, including renal tubuloacinar carcinomas, neuroendocrine carcinomas of the urethra, and phyllodes-like tumors of the seminal vesicle. Here, we describe the histologic and immunohistochemical features of 2 novel extra-prostatic lesions in TRAMP mice: primary anaplastic tumors of uncertain cell origin in the midbrain and poorly differentiated adenocarcinomas of the submandibular salivary gland. These newly characterized tumors apparently result from transgene expression in extra-prostatic locations rather than representing metastatic prostate neoplasms because lesions were identified in both male and female mice and in male TRAMP mice without histologically apparent prostate tumors. In this article, we also calculate the incidences of the urethral carcinomas and renal tubuloacinar carcinomas, further elucidate the biological behavior of the urethral carcinomas, and demonstrate the critical importance of complete necropsies even when evaluating presumably well characterized phenotypes in genetically engineered mice.

HER2 and EGFR gene copy number alterations are predominant in high-grade salivary mucoepidermoid carcinoma irrespective of MAML2 fusion status.

AIMS: In this study, we aimed to investigate the molecular mechanisms underlying the development of mucoepidermoid carcinoma (MEC). METHODS AND RESULTS: In 31 cases, we examined the MAML2 fusion status using reverse transcriptase-polymerase chain reaction, and HER2 and EGFR status using immunohistochemistry and chromogenic in-situ hybridization. MAML2 fusions were detected in 15 (57.7%) of 26 MECs analysed, including 11 of 16 (68.8%) low-grade, two of four (50%) intermediate-grade and two of six (33.3%) high-grade MECs. HER2 gene amplification and an increased EGFR gene copy number (with balanced chromosome 7 high-polysomy) were each detected in four of 28 (14.3%) MECs analysed. Irrespective of MAML2 fusion status, all seven high-grade MECs had an increased gene copy number of either HER2 or EGFR, in a mutually exclusive manner, whereas such abnormalities were extremely rare in low- and intermediate-grade MEC. CONCLUSIONS: These results suggest that HER2 or EGFR gene abnormality could play an important role in the development of high-grade MEC, and also in the progression from MAML2 fusion-positive low-/intermediate-grade to high-grade in a subset of MEC. Furthermore, we suggest that high-grade MEC comprises a heterogeneous group of tumours in terms of molecular pathogenesis, in particular MAML2 fusion status.

Mammary analogue secretory carcinoma: the first submandibular case reported including findings on fine needle aspiration cytology.

We present the first case (male, 35 years old) of a mammary analogue secretory carcinoma occurring in a submandibular gland and document findings on fine needle aspiration cytology. On histology, the tumor displayed characteristic features: circumscribed nodules composed of bland, pink to light red neoplastic cells with low proliferative/mitotic activity arranged in tubular, vaguely cribriform, and microcystic structures containing Periodic acid Schiff-positive, diastase-resistant secretory material. Immunohistochemistry showed strong and diffuse positivity for cytokeratin 7, S100 protein, and vimentin, as well as moderate to strong immunoreactivity for c-kit in the majority of tumor cells. A rearrangement of the ETV6 gene on fluorescence in situ hybridization was documented. The patient underwent an ipsilateral selective (levels I-IV) neck dissection which showed metastasis in 3 out of 36 lymph nodes (levels 1-3). Adjuvant radiotherapy was administered. No local recurrence or metastatic disease has been detected during a follow up period of 28 months.

Therapy-related acute leukemia with mixed phenotype and t(9;22)(q32;q11.2): a case report and review of the literature.

Therapy-related acute leukemia showing mixed phenotype is extremely rare. We report a 49-year-old woman who presented with palpable masses in her neck and back. She had received systemic chemotherapy (adriamycin and cisplatin) and radiotherapy for endometrial adenocarcinoma 7 years before. Her peripheral blood and bone marrow showed increased blasts, which coexpressed myeloid (CD13, CD33, and myeloperoxidase) and B-lymphoid antigens (CD19 and CD79a). Cytogenetic analysis showed a karyotype of 46,XX,dup(1)(q21q32),add(5)(q33),t(9;22)(q34;q11.2)[12]/47,idem,+der(22)t(9;22)[8], and BCR/ABL1 rearrangement was detected. Leukemic infiltration was also confirmed in her back mass. After induction chemotherapy with idarubicin, cytarabine, and imatinib, she achieved complete remission. Only 2 cases of therapy-related acute leukemia with mixed phenotype have been reported so far: one with hyperploidy and the other with t(1;21)(p36;q22). To the best of our knowledge, this is the first case of therapy-related acute leukemia with mixed phenotype and t(9;22) as well as extramedullary leukemic infiltrations.

