NIHRDataInsights/HRCSData huggingface dataset:
Title: PRH/HHEX in intrahepatic cholangiocarcinoma, disease progression and response to chemotherapeutics.
Funder: Medical Research Council

Abstract: Our central hypothesis is that depletion of PRH protein or PRH inactivation by increased protein kinase CK2-dependent phosphorylation is associated with ICC and progression to ICC and that changes in cell behaviour brought about by the loss of PRH activity are of critical importance in the response of ICC to chemotherapeutics. Moreover, we propose that restoration of PRH through pharmacological interventions that decrease PRH inactivation through phosphorylation such as the CK2 inhibitor CX-4945, will be of value in the treatment of ICC patients with elevated levels of phosphorylated PRH. We will use immunohistochemistry to test our hypothesis that PRH levels and/or phosphorylated PRH levels are associated with ICC and to determine whether they are an indicator of poor prognosis in primary sclerosing cholangitis. We will also determine PRH and pPRH protein levels in ICC cell lines and knockdown PRH in these cells using lentiviral constructs expressing PRH shRNA. This will allow us to determine whether PRH knockdown increases the sensitivity of these cells to Cisplatin and the role that PRH plays in mediating the effects of the CX-4945. We will determine whether increased PRH activity in ICC cells results in decreased tumour growth and metastasis in xenograft mouse models. To achieve this we will implant luciferase-labelled PRH over-expressing ICC cells in nude mice and measure tumour growth by repeated in vivo imaging. To examine changes in metastasis we will inject luciferase-labelled PRH knockdown and PRH over-expression ICC cells intravenously into nude mice and monitor the appearance of metastases in internal organs (lymph nodes, lungs, and liver). Finally, we will identify PRH target genes in cholangiocytes and compare the expression of these target genes in cholangiocytes and ICC cells using a genome-wide microarray screen. We will then examine whether the products of these genes are potential serum biomarkers for early diagnosis of ICC.


Research activity codes: 4.1 DISCOVERY AND PRECLINICAL TESTING OF MARKERS AND TECHNOLOGIES
Health categories: CANCER AND NEOPLASMS
Title: Development of Metabolism Radiotracers to Probe Disease Pathology in Human Subjects with Cancer
Funder: Medical Research Council

Abstract: Our group is interested in discovery and development of novel radiotracers for probing disease biology. In this quinquenium, we propose a translational programme that aims to develop the next generation of metabolism imaging tracers for investigation of disease biology in humans. Imaging using positron emission tomography (PET) remains one of the most direct ways of interrogating molecular mechanisms derailed in many diseases including cancer. PET methods are non-destructive and allow tissue biology to be investigated in the species of interest - the human being - non-invasively without associated sampling errors or tissue alterations that occur with biopsy-based approaches. Availability of appropriate probes and their validation in humans remains the major bottleneck in this field of research. Using cancer as the model, we have over the past five years advanced new probes for PET into human imaging to allow associated molecular mechanisms to be investigated. Of current interest is the reprogrammed tumour metabolism - the result of multiple signalling pathways - which is difficult to measure, as the probes require bidirectional transit through the plasma membrane of cells within the target tissue and localisation within the cell only when the specific pathology exists. In the coming 5 years we aim to complete biological validation of a choline kinase radiotracer recently transitioned into humans, as well as introduce two new imaging probes discovered from our preclinical programme for assessing glycogenesis and fatty acid oxidation. The connectivity of this programme - chemistry design, automation, regulatory, mathematical modelling, and clinical imaging-pathology correlative science - will allow us to provide the tools for translating our post-genome understanding of reprogrammed tumour metabolism into scientific investigation of pathology in situ in humans, while developing candidate probes with potential for managing patients.


