NIHRDataInsights/HRCSData huggingface dataset:
Title: Drug Transport and Tumour Metabolism
Funder: Medical Research Council

Abstract: Transporters play a vital role in physiology, regulating not only the uptake of nutrients into cells, but also the flux of metabolites between different cellular compartments. The importance of these proteins is illustrated by the fact that more than 80 monogenic diseases are associated with mutations in transporter-encoding genes. Tumour-specific metabolic changes are essential in order to fuel the increased proliferation rates of cancer cells, while also allowing them to survive in the often-hostile tumour micro-environment. Alterations in the expression patterns of transporters are a key component of these changes, allowing the cell to efficiently scavenge available nutrients and prevent a toxic build-up of waste products. Classically, entry of a drug into cells was thought to be determined by the compound lipophilicity, however it is now becoming clear that many drugs actually enter cells through membrane transporters. This means that the expression level of the relevant transporters will determine the intracellular concentration of a drug, thereby determining its efficacy, specificity, and potentially toxicity. In the case of tumours, altered transporter expression patterns may therefore define the drug sensitivity profile of a tumour, as well as dictating its metabolic profile. By combining analysis of large-scale, publicly available datasets with CRISPR screening/gene-editing, cell biology and mass spectrometry, we will identify specific transporters involved in the uptake of cancer therapeutic compounds in order to fully understand how they affect the efficacy of these drugs. As well as understanding their roles in pharmacology, we will use metabolomics to look at how our transporters of interest influence cancer metabolism, with the aim of exploiting this biology to drive efficient drug uptake. Understanding which transporters are required for cellular uptake of cancer therapeutics could provide functional biomarkers to predict compound efficacy at the level of individual patients. This could allow personalised treatment plans based, in part, on transporter expression patterns, as well as enabling stratification of patients in clinical trials. In order to be able to implement these findings, we will establish robust quantitative proteomics assays to determine transporter expression patterns, initially from cell lines and animal models, and eventually (through collaboration) in patient samples.


Research activity codes: 5.1 PHARMACEUTICALS;1.1 NORMAL BIOLOGICAL DEVELOPMENT AND FUNCTIONING;2.1 BIOLOGICAL AND ENDOGENOUS FACTORS
Health categories: CANCER AND NEOPLASMS;GENERIC HEALTH RELEVANCE
Title: Control of cell cycle entry in health and disease
Funder: Medical Research Council

Abstract: Our group aims to understand how cells control the entry into, and exit from, the cell cycle, in both health and disease. These proliferation-quiescence decisions are poorly understood in mammalian cells, yet getting these decisions right is vital for normal development and tissue homeostasis. Dysregulation of proliferation-quiescence decisions underpins uncontrolled proliferation in tumorigenesis. If we can understand how signalling networks regulating proliferation and quiescence are controlled, then we can design strategies to specifically inhibit cancer cell proliferation or to improve tissue regeneration and improve human health. We have a number of objectives to achieve our aim. First, we will define the mechanisms that maintain reversibility in quiescence. Second, we will characterise how cell intrinsic and extrinsic factors influence CDK activity in proliferation-quiescence decisions. Finally, we will determine how cancer-specific alterations perturb signalling networks controlling cell cycle entry to drive the continuous proliferation of cells. To achieve our objectives, we will use our expertise in quantitative, single-cell, time-lapse imaging of human cell lines that we have engineered to express fluorescently-labelled proteins or biosensors. This strategy allows us to simultaneously measure cell cycle entry and the regulatory signalling events in real-time in the same cell. By generating and integrating proteomic and phosphoproteomic datasets with our imaging data, we will work towards a systems-level understanding of proliferation-quiescence decisions in mammalian cells. We will also use our imaging data to parametrise new mathematical models to achieve a mechanistic understanding of the control of proliferation-quiescence decisions. These models will be tested experimentally using genetic (siRNA, CRISPR knockouts, degron tags) and chemical (small molecule inhibitors) perturbations to test the predictive value of our models.


