Research Renzo Mancuso

Abstract

The discovery of strong microglial genetic risk factors in Alzheimer's Disease (AD) increased the interest towards the functional role of these cells in disease. Recent efforts describe the shift in microglial transcriptional states as they are believed to be crucial in the understanding of pathology. However, the outstanding challenge remains the translation from phenotype to function. Combining cutting-edge TurboID proximity ligation with xenotransplantations of human iPSC-derived microglia into the mouse brain (MIGRATE) opens an exceptional opportunity to study one of the main microglial functions in vivo: the secretion of molecules and its impact on disease progression. I will study the microglial secretome (native and in TREM2R47H) in amyloid and tau models. I already developed an endoplasmic reticulum (ER) baited TurboID that labels proteins destined for conventional secretion, and expressed in iPSC prior to differentiation and transplantation. Employing precipitation and mass spectrometry of biotinylated proteins, I will map the endogenous secretome of human microglia in response to pathology and in clinically relevant genetic mutations, in vivo. I will also validate my findings in human CSF and using iPSC-derived neurons. I expect to infer the implication of the microglial secretome in relation to pathology and deconstruct the microglial response to the main pathological hallmarks in AD. My findings will open avenues to new biomarker and therapeutic applications.

Researcher(s)

• Promoter: Mancuso Renzo
• Fellow: van Hoek Maxim

Research team(s)

• VIB CMN - Microglia and Inflammation in Neurological Disorders (MIND) lab

Project type(s)

• Research Project

Abstract

Soluble transducers of human microglial toxicity in Alzheimer's disease Alzheimer's disease is not just a matter of neurons. Other types of cells in the brain matter as well. Most importantly, the main immune cells of the brain, called microglia, have a critical role in Alzheimer's disease. They express many genes that are linked to Alzheimer's disease and are important for controlling inflammation in the brain. One of the tasks microglia perform to fulfill their immune functions, is the secretion of soluble molecules. A critically understudied issue is how this secretion "landscape" or "secretome" changes during disease. That is the question our research aims to answer. To do this, we use a new technique that labels molecules secreted by microglia with a marker that can be easily retrieved during analysis. By comparing healthy microglia with models that show Alzheimer-like properties, we can find out how Alzheimer's disease changes the brain immune system, which could lead to new biomarkers and therapies.

Researcher(s)

• Promoter: Mancuso Renzo

Research team(s)

• VIB CMN - Microglia and Inflammation in Neurological Disorders (MIND) lab

Project type(s)

• Research Project

Abstract

ABCA7 is one of the most important risk genes for Alzheimer's disease (AD). The ATP-binding cassette (ABC) transporter, subfamily A member 7 (ABCA7) is one of the major risk factors for AD. Many deleterious variants have been reported for ABCA7, decreasing its expression, thus suggesting an overall loss-of-function (LOF) to underlie the genetic risk. Targeted resequencing studies revealed that rare protein truncating (PTC) and missense variants that alter susceptibility to AD (odds ratio (OR) = 2.6). In addition, other mutations such as an intronic pathogenic repeat expansion was identified that decreases ABCA7 expression and strongly increases AD risk (OR = 4.5). Given that that one APOE4 allele renders an OR of 4-5 and TREM2 is associated with an OR between 2-4, the risk associated with ABCA7 pathogenic variants is very high. This puts ABCA7 forward as a key player in AD pathogenesis. Our preliminary data shows that ABCA7 deficiency impairs the transition of human microglia towards activation states and has a strong impact of phagocytic and endolysosomal function both in vitro and in vivo. Here, we aim to fully dissect the role of ABCA7 in microglial endolysosomal biology both in vitro and in vivo. We plan to take a step further and use a systematic approach, where we will investigate the impact of ABCA7 deficiency at all levels of the endolysosomal system, starting from its effect of lipid microdomains and receptor availability in the plasma membrane, and toward the intracellular endolysosomal function and trafficking. We will also investigate the protein interactome of ABCA7 in vivo using unique 3xFlag-ABCA7 iPSC line to determine potential effectors downstream ABCA7 deficiency that could underlie functional alterations and could be targeted to ameliorate disease. We will not only explore the underlying mechanism of one of the strongest risk-increasing AD variants, but we also believe that novel insights gained from this research will be invaluable for understanding the general role of microglia in AD, and therefore for the development of novel therapeutic strategies both in and beyond ABCA7-related backgrounds.

