Investigating the roles of NFATC2 in acute myeloid leukaemia

Acute myeloid leukaemia (AML) is a malignancy affecting the myeloid lineage of haemopoietic cells and has poor patient outcomes. Chemotherapy is poorly tolerated in a high proportion of patients and targeted therapies have limited impact at present. Additionally, relapse and/or resistance to existing therapy are common. High (cyto)genetic heterogeneity within and between AML patients make the development of effective, long-term targeted therapies highly challenging. As such, there is an unmet clinical need for the identification of novel oncogenic mechanisms and pathways which can be targeted therapeutically, in order to improve the clinical outcomes for larger groups of AML patients. Previously, the histone lysine demethylase KDM4A was identified as a putative ‘master regulator’ of leukaemogenic signalling in models of AML, which was not essential for healthy haemopoietic cell survival. One of the key targets of KDM4A identified in AML was the nuclear factor of activated T cells (cytoplasmic) 2 (NFATC2). The wider NFAT family of transcription factors (TFs) has been attributed roles in normal myelopoiesis, in which it is thought to regulate elements of the cell cycle and differentiation, which are key processes that become deregulated in AML. NFATs are also characterised as contributing to oncogenesis and drug resistance in myeloid leukaemias, however much of the evidence focuses on NFATC1. In light of the identification of the KDM4A-NFATC2 axis in AML, it was hypothesised that NFATC2 is essential for the oncogenic function and survival of AML cells. Prior to testing this hypothesis, NFATC2 was first characterised in cell line models of AML. Using established compounds, it was found that the MLL-AF9 TP53mut THP-1 AML cell line was sensitive to depletion/inhibition of calcium and calcineurin, which are both upstream regulators of NFATs in T cells. Next, shRNA knockdown (KD) of NFATC2 led to the loss of colony-forming capacity in a number of AML cell lines, highlighting that numerous subtypes of AML cells are dependent on NFATC2. In addition, increased apoptosis and cell cycle arrest were individually observed in some of these AML models, but not others, indicating that the mechanisms affected by NFATC2 depletion are dependent on the (cyto)genetic landscape. Global transcriptome profiling of THP-1 cells with NFATC2 KD identified a list of deregulated genes, of which a subset was validated in several other cell lines with NFATC2 KD. These included genes involved in intracellular transport and membrane protein function. Enrichment analyses also highlighted targets of oncogene MYC and serine/threonine kinase 33 (STK33) as enriched in the genes perturbed by NFATC2 depletion. Chromatin immunoprecipitation sequencing (ChIP-Seq) found NFATc2 gene binding targets to be enriched with a c-Myc DNA consensus binding sequence. Additionally, a number of novel NFATc2 DNA binding motifs were identified. The expression of NFATC2 was found to stratify patient outcomes in the TARGET-AML dataset, from paediatric AML patients. Of the genes identified by sequencing analyses as putative NFATC2/NFATc2 targets, the expression levels of 13 genes were found to be prognostic for patients in the TARGET-AML dataset, also. Together these data have shown that NFATC2 is essential for the survival of multiple AML cell lines and that it likely regulates elements of the cell cycle and/or apoptosis, depending on the cellular context. Newly-identified transcriptional and binding targets suggest that the oncogene MYC cooperates with NFATC2 and it could be hypothesised that they maintain an oncogenic transcriptional program together in THP-1 cells. These findings require translation into patient cells, which is challenging given the lack of NFATc2-specific inhibitors available. However, findings from open-source patient datasets indicate that NFATC2 and its targets have a significant role to play in clinical outcome and so warrant further investigation to elucidate some of the cellular mechanisms involved

Transcriptional regulation by the NFAT family in acute myeloid leukaemia

Acute myeloid leukaemia (AML) is a haematological cancer with poor outcomes due to a lack of efficacious targeted therapies. The Nuclear Factor of Activated T Cells (NFAT) family of transcription factors is well characterised as a regulator of the cell cycle and differentiation in the myeloid lineage. Recent evidence has demonstrated that NFAT family members may have roles in regulating AML leukemogenesis and resistance to targeted therapy in myeloid leukaemia. Furthermore, gene expression data from patient samples show that some NFATs are more highly expressed in poorly differentiated AML and after disease relapse, implying that the NFAT family may have roles in specific types of AML. This review outlines the evidence for the role of NFAT in healthy myeloid tissue and explores how NFAT might regulate AML pathogenesis, highlighting the potential to target specific NFAT proteins therapeutically in AML