Research activity codes: 1.1 NORMAL BIOLOGICAL DEVELOPMENT AND FUNCTIONING;2.1 BIOLOGICAL AND ENDOGENOUS FACTORS
Health categories: CANCER AND NEOPLASMS;GENERIC HEALTH RELEVANCE
Title: Generating blood stem and progenitor cells from haemogenic endothelium
Funder: Medical Research Council

Abstract: Stem cells are crucial in tissue homeostasis, regeneration and repair. The current interest in stem cell-based therapies has emphasised the importance of understanding how tissue specific stem cells are specified in development. Here, we aim to obtain a mechanistic understanding of the birth of blood stem cells in mouse development. We will build on our previous work in which we have made advances towards unravelling the cis-regulatory complexity of Runx1, a critical regulator of the endothelial-to-haematopoietic transition (EHT), and obtained important new insights on the timeline and process of early haematopoietic commitment. In this programme we will continue our analysis of the Runx1 cis-regulatory logic through application of state-of-the-art technologies, and identify upstream regulators that converge on these cis-elements. In addition, we aim to identify the gene regulatory network that underlies blood stem and progenitor generation in vivo, through genome wide expression and chromatin analyses of primary mouse embryo cells. Together with studies into the developmental origin of the haemogenic endothelium, these experiments will shed light on the spatiotemporal factors and signals that are active in haematopoietic specification and differentiation from gastrulation onwards. This knowledge will inform the development of robust in vitro models for generating definitive haemogenic endothelium and blood stem and progenitor cells from ESC/iPSCs.


Research activity codes: 1.1 NORMAL BIOLOGICAL DEVELOPMENT AND FUNCTIONING;2.1 BIOLOGICAL AND ENDOGENOUS FACTORS
Health categories: CANCER AND NEOPLASMS;GENERIC HEALTH RELEVANCE
Title: Understanding the physiological relevance of selective endoplasmic reticulum autophagy
Funder: Medical Research Council

Abstract: The autophagy of the endoplasmic reticulum (ERphagy) is fundamentally conserved from yeast to higher eukaryotic cells. Clinical mutation of factors involved in ERphagy results in neurodevelopmental defects, pancreatic stress, and cancer, suggesting that ERphagy has pleiotropic roles in cellular homeostasis. Our lab aims to uncover novel factors and pathways that are involved in the regulation of endoplasmic reticulum (ER) stress and ERphagy. Our previous work identified UFMylation (a form of Ubiquitin-like modification) to be involved in ER stress and ERphagy regulation. However, various aspects of UFMylation during ER-phagy remain to be understood. The three main aims that we would like to further address are: (1) What other ER proteins are UFMylated during ER-phagy? (2) Are there specific poly-UFM1 chains that mediate ER-phagy? (3) What are the UFM1-binding proteins that recognize/edit different UFM1 chains? AIM 1- Our previous data suggest that substrate UFMylation (e.g., RPN1 and RPL26) is likely proximity-based rather than unique substrate recognition. To explore additional substrates of UFMylation, we will use denaturing immunoprecipitation of UFM1 coupled with quantitative mass spectrometry (IP-MS) to compare wildtype versus UFMylation defective (CRISPR-based knockout) cells. Follow-up MS experiment enriching for the potential substrates will allow us to identify the target lysine (or other potential amino acids) that are modified by UFMylation. These lysine residues will then be mutated (either endogenously using CRISPR/Cas9 or mutant cDNA rescue experiment) to understand the effect of UFMylation during ERphagy. AIM 2 – We will explore the types of chains formed on two known UFMylation substrates (RPL26 and RPN1). By stably expressing different UFM1 mutant constructs (K-to-R & K-only) in UFM1-knockout cells, we will interrogate the ability of different poly-UFM1 chains to mediate ER-phagy. We will then biochemically verify the UFM1 chains conjugated onto RPL26 and RPN1 (and other potential targets) by co-immunoprecipitation assays and collaborate with the Kulathu group at MRC-PPU for structural characterisation of these different poly-UFM1 chains. AIM 3 - There are currently no reported UFM1-binding proteins or characterised motifs beyond the UFM1-interacting sequence (UIS) reported on the E1 enzyme, UBA5. We will perform tandem IP-MS on HA-RPN1/RPL26 and FLAG-UFM1 to identify potential UFM1 binding proteins. We will then cross-compare the candidates using proximity-based labelling and MS of APEX2-RPN1/RPL26 to identify the interactors that are enriched in the presence of UFMylation. These UFM1-binding partners will provide mechanistic insights on how the UFMylation signal is relayed to ER-phagy regulation.


