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Abstract Background A crucial aspect of continued senior care is the early detection and management of frailty. Developing reliable and secure electronic frailty assessment tools can benefit virtual appointments, a need especially relevant in the context of the COVID-19 pandemic. An emerging effort has targeted web-based software applications to improve accessibility and usage. The objectives of this scoping review are to identify and evaluate web-based frailty assessment tools currently available and to identify challenges and opportunities for future development. Methods We conducted a review with literature (e.g., using MEDLINE databases) and Google searches (last updated on October 10, 2021). Each of the identified web applications were assessed based on eight featured categories and assigned a rating score accordingly. Results Twelve web-based frailty assessment applications were found, chiefly provided by the USA (50%) or European countries (41%) and focused on frailty grading and outcome prediction for specific patient groups (59%). Categories that scored well among the applications included the User Interface (2.8/3) and the Cost (2.7/3). Other categories had a mean score of 1.6/3 or lower. The least developed feature was Data Saving. Conclusions Web-based applications represent a viable option for remote frailty assessments and multidisciplinary integrated care of older adults. Despite the available web-based frailty assessments on the Internet, many missed certain needed features for professional use in healthcare settings. This situation calls for fully comprehensive web-based applications, taking into consideration a number of key functions linking graphical user interface and functionalities, and paying special attention to secure data management.

Remote-sensing data are used extensively to monitor water quality parameters such as clarity, temperature, and chlorophyll-a (chl-a) content. This is generally achieved by collecting in situ data coincident with satellite data collections and then creating empirical water quality models using approaches such as multi-linear regression or step-wise linear regression. These approaches, which require modelers to select model parameters, may not be well suited for optically complex waters, where interference from suspended solids, dissolved organic matter, or other constituents may act as “confusers”. For these waters, it may be useful to include non-standard terms, which might not be considered when using traditional methods. Recent machine-learning work has demonstrated an ability to explore large feature spaces and generate accurate empirical models that do not require parameter selection. However, these methods, because of the large number of included terms involved, result in models that are not explainable and cannot be analyzed. We explore the use of Least Absolute Shrinkage and Select Operator (LASSO), or L1, regularization to fit linear regression models and produce parsimonious models with limited terms to enable interpretation and explainability. We demonstrate this approach with a case study in which chl-a models are developed for Utah Lake, Utah, USA., an optically complex freshwater body, and compare the resulting model terms to model terms from the literature. We discuss trade-offs between interpretability and model performance while using L1 regularization as a tool. The resulting model terms are both similar to and distinct from those in the literature, thereby suggesting that this approach is useful for the development of models for optically complex water bodies where standard model terms may not be optimal. We investigate the effect of non-coincident data, that is, the length of time between satellite image collection and in situ sampling, on model performance. We find that, for Utah Lake (for which there are extensive data available), three days is the limit, but 12 h provides the best trade-off. This value is site-dependent, and researchers should use site-specific numbers. To document and explain our approach, we provide Colab notebooks for compiling near-coincident data pairs of remote-sensing and in situ data using Google Earth Engine (GEE) and a second notebook implementing L1 model creation using scikitlearn. The second notebook includes data-engineering routines with which to generate band ratios, logs, and other combinations. The notebooks can be easily modified to adapt them to other locations, sensors, or parameters.

We present the development and testing of a web application called the historical validation tool (HVT) that processes and visualizes observed and simulated historical stream discharge data from the global GEOGloWS ECMWF streamflow services (GESS), performs seasonally adjusted bias correction, computes goodness-of-fit metrics, and performs forward bias correction on subsequent forecasts. The HVT corrects GESS output at a local scale using a technique that identifies and corrects model bias using observed hydrological data that are accessed using web services. HVT evaluates the performance of the GESS historic simulation data and provides more accurate historic simulation and bias-corrected forecast data. The HVT also allows users of the GEOGloWS historical streamflow data to use local observed data to both validate and improve the accuracy of local streamflow predictions. We developed the HVT using Tethys Platform, an open-source web application development framework. HVT presents data visualization using web mapping services and data plotting in the web map interface while functions related to bias correction, metrics reporting, and data generation for statistical analysis are computed by the back end. We present five case studies using the HVT in Australia, Brazil, Colombia, the Dominican Republic, and Peru. In these case studies, in addition to presenting the application, we evaluate the accuracy of the method we implemented in the HVT for bias correction. These case studies show that the HVT bias correction in Brazil, Colombia, and Peru results in significant improvement in historic simulation across the countries, while bias correction only resulted in marginal historic simulation improvements in Australia and the Dominican Republic. The HVT web application allows users to use local data to adjust global historical simulation and forecasts and validate the results, making the GESS modeling results more useful at a local scale.

