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Background Ten–eleven translocation methylcytosine dioxygenase 1 (TET1) is involved in DNA demethylation and transcriptional regulation, plays a key role in the maintenance of stem cell pluripotency, and is dysregulated in malignant cells. The identification of cancer stem cells (CSCs) driving tumor growth and metastasis is the primary objective of biomarker discovery in aggressive prostate cancer (PCa). In this context, we analyzed TET1 expression in PCa. Methods A large-scale immunohistochemical analysis of TET1 was performed in normal prostate (NOR) and PCa using conventional slides (50 PCa specimens) and tissue microarrays (669 NOR and 1371 PCa tissue cores from 371 PCa specimens). Western blotting, RT-qPCR, and 450 K methylation array analyses were performed on PCa cell lines. Genome-wide correlation, gene regulatory network, and functional genomics studies were performed using publicly available data sources and bioinformatics tools. Results In NOR, TET1 was exclusively expressed in normal cytokeratin 903 (CK903)–positive basal cells. In PCa, TET1 was frequently detected in alpha-methylacyl-CoA racemase (AMACR)–positive tumor cell clusters and was detectable at all tumor stages and Gleason scores. Pearson’s correlation analyses of PCa revealed 626 TET1-coactivated genes (r > 0.5) primarily encoding chromatin remodeling and mitotic factors. Moreover, signaling pathways regulating antiviral processes (62 zinc finger, ZNF, antiviral proteins) and the pluripotency of stem cells were activated. A significant proportion of detected genes exhibited TET1-correlated promoter hypomethylation. There were 161 genes encoding transcription factors (TFs), of which 133 were ZNF-TFs with promoter binding sites in TET1 and in the vast majority of TET1-coactivated genes. Conclusions TET1-expressing cells are an integral part of PCa and may represent CSCs with oncogenic potential.

Objectives Despite improved survival rates of patients with HPV-associated OPSCC, a subset has distant metastasis or develops local recurrence during follow-up. To investigate potential underlying genetic alterations, we analyzed patients with HPV-driven OPSCC who suffered from recurrence in comparison to matching pairs with successful tumor control. Materials and methods We performed chromosomal copy number analyses and targeted next generation sequencing using a custom panel comprising genes that are frequently mutated in HPV-associated OPSCC. Results Specific differences regarding chromosomal aberrations were not observed between both groups. In HPV-driven OPSCC from patients with recurrence we found higher mutation rates compared to patients with successful tumor control. Especially mutation rates of HRAS (p ≤ 0.05) PIK3R1, STK11 and TP63 (p ≤ 0.1 each) were statistically significant or trending towards significance. The respective genes can be linked to transcription factors and signaling pathways involved in cell cycle regulation, proliferation and survival. Additionally, combinations of alterations were observed on chromosomes 16 and 19, which might also influence outcome. Conclusion Patients with HPV-driven OPSCC who develop recurrence or have metastasis may be defined by genetic alterations that might be responsible for poor outcome after standard therapy. This might be of importance for stratification in future de-escalation and targeted therapy.

Background Previous studies of our group revealed an association of the scaffolding protein Protein Tyrosine Phosphatase Interacting Protein 51 (PTPIP51) with the NFκB signaling on the RelA and IκB level. The NFκB signaling plays a pivotal role in many different tumor entities. Especially HER2 amplified breast cancer cells frequently display activation of the NFκB signaling. Here, the NFκB signaling is linked to the initiation, progression, and the metastasis of the breast cancer. Furthermore, a large body of evidence associates the NFκB signaling with the development of therapy resistance. We aimed to clarify the effects of NFκB inhibition on the NFκB- and the MAPK-related interactome of PTPIP51 in HaCat cells and SKBR3 cells and their correlation with cell viability. Results Both NFκB signaling inhibitors, PDTC and IKK-16, reduced the cell viability of SKBR3 cells. IKK-16 selectively reduced the cell viability in SKBR3 cells at 5µM, while the viability of HaCat cells was not affected. PDTC impaired the cell viability of both cell lines and induced a formation of the Raf1/14-3-3/PTPIP51 complex in SKBR3 cells, indicating a shift of PTPIP51 into MAPK signaling. The MAPK-related interactome of PTPIP51 remained unaffected by IKK-16. Conclusion (1)The effectiveness and selectivity of NFκB inhibition in malignant and non-malignant signaling systems depends on the level of the targeted signaling molecule. (2) PTPIP51 might serve as the mediator between the NFκB signaling and the MAPK pathway in SKBR3 cells upon NFκB inhibition.