NUT gene rearrangement in a poorly-differentiated carcinoma of the submandibular gland.

NUT midline carcinomas (NMC) are a rare, recently described class of poorly-differentiated tumors that exhibit rapid onset and highly aggressive clinicopathologic behavior. These tumors are defined by rearrangement of the nuclear protein in testis (NUT) gene on chromosome 15q14, most commonly in a balanced translocation with the BRD4 gene on chromosome 19p13.1, resulting in the characteristic BRD4-NUT fusion gene and protein which blocks epithelial differentiation through chromatin binding. NMC frequently involve midline structures of adolescents and young adults and affect the head and neck region in 50% of cases. To our knowledge, only one case has been previously reported involving a salivary gland. Here, we present a case of a NMC of the salivary gland in an adolescent male presenting with an intermittently painful left submandibular mass of 3 months duration.

The tortoiseshell pattern in one or both sides of the submandibular glands in mucosa-associated lymphoid tissue lymphoma is related to chromosomal aberrations and the disease extent.

OBJECTIVE: Lesions of mucosa-associated lymphoid tissue (MALT) lymphoma in the submandibular glands are localized or a part of systemic involvement in association with chromosomal aberrations. This series was undertaken to investigate the sonographic features of MALT lymphoma in the submandibular glands and their relationships with chromosomal aberrations and the disease extent. METHODS: A total of 5 patients with MALT lymphoma without Sjögren syndrome in the submandibular glands were enrolled in this series. Patients underwent sonography of the submandibular glands with a high-resolution transducer before surgical biopsy of the main lesion. Sonographic characteristics of the lesions were described for their location, presence of a posterior echo, texture, and presence of an internal echo. RESULTS: Sonography in all cases showed hypoechoic and solid masses with increased posterior echo enhancement. There was an arrangement of hypoechoic small compartments demarcated by hyperechoic contour lines, which had a tortoiseshell pattern. This pattern was classified into 2 types according to its location: a lesion in the right or left side and lesions in both sides of the submandibular glands, found in 3 and 2 patients, respectively. The latter 2 cases had chromosomal aberrations of t(11;18)(q23;q23) and t(12;18)(q22;q21), respectively, and were revealed as secondary organ involvement. CONCLUSIONS: The sonographic appearance of MALT lymphoma in the submandibular glands was characterized by the tortoiseshell pattern in both primary and secondary lesions. Detection of this pattern in both sides of the submandibular glands can be an indicator of chromosomal aberrations and systematic involvement of the disease.

[Salivary gland mucoepidermoid carcinoma: unusual variants with detection of the t(11,19)(q21;p13) translocation]. / Mukoepidermoidkarzinom der Speicheldrüse: Ungewöhnliche Varianten mit Nachweis der Translokation t(11,19)(q21;p13).

Mucoepidermoid carcinoma (MEC) represents the most common malignant salivary gland tumour. Based on the proportion of their constituent cell types, MECs may display a wide morphological spectrum, thereby mimicking diverse types of other primary salivary gland carcinomas. The correct diagnosis relies on demonstration of classical MEC features, assisted by histochemical and immunohistochemical stains in equivocal cases. The current article aims to demonstrate, on the basis of own observations, how the detection of the MEC-typical t(11, 19) translocation may be of great value in difficult-to-classify cases. The differential diagnosis includes squamous cell carcinoma, clear cell carcinoma and oncocytic neoplasms, as well as highly malignant adenocarcinomas which cannot be further classified.

Graph models for phenotype and genotype association between oral mucosa and submandibular gland tumorigenesis in rat.

BACKGROUND: In recent years, success of statistics in field of genetics has been the identification of genes that affect the process of disease. Experimental models using animals enable early stages of tumor development to be studied. The aim of this study was to apply graph models to assess the association between the observed phenotypic changes in rat oral mucosa and induced tumorigenesis in the submandibular gland (SMG). MATERIALS AND METHODS: We studied changes in oncogenes TP53 and bcl-2, histopathological and immunomarker variables in samples of oral mucosa and SMG of Wistar male rats, 60 days old and 180 g in weight, in which tumorigenesis was induced in their SMG by a 0.5% solution of 9,10-dimethyl-1,2-benzanthracene in acetone. A set of linear structural equations were defined, with each formula indicating the response variables and the direct influences. In graph models, saliva was considered as a latent variable. The association was analyzed using Graphical Gaussian Markov models and odd ratios. RESULTS: About 40% of animals treated with 9, 10-dimethyl-1, 2-benzanthracene showed histological alterations in the epithelial basal strata of their oral mucosa only at 150 days. Statistical models indicated a relationship between gene alteration in gene bcl-2 in the SMG and histological changes observed in the oral mucosa (P = 0.04). CONCLUSION: Graph statistical model with one latent variable allows to conclude that these results associated with other clinical parameters may be useful in detecting early changes in SMG tumorigenesis. Furthermore, the design of randomized sampling of oral mucosa allows to validate these results and establish a reliable methodology for presumptive diagnosis or screening in the future.