Abstract Background Prostate cancer is a common disease in men and at present there is no effective therapy available due to its recurrence despite androgen deprivation therapy. The epidermal growth factor receptor family (EGFR/HER1, HER2/neu and HER3)/PI3K/Akt signaling axis has been implicated in prostate cancer development and progression. However, Erlotinib, an EGFR tyrosine kinase inhibitor, has less effect on proliferation and apoptosis in prostate cancer cell lines. In this study, we evaluate whether MP470, a novel receptor tyrosine kinase inhibitor alone or in combination with Erlotinib has inhibitory effect on prostate cancer in vitro and in vivo. Methods The efficacy of MP470 or MP470 plus Erlotinib was evaluated in vitro using three prostate cancer cell lines by MTS and apoptosis assays. The molecular mechanism study was carried out by phosphorylation antibody array, immunoblotting and immunohistochemistry. A LNCaP mouse xenograft model was also used to determine the tumor growth inhibition by MP470, Erlotinib or the combination treatments. Results MP470 exhibits low μM IC50 in prostate cancer cell lines. Additive effects on both cytotoxicity and induction of apoptosis were observed when LNCaP were treated with MP470 in combination with Erlotinib. This combination treatment completely inhibited phosphorylation of the HER family members (HER1, 2, 3), binding of PI3K regulatory unit p85 to HER3 and downstream Akt activity even after androgen depletion. Furthermore, in a LNCaP mouse xenograft model, the MP470-Erlotinib combination produced 30–65% dose-dependent tumor growth inhibition (TGI). Conclusion We propose that MP470-Erlotinib targets the HER family/PI3K/Akt pathway and may represent a novel therapeutic strategy for prostate cancer.

Recently, an antibody which inhibits the glycoprotein A repetitions predominant (GARP)-mediated release of active transforming growth factor beta (TGFβ) from the TGFβ propeptide latency-associated peptide (LAP) showed preclinical activity in a murine model of the chronic myeloproliferative neoplasms (MPN). Consequently, we investigated the expression of the immunosuppressive molecules LAP and GARP on peripheral blood lymphocytes from 56 MPN patients and 11 healthy donors (HD). We found that lymphocytes from patients with MPN express higher levels of LAP and GARP with no strong differences found between the different MPN diagnoses. The impact of clinical parameters on the expression of LAP and GARP by lymphocytes showed that patients with calreticulin (CALR)mut MPN have increased expression compared with HD and patients with the Januskinase2 (JAK2) mutation. The fraction of lymphocytes bound to activated platelets (aPLT) strongly correlate to LAP and GARP expression suggesting that it is not the lymphocytes themselves but aPLT, which confer the increased expression of GARP and LAP on MPN patient lymphocytes. Notably, no differences in neither platelet counts nor anti-thrombotic therapy was identified between patients with JAK2- and CALRmut patients. Analysis of platelet gene expression failed to identify differences in expression of relevant genes between JAK2- and CALRmut patients., (© 2024 British Society for Haematology and John Wiley & Sons Ltd.)

T-cell-based immunotherapy has recently evolved as a treatment option for a number of haematological malignancies and is also being developed in solid tumours. A common side effect of chimeric antigen T-cell therapy (CAR-T) and treatment with T-cell engagers is cytokine release syndrome (CRS), which is a potentially life-threatening condition characterized by release of inflammatory mediators. The treatment of CRS is similar to that of other hyper-inflammatory conditions and involves supportive treatment as well as immunosuppressive therapy. The risk of CRS can be mitigated by step-up dosing and immunosuppressive pre-treatment, as argued in this review., (Published under Open Access CC-BY-NC-BD 4.0. https://creativecommons.org/licenses/by-nc-nd/4.0/.)