Influenza A viruses (IAV) can cause lung injury and acute respiratory distress syndrome (ARDS), which is characterized by accumulation of excessive fluid (edema) in the alveolar airspaces and leads to hypoxemia and death if not corrected. Clearance of excess edema fluid is driven mostly by the alveolar epithelial Na,K-ATPase and is crucial for survival of patients with ARDS. We therefore investigated whether IAV infection alters Na,K-ATPase expression and function in alveolar epithelial cells (AECs) and the ability of the lung to clear edema. IAV infection reduced Na,K-ATPase in the plasma membrane of human and murine AECs and in distal lung epithelium of infected mice. Moreover, induced Na,K-ATPase improved alveolar fluid clearance (AFC) in IAV-infected mice. We identified a paracrine cell communication network between infected and noninfected AECs and alveolar macrophages that leads to decreased alveolar epithelial Na,K-ATPase function and plasma membrane abundance and inhibition of AFC. We determined that the IAV-induced reduction of Na,K-ATPase is mediated by a host signaling pathway that involves epithelial type I IFN and an IFN-dependent elevation of macrophage TNF-related apoptosis–inducing ligand (TRAIL). Our data reveal that interruption of this cellular crosstalk improves edema resolution, which is of biologic and clinical importance to patients with IAV-induced lung injury.

Rationale While severe coronavirus infections, including Middle East respiratory syndrome coronavirus (MERS-CoV) cause lung injury with high mortality rates, protective treatment strategies are not approved for clinical use. Objectives We elucidated the molecular mechanisms by which the cyclophilin inhibitors Cyclosporin A (CsA) and Alisporivir (ALV) restrict MERS-CoV to validate their suitability as readily-available therapy in MERS-CoV infection. Methods Calu-3 cells and primary human alveolar epithelial cells (hAEC) were infected with MERS-CoV and treated with CsA or ALV or inhibitors targeting cyclophilin inhibitor-regulated molecules including Calcineurin, NFAT, or MAP kinases. Novel CsA-induced pathways were identified by RNA sequencing and manipulated by gene knockdown or neutralising antibodies. Viral replication was quantified by qRT-PCR and TCID50. Data were validated in a murine MERS-CoV infection model. Results CsA and ALV both reduced MERS-CoV titers and viral RNA replication in Calu-3 and hAEC improving epithelial integrity. While neither Calcineurin nor NFAT inhibition reduced MERS-CoV propagation, blockade of c-Jun N-terminal kinase diminished infectious viral particle release but not RNA accumulation. Importantly, CsA induced interferon regulatory factor 1 (IRF1), a pronounced type-III-interferon (IFNλ) response and expression of antiviral genes. Down-regulation of IRF1 or IFNλ increased MERS-CoV propagation in presence of CsA. Importantly, oral application of CsA reduced MERS-CoV replication in vivo, correlating with elevated lung IFNλ levels and improved outcome. Conclusions We provide evidence that cyclophilin inhibitors efficiently decrease MERS-CoV replication in vitro and in vivo via upregulation of inflammatory, antiviral cell responses, in particular IFNλ. CsA might therefore represent a promising candidate to treat MERS-CoV infection., The cyclophilin inhibitors Cyclosporin A and Alisporivir activate host innate immunity by induction of interferon lambda via activation of IRF1 in human lung epithelial cells and in vivo, resulting in a significant inhibition of MERS-CoV.