Research activity codes: 4.1 DISCOVERY AND PRECLINICAL TESTING OF MARKERS AND TECHNOLOGIES
Health categories: CANCER AND NEOPLASMS
Title: Regulation of the oncogenic potential of signalling through the Ras/ERK pathway by dual-specificity phosphatase 5
Funder: Medical Research Council

Abstract: Tumours in which the Ras/ERK pathway is abnormally activated, such as pancreatic and lung cancers are relatively common, but have few treatment options and a poor prognosis. We recently discovered that DUSP5, a protein phosphatase which "switches off" Ras/ERK signals in the nucleus, plays a key role in preventing tumours in a mouse model of skin cancer, demonstrating that DUSP5 is a bona fide tumour suppressor. We therefore hypothesise that DUSP5 plays an important role in cancer development driven by Ras/ERK signalling in other tissue types in mouse models and in humans. We will test this hypothesis by addressing two main objectives. 1) Firstly, we will establish if DUSP5 influences the genesis and progression of tumours in clinically relevant experimental models of Kras-induced pancreatic and lung cancer. We will analyse both tumours and cells derived from these cancers to determine the molecular mechanisms by which DUSP5 affects the oncogenic potential of Ras/ERK signalling and study the expression of DUSP5 in human tumour samples to establish if the levels of this protein are predictive of disease severity or progression. 2) Our preliminary data comparing the response to DUSP5 loss in cells containing either mutant Ras or mutant Braf suggest that the role of DUSP5 changes depending on the nature of the oncogene that drives cancer and may modify the response of cancer cells to drugs that inhibit the Ras/ERK pathway. Our second objective is therefore to determine how DUSP5 function is altered by Ras or Braf mutation and its implications for mediating drug response and drug resistance. This project will provide new mechanistic insights into the ways that the Ras/ERK signalling pathway is "rewired" in cancer and may reveal new biomarkers which will allow the stratification of patients receiving drugs which inhibit Ras/ERK signalling and reveal potential new strategies to prevent the emergence of drug resistance.


Research activity codes: 4.1 DISCOVERY AND PRECLINICAL TESTING OF MARKERS AND TECHNOLOGIES;2.1 BIOLOGICAL AND ENDOGENOUS FACTORS;2.4 SURVEILLANCE AND DISTRIBUTION
Health categories: CANCER AND NEOPLASMS
Title: Uncovering function and mode of activation of the central Fanconi Anemia FANCD2/FANCI DNA repair protein complex, a potential cancer drug target.
Funder: Medical Research Council

Abstract: Fanconi Anemia (FA) is a genetic disease predisposing to cancer. 18 FA proteins, including BRCA1 and BRCA2, often mutated in breast cancer, compose the FA pathway, which repairs dangerous DNA interstrand crosslinks (ICLs). Mutation in any of these genes can cause FA. Due to the severe cellular toxicity of ICLs, ICL-forming drugs (e.g. cisplatin) are used successfully to treat cancers in non-FA patients. However, patients often develop resistance, which can be caused by an increased ability of the cancer cells to repair ICLs. An inhibitor of the FA pathway would prevent cancer cells from repairing ICLs, thereby sensitizing them to ICL-forming drugs. Two of the central FA proteins, FANCD2 and FANCI, form a complex that is activated by an essential monoubiquitination step. Absence of either of the subunits, or their modification, completely shuts down the pathway, and sensitizes cells to ICL-forming drugs. Despite the importance of this complex in DNA repair, we still do not know what regulates its activation and what its functional activity is in ICL repair. We have recently obtained the structure of full-length human FANCD2/FANCI complex and discovered a novel domain, which we have termed the Tower domain. This domain is essential for function of the complex. We aim to uncover the mechanisms underlying the activation of DNA repair by the FA complex and understand how the Tower domain controls this process. We will undertake a multidisciplinary approach including a) study the structure of the FANCD2/FANCI complex bound to DNA by cryo EM, b) apply various biochemical techniques such as in vitro reconstitution of monoubiquitination c) live-cell imaging to monitor DNA repair in live cells. The information resulting from this research will be valuable for development of drugs targeting the pathway, thereby sensitizing drug-resistant cancers to existing chemotherapeutic drugs, in turn enabling clinicians to treat these currently untreatable patients.