Objective: Given a proposed role for PD-L1 + and IL-10-producing B-cell subsets in promoting certain cancers, we sought to characterize the frequency and phenotype of B cells in patients with chronic myeloproliferative neoplasms (MPNs) and the influence of ruxolitinib and interferon-α2 therapy., Methods: We analyzed B-cell frequencies and phenotype in patients with MPNs (n = 107), before and during treatment with ruxolitinib (n = 29), interferon-α2 (n = 21), or the two drugs in combination (COMBI; n = 42) and healthy donors (HDs; n = 52) using flow cytometry., Results: Myelofibrosis patients had lower lymphocyte counts and proportions of B cells than patients with essential thrombocythemia or polycythemia vera and HDs. The B-cell count correlated inversely with JAK2-V617F allele burden and spleen size and increased after ruxolitinib or COMBI treatment. The proportions of PD-L1 + B cells and PD-1 + B cells were significantly higher in patients with myelofibrosis or polycythemia vera than in HDs and decreased during ruxolitinib and COMBI treatment. The proportions of TNF-α + and IL-6 + B cells were elevated in myelofibrosis patients. The proportion of IL-6 + B cells decreased, and the proportion of IL-10 + B cells increased during ruxolitinib treatment., Conclusion: B-cell frequency and phenotype were altered in MPN patients. Ruxolitinib therapy had marked effects on both frequency and phenotype., (© 2019 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Mutations in exon 9 of the calreticulin gene (CALR) frequently occur in patients with chronic myeloproliferative neoplasms (MPN). Patients exhibit spontaneous cellular immune responses to epitopes derived from the mutant CALR C-terminus, and CALR-mutant-specific T cells recognize autologous CALR-mutant malignant cells. This study investigated whether CALR-mutant-specific T cells occur naturally in CALRwt MPN-patients and in healthy individuals. Specific immune responses against epitopes in the mutant CALR peptide sequence were detected in both CALRwt MPN-patients and in healthy individuals. Healthy donors displayed more frequent and stronger CALR-mutant specific T-cell responses compared to the responses identified in CALR-mutant MPN-patients. Several T-cell responses were identified in healthy donors directly ex vivo. Importantly, by running functional analyses on live-sorted immune cells from healthy donors, we showed that circulating CALR-mutant-specific immune cells are T-memory cells. These findings suggest, that healthy individuals acquire a CALR exon 9 mutation, but the immune system reacts and clears the mutant cells, and during this reaction generates CALR-mutant specific T-memory cells. We believe that these findings provide the evidence for tumor immune surveillance in MPN.

Compelling evidence supports the existence of a profound immune dysregulation in patients with chronic myeloproliferative neoplasms (MPN). Increased Arginase-1 expression has been described in MPN patients and in solid cancers. This increase contributes to an immunosuppressive tumor microenvironment in MPN patients because of L-arginine depletion by Arginase-1-expressing regulatory cells and cancer cells, which subsequently limits the activation of circulating effector cells. In the present study, we demonstrate that Arginase-1-derived peptides are recognized by T cells among peripheral mononuclear blood cells from MPN patients. We characterized the Arginase-1-specific T cells as being CD4 + and found that the magnitude of response to the Arginase-1 peptides depends on disease stage. Activation of Arginase-1-specific T cells by vaccination could be an attractive novel immunotherapeutic approach to targeting malignant and suppressive cells in MPN patients in combination with other immunotherapeutics.

The Chronic Myeloproliferative Neoplasms (MPN) are cancers characterized by hyperinflammation and immune deregulation. Concurrently, the expression of the immune check point programmed death ligand 1 (PD-L1) is induced by inflammation. In this study we report on the occurrence of spontaneous T cell responses against a PD-L1 derived epitope in patients with MPN. We show that 71% of patients display a significant immune response against PD-L1, and patients with advanced MPN have significantly fewer and weaker PD-L1 specific immune responses compared to patients with non-advanced MPN. The PD-L1 specific T cell responses are CD4 + T cell responses, and by gene expression analysis we show that expression of PD-L1 is enhanced in patients with MPN. This could imply that the tumor specific immune response in MPN could be enhanced by vaccination with PD-L1 derived epitopes by boosting the anti-regulatory immune response hereby allowing tumor specific T cell to exert anti-tumor immunity.

The calreticulin (CALR) exon 9 mutations are found in ∼30% of patients with essential thrombocythemia and primary myelofibrosis. Recently, we reported spontaneous immune responses against the CALR mutations. Here, we describe that CALR-mutant (CALRmut)-specific T cells are able to specifically recognize CALRmut cells. First, we established a T-cell culture specific for a CALRmut epitope. These specific T cells were able to recognize several epitopes in the CALRmut C terminus. Next, we established a CALRmut-specific CD4 + T-cell clone by limiting dilution. These CD4 + T cells recognized autologous CALRmut monocytes and hematopoietic stem cells, and T-cell recognition of target cells was dependent on the presence of CALR. Furthermore, we showed that the CALRmut response was human leukocyte antigen (HLA)-DR restricted. Finally, we demonstrated that the CALRmut-specific CD4 + T cells, despite their phenotype, were cytotoxic to autologous CALRmut cells, and that the cytotoxicity was mediated by degranulation of the T cells. In conclusion, the CALR exon 9 mutations are targets for specific T cells and thus are promising targets for cancer immune therapy such as peptide vaccination in patients harboring CALR exon 9 mutations.

Gene expression profiling in Philadelphia-negative chronic myeloproliferative neoplasms (MPNs) have unraveled significant deregulation of several immune and inflammation genes of potential importance for clonal evolution. Other mechanisms might be downregulation of major histocompatibility class I and II genes used by tumor cells to escape antitumor T-cell-mediated immune responses. Several genes encoding human leukocyte antigen (HLA) class I and II molecules have been shown to be significantly downregulated. Upregulation of HLA genes is considered one of the mechanisms of action of interferon (IFN)-alpha2, but regulation of these genes during IFN-alpha2 treatment in MPNs has never been studied. Our findings show a significant upregulation of several HLA genes of importance for tumor immune surveillance by IFN-alpha2 treatment in MPNs. This mechanism might enhance the cytotoxic potential of immune cells against MPNs and explain the induction of minimal residual disease by IFN-alpha2 treatment in these patients.