BACKGROUND Anterior knee pain is the most common complication after intramedullary tibial nailing. The cause is often multifactorial and varies among individuals. Violation of the anterior intermeniscal ligament (AIL) during intramedullary tibial nailing might be a possible source of postsurgical anterior knee pain. Although there is a certain ambiguity regarding the importance and function of the AIL, neural structures in the AIL tissue might play a significant role with respect to functional purposes and pain perception. METHODS We subjected 6 AIL specimens to histologic examination to identify the neural structures that are a mandatory requirement as a source of anterior knee pain. Specifically, we performed three-dimensional immunohistochemical investigation of subtyping, orientation, and detailed characterization of neural structures within the AIL tissue. RESULTS Histologic and three-dimensional immunohistochemical examinations confirmed the presence of neural structures in all 6 AIL specimens. We identified myelinated and unmyelinated nerve fibers, as well as all types of mechanoreceptors. CONCLUSIONS Free nerve endings are a mandatory requirement for pain perception as a result of AIL violation during tibial nailing. Our verification of all different types of mechanoreceptors in the AIL tissue makes a role of the ligament in knee joint function and proprioception highly probable. Further investigations are necessary to clarify possible correlations between neural supply and function of the AIL. Violation of the ligament during operative procedures should be avoided, although the significance of the AIL is still debated.

Breast cancer is the most common female cancerous disease and the second most cause of cancer death in women. About 20–30% of these tumors exhibit an amplification of the HER2/ErbB2 receptor, which is coupled to a more aggressive and invasive growth of the cancer cells. Recently developed tyrosine kinase inhibitors and therapeutic antibodies targeting the HER2 receptor improved the overall survival time compared with sole radio- and chemotherapy. Upcoming resistances against the HER2-targeted therapy make a better understanding of the receptor associated downstream pathways an absolute need. In earlier studies, we showed the involvement of Protein Tyrosine Phosphatase Interacting Protein 51 (PTPIP51) in the mitogen-activated protein kinase (MAPK) pathway. The MAPK pathway is one of the most frequently overactivated pathways in HER2-amplified breast cancer cells. This study is aimed to elucidate the effects of four different TKIs on the interactome of PTPIP51, namely with the receptors EGFR and HER2, 14-3-3/Raf1 (MAPK pathway), its regulating enzymes, and the mitochondria-associated interaction partners in HER2 breast cancer cell lines (SK-BR3 and BT474) by using the Duolink proximity ligation assay, immunoblotting and knockdown of PTPIP51. Inhibition of both EGFR and HER2/ErbB2R shifted PTPIP51 into the MAPK pathway, but left the mitochondria-associated interactome of PTPIP51 unattended. Exclusively inhibiting HER2/ErbB2 by Mubritinib did not affect the interaction of PTPIP51 with the MAPK signaling. Selective inhibition of HER2 induced great alterations of mitochondria-associated interactions of PTPIP51, which ultimately led to the most-effective reduction of cell viability of SK-BR3 cells of all tested TKIs. The results clearly reveal the importance of knowing the exact mechanisms of the inhibitors affecting receptor tyrosine kinases in order to develop more efficient anti-HER2-targeted therapies.

The extracardiac juvenile rhabdomyoma is extremely rare in the field of Otorhinolaryngology. The tumour usually arises from the soft tissue of the face or from mucosal sites, especially the oropharynx and the oral cavity but only sporadic endolaryngeal cases have been described in literature so far with predominance of young males. Here, we describe the very rare case of endolaryngeal extracardiac juvenile rhabdomyoma in a 42-year-old male. Clinical examination showed a mass of the right vocal cord, resembling a cystic lesion. Microlaryngoscopy revealed a non-encapsulated lesion and histopathology including immunohistochemistry which consecutively led to the correct diagnosis. This case suggests that the endolaryngeal extracardiac juvenile rhabdomyoma can be easily confused with a vocal cord cyst. Malignant transformations have not been reported but recurrences have been described. When total excision cannot be accomplished, reoperation or narrow follow-up is indicated to prevent advanced revision surgeries.