Research activity codes: 2.1 BIOLOGICAL AND ENDOGENOUS FACTORS;1.1 NORMAL BIOLOGICAL DEVELOPMENT AND FUNCTIONING;2.2 FACTORS RELATING TO PHYSICAL ENVIRONMENT
Health categories: CANCER AND NEOPLASMS;GENERIC HEALTH RELEVANCE
Title: Endogenous DNA damage and its repair during haematopoiesis
Funder: Medical Research Council

Abstract: We aim to explain the physiological purpose for DNA repair, specifically why certain cell lineages and tissues are dependent on specialised DNA repair pathways. Our foundational discovery is that two simple reactive aldehydes (acetaldehyde and formaldehyde) are ubiquitous drivers of endogenous DNA damage which is repaired by DNA crosslink repair. We established how we are protected from these endogenous genotoxins, firstly by enzymes such as ALDH2 or ADH5 (Tier-1 protection) that detoxify them, and secondly by DNA crosslink repair by the Fanconi anaemia (FA) proteins (Tier-2 protection). This Two-Tier protection mechanism is essential for the renewal of blood because when it is inactivated in humans and mice, haematopoiesis ceases. We will continue to define how Two-Tier protection ensures blood stem cell survival and integrity. We do not know when during the development of these vital cells Two-Tier protection becomes essential, and how it then enables lifelong blood production. We want to better understand how the p53 response impacts aldehyde-damaged stem cells. With our collaborators, we will continue our mechanistic research to explain how FA crosslink repair functions and to define the genomic land-scape of aldehyde-induced DNA damage and mutagenesis in haematopoietic cells. Finally, we will examine how Tier-1 deficiency might have arisen to be a very common genetic trait in humans, despite the fact that such individuals accumulate aldehydes and are therefore completely reliant on Tier-2 protection.


Research activity codes: 1.1 NORMAL BIOLOGICAL DEVELOPMENT AND FUNCTIONING;2.1 BIOLOGICAL AND ENDOGENOUS FACTORS
Health categories: CANCER AND NEOPLASMS;GENERIC HEALTH RELEVANCE
Title: How cells regulate protein levels (and fail to do so in cancer)
Funder: Medical Research Council

Abstract: There is a major discordance between mRNA and protein expression levels in human cells. Why is this so and what mechanisms are behind it? The aim of my lab will be to understand the principles, mechanisms and regulators that shape the proteome at the level of translation and protein degradation. Crucially, anomalies in this protein-level regulation drive or sustain the growth of many types of cancer. Nevertheless, these regulatory processes remain poorly understood, leaving an enormous potential for therapeutic intervention unfulfilled. I will address three key questions: 1. The role of translation and degradation rates in regulating protein levels, e.g. to understand how they buffer gene copy number alterations in cancer. For this I will quantify the change of mRNA and protein levels together with changes in translation and protein degradation rates across many biological conditions and interrogate the data using correlation, regression, clustering and other bioinformatic analyses. 2. Identifying regulators of translation and degradation, e.g. to reveal regulatory networks between E3 ubiquitin ligases and their targets. For this I will computationally re-purpose previously reported proteomics data on a massive scale and apply existing algorithms from the transcriptomics field. 3. Understanding the role of degradation in shaping the nascent proteome, e.g. to identify cancer-specific unconventional translation products. For this I will combine total-transcriptome sequencing with deep profiling of affinity-captured nascent proteins with optional proteasome inhibition. Together, these projects will reveal fundamental insights into how cells regulate protein levels and fail to do so in cancer. This knowledge could lead to new therapeutic strategies exploiting proteotoxic stress in cancer, explain why proteasome inhibitors elicit heterogeneous responses in patients and highlight new therapeutic targets.


