NIHRDataInsights/HRCSData huggingface dataset:
Title: Endothelial cell biology and the initiation of inflammation
Funder: Medical Research Council

Abstract: At sites of inflammation, the surface of the endothelial cells lining blood vessels changes to recruit leukocytes. The regulated appearance of a series of molecules which interact with receptors on leukocytes is critical for the initiation of inflammation. The first membrane protein to appear is P-selectin. The disappearance of P-selectin is equally tightly controlled to avoid inappropriately prolonged inflammation. P-selectin is stored within the membrane of the Weibel-Palade body (WPB) within cells so that using regulated exocytosis of the WPBs, it can be placed on the plasma membrane within a few minutes. After its appearance at the cell surface, P-selectin is internalised and 50% then recycles back to WPBs for re-use, while 50% is targeted to the lysosomes for degradation. The functioning of P-selectin thus depends on a series of membrane trafficking events that control its itinerary. Further, the molecular machinery controlling its trafficking integrates P-selectin functioning with the inflammatory state of the cell by responding to physiologically generated signals. We are characterising the machinery controlling the itinerary of P-selectin within primary human endothelial cells obtained from the umbilical cord (HUVECS). We are determining the sequences within P-selectin that target this protein to the WPBs, that control its internalisation in endothelial cells, and that lead to its delivery to lysosomes or to its recycling. We are screening for proteins that interact with these targeting motifs by genetic as well as biochemical methods in addition to employing a candidate approach in identifying key players that control the behaviour of P-selectin. The response of this machinery to inflammatory signalling will be determined. Moreover, in addition to our analyses at the tissue culture level, we are collaborating with colleagues in Sheffield to analyse the effects of defects in the trafficking machinery on the ability of the endothelium to recruit leukocytes using intravital microscopy in mice. P-selectin storage depends on the formation of the WPB, which in turn depend on the haemostatic protein von Willebrands Factor (VWF). We are examining how VWF drives the formation of the WPBs, and in turn how that is affected in Von Willebrands disease- the commonest genetic haemostatic disorder, as well as in other inherited human bleeding disorders. These experiments use siRNA-mediated knock-down of disease genes followed by analyses in vitro using HUVECs of WPB formation and function. Our work on the WPB is revealing which defects in WPB formation might lead to haemostatic problems, and we collaborate with clinical colleagues at Hammersmith and Sheffield in the clinical interpretation of this data.


Research activity codes: 1.1 NORMAL BIOLOGICAL DEVELOPMENT AND FUNCTIONING;2.1 BIOLOGICAL AND ENDOGENOUS FACTORS
Health categories: GENERIC HEALTH RELEVANCE;BLOOD
Title: Defining the transcriptome of the first functional haematopoietic stem cells in the mouse embryo.
Funder: Wellcome Trust

Abstract: Despite clinical demand for hematopoietic stem cell (HSC) transplantations for patients with blood cancers or hematologic diseases, little progress has been achieved in in vitro generation or expansion of HSCs. Cellular reprogramming by overexpression of critical transcription factors has been impeded by our incomplete understanding of how HSCs develop in the embryo. The very first HSCs are generated from endothelial cells in the aorta-gonad-mesonephros region in the embryo, with only a few functional stem cells hidden among other blood progenitors. Separating true HSCs from other blood progenitor cells in the embryo has proven challenging because they have similar surface protein expression. Moreover, testing function and then sequencing cells is impossible and HSCs differentiate and alter their transcriptome in expansion. Therefore, new methods of identifying functional HSCs are needed to advance our understanding of these clinically relevant cells. I aim to develop a model that predicts HSC function based on the precise levels of gene expression. I will then test the predictions of the model and determine if narrowing the ranges of gene expression can distinguish HSCs from progenitor cells. Accurate predictions will inform future studies into in vitro generation and expansion as precise required gene expression levels will be uncovered.