BACKGROUND Ras association domain family (RASSF) comprises several tumor suppressor genes, which are often epigenetically inactivated in human tumors. Here, we aim to analyze the relevance of the recently identified member RASSF10 in prostate carcinogenesis. METHODS RASSF10 promoter methylation and mRNA expression were investigated by bisulfite-pyrosequencing and qRT-PCR, respectively, in prostate carcinoma (PCa) cell lines (LNCaP, 22Rv1, DU-145) and in 83 primary PCa and 53 primary benign prostatic hyperplasia (BPH) tissues obtained after radical prostatectomy. Histological localization of RASSF10 was done by in situ hybridization. To prove the epigenetic nature of RASSF10 down regulation, PCa cell lines were treated with 5-aza-2-deoxycytidine and trichostatin A. Potential function of RASSF10 was analyzed in LNCaP by colony formation and apoptosis assays. RESULTS RASSF10 mRNA was localized to cells of the basal layer of the prostatic gland. Absence (LNCaP) and decrease (22Rv1, DU-145) of RASSF10 expression was associated with promoter methylation and could be restored by demethylating agents. A link between RASSF10 mRNA reduction and promoter methylation was also detected in primary prostate tissues (P = 0.006), where PCa showed more frequently reduced RASSF10 levels when compared with BPH (33.7% vs. 13.2%, P = 0.009). RASSF10 methylation could be further associated with advanced tumor stage and advanced age (P-values

Scheipl S, Froehlich E V, Leithner A, Beham A, Quehenberger F, Mokry M, Stammberger H, Varga P P, Lazary A, Windhager R, Gattenloehner S & Liegl B (2012) Histopathology 60, 999–1003 Does insulin-like growth factor 1 receptor (IGF-1R) targeting provide new treatment options for chordomas? A retrospective clinical and immunohistochemical study Aims: The overall prognosis of chordoma is poor, and current treatment options are limited. The insulin-like growth factor 1 receptor (IGF-1R) pathway is important for cell signalling, and attractive for selective inhibition. We investigated the expression of IGF-1R and its ligands, IGF-1 and IGF-2, in a series of 50 chordomas, in order to assess whether IGF-1R-signalling could be a potential target for specific inhibition in chordomas. Methods and results: Fifty chordomas (34 primary tumours, 16 recurrences) from 44 patients were evaluated immunohistochemically for the expression of IGF-1R, IGF-1 and IGF-2. Thirty-eight chordomas (76%) expressed IGF-1R, 46 (92%) expressed IGF-1 and 25 (50%) expressed IGF-2. Semi-quantitative analyses revealed a moderate to strong staining intensity in ≥50% of tumour cells for IGF-1R, IGF-1 and IGF-2 in 18 (36%), 32 (64%) and eight (16%) chordomas, respectively. Tumour volume correlated significantly with IGF-1R-staining intensity in primary chordomas (P = 0.042). Conclusions: IGF-1R and IGF-1 are expressed in the majority of chordomas. IGF-1 expression is much stronger than IGF-2 expression. Patients whose chordomas show a moderate to strong staining intensity in ≥50% of tumour cells for IGF-1R (36%) might benefit most from IGF-1R targeting, particularly if they suffer from large and surgically non-resectable chordomas.

Influenza viruses (IVs) cause pneumonia in humans with progression to lung failure. Pulmonary DCs are key players in the antiviral immune response, which is crucial to restore alveolar barrier function. The mechanisms of expansion and activation of pulmonary DC populations in lung infection remain widely elusive. Using mouse BM chimeric and cell-specific depletion approaches, we demonstrated that alveolar epithelial cell (AEC) GM-CSF mediates recovery from IV-induced injury by affecting lung DC function. Epithelial GM-CSF induced the recruitment of CD11b(+) and monocyte-derived DCs. GM-CSF was also required for the presence of CD103(+) DCs in the lung parenchyma at baseline and for their sufficient activation and migration to the draining mediastinal lymph nodes (MLNs) during IV infection. These activated CD103(+) DCs were indispensable for sufficient clearance of IVs by CD8(+) T cells and for recovery from IV-induced lung injury. Moreover, GM-CSF applied intratracheally activated CD103(+) DCs, inducing increased migration to MLNs, enhanced viral clearance, and attenuated lung injury. Together, our data reveal that GM-CSF-dependent cross-talk between IV-infected AECs and CD103(+) DCs is crucial for effective viral clearance and recovery from injury, which has potential implications for GM-CSF treatment in severe IV pneumonia.