Apoptotic effect of CKD-602 (Camtobell) on oral squamous cell carcinoma cell lines.

The purposes of this study were to measure the cytotoxic effect of CKD-602 on oral squamous cell carcinoma (OSCC) cell lines, to evaluate the apoptotic aspect of dead cells, and to identify the signaling molecules involved in apoptosis. The human OSCC cell lines A253, HSC-3 and KB were treated with CKD-602. The apoptotic proportion of the cells was analyzed using flow cytometry. The expression of Bax, Bcl-2, and p53 were detected by western blotting analysis. CKD-602 showed excellent cytotoxicity to the OSCC cell lines. Most cell death was attributed to apoptosis rather than necrosis. CKD-602 induced the down-regulation of Bcl-2 in A253 and HSC-3 cells, and p53 was expressed in the KB cell line after treatment with CKD-602.

Comprehensive loss of heterozygosity analysis and identification of a novel hotspot at 3p21 in salivary gland neoplasms.

OBJECTIVES: We sought to assess loss of heterozygosity (LOH) profiles of 3p, 6q, 8q, 10q, 12q, 13q, and 17p and to identify the tumor suppressor genes involved in salivary gland neoplasms. STUDY DESIGN: LOH analysis was performed using 26 microsatellite markers by polymerase chain reaction-polyacrylamide gel electrophoresis method in 20 benign and 6 malignant salivary gland tumors. RESULTS: Overall, LOH was detected in at least one informative locus in 18 of 20 (90%) of benign tumors and in all of 6 cases of malignant tumors. High LOH frequencies were revealed at the loci D3S1307 (22%, 3p26), D3S966 (41%, 3p21), D6S255 (27%, 6q25), D8S166 (25%, 8q12), D8S199 (21%, 8q24), and D10S1765 (28%, 10q23) in benign tumors, defining the hotspot regions for putative tumor suppressor genes. CONCLUSIONS AND SIGNIFICANCE: The hotspot regions defined by the present study suggest that new tumor suppressor genes related to the development of salivary gland tumors may reside at several chromosomal loci, including loci at 3p, 6q, 8q and 10q.

Early phenotypic and genotypic alterations in submandibular gland oncogenesis in rats.

The present study evaluates the phenotypic and genotypic changes that take place during early oncogenesis. The submandibular glands of male rats were injected with a 0.5% solution of 9,10-dimethyl-1,2-benzanthracene (DMBA) in acetone. Gland samples were taken at 0, 7, 30 and 150 days post-injection and submitted to histological, biochemical, immunocytochemical and PCR evaluation. Histopathological analysis was performed on hematoxylin-eosin stained slides. Total protein content was assessed by Lowry's method and the protein profile was analyzed by 12% SDS-PAGE. Bcl-2 was demonstrated by silver-enhanced gold immunolabeling. p53 immunolabeling was performed using the streptavidin-biotin system. All the treated animals developed carcinoma-like lesions at 30 and 150 days. Total protein concentration rose significantly (p < 0.05) above control values at 7, 30 and 150 days. The treated glands exhibited positive immunolabeling for p53 in the nuclei of neoplastic cells at 30 and 150 days. Treated glands also showed positive cytoplasmic immunolabeling for Bcl-2, exhibiting statistically significant differences between 7, 30 and 150 days (p = 0.0015), and with controls (p < 0.0001). No p53 mutations were observed whereas a point mutation, C-to-A, of the Bcl-2 gene was detected at 7, 30 and 150 days by PCR amplification. This mutation led to a single aminoacid change (thre --> asn) in the protein molecule. Our results suggest that the early histopathological changes correspond to quantitative and qualitative protein changes. The histopathological, biochemical, immunocytochemical and genetic alterations observed during the course of experimental carcinogenesis in the submandibular gland of the rat could constitute reproducible indices of malignant transformation applicable to human oncogenesis, given the high degree of homology between the oncogenes of mice, rats and human beings.