Research activity codes: 1.1 NORMAL BIOLOGICAL DEVELOPMENT AND FUNCTIONING
Health categories: GENERIC HEALTH RELEVANCE;BLOOD;CARDIOVASCULAR;INFLAMMATORY AND IMMUNE SYSTEM
Title: Role of the bone marrow niche in regulating the haematopoietic stem cell response to infection
Funder: Wellcome Trust

Abstract: During infections, such as malaria, which is caused by plasmodium parasites, platelets and red blood cells are destroyed. Hematopoietic stem cells, which are responsible for the production of all the cell types of the blood system, are then put under stress to produce high numbers of platelets and red blood cells. Hematopoietic stem cells are found in the bone marrow, surrounded by many types of other cells. This project aims to investigate how only a specific subset of stem cells becomes activated to replenish the platelet cell counts. I will study the expression and function of specific genes in cells of the environment of stem cells and in stem cells themselves using microscopy-based assays on tissue slices and genetically edited cells implanted into mice. I will use intravital microscopy to study behaviours of stem cells such as cell division and movement, in the bone marrow in the skull. The results of this project could lead to the development of improved treatments for malaria patients and patients with bone marrow transplants.


Research activity codes: 1.1 NORMAL BIOLOGICAL DEVELOPMENT AND FUNCTIONING
Health categories: GENERIC HEALTH RELEVANCE;BLOOD;INFECTION;INFLAMMATORY AND IMMUNE SYSTEM
Title: Role of STK4/MST1 signalling in radiation-mediated G1 checkpoint activation and radiation-resistance
Funder: Worldwide Cancer Research

Abstract: <p>When a cell&rsquo;s DNA gets damaged an emergency signalling programme unfolds that blocks cell cycle progression and instigates damage repair. Genotoxic insult blocks cell cycle passage at multiple positions of the cell cycle and checkpoint activation is recognised as a key event protecting cells from death following DNA damage (DD). In doing so checkpoint activation can adversely influence the response of cancer cells to genotoxic anticancer treatment, including radiotherapy and understanding of signalling involved in checkpoint activation could uncover therapeutic opportunities to improve response. Although the candidate executors for different checkpoints have been identified there is considerable lack of knowledge as to signal transduction required to initiated, support, sustained and terminate signalling in the context of damage repair.</p> <p>In work preceding this application we performed a kinome-spanning siRNA screen to uncover signalling components required for radiation-mediated retinoblastoma protein activation, the recognized executor of radiation-damage-induced G1 checkpoint control. This work identified a group of hitherto unsuspected signalling components including the serine-threonine kinase 4/mammalian sterile-20 like kinase 1 (STK4/MST1), alongside several functions known to be functionally linked to signalling through this kinase, strongly implying that STK4-signalling is involved in the execution of radiation-mediated G1 checkpoint activation.&nbsp; We also observed that knockdown of STK4 reduces radiation survival of cell where G2 cell cycle checkpoint control is experimentally compromised, in keeping with the notion that G1 checkpoint functioning is critical for survival of damaged cells with lesions in other checkpoints.</p> <p>The proposed work seeks to corroborate the hypothesis that STK4/MST1 facilitates G1 radiation checkpoint activation and to clarify how in detail this is achieved. The work proposed should enhance our fundamental understanding of genotoxicity-mediated responses in cells and may yield identification and validation of medicinal targets suitable for the enhancement of response to anticancer radiotherapy.</p>


Research activity codes: 1.1 NORMAL BIOLOGICAL DEVELOPMENT AND FUNCTIONING
Health categories: GENERIC HEALTH RELEVANCE;CANCER AND NEOPLASMS
Title: Novel factors regulating replication origin assembly in embryonic, somatic and cancer cells
Funder: Worldwide Cancer Research

Abstract: <p>The correct duplication of genetic information is essential to maintain genome stability, which is lost in cancer cells. DNA replication begins from replication origins. Cancer cells have a higher number of replication origins that can confer high proliferative potential. Reducing the number of available origins might represent a valid strategy to kill cancer cells. However, the molecular mechanisms regulating replication origin organization in cancer cells are unknown. Cellular transformation and tumour progression are thought to be a regression to the early stages of embryonic development. The Xenopus laevis egg cell free extracts constitute a unique system to study embryonic DNA replication and origin assembly. During early embryonic development of Xenopus laevis complete genome replication occurs faster than in somatic cells and replication origins are set at high density with short regular intervals. Somatic cell DNA replication pattern, which is characterized by fewer origins can be converted to an embryonic one more similar to cancer cells during somatic nuclei reprogramming. This process takes place following incubation of somatic nuclei in egg extract. Recently, we identified SSRP1, a subunit of the heterodimeric FACT chromatin- remodelling complex as main player in the control of replication origin assembly. We demonstrated that SSRP1 protein promotes somatic replication origin reprogramming when added to egg extract. We now propose to identify the molecular mechanism underlying this process. In particular we will study SSRP1&rsquo;s role in replication origins assembly. We will explore the possible direct link between the replication origin recognition complex (ORC) and SSRP1 and we will identify the chromatin modifications induced by SSRP1 leading to replication origin assembly. These studies will lead to a better understanding of replication origin formation and control in normal and cancer cells.</p>