Multiple sclerosis is an inflammatory demyelinating disease of the central nervous system (CNS) that is thought to be caused by a combination of genetic and environmental factors. To date, considerable evidence has associated Epstein-Barr virus (EBV) infection with disease development. However, it remains controversial whether EBV infects multiple sclerosis brain and contributes directly to CNS immunopathology. To assess whether EBV infection is a characteristic feature of multiple sclerosis brain, a large cohort of multiple sclerosis specimens containing white matter lesions (nine adult and three paediatric cases) with a heterogeneous B cell infiltrate and a second cohort of multiple sclerosis specimens (12 cases) that included B cell infiltration within the meninges and parenchymal B cell aggregates, were examined for EBV infection using multiple methodologies including in situ hybridization, immunohistochemistry and two independent real-time polymerase chain reaction (PCR) methodologies that detect genomic EBV or the abundant EBV encoded RNA (EBER) 1, respectively. We report that EBV could not be detected in any of the multiple sclerosis specimens containing white matter lesions by any of the methods employed, yet EBV was readily detectable in multiple Epstein-Barr virus-positive control tissues including several CNS lymphomas. Furthermore, EBV was not detected in our second cohort of multiple sclerosis specimens by in situ hybridization. However, our real-time PCR methodologies, which were capable of detecting very few EBV infected cells, detected EBV at low levels in only 2 of the 12 multiple sclerosis meningeal specimens examined. Our finding that CNS EBV infection was rare in multiple sclerosis brain indicates that EBV infection is unlikely to contribute directly to multiple sclerosis brain pathology in the vast majority of cases.

A compound or collision tumor is a rare occurrence in dermatological findings [1]. The coincidence of malignant melanoma (MM) and basal cell carcinoma (BCC) within the same lesion have only been described in few cases in the literature [2–5]. However, until now the pathogenesis of collision tumors existing of MM and BCC remains unclear [2]. To our knowledge it has not been yet established whether there is a concordant genetic background or independent origin as a possible cause for the development of such a compound tumor. We, therefore, present the extremely rare case of a collision tumor of MM and BCC and the results of a genome-wide analysis by single nucleotide polymorphism array (SNP-Array) for detection of identical genomic aberrations.

Detection of minimal residual disease or micrometastases in rhabdomyosarcoma (RMS) has been an unresolved problem in 70 to 80% of RMS patients. In patients with alveolar type RMS, which harbors chromosomal translocations and produces tumor-specific fusion products, polymerase chain reaction (PCR)-based diagnosis is clear-cut. In the more frequent embryonal RMS, however, no such PCR-based marker has been described. Recently it has been suggested that the PCR-based detection of MyoD1 may be a valuable adjunct in the diagnosis of minimal disease in embryonal RMS. We report here that MyoD1 mRNA is not specific for RMS, but can be amplified from ex vivo samples of many other childhood tumors and some normal tissues. By contrast, simultaneous amplification of α and γ subunit message of the fetal type acetylcholine receptor (AChR), by a novel duplex PCR, and the quantification of both transcripts resulting in a α/γAChR ratio

