C(3)1-TAg in C57BL/6 J background as a model to study mammary tumor development.

Diagnosis and prognosis of breast cancer is based on disease staging identified through histopathological and molecular biology techniques. Animal models are used to gain mechanistic insights into the development of breast cancer. C(3)1-TAg is a genetically engineered mouse model that develops mammary cancer. However, carcinogenesis caused by this transgene was characterized in the Friend Virus B (FVB) background. As most genetic studies are done in mice with C57BL/6 J background, we aimed to define the histological alterations in C3(1)-TAg C57BL/6 J animals. Our results showed that C3(1)-TAg animals with C57BL/6 J background develop solid-basaloid adenoid cystic carcinomas with increased fibrosis, decreased area of adipocytes, and a high proliferative index, which are triple-negative for progesterone, estrogen, and human epidermal growth factor receptor 2 (HER2) receptors. Our results also revealed that tumor development is slower in the C57BL/6 J background when compared with the FVB strain, providing a better model to study the different stages in breast cancer progression.

Phase 1 safety and pharmacodynamic study of lenalidomide combined with everolimus in patients with advanced solid malignancies with efficacy signal in adenoid cystic carcinoma.

BACKGROUND: Purpose: The combination of a mammalian target of rapamycin inhibitor and lenalidomide showed enhanced preclinical cytotoxicity. We conducted a phase 1 study in advanced solid tumour patients to assess safety, efficacy and pharmacodynamic (PD) outcomes. METHODS: We employed a 3+3 dose escalation design to establish the safety and recommended phase 2 doses (RP2D) of daily everolimus and lenalidomide in patients with advanced solid tumours. The starting doses were 5 and 10 mg, respectively, with planned escalation to maximum single-agent doses of 10 and 25 mg in the absence of dose-limiting toxicity. PD endpoints of lymphocyte subsets and immune cytokines were assessed in peripheral blood using multiparameter flow cytometry and LUMINEX assay. Efficacy was evaluated by cross-sectional imaging after every two cycles of treatment. RESULTS: The study enrolled 44 patients, median age of 58 years and 28 males (63.6%). The RP2D was established as 10 and 25 mg daily continuously for everolimus and lenalidomide. Common (>5%) grade ≥3 adverse events included rash (19%), neutropenia (19%), hypokalaemia (11%) and fatigue (9%). Best efficacy outcomes in 36 evaluable patients were partial response in 5 (13.8%), stable disease in 24 (55.8%) and progressive disease in 7 (19.4%) patients. PD assessment revealed significant association of cytokine levels (interleukin-2 (IL2), IL21 and IL17), baseline activated and total CD8+ lymphocytes and change in B cell lymphocytes and activated NK cells with clinical benefit. CONCLUSIONS: The study demonstrated the safety of everolimus and lenalidomide with promising efficacy signal in thyroid and adenoid cystic cancers. CLINICAL TRIAL REGISTRATION: NCT01218555.

Treatment-induced changes of lymphocyte subsets in patients with adenoid cystic carcinoma of the head and neck.

PURPOSE: Adenoid cystic carcinoma (ACC) of the head and neck is a rare and highly malignant tumor, characterized by perineural growth and early distant metastases. The composition of immune cells in the peripheral blood and the tumor microenvironment is critical to tumor growth and control. However, little is known about the frequency and function of the relevant immune cell subsets in this entity. METHODS: In ACC patients (n = 11) and matched healthy donors (n = 11), the frequency of peripheral blood T and B cells was measured by flow cytometry at different treatment stages of disease (24 samples). Cells were further characterized by their expression of CCR7, PD-1, CD39 and CD73. Tumor-infiltrating lymphocytes (TIL) were analyzed by immunohistochemistry for ten patients and for three patients by flow cytometry. RESULTS: CD4+ T cells had significantly lower frequency after radiotherapy (RT). All other cell frequencies, including Treg, were stable through course of the disease. In B cells, CD73 was reduced after RT. CCR7 expression on T and B cells in patients with relapse/metastases (R/M) differed significantly from patients with active disease. PD-1 remained stable. Treg were more present in TIL compared to peripheral blood. CONCLUSION: Composition of lymphocyte subgroups behaves similar to squamous cell carcinoma in the head and neck, except for Treg, which remained stable. Nevertheless, the CD4+/Treg ratio was lower after RT, which could stand for an immunosuppressive effect in these patients. Therefore, it could be beneficial treating ACC with combined RT and immunomodulatory drugs.