Research activity codes: 1.1 NORMAL BIOLOGICAL DEVELOPMENT AND FUNCTIONING;2.1 BIOLOGICAL AND ENDOGENOUS FACTORS
Health categories: CANCER AND NEOPLASMS;GENERIC HEALTH RELEVANCE
Title: Homologous Recombination at Human Centromeres: Friend or Foe?
Funder: Medical Research Council

Abstract: Homologous recombination (HR), which is catalysed by the RAD51 recombinase, plays pivotal roles in preventing human diseases associated with genome instability, including cancer, developmental disorders and premature ageing. Indeed, malfunctions of key RAD51 regulators BRCA2 and PALB2 increase the risks of these diseases. However, HR can also be an entry point for genome instability and hence must be tightly regulated. This is particularly important at highly repetitive genome regions. Among them, centromeres are especially unusual genome regions, consisting of long stretches of repeats and playing a central role in chromosome segregation. It remains enigmatic how HR acts positively or negatively at human centromeres. Our previous work revealed that RAD51 maintains centromere integrity and assists accurate chromosome segregation in mitosis. Building on these findings, we have developed a versatile methodology to detect DNA damage at human centromeres in single cells. This technological breakthrough enabled us to demonstrate that RAD51 protects centromeres from DNA damage even in non-dividing and non-cancerous human cells. We hypothesise that RAD51 ensures the integrity of centromeres, which are susceptible to spontaneous DNA break, while also preventing harmful recombination of their repetitive sequences and, in this way, promotes genome stability. This project will examine: i) how RAD51 protects human centromeres from DNA breakage; ii) how RAD51 is recruited to centromeres; and iii) the mechanisms that lead to the potentially harmful outcomes of HR. The use of light imaging approaches directly detecting DNA damage at centromeres, genetic tools, unbiased genome-wide sequencing technology and quantitative proteomics will be key to examine the role of HR at human centromeres, the main focus of this study. The proposed research represents an important first step towards the development of new treatments to prevent diseases associated with centromere dysfunction.


Research activity codes: 1.1 NORMAL BIOLOGICAL DEVELOPMENT AND FUNCTIONING
Health categories: CANCER AND NEOPLASMS;GENERIC HEALTH RELEVANCE
Title: Structural mechanism of DNA damage response on replicating chromatin
Funder: Medical Research Council

Abstract: Background: Our DNA is constantly damaged by triggers in surrounding environment, lifestyles and even by-products of normal cellular functions. We rely on the DNA damage response (DDR), a highly regulated cellular signalling network, to deal with this damage. Failed DDR can lead to genome instability, cell death and also influences the effectiveness of cancer treatments, which often generate multiple DNA damage events. Complex DNA damage, such as interstrand crosslink (ICL) that blocks the DNA replication machinery on chromatin, invokes a more specialized DDR pathway. The molecular detail of this specialized pathway remains elusive, with an incomplete understanding of what, how and when DDR factors assemble and function. Aim: This proposal aims to elucidate mechanism of this DDR pathway by combining integrated structural biology (cryo-electron microscopy, X-ray crystallography and structural mass spectrometry) and the development of protein tools (Affimers) to reveal how a new protein complex called SLF1-SLF2 (SMC5/6 localization factor 1 and 2) functions in DNA damage response on replicating chromatin. Scientific and Medical Outcomes: I will characterise and determine individual and higher-order structures of SLF1-SLF2 and its interaction network including RAD18, SMC5/6 and the nucleosome. SLF1-SLF2 functions as a reader for new histones incorporated in the replicating chromatin near DNA damage sites and is also a recruiter for a protein complex responsible for chromosome structure maintenance. My research will reveal new understanding of how the DDR functions on replicating chromatin and also create new approaches to discover the unknown mechanism of DDR. These multi-faceted insights from my research will also provide a pipeline for investigating other key proteins within the DDR signalling network and pave the way in the long term for better cancer diagnostics, improved prediction of patient outcomes, and generating new targets for drug development.