Research activity codes: 2.1 BIOLOGICAL AND ENDOGENOUS FACTORS
Health categories: CANCER AND NEOPLASMS
Title: Defining a protein hydroxylase tumour suppressor pathway
Funder: Medical Research Council

Abstract: Deregulated protein modifications have been widely implicated in the pathogenesis of major human diseases. The discovery and characterisation of novel post-translational modifications (PTM) has opened new avenues of biomedical research and stimulated new therapeutic opportunities. Hydroxylation is an under-studied but emerging PTM. Pioneering work on collagen and oxygen sensing highlighted the potential importance of hydroxylation in physiology and pathology, and related oxygenases have since been implicated in a variety of important disease-associated processes. However, several members of the hydroxylase family are poorly characterised and the extent and functions of protein hydroxylation in disease remain unclear. We previously reported that a nuclear JmjC protein called 'Myc-Induced Nuclear Antigen' (MINA) is a histidine hydroxylase. Our functional studies indicate that MINA has tumour suppressor (TS) activity, consistent with the observation that it is mutated and inactivated in a variety of tumour types, including gastrointestinal cancers. However, a detailed understanding of the extent of MINA deregulation in cancer requires knowledge of other potential mechanisms of inactivation. Furthermore, the downstream consequences of MINA inhibition are not known. Here we propose to address these limitations by characterising the regulation of MINA by an oncogenic kinase and identifying novel cancer-relevant substrates and gene targets involved in MINA-dependent transcriptional control. Importantly, the findings will be extended into an independent cohort of several hundred gastric tumour samples in a combined approach involving targeted resequencing, gene expression analyses and histopathology. The work will improve our understanding of protein hydroxylation and its role in fundamental cellular processes, define a novel tumour suppressor pathway, and highlight new therapeutic opportunities for the treatment of cancer and other diseases.


Research activity codes: 2.1 BIOLOGICAL AND ENDOGENOUS FACTORS
Health categories: CANCER AND NEOPLASMS
Title: Genetic and telomere characteristics of soft tissue sarcomas
Funder: Medical Research Council

Abstract: Despite treatment, 35% of patients with high-grade Soft Tissue Sarcomas (STSs) die within 5 years. Crucially, 50% of STSs recur following surgery. There are no biomarkers of this recurrence, and only limited systemic treatments for patients with metastatic disease. Circulating tumour DNA (ctDNA) characteristics correlate with disease stage in some cancers. If assays were developed to characterise ctDNA in STSs, novel biomarkers may be identified. For a mutation to be a useful biomarker it should ideally be present in all cancer cells. A hallmark of tumour cells is their ability to divide an unlimited number of times - a process facilitated by Telomere Maintenance Mechanisms (TMMs). As TMMs are not utilised by differentiated normal cells, the presence genetic mutations associated with a TMM in ctDNA should act as a stable marker of STS cells. Our primary aim is to investigate the potential of ctDNA to monitor STSs patients. Using a bioinformatic approach and published literature we have developed a custom 'STS hotspot panel' for next-generation semiconductor sequencing. Using this panel we have identified ctDNA in metastatic STS patients for the first time. After assessing Telomerase and Alternative Lengthening of Telomere (ALT) activity in recruited STSs we next plan to develop highly sensitive Taqman assays for use in digital droplet PCR to target appropriate TMM associated mutations. Using these, and our refined STS hotspot panel, we will then track and characterise ctDNA in STS patients throughout follow up. By correlating ctDNA characteristics with outcome we will determine their potential as new biomarkers, which (if validated) may allow clinicians to diagnose STS recurrence earlier, and offer curative treatment more often. Our secondary aim is to establish cell lines from primary STS tissue. After frozen stocks are developed, recovery assessed, and cell line DNA analysed, this would create new resources for those testing novel systemic therapies.