Researcher(s)

• Promoter: Mancuso Renzo

Research team(s)

• VIB CMN - Microglia and Inflammation in Neurological Disorders (MIND) lab

Project type(s)

• Research Project

Abstract

Alzheimer's disease is a complex neurodegenerative disease, pathologically defined by accumulation of extracellular amyloid-beta plaques, and intracellular neurofibrillary tau tangles, combined with neuronal loss, (astro)gliosis and inflammation. Although microglia are the hub of inflammatory events, they do not act alone. Activated microglia have been shown to induce the reactive astrocyte phenotype A1, thus pointing to a role for a coordinated multicellular response in the diseased brain. In this project, I will characterize microglia-astrocyte communication at the single cell level to map their molecular interactome and dissect specific changes that could contribute to AD. I will use an AD mouse models where I will: 1) investigate astrocytes transcriptional profiles in the presence of mouse and human microglia and after microglial ablation using PLX3397. 2) investigate physical cellular interaction with RABID-seq and through single-cell multi-omics, to finally bring to light physical interaction patterns of microglia and astrocyte phenotypes. 3) I will compile a large database of microglia-astrocyte interaction patterns that I will validate in human tissue. I will use this one-of-a-kind biological and computational framework to decipher cell-cell interactions and the molecular basis of cellular states, which will be a pivotal work to finally shed light on AD disease mechanism and provide new therapies.

Researcher(s)

• Promoter: Mancuso Renzo
• Fellow: Bijnens Baukje

Research team(s)

• VIB CMN - Microglia and Inflammation in Neurological Disorders (MIND) lab

Project type(s)

• Research Project

Abstract

Microglia are the resident immune cells of the brain, and contribute to a wide range of cellular processes in both homeostasis and disease. They play a major role in the pathogenesis of Alzheimer's disease (AD), the leading cause of dementia and a major cause of death in the elderly. Microglia closely interact with neurons and modulate their function with high regional specificity, making them potential culprits behind the neuronal damage in AD. However, exactly how microglia and neurons interact and communicate, and how these interactions change in disease is not known. I hypothesize that an early hallmark of AD is the changing interactions between microglia and neighboring cells such as neurons, which contributes to progress of the disease and ultimately results in synaptic and neuronal loss. I will test this in human microglia by using a unique human xenotransplantation model, whereby I transplant human PSC-derived microglial progenitors into the cortex of healthy and APP-NLGF mice; microglia will acquire a human transcriptional and morphological signature, in vivo. Using RABID-seq to trace microglia-centric interactions with single-cell resolution, combined with 10X Genomics, I will precisely map the interactions of distinct subsets of microglia and neurons, to determine homeostatic molecular networks and how cell-to-cell communication is altered in the early stages of AD. Ultimately, this will allow me to identify specific molecular pathways which can be manipulated to support a beneficial function in microglia, while blocking detrimental effects, to protect neurons and arrest disease. Given that microglia are involved in multiple neurodegenerative diseases, finally understanding how they interact with other cells of the CNS will be a valuable resource to the greater neuroscience community in development, ageing, and disease.