Research activity codes: 1.1 NORMAL BIOLOGICAL DEVELOPMENT AND FUNCTIONING;2.1 BIOLOGICAL AND ENDOGENOUS FACTORS
Health categories: GENERIC HEALTH RELEVANCE;CANCER AND NEOPLASMS
Title: EZH2 dependent inhibition of p53 tumour suppressor function by the alternative NF-κB pathway
Funder: Worldwide Cancer Research

Abstract: The NF-κB family of transcription factors are known as regulators of the immune and inflammatory responses but are also activated by cellular stress and respond to many of the same inducers as the p53 tumour suppressor. Aberrantly active NF-κB is associated with many types of cancer. There are two major pathways leading to induction of NF-κB subunits. The classical pathway typically leads to the induction of RelA or c-Rel containing complexes and is dependent on IkB kinase (IKK)β. The alternative pathway, involves the inducible processing of p100 to p52, leading to the induction of p52/RelB complexes and is dependent on IKKα. We have discovered a previously unknown regulatory mechanism through which the alternative NF-κB pathway can suppress p53 dependent senescence. A key component of this regulation is the ability of p52/RelB to prevent p53 mediated repression of the polycomb protein EZH2: many of the effects of p52/RelB on p53 regulated gene expression, particularly a large subset of p53 repressed genes, can be explained by the loss of EZH2 expression seen upon siRNA depletion of the NF-κB subunits. We propose that this pathway provides a mechanism through which the alternative NF-κB pathway, whether activated due to cell stresses/chronic inflammatory conditions or due to genetic mutations in cancer cells, can act to suppress the p53 tumour suppressor programme and, in particular, cell senescence. Here we will: (1) Determine the mechanism through which p52/RelB prevents repression of EZH2. (2) Determine the mechanism through which EZH2 regulates a subset of critical p53 target genes and evaluate the role of p52/RelB at these promoters (3) Investigate the effect of targeting the alternative NF-κB pathway with novel IKKα inhibitors in cancer cell lines and cells from CLL patient cells. (4) Investigate the role of other identified, potential regulators of NF-κB and p53 crosstalk


Research activity codes: 1.1 NORMAL BIOLOGICAL DEVELOPMENT AND FUNCTIONING;5.1 PHARMACEUTICALS
Health categories: GENERIC HEALTH RELEVANCE;CANCER AND NEOPLASMS
Title: Activation mechanism of the MLL methyltransferase complex
Funder: Worldwide Cancer Research

Abstract: We wish to establish how interaction with its associated multi-protein complex stimulates the catalytic activity of the mixed lineage leukaemia (MLL1) enzyme. This activity, histone H3K4 methylation, is part of the epigenetic regulatory network that sets the program for gene activation and is particularly critical during the early stages of development. Aberrant epigenetic signalling is linked to disease and mutations that lead to disruption of MLL1 have been specifically associated with aggressive childhood leukaemia. Our preliminary characterisation of the complex indicates that the complex substituent proteins WDR5, RbBP5 Ash2L and Dpy30 act cooperatively to stimulate the activity by inducing a conformational change in the MLL1 SET domain. By combining techniques in biochemistry, biophysics, electron microscopy and X-ray crystallography we will determine the molecular details of this activation process. A critical advance in our preliminary work that underpins this proposal has been the successful purification of the intact mammalian complex co-expressed in insect cells. Our initial characterisation of this purified recombinant complex has confirmed that the components are stoichiometric, the complex is active, behaves well in biophysical analysis and is suitable for EM studies. We are now seeking funding to take these studies further in order to describe the subunit contacts that stimulate methyltransferase activity. We will locate the position of the protein subunits within the molecular envelope defined by EM studies, identify the key events that lead to optimal activity and optimise the recombinant constructs in order to produce a high resolution X-Ray structure. Ultimately this work will provide mechanistic insights into the regulation of one of the major epigenetic activators of gene transcription. In addition to moving forward the molecular epigenetic field we believe that this work can be the foundation for future inhibitor studies that target regulatory protein:protein interactions.