Research activity codes: 4.1 DISCOVERY AND PRECLINICAL TESTING OF MARKERS AND TECHNOLOGIES
Health categories: CANCER AND NEOPLASMS
Title: Tspan6 as a regulator of immune microenvironment in breast cancer
Funder: Medical Research Council

Abstract: The immune cell landscape within tumour microenvironment is controlled by an elaborate network of communications involving chemokines, cytokines and growth factors which are produced by the immune cells themselves, by cancer cells and by other cell types in cancerous tissues (e.g. tumour endothelium, tumour-associated fibroblasts). Whilst specific contribution of various types of tumour-infiltrating T-cells and macrophages to tumour development and host-immune responses is becoming clearer, significantly less is known and understood about the function of tumour-infiltrating B-cells (TIL-B). Whilst studying the immune landscape in breast cancer we discovered that the abundance of TIL-B correlated with the expression of tetraspanin 6 (Tspan6) by tumour cells. Importantly, Tspan6-expressing breast cancer cells stimulated migration of B-lymphocytes in vitro. These observations point towards a novel pathway underlying communication between tumour cells and TIL-B. As the expression of Tspan6 positively correlated with patients survival, we hypothesise that that Tspan6 suppresses breast cancer progression by modulating tumour infiltration of B-cells and, consequently, the immune landscape of tumour tissues. The proposed work aims to investigate the molecular pathways underlying the phenomenon of increased B-cell chemotaxis towards Tspan6-expressing breast cancer cells. We will: -establish the contribution of chemotactic factors differentially produced by Tspan6-expressing breast cancer cells. -delineate Tspan6-dependent pathways in breast cancer cells that control infiltration of B-cells to cancerous tissues. -examine the B-cell profile in breast cancer tissues in relation to expression of Tspan6. -evaluate the role of Tspan6-dependent recruitment of B-cells to cancerous tissues in vivo.


Research activity codes: 4.1 DISCOVERY AND PRECLINICAL TESTING OF MARKERS AND TECHNOLOGIES
Health categories: CANCER AND NEOPLASMS
Title: Examining the relationship between KRAS mutation and immunotherapy resistance in colorectal cancer
Funder: Medical Research Council

Abstract: Aim: To examine the role of KRAS mutation in driving resistance to immunotherapy in colorectal cancer (CRC) and identify successful immunotherapy combinations for the treatment of CRC. Objectives: 1. Characterise the immune infiltrate associated with KRAS mutation in syngeneic organoid and autochthonous mouse models, and human CRC. 2. Explore the factors regulating the variations in immune infiltrate between these mouse models of CRC. 3. Modulate the immune infiltrate in the above organoid models by novel combinations of immunotherapeutics (focussing on CXCR2 inhibition with cytokine inhibitors/checkpoint inhibitors) and determine the effects on organoid tumour growth and survival. 4. Take the best combination into autochthonous mouse models of CRC Methodology: To achieve this we will utilise a human TMA to characterise the immune infiltrate in human CRC in relation to RAS mutational status. We will further explore this relationship and factors controlling the immune composition with experiments in murine models of CRC of appropriate genetic backgrounds. Immune infiltrates will be characterised in all models via FACs, immunofluorescence, immunohistochemistry, RNAscope in situ hybridisation, and cytokine array analysis. Following these analyses, we will use guided combinations of immunotherapy agents in the same mouse models. Scientific and medical opportunities: Immunotherapy is rapidly becoming a powerful tool for the treatment of human cancer. One central unanswered question is which patients are most likely to respond. KRAS mutation is strongly immunosuppressive in human CRC. Our preliminary data suggest that Kras mutant syngeneic murine organoid transplantation models exhibit a similar immunosuppressive phenotype. Therefore we will examine the role mutant Kras plays in the resistance of CRC to immunotherapy agents, which in turn will reveal novel immunotherapy combinations vitally able to target previously resistant patients.