Background: Although an internal tandem duplication (ITD) of the Fms-like receptor tyrosine kinase 3 receptor (FLT3) confers a very poor prognosis in acute myeloid leukemia (AML), it is insufficient to cause AML, suggesting important cooperating genetic events. NFATc1 is a member of the nuclear factor of activated T-cells (NFAT) transcription factor family, whose activation is dependent on nuclear translocation. NFATc1 is well known for its central role in regulating T-cell development and function, but was also shown to contribute to neoplastic transformation in diverse types of cancers. We have recently demonstrated that NFATc1 is overexpressed in primary FLT3-ITD+ AML causing resistance to the FLT3-kinase inhibitor sorafenib in vitro. Here we asked, whether NFATc1 induces AML transformation and drug resistance when expressed in the context of FLT3-ITD+ in vivo. Methods: Using the stem cell like (SCL)-promoter, Cre-recombinase inducible expression of a constitutively nuclear NFATc1 protein (cnNFATc1) was targeted to the early hematopoietic stem cell compartment. The SCL-Cre/cnNFATc1 mice were subsequently crossed with transgenic FLT3-ITD mice to yield Scl-Cre+/-/cnNFATc1+/+/FLT3-ITD+/+ mice (FCN+/+), which co-express cnNFAT and FLT3-ITD in the stem cell compartment. Results: Whereas FLT3-ITD mice developed a non-lethal, mild myeloproliferative disease, co-expression of FLT3-ITD with cnNFATc1 led to rapidly fatal AML. The median survival of FCN mice was only four months (range, 2 to 8 ). In median, FCN mice had 15-fold (range, 10 to 20) increased white blood cell count, a high proportion of circulating peripheral blasts, severe anemia and thrombocytopenia. There was also a significant B-cell developmental block. As opposed to F+/+ animals, FCN+/+ mice revealed a severe spleno- and hepatomegaly secondary to a gross leukemic infiltration, which disrupted the normal anatomical architecture of the involved organs. Fluorescence activating cell sorting and colony forming unit (CFU) assays demonstrated that cnNFATc1, when expressed in the context of FLT3-ITD, leads to a significant expansion of the lin-Sca+c-kit+ (LSK) stem cell-, progenitor cell (LK)-, the Gr-1-/CD11b+ monocytic and Gr-1low/CD11b+ immature myeloid compartments in bone marrow and spleen. FCN+/+-AML was polyclonal, and re-transplantable in secondary wild-type recipient mice. In vivo, cnNFATc1 caused not only potent FLT3-ITD inhibitor resistance to sorafenib and to the more FLT3-ITD-selective compound quizartinib, but also to chemotherapy. Comparative mRNA-sequencing of sorted LK and LSK compartments in wild-type, F+/+, C+/- N+/+, and FCN+/+ mice will reveal the genetic basis of the biological cooperativity between NFATc1 and FLT3-ITD in stem and progenitor cell compartments. Consistent with the murine data, a data set of AML patients (n=163) revealed that NFATc1 overexpression alone or when combined with FLT3-overexpression was associated with a significantly decreased overall survival (hazard ratio [HR] 1.91, CI 1.29-2.18, p=0.001 versus HR 2.57, CI 1.62-4.08, p Conclusions: These results provide for the first time evidence for an important role of nuclear NFATc1 overexpression in the molecular pathogenesis of FLT3-ITD positive AML and the induction of drug resistance. These data also underscore that the mutational landscape in AML insufficiently describes AML biology. Disclosures No relevant conflicts of interest to declare.

Ras association domain family (RASSF) comprises several tumor suppressor genes, which are often epigenetically inactivated in human tumors. Here, we aim to analyze the relevance of the recently identified member RASSF10 in prostate carcinogenesis.RASSF10 promoter methylation and mRNA expression were investigated by bisulfite-pyrosequencing and qRT-PCR, respectively, in prostate carcinoma (PCa) cell lines (LNCaP, 22Rv1, DU-145) and in 83 primary PCa and 53 primary benign prostatic hyperplasia (BPH) tissues obtained after radical prostatectomy. Histological localization of RASSF10 was done by in situ hybridization. To prove the epigenetic nature of RASSF10 down regulation, PCa cell lines were treated with 5-aza-2-deoxycytidine and trichostatin A. Potential function of RASSF10 was analyzed in LNCaP by colony formation and apoptosis assays.RASSF10 mRNA was localized to cells of the basal layer of the prostatic gland. Absence (LNCaP) and decrease (22Rv1, DU-145) of RASSF10 expression was associated with promoter methylation and could be restored by demethylating agents. A link between RASSF10 mRNA reduction and promoter methylation was also detected in primary prostate tissues (P = 0.006), where PCa showed more frequently reduced RASSF10 levels when compared with BPH (33.7% vs. 13.2%, P = 0.009). RASSF10 methylation could be further associated with advanced tumor stage and advanced age (P-values 0.05). Our preliminary functional assays revealed the ability of RASSF10 to inhibit colony formation (P = 0.018) and to increase apoptosis (P = 0.035).This is the first study, which demonstrates the frequent epigenetic inactivation of RASSF10 in PCa and its implication in clinical symptoms of PCa.