Recent studies have shown that a large proportion of patients classified as essential thrombocythemia (ET) actually have early primary prefibrotic myelofibrosis (prePMF), which implies an inferior prognosis as compared to patients being diagnosed with so-called genuine or true ET. According to the World Health Organization (WHO) 2008 classification, bone marrow histology is a major component in the distinction between these disease entities. However, the differential diagnosis between them may be challenging and several studies have not been able to distinguish between them. Most lately, it has been argued that simple blood tests, including the leukocyte count and plasma lactate dehydrogenase (LDH) may be useful tools to separate genuine ET from prePMF, the latter disease entity more often being featured by anemia, leukocytosis and elevated LDH. Whole blood gene expression profiling was performed in 17 and 9 patients diagnosed with ET and PMF, respectively. Using elevated LDH obtained at the time of diagnosis as a marker of prePMF, a 7-gene signature was identified which correctly predicted the prePMF group with a sensitivity of 100% and a specificity of 89%. The 7 genes included MPO, CEACAM8, CRISP3, MS4A3, CEACAM6, HEMGN, and MMP8, which are genes known to be involved in inflammation, cell adhesion, differentiation and proliferation. Evaluation of bone marrow biopsies and the 7-gene signature showed a concordance rate of 71%, 79%, 62%, and 38%. Our 7-gene signature may be a useful tool to differentiate between genuine ET and prePMF but needs to be validated in a larger cohort of "ET" patients., Competing Interests: The authors have declared that no competing interests exist.

Long-term therapy with IFN-α2 is associated with sustained major molecular remissions in JAK2-positive ET and PV. The efficacy of IFN-α2 may be partly mediated by modulation of immune cells, which was investigated in twenty patients with ET (n = 6) and PV (n = 14). The frequency of CD4(+) CD25(+) Foxp3(+) T cells was significantly increased during IFN-α2 treatment in all patients (P < 0.0001). A significant expansion of the CD56(bright) NK cells (P = 0.0002) and a concomitant decrease in the frequency of CD56(dim) NK cells (P < 0.0001) were also detected. Myeloid DCs (mDCs) and plasmacytoid DCs (pDCs) were studied in nine patients, and decreased frequencies of both cell types were observed during the course of treatment. On both mDCs and pDCs, HLA-ABC expression was upregulated (P = 0.003), but decreasing expression levels of HLA-DR was detected on mDCs. The expression of CD40 (P = 0.002), CD83 (P = 0.03), and CD86 (P = 0.01) increased, but was confined to pDCs. Furthermore, PD-L1 expression was reduced on mDC (P = 0.003) and increased on pDCs (P = 0.02). No significant correlations were found between the changes in immune cells and hematological or molecular responses achieved in our cohort of patients. So forth, it remains to be revealed whether the profound changes in circulating immune cells contribute to the beneficial effects of long-term IFN-α2 treatment in some patients., (© 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Essential thrombocythemia (ET) and polycythemia vera (PV) are Philadelphia chromosome-negative chronic myeloproliferative neoplasms (MPNs) characterized by the JAK2 V617F mutation, which can be found in more than 98% of PV patients and in ∼ 50% of ET patients. Assessment of the JAK2 V617F allele burden by a highly sensitive quantitative PCR (qPCR) assay appears to be a useful tool for monitoring minimal residual disease (MRD) and evaluating treatment efficacy. This report expands and substantiates existing data, showing that IFN-alpha2 is a highly potent immunomodulating agent capable of inducing MRD with low-burden JAK2 V617F, major molecular response (MMR), complete hematological remission (CHR) and complete histomorphological normalization of the bone marrow in a sub-set of patients with ET and PV after long-term treatment (≥ 3.5 years). Furthermore, long-lasting hematological, molecular and histomorphological remissions are sustained after discontinuation of IFN-alpha2 for up to ∼ 5-6 years.

Polycythaemia vera, essential thrombocytosis and primary myelofibrosis are closely related, clonal myeloproliferative neoplasms. Our knowledge of the underlying molecular mechanisms driving these diseases has increased dramatically during the latest ten years. Traditionally, treatment of these malignancies has focused on lowering their inherent thromboembolic risk but with the discovery of the JAK2-V617F mutation and most recently the calreticulin mutations new therapeutic options such as interferon-alpha, JAK2-inhibitors and statins are being contemplated. This article reviews these new treatment options.

