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PDF file, 2741KB, YM155 treatment induces survivin migration to nucleus in a time-dependent manner. CAL27-CisR cells were treated with YM155 (10 nM) for different time intervals and survvin localization was examined by staining these cells with survivin (red), β-catenin (green, membrane staining), and DAPI (blue, nucleus).

PDF file, 34KB, Patient tumor characteristics (Group 1).

Cisplatin is one of the commonly used chemotherapeutic drugs for the treatment of head and neck squamous cell carcinoma (HNSCC). However, acquisition of cisplatin resistance is common in patients with HNSCC, and it often leads to local and distant failure. In this study, we showed that survivin expression is significantly upregulated in HNSCC primary tumors and cell lines. In addition, survivin levels were significantly higher in human papilloma virus–negative patients that normally respond poorly to cisplatin treatment. Survivin expression was further increased in cisplatin-resistant cells (CAL27-CisR) as compared with its parent cells (CAL27). Therefore, we hypothesized that targeting of survivin in HNSCC could reverse the resistant phenotype in tumor cells, thereby enhancing the therapeutic efficacy of cisplatin. We used both in vitro and in vivo models to test the efficacy of YM155, a small molecule survivin inhibitor, either as a single agent or in combination with cisplatin. YM155 significantly decreased survivin levels and cell proliferation in a dose-dependent manner. In addition, YM155 pretreatment significantly reversed cisplatin resistance in cancer cells. Interestingly, YM155 treatment altered the dynamic localization of survivin in cells by inducing a rapid reduction in cytoplasmic survivin, which plays a critical role in its antiapoptotic function. In a severe combined immunodeficient mouse xenograft model, YM155 significantly enhanced the antitumor and antiangiogenic effects of cisplatin, with no added systemic toxicity. Taken together, our results suggest a potentially novel strategy to use YM155 to overcome the resistance in tumor cells, thereby enhancing the effectiveness of the chemotherapy in HNSCC. Mol Cancer Ther; 11(9); 1988–98. ©2012 AACR.

Optimally effective antitumor therapies would not only activate immune effector cells but also engage them at the tumor. Folate conjugated to immunoglobulin (F-IgG) could direct innate immune cells with Fc receptors to folate receptor–expressing cancer cells. F-IgG bound to human KB and HeLa cells, as well as murine L1210JF, a folate receptor (FR)–overexpressing cancer cell line, as determined by flow cytometry. Recognition of F-IgG by natural killer (NK) cell Fc receptors led to phosphorylation of the ERK transcription factor and increased NK cell expression of CD69. Lysis of KB tumor cells by NK cells increased by about 5-fold after treatment with F-IgG, an effect synergistically enhanced by treatment with IL2, IL12, IL15, or IL21 (P < 0.001). F-IgG also enhanced the lysis of chronic lymphocytic leukemia cells by autologous NK cells. NK cells significantly increased production of IFNγ, MIP-1α, and RANTES in response to F-IgG–coated KB target cells in the presence of the NK cell–activating cytokine IL12, and these coculture supernatants induced significant T-cell chemotaxis (P < 0.001). F-IgG-coated targets also stimulated FcR-mediated monocyte effector functions. Studies in a murine leukemia model confirmed the intratumoral localization and antitumor activity of F-IgG, as well as enhancement of its effects by IL12 (P = 0.05). The antitumor effect of this combination was dependent on NK cells and led to decreased tumor cell proliferation in vivo. Thus, F-IgG can induce an immune response against FR-positive tumor cells that is mediated by NK cells and can be augmented by cytokine therapy. Cancer Immunol Res; 4(4); 323–36. ©2016 AACR.

Supplementary Figure Legends

PDF file, 1581KB, HNSCC cell line characteristics.

Supplemental Figure 1.F‑IgG and IL-12 mediated NK cell lysis is dependent on the folate receptor availability and does not correlate with FcR genotype. Supplemental Figure 2. MDSC inhibit NK cell lytic activity and IFN-y production. Supplemental Figure 3. CIgG-FITC and FIgG-FITC treated mouse tissues exhibit normal histological features.

PDF file, 828KB, YM155 treatment does not significantly affect mice body weight. Mice weights were measured on days 9, 18, 27 and 36.

PDF file, 2045KB, Patient tumor characteristics (Group 2).

PDF file, 995KB, YM155 inhibits tumor cell proliferation in a dose-dependent manner. Cell proliferation in tumor cells (UM-SCC-74A or CAL27-CisR) or human oral keratinocytes (HOK) was assessed by MTT assay after 72 hrs.

PDF file, 30KB.

PDF file, 5124KB, YM155 and cisplatin combination treatment significantly inhibits tumor cell motility. Tumor cell motility was examined by scratch assay.

PDF file, 839KB, YM155 and cisplatin combination treatment significantly inhibits tumor cell proliferation. UM-SCC-74A cells were treated with YM155 or cisplatin (CDDP) alone or in combination. After 72 hrs, cell proliferation was assessed by MTT assay.