Immunoexpression pattern of TLR3 and TLR7 in minor salivary gland adenoid cystic carcinoma and its role in prognosis.

OBJECTIVES: Adenoid cystic carcinoma (ACC) of the salivary glands has poor long-term prognosis and a high metastatic rate. Toll-like receptors (TLRs), first-line immune activators, have been associated with both tumor progression and suppression. We aimed to study TLR3 and TLR7 behavior in ACC. MATERIALS AND METHODS: We studied TLR3 and TLR7 immunoexpression of 46 minor salivary gland ACCs diagnosed at the Department of Otorhinolaryngology - Head and Neck Surgery, Helsinki University Hospital, Helsinki, Finland over the period 1974-2012. The associations of TLR3 and TLR7 immunoexpression with clinicopathological factors were evaluated by χ2-test and Fisher's exact test. RESULTS: In the majority of samples, both TLR3 and TLR7 were immunoexpressed in cytoplasm. The immunoexpression was heterogeneous between individual tumors. Stronger TLR7 immunoexpression associated with recurrence rate and poorer disease-specific survival (DSS). TLR3 did not associate significantly with survival although we found an inverse correlation between TLR3 and TLR7 immunopositivity. Hence, when TLR3 immunoexpression was negative or mild, TLR7 immunoexpression was moderate to strong, and vice versa. CONCLUSIONS: TLR3 and TLR7 are immunoexpressed in minor salivary gland ACC. TLR7 is potentially an independent prognostic marker for recurrence rate and DSS.

Molecular characterization of the salivary adenoid cystic carcinoma immune landscape by anatomic subsites.

Adenoid cystic carcinoma (AdCC) is a slow-growing salivary gland malignancy that relapses frequently. AdCCs of the submandibular gland exhibit unique differences in prognosis and treatment response to adjuvant radiotherapy compared to other sites, yet the role of tumor anatomic subsite on gene expression and tumor immune microenvironment (TIME) composition remains unclear. We used 87 samples, including 48 samples (27 AdCC and 21 normal salivary gland tissue samples) from 4 publicly available AdCC RNA sequencing datasets, a validation set of 33 minor gland AdCCs, and 39 samples from an in-house cohort (30 AdCC and 9 normal salivary gland samples). RNA sequencing data were used for single sample gene set enrichment analysis and TIME deconvolution. Quantitative PCR and multiplex immunofluorescence were performed on the in-house cohort. Wilcoxon rank-sum, nonparametric equality-of-medians tests and linear regression models were used to evaluate tumor subsite differences. AdCCs of different anatomic subsites including parotid, submandibular, sublingual, and minor salivary glands differed with respect to expression of several key tumorigenic pathways. Among the three major salivary glands, the reactive oxygen species (ROS)/nuclear factor erythroid 2-related factor 2 (NRF2) pathway signature was significantly underexpressed in AdCC of submandibular compared to parotid and sublingual glands while this association was not observed among normal glands. Additionally, the NRF2 pathway, whose expression was associated with favorable overall survival, was overexpressed in AdCCs of parotid gland compared to minor and submandibular glands. The TIME deconvolution identified differences in CD4+ T cell populations between AdCC of major and minor glands and natural killer (NK) cells among AdCC of minor, submandibular, and parotid glands while plasma cells were enriched in normal submandibular glands compared to other normal gland controls. Our data reveal key molecular differences in AdCC of different anatomic subsites. The ROS and NRF2 pathways are underexpressed in submandibular and minor AdCCs compared to parotid gland AdCCs, and NRF2 pathway expression is associated with favorable overall survival. The CD4+ T, NK, and plasma cell populations also vary by tumor subsites, suggesting that the observed submandibular AdCC tumor-intrinsic pathway differences may be responsible for influencing the TIME composition and survival differences.

Molecular Profiling and the Impact of Treatment on Outcomes in Adenoid Cystic Carcinoma Type I and II.