In recent years, major molecular remissions have been observed in patients with JAK2-positive chronic myeloproliferative neoplasms (MPNs) after therapy with IFN-α. IFN-α is known to have altering effects on immune cells involved in immune surveillance and might consequently enhance anti-tumor immune response against the JAK2-mutated clone. The objective of this study was to investigate circulating levels and phenotype of natural killer cells in 29 JAK2-positive MPN patients during IFN-α treatment. Furthermore, functional studies of NK cells upon target-cell recognition and cytokine stimulation were performed. The CD56(bright) and CD56(dim) NK cell subtypes display different properties in terms of cytokine production and cytotoxicity, respectively. Our results show a significant increase in the proportion of CD56(bright) NK cells and a decreasing CD56(dim) population during treatment with IFN-α compared to patients that are untreated, treated with hydroxyurea and healthy controls, P < 0.0001. Furthermore, an overall increase in cytokine-dependent (IL-12 and IL-15) IFN-γ expression by CD56(dim) NK cells during IFN-α treatment was observed. In contrast, our data indicate a compromised NK cell response to target-cell recognition during treatment with IFN-α in four patients. We also report low levels of circulating NK cells in untreated patients compared to healthy donors, patients treated with hydroxyurea and IFN-α, P = 0.02. Based on our findings, one might speculate whether treatment with IFN-α skews the human NK population toward a helper type that may assist in CD8(+) T cell priming in lymphoid tissues at the expense of their immediate cytotoxic functions in peripheral blood and tissues., (© 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

The Philadelphia-negative chronic myeloproliferative neoplasms - essential thrombocythemia (ET), polycythemia vera (PV), and myelofibrosis (MF) (MPNs) - have recently been shown to be associated with chronic inflammation, oxidative stress and accumulation of reactive oxygen species (ROS). Using whole blood transcriptional profiling, we report that several oxidative stress and anti-oxidative stress genes are significantly deregulated in MPNs. Among the twenty most up- and downregulated genes, ATOX1, DEFB122, GPX8, PRDX2, PRDX6, PTGS1, and SEPP1 were progressively upregulated from ET over PV to PMF, whereas AKR1B1, CYBA, SIRT2, TTN, and UCP2 were progressively downregulated in ET, PV and PMF (all FDR <0.05). The gene Nrf2, encoding the transcription factor nuclear factor erythroid 2-related factor 2 (NFE2L2 or Nrf2) was significantly downregulated in all MPNs. Nrf2 has a key role in the regulation of the oxidative stress response and modulates both migration and retention of hematopoietic stem cells (HSCs) in their niche. The patogenetic importance of Nrf2 depletion in the context of expansion of the hematopoietic progenitor pool in MPNs is discussed with particular focus upon the implications of concomitant downregulation of Nrf2 and CXCR4 for stem cell mobilization.

Gene expression profiling studies in the Philadelphia-negative chronic myeloproliferative neoplasms have revealed significant deregulation of several immune and inflammation genes that might be of importance for clonal evolution due to defective tumor immune surveillance. Other mechanisms might be down-regulation of major histocompatibility (MHC) class I and II genes, which are used by tumor cells to escape antitumor T-cell-mediated immune responses. We have performed whole blood transcriptional profiling of genes encoding human leukocyte antigen (HLA) class I and II molecules, β2-microglobulin and members of the antigen processing machinery of HLA class I molecules (LMP2, LMP7, TAP1, TAP2 and tapasin). The findings of significant down-regulation of several of these genes may possibly be of major importance for defective tumor immune surveillance. Since up-regulation of HLA genes is recorded during treatment with epigenome modulating agents (DNA-hypomethylators and DNA-hyperacetylators [histone deacetylase inhibitors]) and interferon-α2, our findings call for prospective transcriptional studies of HLA genes during treatment with these agents.

Within recent years data has accumulated demonstrating the efficacy of recombinant interferon alpha2 (rIFN-alpha2) in the treatment of chronic myeloproliferative neoplasms (MPNs). We report on clinical and molecular data in the largest cohort of JAK2 V617F mutant MPN Danish patients (n=102) being treated long-term with rIFN-alpha2 (rIFN-alpha2a and rIFN-alpha2b in a non-clinical trial setting. The median follow-up was 42 months. We substantiate the capacity of rIFN-alpha2 to induce complete hematologic remissions (ET 95%, PV 68%) and molecular response. In total 76 patients (74.5%) had a decline in JAK2 V617F allele burden with a median reduction from baseline of 59% (95% c.i. 50-73%, range 3-99%). A decline in JAK2 V617F allele burden was recorded in both ET (median 24-10% (95% c.i.: 8-16%), and PV (median 59-35% (95% c.i.: 17-33%). Patients with the lowest pre-treatment JAK2 V617F allele burdens tend to achieve the most favourable responses on long term treatment with rIFN-alpha2. Eleven patients (10%) had deep molecular remissions with ≤ 2% JAK2 V617F mutant DNA. Finally, long term treatment with rIFN-alpha2 was associated with a very low thrombosis rate. Our observations are supportive of the concept of early up-front treatment with rIFN-alpha2., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