The tumor immune microenvironment of oral tongue squamous cell carcinoma may be accountable for differences in clinical behavior, particularly between different age groups. We performed RNA expression profiling and evaluated tumor infiltrating lymphocytes (TILs) and their T-cell subsets in order to assess the functional status of oral tongue squamous cell carcinoma tumor microenvironment and detect potentially clinically useful associations. Archival surgical pathology material from sixteen oral tongue squamous cell carcinoma patients was microscopically evaluated for TIL densities. RNA was extracted from macrodissected whole tumor sections and normal controls and RNA expression profiling was performed by the NanoString PanCancer IO 360 Gene Expression Panel. Immunostains for CD4, CD8 and FOXP3 were evaluated manually and by digital image analysis. Oral tongue squamous cell carcinomas had increased TIL densities, numerically dominated by CD4 + T cells, followed by CD8 + and FOXP3 + T cells. RNA expression profiling of tumors versus normal controls showed tumor signature upregulation in inhibitory immune signaling (CTLA4, TIGIT and PD-L2), followed by inhibitory tumor mechanisms (IDO1, TGF-β, B7-H3 and PD-L1). Patients older than 44 years showed a tumor microenvironment with increased Tregs and CTLA4 expression. Immunohistochemically assessed CD8% correlated well with molecular signatures related to CD8 + cytotoxic T-cell functions. FOXP3% correlated significantly with CTLA4 upregulation. CTLA4 molecular signature could be predicted by FOXP3% assessed by immunohistochemistry (R(2) = 0.619, p = 0.026). Oral tongue squamous cell carcinoma hosts a complex inhibitory immune microenvironment, partially reflected in immunohistochemically quantified CD8 + and FOXP3 + T-cell subsets. Immunohistochemistry can be a useful screening tool for detecting tumors with upregulated expression of the targetable molecule CTLA4. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (10.1007/s12105-020-01229-w) contains supplementary material, which is available to authorized users.

Objectives: The aim of the study was to investigate the association of surgical margin conditions, including positive specimen margins revised to negative relative to local recurrence, disease-free survival, and overall survival (OS) within a cohort of HPV-mediated oropharyngeal squamous cell carcinoma (OPSCC) who underwent en bloc resection via transoral robotic surgery (TORS). Materials and Methods: Retrospective cohort of patients with untreated HPV-mediated OPSCC cT1 or T2 undergoing TORS resection between October 2014 and March 2020. The methodologic description of our interdisciplinary institutional approach, number of cut-through margins (CTMs) during intraoperative consultation, percentage of final positive margin cases, and disease-free survival and OS stratified by margin status and margin tumor-free distance is identified. Results: 135 patients with primary cT1/T2 HPV-mediated OPSCC met inclusion criteria. Twenty-eight of 135 (20.7%) specimens revealed CTM and were revised during the same operative setting. Three of 135 (2.2%) surgical cases had positive final margin status. Local control rate was 97%. On univariate analysis, margin distance did not impact OS. CTM and final positive margins had lower OS than initially negative margins (p = 0.044). Pathologic N-stage significantly impacted OS (p < 0.001). Conclusions: High local control rate and low final positive margin status confound the study of specimen margin-based techniques in HPV-mediated OPSCC resected en bloc with TORS. Pathologic N-stage may impact OS more than margin status. Larger numbers are needed to confirm differences.

Background Lymphoid enhancer binding factor 1 (LEF-1) has recently been reported as a potential immunohistochemical (IHC) marker for basal cell adenoma (BCA) and other salivary gland tumors, which may contribute to an increased accuracy in differentiating basaloid salivary gland neoplasms. We evaluated the utility of LEF-1 in fine needle aspiration (FNA) and resection specimens to distinguish pleomorphic adenoma (PA), BCA, basal cell adenocarcinoma (BCAC), and adenoid cystic carcinoma (ACC) as well as in non-neoplastic salivary gland (NNSG). Methods Cases including 66 PA (35 FNA, 31 resections), 12 BCA (5 FNA, 7 resections), 42 ACC (11 FNA, 31 resections), 1 BCAC FNA, and 10 NNSG (5 FNA, 5 resections) were obtained and stained for LEF-1. Results On cell block (CB), 51% of PA and 60% of BCA were LEF-1 positive while 91% of ACC were LEF-1 negative. Among resections, there was a higher percentage of LEF-1 positive PA (84%) and BCA (86%), and a higher percentage of LEF-1 negative ACC (97%). LEF-1 staining had a low to moderate sensitivity for detecting benign basaloid neoplasms on FNA CB and resection specimens (52.5% and 84%, respectively), but a higher specificity (92% and 97% respectively), and positive predictive value (95% and 97% respectively). Conclusion When comparing benign (PA and BCA) and the most common malignant basaloid salivary gland tumor (ACC), positive LEF-1 favors a benign neoplasm. Additional studies with LEF-1, specifically including other rare basaloid salivary gland neoplasms are needed to further clarify the role of LEF-1 in diagnosing these lesions on FNA.