PURPOSE: Adenoid cystic carcinoma (ACC) is an uncommon salivary gland cancer with no approved therapies available to treat advanced, incurable disease. Recent molecular profiling efforts have identified two important subtypes: the more aggressive ACC-I is characterized by Notch pathway alterations and MYC amplification whereas ACC-II demonstrates a more indolent phenotype and TP63 overexpression. EXPERIMENTAL DESIGN: This retrospective observational cohort study involved de-identified samples from 438 patients with ACC with tumor samples sent for commercially-available molecular profiling (Caris Life Sciences). Next-generation whole-exome and whole-transcriptomic sequencing was performed on primary and metastatic samples. Immunostaining for PD-L1 and RNA deconvolution (quanTIseq) was used to explore the tumor immune microenvironment (TME). Real-world clinical and survival outcome metrics were extracted from insurance claims data. RESULTS: MYC expression was 1.61-fold higher (39.8 vs. 24.7; P < 0.0001) among NOTCH1-mutant ACC-I tumors, whereas MYB/L1 fusion rates were similar among ACC-I/II. The median B-cell fraction in the TME was higher among ACC-II (7.1% vs. 5.8%; P < 0.01), although infiltrating T cells subsets were low among either ACC subgroup (both <1%). When pooling systemic treatment categories, ACC-I patients had worse outcomes with available therapies (HR, 3.06; 95% confidence interval, 1.65-5.68; P < 0.01), with no significant difference in overall survival between ACC-I/II based on chemotherapy or VEGFR tyrosine kinase inhibitor exposure in smaller subsets. CONCLUSIONS: We confirmed the previously reported associations with MYC and TP63 in the prognostically relevant subgroups of ACC-I and -II, respectively, and report immunologic differences among these subtypes. Survival outcomes are comparatively worse in ACC-I regardless of treatment type.

The Immune Landscape of Chinese Head and Neck Adenoid Cystic Carcinoma and Clinical Implication.

Novel systemic agents and effective treatment strategies for recurrence adenoid cystic carcinoma (ACC) of the head and neck are still worthy of further exploration. Here, we analyzed the mutations and expression profiles of 75 Chinese ACC patients, characterized the prognostic value of the immune signature for recurrence or distant metastasis, and explored the potential of immunotherapeutic biomarkers in ACC. In general, MYB fusion and somatic mutations accounted for a high proportion, which was 46.7% (35/75). ACCs displayed an overall low mutation burden and lack of programmed cell death ligand-1 (PD-L1) expression. The antigen-presenting machinery (APM) expression score and immune infiltration score (IIS) were the lowest among ACC patients, compared with other cancer types. For 61 primary cases, the locoregional recurrence-free survival (LRRFS) was statistically significantly correlated with the IIS [univariate analysis; hazard ratio (HR) = 0.32; 95% CI, 0.11-0.92; p = 0.035] and T-cell infiltration score (TIS) (univariate analysis; HR = 0.33; 95% CI, 0.12-0.94; p = 0.037]. Patients with lower IIS (log-rank p = 0.0079) or TIS (log-rank p = 0.0079) had shorter LRRFS. Additionally, solid pattern was also a prognostic factor related to locoregional recurrence, whereas postoperative radiotherapy (PORT) exerted its beneficial effects. We further evaluated the pretreatment immune profile of five ACC patients treated with PD-1 inhibitors. Patients who responded to camrelizumab or pembrolizumab observed elevated APM and TIS, compared with patients with progressive disease. Our study highlights the immune infiltration pattern and messenger RNA (mRNA) signatures of Chinese ACC patients, which has the potential value for prognosis and immunotherapy.

Immunophenotypic aspects of cylindroma and nodular hidradenoma.

BACKGROUND: Some adnexal tumours have many controversies about their histogenesis. OBJECTIVES: To evaluate the eccrine and/or apocrine differentiation phenotype in cases of cylindroma and clear cell hidradenoma with CD15 and p63 antibodies. METHODOLOGY: Slides and blocks of six cases of cylindroma and seven cases of nodular hidradenoma (clear cells) were analyzed by the technique of immunohistochemistry with CD15 and p63 antibodies. RESULTS: In all cases of cylindroma we obtained negative results for CD15 antibody and positive for p63 antibody. In five of seven cases of nodular hidradenoma (clear cell), we could easily observe clear cells between 20% and 50% of tumour cells. In the two other cases, cystic lesions were present and occasional clear cells could be seen. The reaction with CD15 antibody was positive in granular and cytoplasmic pattern in six of seven cases, especially in cells with suggestive clear cytoplasm in lower proportion than this clear cells could be seen in haematoxylin and eosin. The positivity for p63 antibody, nuclear pattern, was observed in six of seven cases, in the major part of tumour cells. In only one case, the positivity was in 20% of cells. Limitation Samples are in small number because these are relatively rare tumours. CONCLUSIONS: The present study suggests eccrine origin for both tumours: cylindroma and clear cell hidradenoma.

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.

Immunophenotypical alterations with impact on the epithelial-mesenchymal transition (EMT) process in salivary gland adenoid cystic carcinomas.