Essential thrombocythemia (ET), polycythemia vera (PV) and primary myelofibrosis (PMF) are hematopoietic stem cell neoplasms that may be associated with autoimmune or chronic inflammatory disorders. Earlier gene expression profiling studies have demonstrated aberrant expression of genes involved in inflammatory responses, mainly being performed on granulocytes or CD34+ cells. Using gene expression profiling of whole blood from patients with ET (n=16), PV (n=36), and PMF (n=9), several genes involved in inflammation and immune regulation were found to be significantly deregulated. Our findings may reflect chronic inflammation to be of pathogenetic importance for the progression of these neoplasms toward the myelofibrotic end-stage and may also account for the increased frequency of second cancer in these diseases., (Copyright © 2012 Elsevier Ltd. All rights reserved.)

The recent discovery of the Janus activating kinase 2 V617F mutation in most patients with polycythemia vera (PV) and half of those with essential thrombocythemia (ET) and primary myelofibrosis (PMF) has favored the hypothesis of a biological continuum from ET over PV to PMF. We performed gene expression profiling of whole blood from control subjects (n = 21) and patients with ET (n = 19), PV (n = 41), and PMF (n = 9) using DNA microarrays. Applying an unsupervised method, principal component analysis, to search for patterns in the data, we demonstrated a separation of the four groups with biological relevant overlaps between the different entities. Moreover, the analysis separates Janus activating kinase 2-negative ET patients from Janus activating kinase 2-positive ET patients. Functional annotation analysis demonstrates that clusters of gene ontology terms related to inflammation, immune system, apoptosis, RNA metabolism, and secretory system were the most significantly deregulated terms in the three different disease groups. Our results yield further support for the hypothesis of a biological continuum originating from ET over PV to PMF. Functional analysis suggests an important implication of these gene ontology clusters in the pathogenesis of these neoplasms and in disease evolution from ET over PV to PMF., (Copyright © 2012 ISEH - Society for Hematology and Stem Cells. Published by Elsevier Inc. All rights reserved.)

Myeloproliferation, myeloaccumulation (decreased apoptosis), inflammation, bone marrow fibrosis and angiogenesis are cardinal features of the Philadelphia-negative chronic myeloproliferative neoplasms: essential thrombocythemia (ET), polycythemia vera (PV) and primary myelofibrosis (PMF). Histone deacetylases (HDACs) have a critical role in modulating gene expression and, accordingly, in the control of cell pathobiology and cancer development. HDAC inhibition has been shown to inhibit tumor growth (impaired myeloproliferation), to modulate the balance between pro- and antiapoptotic proteins in favor of apoptosis (enhanced apoptosis) and also to inhibit angiogenesis. Recently, enhanced HDAC enzyme activity has been found in CD34+cells from patients with PMF, enzyme activity levels highly exceeding those recorded in other chronic myeloproliferative neoplasms (CMPNs). The raised levels correlated to the degree of splenomegaly, suggesting that HDAC might be recruited as ET or PV progresses into myelofibrosis or PMF progresses into a more advanced stage. Accordingly, HDAC inhibition is an obvious novel therapeutic approach in these neoplasms. Using global gene expression profiling of whole blood from patients with CMPNs, we have found a pronounced deregulation of HDAC genes, involving significant up-regulation of the HDAC genes 9 and 11, with the highest expression levels being found in patients with ET (HDAC9 and 11), PMF (HDAC9) and CMPNs (both HDAC9 and HDAC11). Furthermore, we have identified that the HDAC6 gene is progressively expressed in patients with ET, PV and PMF, reflecting a steady accumulation of abnormally expressed HDAC6 during disease evolution. Our results lend further support to HDACs as important epigenetic targets in the future treatment of patients with CMPNs. Since the highest expression levels of HDAC genes were recorded in ET, in PMF and in the entire CMPN group, their down-regulation by HDAC inhibitors might be associated with decreased disease activity, including reduction of splenomegaly.

