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BackgroundMammary phyllodes tumours (PT) are rare biphasic neoplasms but have important clinical significance. Both epithelial and stromal components participate in PT development. Despite a number of studies on stromal c-kit in PT, little is known about the role of its epithelial expression.ObjectiveTo further evaluate the stromal and epithelial expression of c-kit in a cohort of patients with PT.Method and resultsExpression of c-kit in both epithelial and stromal components was examined and correlated with histological features in PT. Stromal c-kit expression was associated positively with stromal cellularity (median expression=10.0, 30.0 and 50.0 from mild to severe cellularity; p=0.019). Conversely, a significant negative trend between epithelial c-kit expression with stromal pleomorphism (median expression=55.0, 30.0 and 2.5 from mild to severe pleomorphism; p=0.043) and mitosis (median expression=70.0 and 20.0 for low and high mitosis respectively; p=0.003); and a trend of negative correlation with increased PT grade was found. Despite these reverse associations, epithelial and stromal c-kit expressions were positively correlated with each other. Notably, the correlation of stromal c-kit expression with malignant histological features appeared to be stronger in cases with low epithelial c-kit expression but not in those with high epithelial c-kit expression.ConclusionsThis study demonstrated the association of epithelial c-kit expression with stromal histological features and stromal c-kit. Interestingly, epithelial c-kit expression affected the strength of the correlation of stromal c-kit with these histological features. These findings provide further evidence of the interaction between the epithelial and stromal components in PT.

As a distinct type of head and neck cancer, non-keratinizing nasopharyngeal carcinoma (NPC) is closely associated with EBV infection and massive lymphoid infiltration. The unique histological features suggest that local inflammation plays an important role in NPC tumourigenesis. We comprehensively characterized NF-κB signalling, a key inflammatory pathway which might contribute to the tumourigenesis of this EBV-associated cancer. By EMSA, western blotting, and immunohistochemical staining, constitutive activation of distinct NF-κB complexes, either p50/p50/Bcl3 or p50/RelB, was found in almost all EBV-positive NPC tumours. siRNA or chemical inhibition of NF-κB signalling significantly inhibited the growth of EBV-positive NPC cells C666-1. Gene expression profiling identified a number of NF-κB target genes involved in cell proliferation, apoptosis, immune response, and transcription. We further confirmed that p50 signals modulate the expression of multiple oncogenes (MYB, BCL2), chemokines, and chemokine receptors (CXCL9, CXCL10, CX3CL1, and CCL20). The findings support a crucial role of these constitutively activated NF-κB signals in NPC tumourigenesis and local inflammation. In addition to expression of the viral oncoprotein LMP1, genetic alteration of several NF-κB regulators (eg TRAF3, TRAF2, NFKBIA, A20) also contributes to the aberrant NF-κB activation in EBV-associated NPC. Except for LMP1-expressing C15 cells, all NPC tumour lines harbour at least one of these genetic alterations. Importantly, missense mutations of TRAF3, TRAF2, and A20 were also detected in 3/33 (9.1%) primary tumours. Taken together with the reported LTBR amplification in 7.3% of primary NPCs, genetic alterations in NF-κB pathways occurred in at least 16% of cases of this cancer. The findings indicate that distinct NF-κB signals are constitutively activated in EBV-positive NPC cells by either multiple genetic changes or EBV latent genes.

The association of Epstein–Barr virus (EBV) infection with the development of nasopharyngeal carcinoma (NPC) is well established. Latent membrane protein 1 (LMP1), the major oncogene encoded by EBV, is believed to play a crucial role in NPC pathogenesis by virtue of its ability to constitutively activate multiple cell signalling pathways. The LKB1–AMPK pathway is a master regulator of cellular metabolism that, via modulation of energy metabolism, has tumour suppressor activity. In this study we identify a novel ability of LMP1 to inhibit the LKB1–AMPK pathway through phosphorylation of LKB1 at serine 428 with subsequent suppression of the phosphorylation of AMPK and its substrates, ACC and Raptor. We show that MEK/ERK–MAPK signalling, activated by the CTAR1 domain of LMP1, is responsible for LKB1–AMPK inactivation. In addition, reactivation of AMPK signalling by AMPK activator, AICAR, abolished LMP1-induced cellular transformation (proliferation and anchorage-independent growth) in nasopharyngeal epithelial cells. Immunohistochemical staining revealed that a low level of phosphorylated AMPK is common in primary NPC specimens, and that this correlated significantly with the expression of LMP1. AICAR treatment inhibited the proliferation and anchorage-independent growth of NPC cells as well as potentiating the cytotoxic effect of the chemotherapeutic drug 5-fluorouracil. The current findings demonstrate that LMP1-mediated AMPK inactivation contributes to the proliferation and transformation of epithelial cells, thereby implicating the LKB1–AMPK pathway in the EBV-driven pathogenesis of NPC. Our findings also suggest that AMPK activators could be used to enhance the efficacy of conventional chemotherapeutic agents in the treatment of local and metastatic NPC. Copyright © 2013 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Aims MicroRNAs (miRs) have been shown to play important roles in tumour progression. Their expression pattern can be useful for cancer classification. However, little is known about miRs in mammary phyllodes tumours (PT). Methods and results In this study, polymerase chain reaction (PCR)-based miR profiling was performed in a small PT cohort to identify deregulated miRs in malignant PT. The purported roles and targets of these miRs were further validated. Unsupervised clustering of miR expression profiling segregated PT into different grades, implicating the miR profile in PT classification. Among the deregulated miRs, miR-21, miR-335 and miR-155 were validated to be higher in malignant than in lower-grade PT in the independent cohort by quantitative PCR (qPCR) (P ≤ 0.032). Their expression correlated with some of the malignant histological features, including high stromal cellularity, nuclear pleomorphism and mitosis. Subsequent analysis of their downstream proteins, namely PTEN for miR-21/miR-155 and Rb for miR-335, also showed an independent significant negative association between miR and protein expression. Conclusions Differential expression of miRs in PT could be useful in diagnosis and grading of PT. Their deregulated expression, together with the altered downstream targets, implicated their active involvement in PT malignant transformation.