BACKGROUND Salivary gland aspiration cytology is useful in the preoperative management of patients but remains challenging, because of the extensive morphologic overlap of some tumors limits the ability to always determine the presence of malignancy. In response to this challenge, there has been increasing drive to develop a risk-based categorization scheme for salivary gland aspirates. Herein, the authors examine the interobserver variability of 1 such pattern and risk-based system. METHODS Select smears and cell-block sections of 50 salivary gland aspirates from 2 large academic centers were digitally imaged. These scanned slides were independently and blindly reviewed by 4 cytopathologists, and each aspirate was assigned to 1 of the proposed pattern-based categories if it was considered neoplastic by the observer. Interobserver agreement was scored and aggregated risks of malignancy were calculated for cases with available surgical follow-up. RESULTS In total, 42 samples (84%) were considered neoplastic by at least 2 observers and were scored for interobserver agreement: 10 of 42 (23.8%) had uniform agreement, 14 of 42 (33.3%) had majority agreement, and 5 of 42 (11.9%) had divided agreement. Only 9 of 42 samples (21.4%) had minimal agreement, and 4 of 42 (9.5%) had no agreement. Condensation of similar categories was able to improve interobserver agreement and still maintain stratified risk of malignancy. CONCLUSIONS The proposed pattern-based risk-stratification scheme, which could be implemented with the forthcoming Milan System, has good overall interobserver agreement and successfully stratifies the risk of malignancy. Some simplification is possible to make the system easier to use and improve interobserver agreement while maintaining stratification of risk. Cancer Cytopathol 2017. © 2017 American Cancer Society.

Fine needle aspiration of salivary gland tumors is a common preoperative triage as it is useful in determining which patients should undergo surgical resection and in guiding the extent of surgery in those cases deemed appropriate for resection. While a specific diagnosis can be achieved on a cytologic specimen in many cases, there is also a considerable amount of morphologic diversity that prevents such confident preoperative classification and in these cases it can be a challenge to confidently determine if a tumor is benign or malignant. Recently, a pattern based risk stratification approach was proposed for salivary gland cytology in which basaloid neoplasms are separated by stromal characteristics and oncocytoid neoplasms are separated primarily by background material such as mucus. In addition to potentially providing a stratification in risk of malignancy for salivary gland tumors, this approach is also useful to narrow differential diagnostic considerations and guide ancillary testing. In this review we use this proposed pattern based approach as a framework to discuss immunostains and fluorescence in situ hybridization studies which we find useful in our practice.

Accurate diagnosis of salivary gland tumors can be challenging because of the many diagnostic entities, the sometimes extensive morphologic overlap, and the rarity of most tumor types. Ancillary testing is beginning to ameliorate some of these challenges through access to newer immunohistochemical stains and fluorescence in situ hybridization probes, which can limit differential diagnostic considerations in some cases. These ancillary testing strategies are especially useful in small biopsy samples, including aspiration cytology. Molecular techniques are also expanding our understanding of salivary gland tumor pathology and are helping to identify potential targets that may improve treatment for some of these tumors. Here, we summarize the clinical use of new immunohistochemical markers in our practice and review the current understanding of chromosomal rearrangements in salivary gland tumor pathology, emphasizing the prospects for exploiting molecular alterations in salivary gland tumors for diagnosis and targeted therapy. We find that immunohistochemistry and fluorescence in situ hybridization are powerful tools toward the diagnosis of salivary gland tumors, especially when used in a systematic manner based on morphologic differential-diagnostic considerations. As new targeted therapies emerge, it will become increasingly vital to incorporate appropriate molecular testing into the pathologic evaluation of salivary gland cancers.

// Bhavna Kumar 1, 2 , Arti Yadav 2 , Nicole V. Brown 3 , Songzhu Zhao 3 , Michael J. Cipolla 1 , Paul E. Wakely 4 , Alessandra C. Schmitt 4, 6 , Robert A. Baiocchi 5 , Theodoros N. Teknos 1, 2 , Matthew Old 1, 2, * , Pawan Kumar 1, 2, * 1 Department of Otolaryngology-Head and Neck Surgery, The Ohio State University, Columbus, OH 43210 USA 2 The Ohio State University Comprehensive Cancer Center, Columbus, OH 43210 USA 3 Center for Biostatistics, The Ohio State University, Columbus, OH 43210 USA 4 Department of Pathology, The Ohio State University, Columbus, OH 43210 USA 5 Department of Internal Medicine, The Ohio State University, Columbus, OH 43210 USA 6 Department of Pathology and Laboratory Medicine, Emory University School of Medicine, Atlanta, GA 30303 USA * These authors contributed equally to this work Correspondence to: Pawan Kumar, email: Pawan.Kumar@osumc.edu Keywords: PRMT5, OPSCC, HPV, cyclin D1, IL-6 Received: July 22, 2016 Accepted: December 27, 2016 Published: January 17, 2017 ABSTRACT Protein arginine methyltransferase-5 (PRMT5) plays an important role in cancer progression by repressing the expression of key tumor suppressor genes via the methylation of transcriptional factors and chromatin-associated proteins. However, very little is known about the expression and biological role of PRMT5 in head and neck cancer. In this study, we examined expression profile of PRMT5 at subcellular levels in oropharyngeal squamous cell carcinoma (OPSCC) and assessed its correlation with disease progression and patient outcome. Our results show that nuclear PRMT5 was associated with poor overall survival ( p 10 pack-years ( p = 0.013). In addition, nuclear PRMT5 was directly correlated with cyclin D1 ( p = 0.0101) and IL-6 expression ( p < 0.001). In a subgroup survival analysis, nuclear PRMT5-positive/IL-6-positive group had worst survival, whereas nuclear PRMT5-negative/IL-6-negative group had the best survival. Similarly, patients with p16-negative/nuclear PRMT5-positive tumors had worse survival compared to patients with p16-positive/nuclear PRMT5-negative tumors. Our mechanistic results suggest that IL-6 promotes nuclear translocation of PRMT5. Taken together, our results demonstrate for the first time that nuclear PRMT5 expression is associated with poor clinical outcome in OPSCC patients and IL-6 plays a role in the nuclear translocation of PRMT5.