Inducible cAMP early repressor (ICER) is a transcriptional repressor, which, because of alternate promoter use, is generated from the 3′ region of the cAMP response modulator ( Crem ) gene. Its expression and nuclear occurrence are elevated by high cAMP levels in naturally occurring regulatory T cells (nTregs). Using two mouse models, we demonstrate that nTregs control the cellular localization of ICER/CREM, and thereby inhibit IL-2 synthesis in conventional CD4 + T cells. Ablation of nTregs in depletion of regulatory T-cell (DEREG) mice resulted in cytosolic localization of ICER/CREM and increased IL-2 synthesis upon stimulation. Direct contacts between nTregs and conventional CD4 + T cells led to nuclear accumulation of ICER/CREM and suppression of IL-2 synthesis on administration of CD28 superagonistic (CD28SA) Ab. In a similar way, nTregs communicated with B cells and induced the cAMP-driven nuclear localization of ICER/CREM. High levels of ICER suppressed the induction of nuclear factor of activated T cell c1 ( Nfatc1) gene in T cells whose inducible Nfatc1 P1 promoter bears two highly conserved cAMP-responsive elements to which ICER/CREM can bind. These findings suggest that nTregs suppress T-cell responses by the cAMP-dependent nuclear accumulation of ICER/CREM and inhibition of NFATc1 and IL-2 induction.

The JAK2 V617F mutation is an essential oncogenic event in Philadelphia negative chronic myeloproliferative disorders (Ph-cMPD). It is still unclear how a unique tyrosine kinase mutation can give rise to the broad clinical and morphologic spectrum of Ph-cMPD. One possible explanation could be differences in the JAK2 V617F gene dosage, or different maturation stages on which myeloid lineages are affected by the mutation. The extent of lymphoid lineage involvement in JAK2 F617F -positive cMPD is still controversial. We comparatively studied the zygosity status of microdissected megakaryocytes, nonmegakaryocytic hematopoietic cells, and reactive as well as neoplastic lymphoid nodules from bone marrow trephines of 61 patients with Ph-cMPD. The presence of the mutation and mutant gene dosage were determined by allele-specific polymerase chain reaction and TaqMan analysis, respectively. The mutation was detected in 22/32 (68%) cases of essential thrombocythemia, all cases of polycythemia vera, and 4/8 (50%) idiopathic myelofibrosis. Comparison of whole bone marrow sections and the different myeloid lineages showed similar percentages of the mutated allele. Restriction to a particular lineage or major differences in allele dosage were not observed, except for 2 cases in which megakaryocytes revealed a higher frequency of the mutated allele. A heterozygous JAK2 V617F mutation was detected in 3/8 "reactive" lymphoid nodule in patients with Ph-cMPD, whereas all concomitant non-Hodgkin lymphoma of B-cell type were negative. These results demonstrate that different myeloid lineages usually show similar frequencies of the JAK2 V617F allele. The occasional detection of JAK2 V617F in benign lymphocytes points to involvement of the lympho-myeloid stem cell.

CD56high acute myeloid leukemias (AMLs) have a poor prognosis, but it has been unclear how CD56 expression is controlled and how it relates to clinical aggressiveness. We show that CD56 expression on AML cells correlates with an abnormal expression pattern of runt-related transcription factor 1 (RUNX1) isoforms. Whereas full-length p48 RUNX1 (p48) up-regulated CD56 in AML cells, 3 previously unknown shorter RUNX1 isoforms, p38a, p30, and p24, suppressed CD56 expression. Both p48 and CD56 induced nuclear translocation of nuclear factor (NF)–κB and increased bcl2L12 expression, and inhibition of this pathway by small inhibitory RNA-mediated p48 knock down or NF-κB blockade substantially increased apoptosis in CD56+ AML cell lines. These findings indicate the potential for new therapy of CD56high AML by suppression of the “overactive” RUNX1/CD56/NF-κB signaling pathway(s).