Adenoid cystic carcinoma (ACC) is one of the most common malignant salivary glands neoplasms with an indolent clinical course, slow-growing but locally aggressive and quite often with delayed recurrence and distant metastasis. In order to elucidate this tumoral behavior, we conducted an immunohistochemical study investigating the alterations of epithelial phenotype with anti-cytokeratin (CK) AE1∕AE3 and anti-E-cadherin antibodies, and the acquisition of mesenchymal phenotype with vimentin, fibronectin, N-cadherin and P-cadherin in salivary ACCs. Thus, we recorded a reduction of CK AE1∕AE3, E-cadherin, P-cadherin and fibronectin reactivity in the solid variant and especially in the cells from the periphery of invasive neoplastic proliferations, regardless histological type. These phenotypical alterations suggest the involvement of the epithelial-mesenchymal transition (EMT) process in the progression of salivary ACCs.

Morphometric analysis of CD34-positive vessels in salivary gland adenoid cystic and mucoepidermoid carcinomas.

BACKGROUND: Carcinomas of the salivary glands are uncommon and morphologically a diverse group of malignancies. To evaluate the prognostic value of CD34 immunostaining of the vessels in adenoid cystic carcinoma (AdCC) and mucoepidermoid carcinoma (MEC), an automated image analysis method was used. METHOD: In a nationwide study, covering salivary gland cancer (SGC) patients in Finland 1991-1996, 37 AdCC and 18 MEC patients (M 25, F 30, age 25-90, mean 63) were included. In addition to clinical characteristics the size, shape, staining intensity and vessel density in CD34 immunostained histologic samples were measured. RESULTS: Altogether 4433 vessels were measured from AdCC and 2615 from MEC tumor. Of the total tumor vessels measured, 2651 were from patients who deceased with disease (Group I) and 4397 were from specimens derived from those who did not die of disease (Group II) during the 10-year follow-up. The staining intensity was significantly higher in MEC than in AdCC tumor (P = 0.0005). In MEC, the Group I patients had a higher staining intensity among high-grade patients compared with patients with low grade disease, whereas the tumors in Group II had a lower staining intensity among the high-grade compared with the low grade tumors (P = 0.018). A higher vessel density was found in patients with MEC in group II compared with group I (P = 0.017). CONCLUSIONS: The staining intensity of CD34 positive vessels in MEC was higher than in AdCC. In MEC, higher staining intensity of vessels in high-grade tumors and lower vessel density in all MEC patients, predicted poor survival.

Utility of p63 and p40 in Distinguishing Polymorphous Adenocarcinoma and Adenoid Cystic Carcinoma.

OBJECTIVE: Adenoid cystic carcinoma and polymorphous adenocarcinoma are primarily the tumor of minor salivary glands. Both show certain morphological similarities, which limit their proper diagnosis in settings where there are obscuring factors and limited biopsy material. However, there is a considerable difference in treatment and prognosis, which raises the need to distinguish these two entities. In this study, we discuss the utility of two immunohistochemical stains, p63 and p40, in different combinations for distinguishing polymorphous adenocarcinoma from adenoid cystic carcinoma. MATERIALS AND METHODS: Two immunohistochemical stains, p63 and p40, were performed on 47 cases of adenoid cystic carcinoma and 23 cases of polymorphous adenocarcinoma. RESULTS: 36 out of 47 cases of adenoid cystic carcinoma showed p63+ve/p40+ve immunoprofile, followed by p63-ve/p40-ve immunoprofile, which is seen in10 cases of adenoid cystic carcinoma. However, 22 out of 23 cases of polymorphous adenocarcinoma displayed p63+ve/ p40-ve immunoprofile. p63-ve/p40+ve is the least frequent observed immunoprofile, which is seen in only one case of adenoid cystic carcinoma. CONCLUSION: On combining all possible immunoprofile combinations, p63+ve/p40-ve immunoprofile appears to be the most sensitive profile for distinguishing polymorphous adenocarcinoma from adenoid cystic carcinoma.

Immune microenvironment and evasion mechanisms in adenoid cystic carcinomas of salivary glands.