Researcher(s)

• Promoter: Mancuso Renzo
• Fellow: Mohebiany Alma Nazlie

Research team(s)

• VIB CMN - Microglia and Inflammation in Neurological Disorders (MIND) lab

Project type(s)

• Research Project

Abstract

Alzheimer's disease (AD) is a neurodegenerative disorder marked by accumulation of extracellular amyloid beta (A?) and intracellular neurofibrillary tau tangles. It is the most common form of dementia, the fifth leading cause of death, and a huge societal and economic burden. Disease-modifying drugs are still limited. Clinical trials for AD focus on clearing A?, the disease-causing agent according to the "amyloid cascade hypothesis". However, these therapies do not consider the downstream contribution of microglia, the immune cells of the brain. Microglia are key players in the "cellular phase theory", which claims that while A? functions as an initial trigger, it is the multitude of cellular changes that lead to disease. I will combine both theories in a new paradigm inspired by one of the most promising treatments in cancer, CAR-T cells, and develop a CAR-microglia system for AD. I will perform a CRISPR-Cas KO screen coupled with an A? phagocytosis assay to study the interactome between A? and human microglia. Hits that lead to altered A? phagocytosis will be used to engineer CAR-microglia cells with high affinity for A?. I will test the efficiency of these cells in clearing A? in vivo using a humanized AD mouse model. Finally, I will provide target engagement readouts of the most promising CAR-microglia with amyloid-PET imaging and CSF analysis. This project will provide new insights on the A?-microglia interactome and deliver an entirely new therapeutic approach for AD.

Researcher(s)

• Promoter: Mancuso Renzo
• Fellow: Jutz Lena

Research team(s)

• VIB CMN - Microglia and Inflammation in Neurological Disorders (MIND) lab

Project type(s)

• Research Project

Abstract

TMEM106B haplotypes have been identified as risk factors for several neurodegenerative diseases such as Frontotemporal Lobar Degeneration with TDP-43 aggregates (FTLD-TDP) and Alzheimer's Disease (AD) and healthy aging, suggesting that they determine neuronal vulnerability. These haplotypes regulate expression of TMEM106B, a lysosomal type-II transmembrane protein, being the risk haplotype causative of a slight expression increase. In this project I hypothesize that subtle changes in TMEM106B expression condition neuronal fitness by dysregulating lysosomal physiology. To study this, I will generate isogenic PSC-derived cortical neurons with different TMEM106B expression levels: a full knockout (TMEM106B-/-) and an inducible-reversible overexpression (TMEM106BOE) model. These will be transplanted in the brains of AppNL-G-F and Grn-/- mice as models of AD and FTLD-TDP, respectively, to analyze how a neurodegenerative environment affects neurons in a TMEM106B expression-dependent manner. Lastly, I also aim to perform an in depth characterization of the lysosomes in these neurons in vitro by analyzing their proteome, trafficking, activity, size and localization and validating these results in the transplanted brain slices. Overall, this project aims to shed light into the molecular mechanism through which TMEM106B expression regulates neuronal vulnerability to disease by integrating neuropathological outcomes observed in the transplanted neurons with lysosomal dysfunctions.

Researcher(s)

• Promoter: Rademakers Rosa
• Co-promoter: Mancuso Renzo
• Fellow: Lastra Osua Miranda

Research team(s)

• VIB CMN - Applied and Translational Neurogenomics

Project type(s)

• Research Project

Abstract

Microglia – the resident immune cells of the brain - are one of the central players in the pathogenesis of Alzheimer's disease (AD). Despite intense efforts in characterizing the transcriptomic and epigenetic response of microglia in AD, it is yet to be defined how they communicate with other cells in the brain and by which mechanism they lead to neuronal alterations. Our overall objective is to define the contribution of human microglia (hMG) to synaptic dysfunction and neurodegeneration in vivo. There are crucial differences between human and mouse microglia, especially in terms of expression of specific AD linked genes, which highlight the necessity of working in humanised systems. In addition, many aspects of the biology of these cells cannot be studied in vitro – for example complex interactions with other cell types. We will use a novel hMG xenotransplantation model that we recently established, and study two of the main functional aspects by which microglia can contribute to the neuropathology of AD, i.e. synaptic elimination and release of soluble inflammatory factors that cause neuronal dysfunction or degeneration. We will apply cutting-edge proximity and metabolic labelling strategies (TurboID) combined with my xenotransplantation model to decipher the precise interactions between hMG and neurons, as well as the complexity of the microglial secretome in AD. We will 1) map the landscape of the microglia-synapse interactions in the healthy and AD brain, using my hMG xenotransplantation model and proximity labelling. 2) Decipher the molecular interactome of hMG and human neurons in AD in vivo, by co-transplanting hMG and neurons and determining their surface interactome. 3) Determine the hMG specific secretome in AD by metabolic labelling of all proteins produced by the transplanted hMG. This project will be the first of its kind to directly study the role of hMG in the AD brain at cellular and molecular level.