Primary myelofibrosis (PMF) is characterized by leukoerythroblastic anemia with circulating immature myeloid cells, including CD34+ cells, progressive splenomegaly and accumulation of connective tissue and neoangiogenesis in the bone marrow. Altered bone marrow stroma and cell adherence account for the egress of CD34+ cells from the bone marrow. Carcinoembryonic antigen-related cell adhesion molecule (CEACAM) 6 has been implicated in cell adhesion, cellular invasiveness, angiogenesis, and inflammation, which are all key processes in the pathophysiology of PMF. Accordingly, CEACAMs may play an important role in these processes in patients with myelofibrosis as well. Using a genome-wide approach, several genes of the CEACAM family were found to be deregulated in patients with PMF and related myeloproliferative neoplasms (n=69). In PMF patients, the CEACAM6 and 8 were significantly upregulated (fold change (FC) 12.5 and 14.0, respectively and FDR adjusted p values 7.71×10(-7) and 1.48×10(-5), respectively). The elevated expression of CEACAM genes may reflect clonal myeloid expansion and neutrophil activation being most exaggerated in patients with PMF. Alternatively, the highly elevated gene expression of CEACAM6 and 8 in PMF may be molecular markers of myelofibrotic transformation, implying enhanced proteolytic activity, egress of CD34+ cells into the circulation, and neoangiogenesis., (Copyright © 2011 Elsevier Ltd. All rights reserved.)

Recent reports have described complete or major molecular remission in patients with polycythemia vera after long-term treatment with the immunomodulatory agent IFN-α2. Accordingly, there are reasons to believe that the immune system is a key player in eradicating the JAK2 mutated clone in these patients. Foxp3(+) regulatory T cells play a pivotal role in maintaining immune homeostasis and, importantly, preventing immune reactivity to self-antigens; however, their suppressive activity can compromise an effective antitumor immune response, and high frequencies of regulatory T cells in peripheral blood have been reported in both hematologic and solid cancers. We have analyzed the number, phenotype, and function of circulating CD4(+)CD25(+)Foxp3(+) T cells in patients with chronic myeloproliferative neoplasms. Surprisingly, we found a marked expansion of this subset of lymphocytes in patients treated with IFN-α2 (13.0%; 95% confidence interval [CI] 10.8% to 15.2%) compared with healthy donors (6.1%; 95% CI 4.9% to 7.2%), patients with untreated chronic myeloproliferative neoplasms (6.9%; 95% CI 5.8% to 7.4%), or patients treated with hydroxyurea (5.8%; 95% CI 4.3% to 7.4%; P < .0001).

Gene expression profiling studies have unraveled deregulation of several genes that might be of pathogenetic importance for the development and phenotype of the Philadelphia-negative chronic myeloproliferative neoplasms. In the context of interferon-alpha2 as a promising therapeutic agent, we focused upon the transcriptional profiling of interferon-associated genes in patients with essential thrombocythemia (ET) (n = 19), polycythemia vera (PV) (n = 41), and primary myelofibrosis (PMF) (n = 9). Using whole-blood transcriptional profiling and accordingly obtaining an integrated signature of genes expressed in several immune cells (granulocytes, monocytes, B cells, T cells, platelets), we have identified a number of interferon-associated genes to be significantly deregulated but with a highly significant deregulation of interferon-inducible gene 27 (IFI27) (ET, PV, and PMF, fold change 8, 16, and 30, respectively). The striking deregulation of IFI genes may reflect a hyperstimulated but insufficient immune system being most enhanced in patients with advanced myelofibrosis, in whom the IFI27 gene displayed an exceedingly high expression. The interferon signature may reflect primary myelofibrosis as the burn-out phase of chronic inflammation which ultimately elicits clonal evolution and expansion owing to an exaggerated but incompetent antitumor immune response. Finally, IFI27 may be a novel biomarker of disease activity and tumor burden in patients with CMPNs., (© 2011 John Wiley & Sons A/S.)

The Philadelphia-negative chronic myeloproliferative neoplasms encompass essential thrombocythemia (ET), polycythemia vera (PV) and primary myelofibrosis (PMF). A major break-through in the understanding of the pathogenesis of these neoplasms occurred in 2005 by the discovery of the JAK2 V617F mutation in the large majority of patients with PV and in half of those with ET and PMF. A number of studies have shown that the "tumor burden" may be monitored at the molecular level in JAK2-positive patients using highly sensitive real-time quantitative PCR. During the last 25 years several studies have shown that interferon-alpha (IFN-alpha) induces complete haematological remissions in a large proportion of the patients. However, its use in clinical practice has unfortunately been limited due to side effects with high drop-out rates in most studies. Recently, IFN-alpha2 has been shown to induce deep molecular remissions and also normalization of the bone marrow in PV, which may be sustained even after discontinuation of IFN-alpha2 therapy. Accordingly, in the coming years we are most likely facing a new era of increasing interest for using IFN-alpha2 in the treatment of patients with PV, ET and the hyperproliferative phase of PMF. This paper reviews the history of IFN - in principle IFN-alpha2 - and its present status in the treatment of PV and related diseases. The role of IFN-alpha2 as immune therapy in the future treatment of CMPNs is highlighted and the rationale for the concept of minimal residual disease and potentially cure after long-term immune therapy with IFN-alpha2 is discussed and foreseen as an achievable goal in the future.

