Harnessing single-cell datasets to enable human disease modelling and drug discovery
During differentiation, progenitor cells respond to exogenous signals by activating transcription factors (TFs) to establish new cell identities. Experimentally over-expressing one or more TFs can be sufficient to reprogram cell fate, as famously shown when somatic cells are reprogrammed into induced pluripotent stem cells (iPSCs), or when iPSCs are exposed to other TFs to “forward program” them to different cell types, such as neurons. Our group is interested in hypothalamic neuron populations that regulate critical behavioural and physiological processes such as eating, drinking, reproduction, stress, and sleep, and are therefore highly disease-relevant. We routinely generate human hypothalamic neurons from iPSCs, including pro-opiomelanocortin (POMC) neurons that regulate appetite and energy expenditure and are relevant to obesity and Type 2 diabetes. In this PhD project, you will use forward programming to more quickly and efficiently generate POMC and other hypothalamic neuron populations to facilitate disease modelling, CRISPR screens, and image-based drug screens that would be difficult to carry out in animal models. You will work collaboratively with other team members who have already used CRISPR/Cas9-based gene editing to make knock-in fluorescent reporters, allowing cell types of interest to be purified and studied in detail using data-rich methods such as transcriptomics, peptidomics, and high-content imaging, and functional methods such as calcium imaging. We have also harnessed mouse and human single-cell sequencing resources such as the human cell atlas (HCA) to identify candidate TFs and cloned libraries of these candidates that can be expressed in reporter iPSCs. You will help carry out TF screens to generate target cell types and then functionally characterise them using the methods described above, creating exciting opportunities for understanding the function of these cells in health and disease. In addition to providing training in cutting-edge technologies, this project presents a unique opportunity to make a substantial impact on the field since most of these human cell types have never been studied, despite their importance.
Axon degeneration mechanisms in human disease
“Programmed axon death is a preventable and widespread mechanism of axon loss in injury and disease. It is regulated by NAD and related metabolites, through opposing actions of a pro-death protein, SARM1 NADase, and pro-survival NMNAT2, which prevents SARM1 activation in axons. Many details of the regulation are now known, along with endogenous regulators of SARM1 and genetic and environmental activators linked to axonal disorders.
Genetic association has been established to two human disorders of long axons: ALS and polyneuropathies. However, animal data indicate much wider relevance including to Parkinson’s disease, glaucoma, multiple sclerosis and peripheral neuropathies. One key to studying this in humans is to identify further naturally-occurring gene variants that alter NMNAT2 or SARM1 function.
This project will contribute to a comprehensive mutation map of these proteins, testing natural human mutations for their ability to confer loss- or gain-of-function, investigating the underlying mechanism, and exploring any association with human disease.
The student will gain extensive experience of molecular biology (DNA cloning, site-directed mutagenesis), cell culture (murine primary neuronal cultures and hiPSC-derived neurons), microscopy (fluorescent, phase contrast), enzyme and metabolite analysis (enzyme assays, NAD-Glo, collaborative use of HPLC and mass spectrometry), and bioinformatics (UK Biobank, 100,000 Genomes Project, etc). The results will help to identify the most appropriate disorders and the most relevant patients for effective clinical trials of drugs already under development. We collaborate with researchers in Italy, USA, UCL, KCL, Oxford, Glasgow and Cambridge, and our highly supportive team culture aims always to value and develop our colleagues as the best way to do great science.
Contact for enquiries: email@example.com
Coleman and Hoke (2020) Nat Rev Neurosci 21: 183-196
Coleman (2022) Neurotherapeutics In Press
Identifying novel genes and developing treatments for children with inherited neuromuscular diseases
Inherited neuromuscular disorders are disabling, progressive, often fatal conditions, representing an enormous unmet medical need with devastating impacts on affected families, the healthcare system, and the economy. There are no cures and the limited therapies available treat symptoms without addressing the underlying disease.
Next-generation sequencing has facilitated a molecular diagnosis for many inherited neurological disorders, such as mitochondrial diseases and other neuromuscular diseases, which are the focus of this research. The development of targeted therapies requires detailed laboratory investigation of molecular and mutational mechanisms, and a systematic evaluation of well-chosen agents as well as gene and transcript directed strategies using standardized experimental systems. Our research is focusing on understanding the molecular pathogenesis of childhood onset inherited neuromuscular diseases, such as mitochondrial disease and other neuromuscular diseases to develop targeted therapies.
Using a translational approach, we aim to
1. understand the clinical course of patients in relation to the underlying disease mechanism
2. delineate the mutational and molecular mechanisms of the molecular defect in the appropriate cell types by developing model systems such as induced neuronal progenitor cells (in vitro) and zebrafish (in vivo)
3. improve the treatment options for patients by developing novel therapies that are directed at these mechanisms, including directly at the genetic mutation or resulting transcript.
We use a combination of exome sequencing, genome sequencing, and other omics technologies to identify novel disease genes and disease mechanisms. By functional evaluation in vitro (induced neuronal progenitor cells) and in vivo (zebrafish) we confirm pathogenicity and uncover molecular mechanisms of disease. To address the mutational mechanisms, we use gene transfer, splice modulation, allele silencing and CRISPR/cas systems.