Research activity codes: 1.1 NORMAL BIOLOGICAL DEVELOPMENT AND FUNCTIONING
Health categories: GENERIC HEALTH RELEVANCE;CANCER AND NEOPLASMS
Title: TNF-receptor trafficking and the regulation of inflammation, cell death and cancer
Funder: Worldwide Cancer Research

Abstract: Deregulated cell death and inflammation can drive chronic remodelling and tissue repair, which can contribute to cancer and treatment failure. Mediators and modulators of the cell death and inflammatory signalling cascades play decisive roles at different stages of tumour development, and likely contribute to the biological basis for the individual variation in clinical behaviour. The principle idea addressed by this application is that cell survival and adaptation mechanisms of tumours are supported, at least in part, by cancer-related inflammation, and that this can be successfully targeted by switching the response from pro-inflammation to cell death, causing tumours to permanently regress. Despite clear evidence that cell death and inflammation are linked, the mechanisms that can switch between the distinct biological outcomes remain elusive. This is important as its resolution, and putative therapeutic intervention, would allow the diversion of cancer-related inflammation into activation of cell death. Our starting point for this proposal is substantial new data on the regulation of cell death and inflammation. We have identified two new regulators of cell death and inflammation, which when depleted turn pro-inflammatory signalling pathways into cytotoxic pathways. Thus, these newly identified proteins dictate whether proinflammatory signals induce cell survival or death. We propose to use transplantation-based cancer models, patient-derived xenografts as well as single-molecule super-resolution imaging, quantitative mass spectrometry and biochemical in vitro approaches to decipher how these proteins regulate survival/death signalling, and how this relates to tumour formation and metastatic success. Given that inflammatory and cell death signal transduction pathways play important roles in diverse physiological and pathological processes at the core of human diseases, our findings will provide new avenues for therapeutic intervention strategies.


Research activity codes: 1.1 NORMAL BIOLOGICAL DEVELOPMENT AND FUNCTIONING;2.1 BIOLOGICAL AND ENDOGENOUS FACTORS
Health categories: GENERIC HEALTH RELEVANCE;CANCER AND NEOPLASMS
Title: How do cells shape and interpret PIP3 signals?
Funder: Biotechnology and Biological Sciences Research Council

Abstract: This proposal is a collaboration between biochemistry groups at BI and mathematical biologists at the EBI to achieve a detailed and quantitative understanding of a major mammalian signal transduction pathway, the PI3K network. Several PI3K isoforms exist in cells that can be selectively engaged by a variety of cell surface receptors to generate the membrane phospholipid PIP3. PIP3 is the initial signal, which is then transduced by 10-50 effector proteins into the regulation of complex cell responses, such as cell growth and movement. Our strategy is to focus on collecting robust, high quality data sets in a panel of isogenic, non-transformed breast cell lines (MCF10a) in which key endogenous components of the pathway can be manipulated and to embed iteration between experiment and modelling to arrive at a more satisfactory explanation of: 1) the key factors which shape the magnitude and spatiotemporal properties of PIP3 signals in response to hormonal stimulation (EGF, insulin, LPA) and oncogenic mutation; 2) The way in which different PIP3 effectors interpret these PIP3 signals and 3) the relative importance of individual PIP3 effectors in delivering regulation of chemokinesis, growth and global transcription. We plan to use homologous gene targetting, siRNA suppression and pharmacological inhibition of pathway components and measure the impact of these perturbations, in several relevant cellular contexts, on i) the levels of PIP3 and other phosphoinositides measured by a novel, quantitative mass spectrometry assay that allows systematic analysis of fatty acid composition; ii) the activity and spatial distribution of several PIP3 effectors (in some cases via knock-in of endogenous GFP-fusion proteins); iii) chemokinesis, markers of growth and global transcription (using next generation sequencing). Models will be built at several levels in the pathway and integrated to allow a deeper understanding of this network and guide more effective therapeutic intervention