Researcher(s)

• Promoter: Mancuso Renzo
• Fellow: Mancuso Renzo

Research team(s)

• VIB CMN - Microglia and Inflammation in Neurological Disorders (MIND) lab

Project type(s)

• Research Project

Abstract

Gene therapy offers the promise of an efficient, single-dose therapeutic solution for many incurable diseases. In reality, despite large efforts of the scientific community and a strong industrial interest, bringing gene therapy to the market has proven challenging. The major hurdles faced by gene therapy come in the form of its safety and efficacy with off- target effects caused by low specificity or inappropriate transgene expression levels. The use of well-designed synthetic regulatory regions, called enhancers, could provide a solution to reach cell type-specificity and high levels of transgene expression. This SBO project proposes to develop new computational and experimental tools, and combine them in a pipeline to identify enhancers in complex tissues. This pipeline will exploit recent advances in single-cell multi-omics, gene regulatory network (GRN) inference, and deep learning. We will use this pipeline to design enhancers that are specifically active in three chosen brain cell types that are of high relevance for gene therapy. Practically, we will: 1) use mouse and human brain samples to generate a single-cell multi- omic atlas; 2) select unique regulatory regions to each cell type and train enhancer models; and 3) design and validate synthetic enhancers in vivo, using massively parallel reporter assays. As a clinically relevant case study, we will focus on microglia enhancers in the context of Alzheimer's disease. In parallel, we will use the AI-based GRN and enhancer models to interpret and prioritize regulatory variation in whole genome sequences, in order to improve diagnosis and prediction of risk and progression for neurodegenerative disease. Our project will yield licensable enhancers, software tools, and diagnostic AI models with direct industrial applicability, and will demonstrate our ability to interpret and generate enhancers specific to any cell type, which can find application in a wide range of diseases, even beyond the scope of our project.

Researcher(s)

• Promoter: Mancuso Renzo

Research team(s)

• VIB CMN - Microglia and Inflammation in Neurological Disorders (MIND) lab

Project type(s)

• Research Project

Abstract

Microglia are central players in Alzheimer's disease (AD). It is yet to be defined by which mechanisms they lead to neuronal alterations. In the healthy brain, microglia secrete trophic molecules that maintain brain homeostasis. Upon activation they release of soluble factors that induce neuronal dysfunction and degeneration. For example, tau pathology and cell death can be induced by microglial inflammatory cytokines. Microglia derived molecules are also found in cerebrospinal fluid of AD patients and correlate with disease progression and outcome. We aim to address a crucial aspect of microglia biology–the secretion of soluble factors that induce or contribute to AD, in vivo. We will use our human microglia xenotransplantation model, combined with cutting-edge proximity labelling strategies to specifically and efficiently label and pool down of secreted molecules. We expect to define the microglial specific secretome in AD, and provide new therapeutic targets and biomarkers for AD.

Researcher(s)

• Promoter: Mancuso Renzo

Research team(s)

• VIB CMN - Microglia and Inflammation in Neurological Disorders (MIND) lab

Project type(s)