Cambridge Stroke: Cerebral small vessel disease (SVD)
Cerebral small vessel disease (SVD) causes a quarter of all strokes and is the most common cause of vascular dementia. Despite its importance we have few treatments for SVD. Increasing evidence implicates inflammation, both systemic (in the blood vessels) and central nervous system, are risk factors in SVD, but which aspects of the dysregulated immune response relate to progression and could be targeted therapeutically remains undetermined. The student will join on ongoing programme looking at the role of a dysregulated immune response and increased CNS inflammation in pathogenesis of SVD and cognitive decline. The programme uses advanced brain imaging to assess CNS inflammation (PK1195 positron emission tomography) and white matter damage (diffusion tensor imaging). It corelates this with detailed immunophenotyping of the blood and CSF, in collaboration with Prof Ziad Mallat in Cardiovascular Science. Systemic immune responses are being determined by a comprehensive blood phenotype programme including cytokine measurement, flow cytometry, cytokine production capacity and single cell RNA sequencing. The relationships between peripheral and central inflammation will be identified, and whether they predict future brain damage (as assessed by diffusion tensor MRI) determined. Much of the data will have been acquired but there is an opportunity for involvement in further MRI and immune phenotyping data collection.
Walsh J, Tozer DJ, Sari H, Hong YT, Drazyk A, Williams G, Shah NJ, O’Brien JT, Aigbirhio FI, Rosenberg G, Fryer TD, Markus HS. Microglial activation and blood-brain barrier permeability in cerebral small vessel disease. Brain. 2021 Jun 22;144(5):1361-1371.
Disease pacemaker Stem Cells in Neurodegenerative Disease
The presence and role of neural stem cells (NSCs) in the adult human brain is a long-debated issue in neuroscience. Recent work has demonstrated that stem-like cells exist in the embryonic, foetal, and human adult brain where they persist well into adulthood and can even contribute to neurogenesis. However, their role in neurodegenerative disease is unknown. Ongoing work in the lab has led to the hypothesis that NSCs may become dysfunctional in neurodegenerative disease resulting in senescence chronic inflammation, and thereby acting as pacemaker cells driving neuronal demise. This ambitious project aims to identify disease-associated NSCs and their phenotype in the context of human neurodegeneration using spatial biology approaches, including imaging mass cytometry, RNA scope and single nuclear RNA sequencing. Relying on post-mortem brain tissue of different stages of Alzheimer’s disease, traumatic brain injury, vascular dementia and chronic stroke, this project will study NSCs in a range of human diseases characterised by neurodegeneration and neuronal injury. Ongoing work in the lab identifies NSC-specific markers based on transcriptomics and protein profiling experiments in brains with progressive multiple sclerosis, enabling to investigate the distribution of NSCs in a wide range of diseases. Spatial transcriptomics and proteomic approaches will allow to study their phenotype and dysfunction in relation to other cell types and local pathology. This project will shed light on the role of NSCs in neurodegeneration and has the potential to identify an entirely novel mechanism of neurodegeneration in human disease.
Molecular mechanisms of microglia-driven neurodegeneration in human tauopathies
Neuroinflammation is associated with many neurodegenerative disorders. In experimental animals of tauopathies, microglia were shown to play a role in tau propagation and synapse loss, however the molecular and cellular processes driving tau accumulation and neurodegeneration in the context of human disease remain poorly understood. We are investigating the role of microglial responses in human tauopathies with progressive supranuclear palsy (PSP) as the demonstrator condition. Using advanced spatial biology and high-plex imaging techniques we will characterise the molecular signatures of microglia and their interactions with tau aggregates and other cell types. This project relies on the large collection of well-characterised PSP brains in the Cambridge Brain Bank, enabling to study microglia in a large range of disease stages. With this strategy we anticipate to better understand the role of microglia in PSP and tauopathies, and hence, to guide the design of novel neuroimmune therapies and disease biomarkers.
Methods for quantitative MRI
The contrast of magnetic resonance datasets is conventionally usually dimensionless; in order to derive meaningful quantitative measures (such as T1, T2, diffusion metrics etc) longer acquisitions and further modelling are required. However, for longitudinal studies and for multi-site studies (including different hardware platforms), repeatable and meaningful metrics are increasingly required (quantitative MRI; qMRI). This project is to study current strategies for robust qMRI including faster data acquisition and/or synthetic approaches. It will benefit from ongoing local longitudinal studies and the availability of multi-site datasets for comparison. The project will be appropriate for a candidate with a background in Physics and strong mathematical skills, and an interest in translating methodology to provide novel and practical research tools.