Optimally effective antitumor therapies would not only activate immune effector cells but also engage them at the tumor. Folate conjugated to immunoglobulin (F-IgG) could direct innate immune cells with Fc receptors to folate receptor–expressing cancer cells. F-IgG bound to human KB and HeLa cells, as well as murine L1210JF, a folate receptor (FR)–overexpressing cancer cell line, as determined by flow cytometry. Recognition of F-IgG by natural killer (NK) cell Fc receptors led to phosphorylation of the ERK transcription factor and increased NK cell expression of CD69. Lysis of KB tumor cells by NK cells increased by about 5-fold after treatment with F-IgG, an effect synergistically enhanced by treatment with IL2, IL12, IL15, or IL21 (P < 0.001). F-IgG also enhanced the lysis of chronic lymphocytic leukemia cells by autologous NK cells. NK cells significantly increased production of IFNγ, MIP-1α, and RANTES in response to F-IgG–coated KB target cells in the presence of the NK cell–activating cytokine IL12, and these coculture supernatants induced significant T-cell chemotaxis (P < 0.001). F-IgG-coated targets also stimulated FcR-mediated monocyte effector functions. Studies in a murine leukemia model confirmed the intratumoral localization and antitumor activity of F-IgG, as well as enhancement of its effects by IL12 (P = 0.05). The antitumor effect of this combination was dependent on NK cells and led to decreased tumor cell proliferation in vivo. Thus, F-IgG can induce an immune response against FR-positive tumor cells that is mediated by NK cells and can be augmented by cytokine therapy. Cancer Immunol Res; 4(4); 323–36. ©2016 AACR.

Introduction Lymphoepithelial carcinoma of the salivary gland is an extremely rare neoplasm and is challenging to diagnose by fine needle aspiration (FNA). There are rare reports on the cytopathologic features of lymphoepithelial carcinoma, which may be mistaken for other high-grade salivary gland neoplasm or a metastasis to the salivary gland. Materials and methods A retrospective review was undertaken of 7 cases of lymphoepithelial carcinoma of the parotid diagnosed on FNA with histologic confirmation from 4 major medical centers. Results Cytomorphologic features of lymphoepithelial carcinoma include smears with moderate cellularity displaying a rich nonneoplastic population of lymphoplasmacytic cells admixed with tissue fragments of high grade, malignant undifferentiated epithelial cells with high nuclear to cytoplasm ratio, hyperchromasia, prominent nucleoli, and scant to abundant, indistinct cytoplasm. Discussion Diagnostic pitfalls of lymphoepithelial carcinoma include metastatic squamous cell carcinoma, metastatic nasopharyngeal carcinoma, and other high grade primary salivary gland neoplasms. Recognizing this lesion may help guide clinicians to perform additional imaging studies to exclude a primary from other sites.

// Bhavna Kumar 1, 2 , Nicole V. Brown 3 , Benjamin J. Swanson 4 , Alessandra C. Schmitt 4, 5 , Matthew Old 1, 2 , Enver Ozer 1, 2 , Amit Agrawal 1, 2 , David E. Schuller 1, 2 , Theodoros N. Teknos 1, 2 , Pawan Kumar 1, 2 1 Department of Otolaryngology-Head and Neck Surgery, The Ohio State University, Columbus, OH 43210, USA 2 Comprehensive Cancer Center, The Ohio State University, Columbus, OH 43210, USA 3 Center for Biostatistics, The Ohio State University, Columbus, OH 43210, USA 4 Department of Pathology, The Ohio State University, Columbus, OH 43210, USA 5 Current affiliation: Department of Pathology and Laboratory Medicine, Emory University School of Medicine, Atlanta, GA 30303, USA Correspondence to: Pawan Kumar, e-mail: Pawan.Kumar@osumc.edu Keywords: myoferlin, OPSCC, HPV, IL-6, nanog Received: December 22, 2015 Accepted: February 11, 2016 Published: February 23, 2016 ABSTRACT Myoferlin (MYOF) is a member of ferlin family of membrane proteins that was originally discovered as a muscle specific protein. Recent studies have shown that myoferlin is also expressed in other cell types including endothelial cells and cancer cells. However, very little is known about the expression and biological role of myoferlin in head and neck cancer. In this study, we examined expression profile of myoferlin in oropharyngeal squamous cell carcinoma (OPSCC) and assessed its correlation with disease progression and patient outcome. In univariate analyses, nuclear MYOF was associated with poor overall survival (p