Research activity codes: 4.1 DISCOVERY AND PRECLINICAL TESTING OF MARKERS AND TECHNOLOGIES
Health categories: CANCER AND NEOPLASMS
Title: Mechanistic investigations of transcriptional regulation by bZIP transcription factors
Funder: Medical Research Council

Abstract: A significant proportion of eukaryotic proteins contain long disordered regions, but are still functional within the cell. The particular over-representation of protein disorder within transcription factors suggests an important biological role for their disorder. How functionally important protein disorder is for these critical regulatory proteins, why, and through what mechanism(s)? Several hypotheses based upon bioinformatics observations and theoretical considerations have been put forward; but most have not been tested experimentally. This project combines computational, structural, biophysical and cellular studies of a highly disordered member of the basic leucine zipper family of transcription factors (bZIPs) to address this fundamental question. bZIPs are highly disordered and contain several modules that fold into alpha-helices upon binding to their partner macromolecules (DNA, other bZIP proteins, co-activator proteins) during assembly of the transcriptional apparatus. I will uncover and quantify the roles of protein disorder within the well biochemically characterised mammalian bZIP CREB. Most experimental studies focus on binding affinities of transcription factors for their isolated cognate sites, however there are reasons to think that being disordered/flexible may be advantageous in kinetic terms within the nucleus (a crowded environment with billions of base pairs of DNA). My experiments will test this hypothesis by measuring rates for 3D diffusion, 1D sliding, dissociation and intersegmental transfer separately for a characterized library of mutant transcription factors with known disorder contents. I will also quantify disorder-mediated allosteric communication between CREB modules to further understanding of transcriptional assembly. Together these mechanistic insights will be used to design peptidomimetic inhibitors of CREB, which are putative anti-cancer treatments, and will be tested on living cells.


Research activity codes: 2.1 BIOLOGICAL AND ENDOGENOUS FACTORS
Health categories: CANCER AND NEOPLASMS
Title: Pathfinder: Defining interactions between the cytokine IL-22 and oncogenic KRAS as a new therapeutic target in colorectal cancer
Funder: Medical Research Council

Abstract: Inflammation driven by the IL-23/Th17 cell axis is related to poor outcome in colorectal cancer. Cytokines produced during Th17 responses, such as IL-22, promote CRC development in animal models by acting directly on intestinal epithelial cells to promote growth and survival. The KRAS oncogene is mutated in roughly 45% of colorectal cancer. No targeted therapies currently in development have shown any activity in KRAS mutant CRC and finding novel approaches is a high priority. The available literature suggests that KRAS mutations have little prognostic impact, but our recent studies have shown that among tumours with evidence of heightened IL-22 signalling, KRAS mutations are strongly associated with poor clinical outcome. (McCuaig et al., submitted). Specifically, using tumour transcriptomic datasets totalling nearly 2000 patients (GSE39582, PETACC3, TCGA, ALMAC), we have identified that in patients with high tumoural expression of either or both subunits of the heterodimeric IL-22 receptor (IL22RA1, IL10RB), KRAS mutation dramatically worsens prognosis. In contrast, KRAS has no prognostic relevance in tumours with low expression of IL-22 receptor subunits. Based on this data we hypothesize that modulation of the IL-22 pathway in IL22RA1-high, KRAS mutant tumours may have therapeutic benefit. Though attractive as a possible therapeutic target for CRC, direct evidence supporting a tumourigenic role for IL-22 in humans is lacking. Furthermore, as KRAS is widely considered an "undruggable" target, methods of improving clinical outcome for patients with KRAS-mutant CRC are urgently needed. We will address both knowledge gaps by studying the role of IL-22 signalling in KRAS wild type versus mutant CRC in order to 1) establish the functional relevance of IL-22 signalling in human CRC; and 2) test the biological impact of IL-22 or IL-23 blockade in molecularly stratified CRC patient subsets in a Phase 0 window trial.


Research activity codes: 2.1 BIOLOGICAL AND ENDOGENOUS FACTORS
Health categories: CANCER AND NEOPLASMS