Research activity codes: 1.1 NORMAL BIOLOGICAL DEVELOPMENT AND FUNCTIONING;2.1 BIOLOGICAL AND ENDOGENOUS FACTORS
Health categories: GENERIC HEALTH RELEVANCE;CANCER AND NEOPLASMS
Title: Activation of the heat shock response by sulfhydryl-reactive chemoprotective agents
Funder: Biotechnology and Biological Sciences Research Council

Abstract: Context: The heat shock response and the phase 2 response represent two highly conserved mechanisms that determine the ability of cells and organisms to adapt and survive under conditions of stress. Both pathways do not normally operate at maximal capacity, but can be induced by small-molecule chemoprotective agents. Induction results in enhanced transcription of a battery of cytoprotective genes under the control of transcription factors heat shock factor 1 (HSF1) and nuclear factor-erythroid 2-related factor 2 (Nrf2). Small molecule activators of Nrf2 have a common chemical signature, that of sulfhydryl reactivity. They react with cysteine residues of the sensor protein Keap1, leading to Nrf2 activation. Emerging evidence suggests that sulfhydryl-reactive molecules also induce the heat shock response. Anti-inflammatory activity is a consistent property of Nrf2 activators, indicating that suppression of inflammation is a central aspect of their chemoprotective actions. However, this property is only partially Nrf2-dependent. Critically, HSF1 activation has anti-inflammatory effects. Aims and Objectives: To test the hypothesis that the cytoprotective and anti-inflammatory activities of sulfhydryl-reactive chemoprotective agents are mediated by activation of Nrf2 and HSF1, through a common chemical signal, that of sulfhydryl reactivity. To establish whether activation of HSF1 is a common property of sulfhydryl-reactive chemoprotective agents. To elucidate the importance of sulfhydryl reactivity in activation of HSF1. To define the individual roles of HSF1 and Nrf2 in protection against pro-inflammatory stimuli. Potential applications and benefits: The proposed studies will establish whether activation of HSF1 mediates the Nrf2-independent portion of the anti-inflammatory activity of sulfhydryl-reactive inducers and will provide essential mechanistic understanding for the development of such molecules as chemoprotective agents to improve human health.


Research activity codes: 1.1 NORMAL BIOLOGICAL DEVELOPMENT AND FUNCTIONING;2.1 BIOLOGICAL AND ENDOGENOUS FACTORS
Health categories: GENERIC HEALTH RELEVANCE;CANCER AND NEOPLASMS
Title: Family carers’ perceptions of their educational needs when providing end-of-life care: a systematic review of qualitative research
Funder: Marie Curie

Abstract: Family carers play a significant and important role in ensuring that people with cancer and other life threatening diseases can be cared for in their own home. Carers often need to develop new skills such as: how to provide physical care, how to manage symptoms such as pain or breathlessness, in addition to maintaining a balance between their own needs and those of the person they are caring for in order to remain well. Carers also need to cope with the idea that their caring role will end with the death of the person they look after. Several different ways of helping carers with their caring role have been devised. These have been shown to enhance their ability to continue their role and reduce their distress. No one has drawn this literature together to understand what it says about developing education interventions, which family carers would find helpful and supportive. A particular type of research called qualitative research – which is concerned with how to make sense of what is happening to them - can help in understanding what carers find important. Bringing research together systematically helps to develop a more complete picture than looking at individual studies. This piece of research will draw together the qualitative research exploring carers’ views about how they are taught to care for people with cancer and other life threatening diseases. Doing this will help develop a more full and extensive understanding of these views, enabling improvements to be made in the way education for carers is planned and taught in the future. This in turn, will ensure the most appropriate care is available for the patient who is ill, in addition to ensuring policy and practice can best support the needs of all members of the family involved in end of life care.


Research activity codes: 7.1 INDIVIDUAL CARE NEEDS;7.2 END OF LIFE CARE
Health categories: GENERIC HEALTH RELEVANCE;CANCER AND NEOPLASMS