Noninvasive follicular thyroid neoplasm with papillary-like nuclear features (NIFTP) was recently proposed as a designation for a subset of follicular variant papillary thyroid carcinoma (FVPTC). Encapsulated FVPTC has been shown to be a fairly indolent tumor, and NIFTP are expected to represent the most indolent subset of these tumors. Many of the exclusion criteria for NIFTP related to architecture and a lack of psammoma bodies are designed to preclude the inclusion of more aggressive non-FVPTC tumors in this indolent group and also exclude the diagnosis of FVPTC. In addition to strict application of histologic features to ensure that NIFTP represents a subset of encapsulated FVPTC without invasion, other exclusion criteria including high mitotic activity and necrosis may also lead to a lack of one-to-one correlation between the diagnosis of NIFTP and encapsulated FVPTC without invasion. In this series, 50 cases previously diagnosed as FVPTC over a 2-year period from a large academic center are retrospectively reviewed for reclassification as NIFTP. Additionally, cases not meeting criteria for NIFTP are more accurately classified using the most up to date WHO criteria. Prior BRAF V600E mutation testing was examined for these tumors when available. Seventeen of 50 (34%) tumors met criteria for classification as NIFTP and, 17 (34%) were classified as encapsulated FVPTC with invasion. Strict application of architectural features led to classification of 12 (24%) tumors as non-FVPTC with a variety of more aggressive designations. Tumors classified as NIFTP and encapsulated FVPTC with invasion lacked lymph node metastases (0/4; 0/7, respectively) and BRAF mutations (0/12; 0/13, respectively). In contrast, infiltrative FVPTC, encapsulated PTC with or without invasion, and conventional PTC showed more aggressive features with lymph node metastases and BRAF V600E mutations. One case not meeting criteria for NIFTP maintained the diagnosis of encapsulated FVPTC without invasion but demonstrated significant mitotic activity (three mitoses/ten HPF) and lacked lymph node metastases and BRAF V600E mutation. These findings demonstrate the importance of using strict criteria, especially the lack of true papillary architecture, for the diagnosis of NIFTP and encapsulated FVPTC to ensure that only truly indolent tumors will be included in these diagnoses and to allow tumors with potential for more aggressive behavior to be appropriately treated.

Objectives: We evaluated SOX10 (SRY-related HMG-box 10) in differentiating acinic cell carcinoma (AciCC) from other salivary gland neoplasms with oncocytic features on fine-needle aspiration cell blocks (FNA CB) and compared its performance to DOG1 (discovered on gastrointestinal stromal tumor 1). Material and Methods: 35 FNA CB of oncocytic salivary gland neoplasms, i.e. 13 cases of AciCC, 16 of Warthin tumor (WT), 3 of mucoepidermoid carcinoma (MEC) and 3 of oncocytoma (ONC), and 75 salivary gland resections, i.e. 26 AciCC, 7 WT, 36 MEC, 3 ONC, 2 mammary analog secretory carcinomas (MASC) and 1 papillary cystadenoma were stained for SOX10 and DOG1. Results: None of the benign oncocytic neoplasms were immunoreactive for SOX10 on CB or resection, similar to DOG1. On CB, 61.5 and 77% of AciCC were positive for SOX10 and DOG1, respectively. All surgically resected AciCC cases were positive for SOX10 and DOG1; other malignant oncocytic lesions such as MEC and MASC demonstrated variable SOX10 and DOG1 staining. Conclusion: The use of SOX10 may increase the diagnostic accuracy of oncocytic lesions on FNA. In this context, SOX10 is equivalent to DOG1 in ruling out benign lesions such as WT and ONC; however, negative results for SOX10 as well as DOG1 do not favor a benign diagnosis since MEC is often negative for both markers.

Fine needle aspiration of salivary gland tumors is a common preoperative triage as it is useful in determining which patients should undergo surgical resection and in guiding the extent of surgery in those cases deemed appropriate for resection. While a specific diagnosis can be achieved on a cytologic specimen in many cases, there is also a considerable amount of morphologic diversity that prevents such confident preoperative classification and in these cases it can be a challenge to confidently determine if a tumor is benign or malignant. Recently, a pattern based risk stratification approach was proposed for salivary gland cytology in which basaloid neoplasms are separated by stromal characteristics and oncocytoid neoplasms are separated primarily by background material such as mucus. In addition to potentially providing a stratification in risk of malignancy for salivary gland tumors, this approach is also useful to narrow differential diagnostic considerations and guide ancillary testing. In this review we use this proposed pattern based approach as a framework to discuss immunostains and fluorescence in situ hybridization studies which we find useful in our practice.

BACKGROUND Human papillomavirus 16 (HPV16) is a major risk factor for the development of head and neck squamous cell carcinoma (HNSCC), particularly the development of oropharyngeal squamous cell carcinoma (OPSCC). Cancer stem cells (CSCs) are resistant to conventional therapies, and it is postulated that they are responsible for disease recurrence and/or progression. Because the prognoses of patients with HPV16-positive and HPV-negative HNSCC are distinct, the authors sought to determine whether differences in the number of CSCs could account for this clinical observation. METHODS CSC populations in HPV16-positive and HPV-negative HNSCC were assessed using a proprietary assay based on expression of the enzyme aldehyde dehydrogenase (ALDH), an in vitro tumorsphere formation assay, and an in vivo limiting cell dilution in nonobese diabetic/severe combined immunodeficiency mice. A high-density tissue microarray was stained with ALDH1, a CSC marker, to determine the association between CSCs and HPV16-positive/HPV-negative OPSCC. RESULTS HPV16-positive HNSCC had a greater intrinsic CSC pool than HPV-negative HNSCC. Inactivation of p53 has been identified as a major mechanism for the elevated CSC population in HPV16-positive HNSCC. In vivo limiting cell dilution experiments using tumors from patients with HPV16-positive and HPV-negative OPSCC indicated that the CSC frequency was 62.5-fold greater in an HPV16-positive OPSCC tumor than in an HPV-negative OPSCC tumor. Primary tumors from patients with HPV16-positive OPSCC were associated with elevated tumor ALDH1 staining, further extending the association between HPV16 and CSCs. CONCLUSIONS The current data and the clinical observation that patients with HPV16-positive HNSCC respond more favorably to current treatment paradigms than patients with HPV-negative HNSCC support the suggestion that CSC phenotype is not homogeneous. Therefore, the reliance on the CSC number may be insufficient to accurately assess the potential of a particular tumor for disease recurrence and/or progression. Cancer 2014;120:992–1001. © 2014 American Cancer Society.