Investigating the Immune Basis of Parkinson’s Disease
It is now well established that immune activation occurs in Parkinson’s disease, but whether it plays a critical role in disease onset and progression remains uncertain. This is a critical question to address given that the immune system is a tractable target for disease-modifying therapy. The Williams-Gray lab investigates the immune component of Parkinson’s disease through neuropathological studies on post-mortem human brain, PET brain imaging studies, analysis of blood and cerebrospinal fluid samples, and interventional clinical studies. The project will extend this work to include investigation of cases at high risk of developing Parkinson’s in order to determine whether immune activation precedes and predicts disease onset, and will involve characterising how the immune response changes with evolution from early to mid and late-stage disease. Studies will involve measuring immune activation and dysregulation in patient-derived biological samples, determining drivers of Parkinson’s related immune activation in-vitro, and investigating how immune responses are linked to longitudinal clinical measures and outcomes. Laboratory techniques will include immunophenotyping using flow cytometry, multiplex immunoassays, and in vitro assays of immune cell function. Brain imaging markers of neuroinflammation may also be incorporated. Ultimate aims of the project will be to identify the most critical immune pathways for therapeutic targeting in Parkinson’s, as well as to identify immune biomarkers which will allow patient selection for future clinical trials of immune-based therapies, and the longitudinal monitoring of such therapies.
Mapping the hypothalamic architecture underlying appetite control in the obese human brain
Obesity is a growing public health problem, however genetic studies point to the brain, and in particular the hypothalamus, as having a crucial role in modulating appetitive behaviour, which has limited the mechanistic insights achievable from human research. Until recently, the major stumbling block has been inaccessibility of the human hypothalamus, which has, to date, meant our understanding of circuitry controlling food intake has emerged primarily from murine studies. However, a recent collaboration with the MRC Brain Bank Network, has allowed us access to fresh and fixed human donor brain samples. These precious samples, coupled with developments in droplet single-cell/nucleus sequencing technologies, spatial transcriptomics, and single-molecule fluorescent in situ hybridization, have provided us the opportunity to map the functional architecture of the human hypothalamus underlying appetitive behaviour. Over the past three years, we have generated a database of more than 350,000 human hypothalamic cells from eight normal weight donors. In this PhD project, we are proposing to explore the transcriptional landscape of the hypothalamus in energy imbalance. Building on our growing database, as well as the methodologies and expertise we have accrued, the successful candidate will profile the hypothalami of donors with obesity from the MRC Brain Bank Network. Our goal for a comprehensive human hypothalamic atlas across the weight spectrum will be of utility to basic and translational researchers attempting to better understand the fundamental nature of regulation of energy balance by the hypothalamus and to manipulate these systems to improve the health of the population.
Using big data to tackle the most significant clinical problems in dementia
Genetics of macroscale brain networks
Macroscale brain networks can be derived from multiple structural, functional and diffusion-weighted MRI phenotypes using multiple different methods. These networks are thought to underlie various aspects of human cognition and behaviour, and have been linked to neurodevelopmental, psychiatric, and neurodegenerative conditions. Yet, a few gaps remain. First, due to the multiple ways by which macroscale brain networks can be generated and characterised, it is unclear precisely what information these networks capture and how they differ from each other. Second, it is also unclear what macroscale brain networks add above and beyond global values for the MRI phenotypes using which they were generated. Third, it is unclear how both common and rare genetic variants influence these macroscale networks, including genes that are linked to psychiatric and neurodevelopmental conditions, and genes that are constrained due to natural selection. Finally, it is unclear if individuals can be meaningfully subgrouped based on multiple macroscale networks and if this offers a model for identifying individuals with increased likelihood for various mental health conditions.
The PhD candidate working on this project will collaborate with me and other research groups in Cambridge to address many of these issues. They will develop skills in analysing and interpreting MRI data, generating networks, large-scale data analysis, and genetic association and heritability analyses alongside other transferable skills gained during a PhD. An ideal candidate for this position must have previous experience in human/statistical genetics, with an interest in applying it to neurodevelopment or previous experience in neuroimaging, with some understanding of and considerable interest in human/statistical genetics. They would need to have good coding skills (e.g., R, Matlab), and a good understanding of statistics.
Using Photobiomodulation therapy as an otoprotective agent
“Near-infrared light has been shown to increase cell resilience, by increasing ATP production though its actions on the mitochondrial cytochromes. This has been termed Photobiomodulation Therapy (PBMT), and used in various systems to protect tissues.
It is a very attractive non-contact technology, that can penetrate bone, and reach deeper structures. We aim to develop a system to use PBMT to protect the inner ear from various damaging insults, including noise damage, ototoxicity from aminoglyosides, and cochlear implantation trauma. The PhD student will join a team of Otologic clinicians (Prof Bance, Mr Matt Smith), cell biologists (Prof Howarth), and an optical physicist (Dr. Gemma Bale) to develop, validate and test this technology in subjects with cystic fibrosis undergoing aminglycoside therapy.
The student will be embedded in a lab (SENSE Lab) with clinicians, cell biologists and engineers. The work will involve cell culture and ATP assays, development (with assistance of engineering professors and students) of models, 3D printing of models of the ear, and experience in clinical trial design of therapies.
This technology has a broad range of applications, and we will be developing a platform technology that could be used in other systems as well.