OBJECTIVES: The objective of the present study was to investigate the expression of immune checkpoints (PD-L1, PD-L2, PD-1 and CTLA-4), immune inhibitory molecule HLA-G, markers of tumor-infiltrating lymphocytes (TIL) and dendritic cells (DC), as well as its association with clinicopathological features of adenoid cystic carcinomas (ACC) of the salivary glands. MATERIALS AND METHODS: Thirty-six samples from patients with ACC were analyzed immunohistochemically for the expression of PD-L1, PD-L2, PD-1, CTLA-4, HLA-G, CD8, GrB, CD1a and CD83. Positivity of HLA-G, PD-L1 and PD-L2 expression was defined by cut-offs values. CD8+ TIL was measured semiquantitatively and also using cut-off values obtained by the ROC curve considering recurrence of the lesion. RESULTS: ACC showed low CD8+, GrB+  TIL, CD1a and CD83 populations, as well as scarce positivity for CTLA-4 and PD-1. In contrast, PD-L2 and HLA-G expression was increased, while no PD-L1 expression was detected. Interestingly, cases with lower CD8+ TIL density presented greater recurrence rates. CONCLUSION: Our findings suggest that the ACC microenvironment exhibits low immunogenicity, represented by low TIL and DC density. Moreover, there seems to be activation of the immune inhibitory proteins/PD-L2 and HLA-G, a scenario that may favor tumor escape from the immune system and partially explain the poor prognosis of ACC.

Relations among expression of CXCR4, histological patterns, and metastatic potential in adenoid cystic carcinoma of the head and neck.

Adenoid cystic carcinoma (ACC) may acquire a chemokine-mediated mechanism during the process of metastasis. To investigate the involvement of chemokines in metastasis from ACC, expression of CXCR4 in surgical specimens of ACC and two tumor lines transplantable to nude mice was examined immunohistochemically. In addition, the expression levels of CXCR4 protein and mRNA were examined by Western blotting and reverse-transcription polymerase chain reaction. Our results showed that patients whose tumors expressed high levels of CXCR4 had metastases to the regional lymph nodes and the lung, resulting in poor outcomes. ACCs showing a solid or cribriform pattern with distant metastasis were strongly positive for CXCR4, while those showing a tubular or cribriform pattern without metastasis were weakly positive for CXCR4. In the in vivo model, ACCY tumor showed increasing expression levels of CXCR4 with tumor growth, and the histological pattern changed from cribriform to solid. The histological pattern of ACCI, associated with spontaneous metastasis to the neck, changed from cribriform to undifferentiated carcinoma and was highly metastatic to the lung. This tumor showed high levels of CXCR4 protein and mRNA. These results suggest that CXCR4 expression, histological patterns, and metastatic potential are closely related in ACC.

Assessment of CTNNB1 gene mutations and ß-catenin immunoexpression in salivary gland pleomorphic adenomas and adenoid cystic carcinomas.

ß-Catenin exerts multiple functions in several neoplasms, playing a major role in cell signaling and tumor progression. This study analyzed possible CTNNB1 mutations in salivary gland pleomorphic adenomas (PAs) and adenoid cystic carcinomas (ACCs), and determined possible differences in ß-catenin immunoexpression in relation to these mutations, as well as histopathological aspects of these tumors. Twenty-four PAs (15 cell-rich and 9 cell-poor tumors) and 24 ACCs (10 tubular, 8 cribriform, and 6 solid tumors) were selected for the analysis of ß-catenin distribution and cellular localization. Furthermore, ß-catenin expression was evaluated using the H-score scoring system. Mutations in CTNNB1 exon 3 were investigated by the single-strand conformational polymorphism test. Diffuse ß-catenin expression was more frequently observed in ACCs compared to PAs (P = 0.008). No significant difference in ß-catenin cellular localization was observed between these tumors (P = 0.098). Comparisons between PA and ACC cases revealed a higher median H-score in the latter (P = 0.036). Cell-rich PAs exhibited a trend for higher H-score than cell-poor tumors (P = 0.060), whereas lower H-scores were observed in cribriform ACCs when compared to tubular and solid ACCs (P = 0.042). Mutations in CTNNB1 were observed in 6 PAs and 7 ACCs, with no significant difference in H-scores for ß-catenin according to mutation status (P = 0.135). ß-Catenin is important in the pathogenesis of salivary gland PAs and ACCs. In addition, CTNNB1 exon 3 mutations do not seem to significantly influence ß-catenin cytoplasmic/membranous expression or nuclear translocation in these tumors.

Autoimmune Granulomatous Inflammation of Lacrimal Glands and Axonal Neuritis Following Treatment With Ipilimumab and Radiation Therapy.