Herein we describe the development and implementation of a nanoporous cell-therapy device with controllable biodegradation. Dopamine-secreting PC12 cells were housed within newly formulated alginate-glutamine degradable polylysine (A-GD-PLL) microcapsules. The A-GD-PLL microcapsules provided a 3-D microenvironment for good spatial cell growth, viability and proliferation. The microcapsules were subsequently placed within a poly(ethylene glycol) (PEG)-coated poly(ε-caprolactone) (PCL) chamber covered with a PEG-grafted PCL nanoporous membrane formed by phase inversion. To enhance PC12 cell growth and to assist in controlled degradation of both the PC12 cells and the device construct, small PCL capsules containing neural growth factor (PCL-NGF) and a poly(lactic-co-glycolic acid) pellet containing glutamine (PLGA-GLN) were also placed within the PCL chamber. Release of NGF from the PCL-NGF capsules facilitated cell proliferation and viability, while the controlled release of GLN from the PLGA-GLN pellet resulted in A-GD-PLL microcapsule degradation and eventual PC12 cell death following a pre-specified period of time (4 weeks in this study). In vivo, our device was found to be well tolerated and we successfully demonstrated the controlled release of dopamine over a period of four weeks. This integrated biodegradable device holds great promise for the future treatment of a variety of diseases.

Cisplatin is one of the commonly used chemotherapeutic drugs for the treatment of head and neck squamous cell carcinoma (HNSCC). However, acquisition of cisplatin resistance is common in patients with HNSCC, and it often leads to local and distant failure. In this study, we showed that survivin expression is significantly upregulated in HNSCC primary tumors and cell lines. In addition, survivin levels were significantly higher in human papilloma virus–negative patients that normally respond poorly to cisplatin treatment. Survivin expression was further increased in cisplatin-resistant cells (CAL27-CisR) as compared with its parent cells (CAL27). Therefore, we hypothesized that targeting of survivin in HNSCC could reverse the resistant phenotype in tumor cells, thereby enhancing the therapeutic efficacy of cisplatin. We used both in vitro and in vivo models to test the efficacy of YM155, a small molecule survivin inhibitor, either as a single agent or in combination with cisplatin. YM155 significantly decreased survivin levels and cell proliferation in a dose-dependent manner. In addition, YM155 pretreatment significantly reversed cisplatin resistance in cancer cells. Interestingly, YM155 treatment altered the dynamic localization of survivin in cells by inducing a rapid reduction in cytoplasmic survivin, which plays a critical role in its antiapoptotic function. In a severe combined immunodeficient mouse xenograft model, YM155 significantly enhanced the antitumor and antiangiogenic effects of cisplatin, with no added systemic toxicity. Taken together, our results suggest a potentially novel strategy to use YM155 to overcome the resistance in tumor cells, thereby enhancing the effectiveness of the chemotherapy in HNSCC. Mol Cancer Ther; 11(9); 1988–98. ©2012 AACR.

Glomus tumors are uncommon soft tissue tumors that usually occur in the hands or feet but rarely have been described to appear in the tracheobronchial tree. Tracheal glomus tumors present with symptoms including cough, dyspnea, and wheezing that may be mistaken for other pulmonary disorders. Imaging and pulmonary function testing can detect tracheal obstruction, but pathology is necessary to differentiate glomus tumors from other airway tumors. On pathology, glomus tumors are made up of glomus cells, blood vessels, and smooth muscle and are classified based on the predominant cell type. The differential for this tumor includes carcinoid tumors, paragangliomas, and hemangiomas, and immunohistochemical stains can be used to obtain the correct diagnosis. The most common modality for treatment of these tracheal tumors has been surgical resection. However, there have been reported cases of successful removal with rigid or flexible bronchoscopy. We present a case of a tracheal glomus tumor that was successfully resected using electrocautery snare during flexible bronchoscopy. Our case adds to the evidence that flexible bronchoscopy is a safe, less invasive approach to management of tracheal glomus tumors in select patients.

In the original publication, the author name Kimberly Point du Jour was incorrectly captured. The correct Given name should be Kimberly and the Family name should be Point du Jour. The correct author name is presented in this correction article.