As a distinct type of head and neck cancer, non-keratinizing nasopharyngeal carcinoma (NPC) is closely associated with EBV infection and massive lymphoid infiltration. The unique histological features suggest that local inflammation plays an important role in NPC tumourigenesis. We comprehensively characterized NF-κB signalling, a key inflammatory pathway which might contribute to the tumourigenesis of this EBV-associated cancer. By EMSA, western blotting, and immunohistochemical staining, constitutive activation of distinct NF-κB complexes, either p50/p50/Bcl3 or p50/RelB, was found in almost all EBV-positive NPC tumours. siRNA or chemical inhibition of NF-κB signalling significantly inhibited the growth of EBV-positive NPC cells C666-1. Gene expression profiling identified a number of NF-κB target genes involved in cell proliferation, apoptosis, immune response, and transcription. We further confirmed that p50 signals modulate the expression of multiple oncogenes (MYB, BCL2), chemokines, and chemokine receptors (CXCL9, CXCL10, CX3CL1, and CCL20). The findings support a crucial role of these constitutively activated NF-κB signals in NPC tumourigenesis and local inflammation. In addition to expression of the viral oncoprotein LMP1, genetic alteration of several NF-κB regulators (eg TRAF3, TRAF2, NFKBIA, A20) also contributes to the aberrant NF-κB activation in EBV-associated NPC. Except for LMP1-expressing C15 cells, all NPC tumour lines harbour at least one of these genetic alterations. Importantly, missense mutations of TRAF3, TRAF2, and A20 were also detected in 3/33 (9.1%) primary tumours. Taken together with the reported LTBR amplification in 7.3% of primary NPCs, genetic alterations in NF-κB pathways occurred in at least 16% of cases of this cancer. The findings indicate that distinct NF-κB signals are constitutively activated in EBV-positive NPC cells by either multiple genetic changes or EBV latent genes.

Aims Nerve growth factor receptor (NGFR) is a transmembrane receptor for the neurotrophin family. It acts either as tumour suppressor or oncogene depending on cellular context. Its role in breast cancers remained conflicting, possibly due to the heterogeneity of breast cancer subtypes. Methods In this study, we have analysed NGFR expression in 602 cases of breast cancers by immunohistochemistry. Its expression was correlated with biomarker expression and different breast cancer subtypes. Results NGFR expression was found to be positively correlated with basal markers, including Ki67, Cytokeratin (CK5/6), CK14, p63, c-kit and Epidermal growth factor receptor (EGFR) , but negatively with hormonal receptors. Among different molecular subtypes, it was negatively associated with luminal A, but positively with luminal B, and basal-like breast cancer BLBC subtypes. When comparing NGFR with other basal markers in BLBC, though less sensitive, its specificity was comparable to or better than other basal markers. For luminal B cancers, NGFR showed a high specificity which was also comparable to or better than the defining markers (estrogen receptor (ER), progesterone receptor (PR), Human epidermal growth receptor 2 (HER2) and Ki-67) for the subtype. Conclusions Overall, these findings suggested that NGFR expression could be indicative for the BLBCs or luminal B subtypes. It may represent a potential adjunct marker for these two subtypes.