Immune checkpoint inhibitors such as anti-CTLA-4 (cytotoxic T-lymphocyte-associated protein 4), anti PD-1 (programmed cell death protein 1) and PD-L1 (programmed cell death protein-ligand 1) monoclonal antibodies are emerging as standard oncology treatments in various tumor types. The indications will expand as immunotherapies are being investigated in various tumors with promising results. Currently, there is inadequate identification of predictive biomarkers of response or toxicity. Unique response patterns include pseudoprogression and delayed response. The use of immune checkpoint inhibitors exhibit an unique toxicity profile, the immune-related adverse events (irAEs). The most notable immune reactions are noted in skin (rash), gastrointestinal track (colitis, hepatitis, pancreatitis), lung (pneumonitis), heart (myocarditis), and endocrine system (thyroiditis, hypophysitis). We present a patient with metastatic adenoid cystic carcinoma of the left submandibular gland with granulomatous inflammation of the lacrimal glands and axonal neuritis of the cervical and paraspinal nerves following treatment with ipilimumab and radiation therapy.

Zebrafish blastomere screen identifies retinoic acid suppression of MYB in adenoid cystic carcinoma.

Pluripotent cells have been used to probe developmental pathways that are involved in genetic diseases and oncogenic events. To find new therapies that would target MYB-driven tumors, we developed a pluripotent zebrafish blastomere culture system. We performed a chemical genetic screen and identified retinoic acid agonists as suppressors of c-myb expression. Retinoic acid treatment also decreased c-myb gene expression in human leukemia cells. Translocations that drive overexpression of the oncogenic transcription factor MYB are molecular hallmarks of adenoid cystic carcinoma (ACC), a malignant salivary gland tumor with no effective therapy. Retinoic acid agonists inhibited tumor growth in vivo in ACC patient-derived xenograft models and decreased MYB binding at translocated enhancers, thereby potentially diminishing the MYB positive feedback loop driving ACC. Our findings establish the zebrafish pluripotent cell culture system as a method to identify modulators of tumor formation, particularly establishing retinoic acid as a potential new effective therapy for ACC.

Primary intraosseous adenoid cystic carcinoma of the mandible: a comprehensive review and analysis of four new cases with emphasis on morphologic, immunophenotypic, and molecular characteristics.

OBJECTIVE: To comprehensively review the clinical manifestations, imaging, diagnosis, treatment, and pathologic features of primary intraosseous adenoid cystic carcinoma (PIACC) of the mandible and analyze PIACC histopathology and molecular features in four cases. STUDY DESIGN: We reviewed the literature and retrospectively studied four cases of PIACC. RESULTS: The clinical and imaging findings of PIACC are similar to other malignant or borderline-malignant mandible tumors. The four cases of PIACC included three males and one female (aged between 41 and 58 years). The histopathologic features of the tumors resembled those of ACC. We observed abundant osteoclasts resorbing bone at the leading edge of the tumors characterized by solid structure histology but not by the cribriform subtype. Additionally, all four cases showed abnormalities in the MYB gene and high expression of MYB protein. All patients survived for the duration of follow-up, and two patients had distant metastases (followed up for 3 to 36 months). CONCLUSIONS: PIACC is extremely rare and is often clinically misdiagnosed. Different histologic subtypes could show different mechanisms of invasion of the mandible. MYB gene and protein expression abnormalities can be used as indicators for the precise diagnosis of PIACC.

Immune Profiling of Adenoid Cystic Carcinoma: PD-L2 Expression and Associations with Tumor-Infiltrating Lymphocytes.

Adenoid cystic carcinoma (ACC) is among the most lethal salivary gland tumors, with no treatments for metastatic disease that prolong survival. We examined tissue from 28 primary and metastatic ACC deposits obtained from 21 patients for infiltrating immune cells and PD-L1/PD-L2 expression and determined mRNA profiles of over 1,400 oncogenic and immune-related genes. We also assessed the effect of chemoradiation on immune mediators in a patient who had serial biopsies available. Most tumors expressed PD-L2 but had few infiltrating immune cells. Lack of immune-cell infiltrate was associated with expression of genes in the ß-catenin/Wnt and PI3K pathways. Additionally, certain transcripts linked to growth and invasion were differentially expressed among primary and metastatic deposits. Chemoradiation appeared to increase CD8(+) effector T cells, decrease regulatory T cells, and promote a systemic humoral response. These data suggest a potential role for PD-L2 inhibition and immune modulation as treatment for patients with ACC. Cancer Immunol Res; 4(8); 679-87. ©2016 AACR.

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.