Gastrointestinal stromal tumors (GIST) usually occur within the bowel wall, and most contain KIT - or PDGFRA - activating mutations. The routine diagnosis of GIST relies on C-kit immunohistochemical detection; however, up to 15% of GISTs are C-kit negative. Antibodies with increased sensitivity and specificity in the detection of C-kit–negative GIST cases could be of value because these cases could also benefit from imatinib mesylate therapy. We investigated two DOG-1 antibody clones, SP31 and K9, to determine which would be more useful in the diagnosis of GIST. Immunohistochemistry was performed on 40 GISTs, five adenoid cystic carcinomas, five mastocytomas, and five seminomas. Two DOG-1 antibody clones (SP31 and K9), two C-kit antibody clones (polyclonal C-kit and monoclonal C-kit), and monoclonal protein kinase C theta (PKCθ) antibody were analyzed. Both DOG-1 antibodies were found to have a high sensitivity and specificity in diagnoses of GIST. They were also both more sensitive and more spec...

BACKGROUND: Paired box gene 8 (PAX8), a member of the thyroid-specific transcription factors, has recently been investigated as a useful marker of thyroid epithelial neoplasms. To the authors' knowledge, its utility in the immunohistochemistry of papillary thyroid carcinoma (PTC) has not been well studied to date. The goal of the current study was to compare the immunohistochemical expression of PAX8 with HBME-1 and cytokeratin 19 (CK19) expression on cell block preparations of PTC and benign thyroid nodules (BTN). METHODS: Thirty-two cases of PTC and 20 cases of BTN diagnosed on fine-needle aspiration were included. The cell blocks were immunohistochemically stained for PAX8, HBME-1, and CK19. RESULTS: PAX8 demonstrated nuclear positivity in >90% of cases with PTC (31 of 32 cases; 96.9%) and BTN (20 of 20 cases; 100%), with a high sensitivity (0.97), but low specificity (0). HBME-1 and CK19 were found to have high sensitivity (0.78 and 0.75, respectively) and specificity (0.95 for both) for PTC. Statistically significant differences in staining intensity and distribution were noted for the PAX8 antibody (negative [1 of 32 cases] to weakly positive [20 of 32 cases] in 65.6% of PAX8-positive PTCs, and intermediate to strong in 100% of BTNs). CONCLUSIONS: In contrast to PAX8, HBME-1 and CK19 were found to have a high degree of both sensitivity and specificity for PTC. The results of the current study demonstrate that strong PAX8 staining appears to be typical of BTNs, whereas negative to weakly positive staining is noted in PTC. PAX8, HMBE-1 and CK19 may all play a role, particularly as a panel; however, the interpretation of PAX8 should consider the intensity and distribution of staining. Cancer (Cancer Cytopathol) 2010. © 2010 American Cancer Society.

Background: The Wnt/β-catenin signaling pathway has been noted to be upregulated in head and neck cancers, including oropharyngeal squamous cell carcinoma (OSCC). This study compared the efficacy of β-catenin immunohistochemistry (IHC), p16 IHC and automated human papillomavirus (HPV) in situ hybridization (ISH) in OSCC. Methods: Sixty-eight OSCCs (48 surgical specimens and 20 fine-needle aspirations) were evaluated. Nuclear staining only of β-catenin was assessed as 0-3+ intensity (relative to controls of benign squamous mucosa). p16 was interpreted as positive if 70% of tumor cells showed brown nuclear and cytoplasmic staining. HPV ISH was interpreted as positive if a minimum of one tumor cell showed brown punctate dot-like nuclear positivity. p16 IHC and HPV ISH were then correlated with β-catenin staining. HPV ISH was used as the gold standard. Results: Twenty-five of 48 surgical specimens (52.1%) and 11 of 20 cell blocks (55%) stained positively for β-catenin, making a total of 36 of 68 (52.9%) staining positively for β-catenin, as compared to 61.7% positive for p16 IHC and 70.6% positive by automated HPV ISH, the gold standard method for OSCC diagnosis. χ2 analysis revealed no significant correlation between β-catenin and HPV ISH (p > 0.05) and demonstrated a strong correlation between p16 and HPV ISH (p < 0.05). Conclusion: β-Catenin IHC is not a sensitive or specific marker of HPV and is unlikely to be a useful adjunct to p16 IHC or HPV ISH in the setting of advanced OSCC. However, as this study focused on samples of advanced OSCC, β-catenin IHC may still find some use in the diagnosis of early-stage OSCC.

Introduction DOG1 is a calcium-activated chloride channel protein that may have a potential role in secretory cells of salivary glands and tumors derived from them. Its role in cytologic specimens is not well documented. This study was performed to evaluate its utility in separating acinic cell carcinoma (AciCC) from other closely related differential diagnoses on cytologic samples. In addition, an immunohistochemical panel consisting of DOG1, p63, and S100 protein to assist in the subclassification of these salivary gland neoplasms with oncocytic differentiation was also investigated. Materials and methods Thirty-one fine-needle aspiration cell blocks (CBs) of oncocytic salivary gland neoplasms (16 Warthin tumors [WTs], 10 AciCCs, 3 mucoepidermoid carcinomas [MECs], and 2 oncocytomas [ONCs]), and 75 salivary gland resections (7 WTs, 27 AciCCs, 36 MECs, 2 high-grade adenocarcinomas, 2 ONCs, 1 papillary cystadenoma) were immunostained for DOG1, p63, and S100. Results DOG1 and p63 were very useful in distinguishing AciCC from WT on CB, because 100% of WTs were DOG1-negative and 87.5% were p63-positive, whereas 70% of AciCCs were DOG1-positive and 100% were p63-negative. The resection results correlated with those on CBs: 100% of WTs were DOG1-negative and 86% were p63-positive, whereas 93% of AciCCs were DOG1-positive and 89% were p63-negative. S100 and DOG1 were negative in both WTs and ONCs, with Conclusions DOG1 was very helpful in separating AciCC from WT, MEC, and ONC. In summary, an immunohistochemical panel including DOG1, p63, and S100 can significantly improve the accuracy of diagnosing oncocytic salivary gland neoplasms on CBs.