Forecasting substantial data revisions in the presence of model uncertainty

A recent revision to the preliminary measurement of GDP(E) growth for 2003Q2 caused considerable press attention, provoked a public enquiry and prompted a number of reforms to UK statistical reporting procedures. In this article, we compute the probability of 'substantial revisions' that are greater (in absolute value) than the controversial 2003 revision. The predictive densities are derived from Bayesian model averaging over a wide set of forecasting models including linear, structural break and regime-switching models with and without heteroscedasticity. Ignoring the nonlinearities and model uncertainty yields misleading predictives and obscures recent improvements in the quality of preliminary UK macroeconomic measurements

Sequential Infection with Influenza A Virus Followed by Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) Leads to More Severe Disease and Encephalitis in a Mouse Model of COVID-19

COVID-19 is a spectrum of clinical symptoms in humans caused by infection with SARS-CoV-2. The coalescence of SARS-CoV-2 with seasonal respiratory viruses, particularly influenza viruses, is a global health concern. To understand this, transgenic mice expressing the human ACE2 receptor (K18-hACE2) were infected with influenza A virus (IAV) followed by SARS-CoV-2 and the host response and effect on virus biology was compared to K18-hACE2 mice infected with IAV or SARS-CoV-2 alone. The sequentially infected mice showed reduced SARS-CoV-2 RNA synthesis, yet exhibited more rapid weight loss, more severe lung damage and a prolongation of the innate response compared to the singly infected or control mice. Sequential infection also exacerbated the extrapulmonary encephalitic manifestations associated with SARS-CoV-2 infection. Conversely, prior infection with a commercially available, multivalent live-attenuated influenza vaccine (Fluenz Tetra) elicited the same reduction in SARS-CoV-2 RNA synthesis, albeit without the associated increase in disease severity. This suggests that the innate immune response stimulated by IAV inhibits SARS-CoV-2. Interestingly, infection with an attenuated, apathogenic influenza vaccine does not result in an aberrant immune response and enhanced disease severity. Taken together, the data suggest coinfection (‘twinfection’) is deleterious and mitigation steps should be instituted as part of the comprehensive public health and management strategy of COVID-19

SARS-CoV-2 infects an upper airway model derived from induced pluripotent stem cells

As one of the primary points of entry of xenobiotic substances and infectious agents into the body, the lungs are subject to a range of dysfunctions and diseases that together account for a significant number of patient deaths. In view of this, there is an outstanding need for in vitro systems in which to assess the impact of both infectious agents and xenobiotic substances of the lungs. To address this issue, we have developed a protocol to generate airway epithelial basal-like cells from induced pluripotent stem cells, which simplifies the manufacture of cellular models of the human upper airways. Basal-like cells generated in this study were cultured on transwell inserts to allow formation of a confluent monolayer and then exposed to an air-liquid interface to induce differentiation into a pseudostratified epithelial construct with a marked similarity to the upper airway epithelium in vivo. These constructs contain the component cell types required of an epithelial model system, produce mucus and functional cilia, and can support SARS-CoV-2 infection/replication and the secretion of cytokines in a manner similar to that of in vivo airways. This method offers a readily accessible and highly scalable protocol for the manufacture of upper airway models that could find applications in development of therapies for respiratory viral infections and the assessment of drug toxicity on the human lungs

The Golden Rule:Interfaith Peacemaking and the Charter for Compassion

The Charter for Compassion has been signed by over two million people from around the world and partnered with hundreds of interfaith organizations and cities seeking to put into practice the Golden Rule, common to the main faith traditions, of doing unto others as you would be done by. This article sets the Charter within the context of a post secular international society and faith-based diplomacy, in which religious interreligious initiatives emerge as serious, rather than peripheral, actors in developing sustainable peace making through bottom-up approaches. The article critically engages with the Charter's claim that ‘any interpretation of scripture that breeds violence, hatred or disdain is illegitimate’ while accepting that peaceful interpretations of scriptures are helpful to peace processes where religious actors are involved. The article explores the claims of the Charter for Compassion International as they seek to make peace through compassion, before concluding that the Charter for Compassion is a long-term project aimed at changing hearts and minds but has had limited substantive impact to date

Remdesivir-ivermectin combination displays synergistic interaction with improved in vitro activity against SARS-CoV-2

A key element for the prevention and management of COVID-19 is the development of effective therapeutics. Drug combination strategies of repurposed drugs offer several advantages over monotherapies, including the potential to achieve greater efficacy, the potential to increase the therapeutic index of drugs and the potential to reduce the emergence of drug resistance. Here, we report on the in vitro synergistic interaction between two FDA approved drugs, remdesivir and ivermectin resulting in enhanced antiviral activity against SARS-CoV-2. Whilst the in vitro synergistic activity reported here does not support the clinical application of this combination treatment strategy, due to insufficient exposure of ivermectin in vivo, the data do warrant further investigation. Efforts to define the mechanisms underpinning the observed synergistic action, could lead to the development of novel therapeutic treatment strategies