The association of Epstein-Barr virus (EBV) infection with the development of nasopharyngeal carcinoma (NPC) is well established. Latent membrane protein 1 (LMP1), the major oncogene encoded by EBV, is believed to play a crucial role in NPC pathogenesis by virtue of its ability to constitutively activate multiple cell signalling pathways. The LKB1-AMPK pathway is a master regulator of cellular metabolism that, via modulation of energy metabolism, has tumour suppressor activity. In this study we identify a novel ability of LMP1 to inhibit the LKB1-AMPK pathway through phosphorylation of LKB1 at serine 428 with subsequent suppression of the phosphorylation of AMPK and its substrates, ACC and Raptor. We show that MEK/ERK-MAPK signalling, activated by the CTAR1 domain of LMP1, is responsible for LKB1-AMPK inactivation. In addition, reactivation of AMPK signalling by AMPK activator, AICAR, abolished LMP1-induced cellular transformation (proliferation and anchorage-independent growth) in nasopharyngeal epithelial cells. Immunohistochemical staining revealed that a low level of phosphorylated AMPK is common in primary NPC specimens, and that this correlated significantly with the expression of LMP1. AICAR treatment inhibited the proliferation and anchorage-independent growth of NPC cells as well as potentiating the cytotoxic effect of the chemotherapeutic drug 5-fluorouracil. The current findings demonstrate that LMP1-mediated AMPK inactivation contributes to the proliferation and transformation of epithelial cells, thereby implicating the LKB1-AMPK pathway in the EBV-driven pathogenesis of NPC. Our findings also suggest that AMPK activators could be used to enhance the efficacy of conventional chemotherapeutic agents in the treatment of local and metastatic NPC.

Nasopharyngeal carcinoma (NPC) is a unique EBV-associated epithelial malignancy, showing highly invasive and metastatic phenotype. Despite increasing evidence demonstrating the critical role of cancer stem-like cells (CSCs) in the maintenance and progression of tumors in a variety of malignancies, the existence and properties of CSC in EBV-associated NPC are largely unknown. Our study aims to elucidate the presence and role of CSCs in the pathogenesis of this malignant disease. Sphere-forming cells were isolated from an EBV-positive NPC cell line C666-1 and its tumor-initiating properties were confirmed by in vitro and in vivo assays. In these spheroids, up-regulation of multiple stem cell markers were found. By flow cytometry, we demonstrated that both CD44 and SOX2 were overexpressed in a majority of sphere-forming C666-1 cells. The CD44+SOX2+ cells was detected in a minor population in EBV-positive xenografts and primary tumors and considered as potential CSC in NPC. Notably, the isolated CD44+ NPC cells were resistant to chemotherapeutic agents and with higher spheroid formation efficiency, showing CSC properties. On the other hand, microarray analysis has revealed a number of differentially expressed genes involved in transcription regulation (e.g. FOXN4, GLI1), immune response (CCR7, IL8) and transmembrane transport (e.g. ABCC3, ABCC11) in the spheroids. Among these genes, increased expression of CCR7 in CD44+ CSCs was confirmed in NPC xenografts and primary tumors. Importantly, blocking of CCR7 abolished the sphere-forming ability of C666-1 in vitro. Expression of CCR7 was associated with recurrent disease and distant metastasis. The current study defined the specific properties of a CSC subpopulation in EBV-associated NPC. Our findings provided new insights into developing effective therapies targeting on CSCs, thereby potentiating treatment efficacy for NPC patients.

Although recent molecular investigations have identified a number of genetic alterations that are associated with the development of pituitary adenomas, the exact pathogenesis mechanism of these tumors remains largely unknown. In this study, we used a genome-wide survey to detect specific genetic changes within the genome of pituitary adenomas. A series of 10 growth hormone-secreting adenomas were analyzed for their genetic imbalances on all 22 autosomes by comparative genomic hybridization (CGH). Chromosomal imbalances were detected in 8 GH-secreting adenomas, whereas 2 tumors had no detectable genetic abnormalities. Chromosome gains were more frequent than losses. Overrepresentation of whole or parts of chromosomes were detected in 5/10 (50%) in 19, 3/10 (30%) in each of 5, 9, and 22q, 2/10 (20%) in 17p12-q21, whereas DNA loss were 3/10 (30%) in 13q and 2/10 (20%) in 18. No detectable gain or loss of genetic material was observed in chromosomes 7, 8, 10, 12, 15, and 20. The findings of overrepresentation of chromosomes 5q, 9p, 17q and DNA loss of chromosome 18 were consistent with those detected in nonfunctioning adenomas (Daniely M, Aviram A, Adams EF, et al:J Clin Endocrinol Metab 83:1801-1805, 1998) suggesting that the development of pituitary tumors, at least in somatotroph and nonfunctioning adenomas, may share common pathway. Frequent amplifications in chromosomes 19 and 22q imply that candidate genes residing in these chromosomal regions may be involved in the pathogenesis of GH-secreting adenomas.