The broad morphologic spectrum, inherent immunophenotypic heterogeneity of malignant melanoma and its rarity in the sinonasal tract are major challenges in eliciting the correct diagnosis, which may lead to misclassification and inadequate medical management. Herein, we describe a single case of a 70 year-old male with sinonasal mucosal melanoma, exhibiting varying histologic phenotypes including small round blue cell morphology, epithelioid and focal rhabdoid morphology and strong, diffuse desmin immunoreactivity. These constellation of features initially prompted the diagnosis of rhabdomyosarcoma. The differential diagnosis in this anatomic area includes other malignant small round blue cell tumors of the sinonasal mucosa such as rhabdomyosarcoma, olfactory neuroblastoma, sinonasal undifferentiated carcinoma, and lymphoma. We reviewed precedent literature and further discuss the potential pitfalls to which pathologists may be prone.

SOX10 is a transcription factor expressed from the developmental stage to adulthood in acinar and intercalated ducts of major salivary glands, and in some salivary gland tumors. Its role in cytologic specimens has been rarely documented. This study was performed to evaluate its utility in the diagnosis of salivary gland oncocytic tumors on fine …

Introduction: Malignant Mixed Mullerian Tumor/Carcinosarcoma (MMMT) is a clinically aggressive endometrial malignancy composed of mixture of carcinomatous and sarcomatous elements usually affecting postmenopausal women. As this tumor is relatively rare and morphologically diverse, the findings on cervicovaginal cytology, specifically liquid based “SurePath” cytology samples, are not well described. Here we describe the cytological features and correlation with histopathological and clinical findings of patients with MMMT. Material and Methods: A retrospective search (2001-2012) of patients with MMMT was performed. Available PAP smears, biopsies and surgical excisions were reviewed. Tumor characteristics including tumor size presence of heterologous elements, cervical involvement, lympho-vascular invasion (LVI) and tumor pathology staging were extracted from pathology reports and analyzed. Results: There were a total of 30 MMMT patients with available PAPs preceding surgery. All except two patients (28/30) were African American women with average age of 63.03. Ten patients (33.3%) had negative PAPs, eleven (36.7%) were diagnosed as malignant (carcinoma or adenocarcinoma) and nine (30%) as atypical (AGUS, ASCUS or squamous dysplasia). Patients with positive PAP smears had larger tumors and in almost all (90.9%) of the cases tumor showed cervical stromal involvement, and LVI (90.9%). Group with atypical pap smear result did not show correlation with any tumor characteristics or tumor stage. Negative results on PAP smears were seen more commonly in patients without cervical involvements (80%). Common cytology features were tumor diathesis, glandular atypia, squamous dysplasia, presence of nucleoli, multinuclear and anaplastic cells, while sarcomatous elements were rarely encountered. Conclusions: Two thirds of patients with MMMT had either positive or atypical PAPs prior to biopsy or surgery. Patients with cervical involvement of MMMT were more likely to have positive PAP smear independent of tumor staging.

Summary Background Human papillomavirus (HPV) is a cause of oropharyngeal cancer, but a role for HPV in the etiology of oral cavity squamous cell carcinomas (OCSCC) remains uncertain. Methods We sought to estimate the etiologic fraction for HPV among consecutive, incident OCSCC diagnosed from 2005 to 2011 at four North American hospitals. DNA and RNA purified from paraffin-embedded tumors were considered evaluable if positive for DNA and mRNA control genes by quantitative PCR. Fifteen high-risk (HR) HPV types were detected in tumors by consensus PCR followed by type-specific HR-HPV E6/7 oncogene expression by quantitative reverse-transcriptase PCR. P16 expression was evaluated by immunohistochemistry (IHC). A study of 400 cases allowed for precision to estimate an etiologic fraction of as low as 0% (97.5% confidence interval, 0–0.92%). Results Of 409 evaluable OCSCC, 24 (5.9%, 95%CI 3.6–8.2) were HR-HPV E6/7 expression positive; 3.7% (95%CI 1.8–5.5) for HPV16 and 2.2% (95%CI 0.8–3.6) for other HR-HPV types. HPV-positive tumors arose from throughout the oral cavity (floor of mouth [ n = 9], anterior tongue [6], alveolar process [4], hard palate [3], gingiva [1] and lip [1]) and were significantly associated with male gender, small tumor stage, poor tumor differentiation, and basaloid histopathology. P16 IHC had very good-to-excellent sensitivity (79.2%, 95%CI 57.9–92.9), specificity (93.0%, 95%CI 90.0–95.3), and negative-predictive value (98.6%, 95%CI 96.8–99.6), but poor positive-predictive value (41.3%, 95%CI 27.0–56.8) for HR-HPV E6/7 expression in OCSCC. Conclusion The etiologic fraction for HR-HPV in OCSCC was 5.9%. p16 IHC had poor positive predictive value for detection of HPV in these cancers.