Research activity codes: 1.1 NORMAL BIOLOGICAL DEVELOPMENT AND FUNCTIONING;2.1 BIOLOGICAL AND ENDOGENOUS FACTORS
Health categories: CANCER AND NEOPLASMS;GENERIC HEALTH RELEVANCE
Title: A framework for multi-indication evidence synthesis in oncology Health Technology Assessment
Funder: Medical Research Council

Abstract: Health Technology Assessment (HTA) evaluates the health and cost consequences of competing health care interventions. Typically, this evaluation is focused on evidence of the effects of a therapy in a specific indication. However, an increasing number of health technologies, particularly in oncology, are multi-indication gradually attaining license extensions to other indications as trial evidence emerges. Indication-by-indication HTA typically ignores evidence for other indications on the same treatment and therefore commonly presents high levels of evidential uncertainty. The aim of this project is to make more efficient use of the entirety of evidence, within and across indications, thus reducing decision uncertainty and improving decisions. We plan a methodological programme of work, grounded on a case study in oncology. We will extend methodology for evidence maps to allow summarising evidence in multi-indication oncology HTA. We will extend and adapt two existing methodologies (mixture prior and mixture hierarchical models), which enable different ways for information to be shared across indications, to multi-indication network meta-analysis. We will conduct a simulation study to generalise the case study findings and help us make appropriate recommendations for information sharing in this context. We will develop methods for eliciting expert judgements on the appropriateness of information sharing. Finally, we will identify the implications of multi-indication evidence synthesis to decisions, aiming to demonstrate how the fuller use of evidence contributes to the understanding of the value of the treatment across indications. We will engage with relevant stakeholders to develop a policy framework to facilitate rapid translation into the methods and processes of HTA agencies.


Research activity codes: 8.4 RESEARCH DESIGN AND METHODOLOGIES (HEALTH SERVICES)
Health categories: CANCER AND NEOPLASMS;GENERIC HEALTH RELEVANCE
Title: Structural Mechanisms of Nucleotide Excision Repair
Funder: Medical Research Council

Abstract: The genomes of our cells are constantly exposed to DNA-damaging agents and processes. To preserve genomic integrity and cellular function, numerous DNA repair pathways that specialize in repair of specific types of DNA damage have evolved. Nucleotide excision repair (NER) serves to repair DNA lesions induced by UV light or by chemical modification of DNA bases. Because NER is critical for cellular function, it is important for human health. Mutations affecting the molecular components of the pathway cause human diseases, including xeroderma pigmentosum, Cockayne syndrome, and trichothiodystrophy. Additionally, NER is able to remove DNA lesions caused by cancer chemotherapeutics and may reduce efficacy of treatment. A mechanistic understanding of NER, its regulation, and disease-causing mechanisms is therefore critical. This requires structural information on the complex molecular assemblies that perform the repair reaction. I aim to elucidate the structural mechanism of NER by using cryo-electron microscopy to determine the structures of repair intermediates along the NER pathway. Additionally, I aim to perform structural studies of disease mutations that affect the crucial NER factor TFIIH. To this end, I am reconstituting the pathway from recombinantly expressed and purified components, which can be obtained in larger amounts than endogenously purified material and can harbour engineered mutations if required. The cryo-electron microscopy experiments will be complemented by structure-guided functional investigation of NER in a collaboration with the laboratory of Prof. Wojciech Niedzwiedz. The combined structural and functional studies will enable a mechanistic understanding of the function and dysfunction of NER.


Research activity codes: 1.1 NORMAL BIOLOGICAL DEVELOPMENT AND FUNCTIONING;2.1 BIOLOGICAL AND ENDOGENOUS FACTORS
Health categories: CANCER AND NEOPLASMS;GENERIC HEALTH RELEVANCE