Introduction: The diagnosis of hereditary spherocytosis (HS) is based upon clinical presentation, typical laboratory findings of haemolysis with an increased mean corpuscular haemoglobin concentration (MCHC) combined with a positive osmotic fragility result. The disorder is caused by structural defects in red cell cytoskeletal proteins. The dye eosin-5'-maleimide (EMA) binds to band three of the red cell membrane. The fluorescence intensity of EMA-labelled red cells can be quantified by flowcytometric analysis. Decreased fluorescence is found in patients with HS. We have evaluated this method by comparing flowcytometric analysis of red cells from patients with HS and patients with other haemolytic disorders., Material and Methods: We included 21 patients with HS and 27 patients with other haemolytic disorders. The red cells were incubated and labelled with EMA followed by flowcytometric analysis measuring the mean-fluorescence-intensity expressed as EMA percentage., Results: Based on the overall results, we assess an EMA percentage threshold of 15 or above to indicate HS. We found a sensitivity of 95% and a specificity of 93%., Conclusion: The osmotic fragility test does not have the same high degree of sensitivity and specificity and the test is time-consuming in the laboratory setting. Flowcytometric analysis with quantification of fluorescence intensity of red cells labelled with the EMA dye has proven to be a rapid and user-friendly method available to any laboratory with a flowcytometer. The method has a high sensitivity and specificity and can be recommended as a diagnostic tool for HS.

The Ph-negative chronic myeloproliferative disorders (CMPDs) polycythaemia vera, essential thrombocytosis and idiopathic myelofibrosis are acquired stem cell disorders, which pathophysiologically are featured by clonal myeloproliferation and accumulation of myeloid cells, the latter being consequent to decreased apoptosis. Myelofibrosis and neoangiogenesis in the bone marrow and spleen are the histopathological hallmarks of idiopathic myelofibrosis but may develop in the other diseases as well. In patients with myelofibrosis elevated levels of circulating CD34+ cells are highly characteristic being partly explained by a proteolytic bone marrow mileu owing to excessive release of various proteases with ensuing extracellular matrix degradation and constitutive mobilisation of CD34+ cells into the peripheral blood. Thrombohaemorrhagic complications are major clinical problems contributing significantly to morbidity and mortality. Based upon in vitro and in vivo studies of the effects of statins (antithrombotic, antiproliferative, antiangiogenic, antiproteolytic) and zoledronic acid (antiproliferative antiproliferative, antiangiogenic, antiproteolytic) this review focusses on the translation of these effects into potential clinical benefits of combinational therapy with these agents in patients with CMPDs.

Thrombohaemorrhagic complications are major clinical problems in the classical chronic Ph-negative myeloproliferative disorders (CMPDs), polycytaemia vera (PV), essential thrombocythaemia (ET) and idiopathic myelofibrosis (IMF), contributing significantly to morbidity and mortality. Pathophysiologically these disorders are characterized by clonal myeloproliferation, myeloaccumulation and a propensity to develop myelofibrosis and neoangiogenesis in both the bone marrow and spleen. Based upon in vitro and in vivo studies of the effects of statins (antithrombotic, antiproliferative, proapoptotic and antiangiogenic), this review focuses on the translation of these effects into potential clinical benefits of statin therapy in patients with CMPDs.

Purpose: The present investigation aimed to identify factors that predict reduced visual acuity in keratoconus from a prospective, longitudinal study., Methods: This report from the Collaborative Longitudinal Evaluation of Keratoconus (CLEK) Study used 7 years of follow-up data from 953 CLEK subjects who did not have penetrating keratoplasty in either eye at baseline and who provided enough data to compute the slope of the change over time in high- or low-contrast best-corrected visual acuity (BCVA). Outcome measures included these slopes and whether the number of letters correctly read decreased by 10 letters or more in at least one eye in 7 years., Results: Mean age of the subjects at the first follow-up visit was 40.2 +/- 11.0 years (mean +/- SD). Overall, 44.4% were female, and 71.9% were white. The slope of the change in high- and low-contrast BCVA (-0.29 +/- 1.5 and -0.58 +/- 1.7 letters correct/year, respectively) translated into expected 7-year decreases of 2.03 high- and 4.06 low-contrast letters correct. High- and low-contrast visual acuity decreases of 10 or more letters correct occurred in 19.0% and 30.8% of subjects, respectively. Independent predictors of reduced high- and low-contrast BCVA included better baseline acuity, steeper first definite apical clearance lens (FDACL), and fundus abnormalities. Each diopter of steeper baseline FDACL predicted an increased deterioration of 0.49 high- and 0.63 low-contrast letters correct., Conclusions: CLEK Study subjects with keratoconus exhibited a slow but clear decrease in BCVA during follow-up, with low-contrast acuity deteriorating more rapidly than high-contrast. Better baseline BCVA, steeper FDACL, and fundus abnormalities were predictive of greater acuity loss with time.