• Research Project

Abstract

The goal of this project is to decipher the exact cellular and molecular mechanisms by which human microglia transduce toxicity to neurons in Alzheimer's disease (AD). Genetics revealed that microglia are one of the central players in the pathogenesis of AD. I and others have characterized the phenotypic response of microglia in AD using transcriptomic and epigenetic tools. Nevertheless, it is yet to be defined how microglia communicate with other cells in the brain, by which mechanism they lead to neuronal alterations, and what the particular role of different microglial activation states is in this process. I will investigate the two major functional outputs of microglia - namely physical interaction with synapses and release of inflammatory factors that induce neuronal dysfunction and degeneration - in AD. Determining the role of human microglia in AD comes with technical challenges, including lack of homology between mouse and humans, and limited expression of AD risk genes in mouse microglia. I will use a human microglia xenograft model I pioneered where iPSC-derived cells are transplanted into the mouse brain, and in which they adopt a brain resident phenotype and can be exposed to amyloid-b plaques. I plan to combine xenotransplantation of genetically engineered iPSC-derived microglia and neurons, with stateof- the-art strategies to define the human microglia-synapse protein interactome as well as the cell-specific proteome/secretome. I aim to 1) define the surface protein interactome between human microglia and mouse synapses in vivo, 2) co-transplant human microglia and human neurons in the mouse brain to explore human to human specific aspects of this interaction, and 3) investigate the human microglia specific secretome in AD. This project will be the first of its kind to directly study the role of human microglia in the AD brain at cellular and molecular levels, opening new avenues for the development of new therapeutics and biomarkers to tackle disease.

Researcher(s)

• Promoter: Mancuso Renzo

Research team(s)

• VIB CMN - Microglia and Inflammation in Neurological Disorders (MIND) lab

Project type(s)

• Research Project

Abstract

In the most recent meta-analysis of Alzheimer's disease (AD) genome-wide association studies by Sleegers Lab and collaborators, 39 known and 44 novel variants were reported to have significant genome-wide signals. Pathway- and single cell enrichment analysis points to microglia as the major responsible for this genetic risk. However, none of these risk factors - not even APOE e4 - can explain alone the development of AD. We hypothesized that unknown combinations of these variants may significantly modify the impact of APOE e4 and have the potential to better stratify AD patients and controls. To test this hypothesis, as a proof of concept, we first used multifactor dimensionality reduction and logistic regression over the AD-Belgian-Flemish Cohort to search for the putative combinations (combos) of 85 genetic risk factors. The preliminary analysis identified numerous high-risk statistically significant combos, composed by APOE e4 and low-risk genetic factors. In this project, we aim to validate these fundings in a larger cohort, and provide functional evidence to link these genetic combos to microglial functional alterations. To this end, we are currently running the same pipeline in the largest European cohort of AD patients, the European AD Biobank consortium, where we will at the same time correlate several clinical parameters with the presence of the combos described. To validate the in silico analysis from a functional perspective, we will use cutting-edge CRISPR technology to A) recapitulate top 3 combos' effect in human microglia by multiplex gene editing and B) xenotransplant the edited microglia into the brain of AD-like mice following the MIGRATE protocol recently published by the Mancuso Lab. We believe that the unique combination of GWAS-based significant combos with microglia xenotransplantation technology offers us a way to peer deeper than ever before into the complex genetic etiology of Alzheimer's disease, charting new genetic interactions with APOE and investigating in vivo phenotypical changes in the AD-like brain.

Researcher(s)

• Promoter: Mancuso Renzo
• Co-promoter: Sleegers Kristel

Research team(s)

• VIB CMN - Microglia and Inflammation in Neurological Disorders (MIND) lab

Project type(s)

• Research Project

Abstract

Alzheimer's disease (AD) patients lose their neurons and synapses, followed by their memory, cognition, and grasp on reality. Microglia, the brain's immune cells and mediators of neuroinflammation, eliminate synapses during development and could be key players in the synaptic loss observed in AD. Many AD risk genes are linked to microglial function or are highly expressed in microglia, and microglia respond to, and become activated by, the extracellular Amyloid-ß (Aß) plaques characteristic of AD brains. However, how microglia molecularly interact with neurons and synapses, and how these interactions are affected in AD, is poorly understood, in part because our knowledge on microglia is largely based on rodent models, which do not accurately reflect the role of human microglia in AD. We hypothesize that human microglia interact with synapses through specific ligand-receptor pairs, and that alterations of these interactions are an integral part of the synaptic pathology observed in AD. Here, we will use a novel xenotransplantation model in combination with split-TurboID proximity labeling to identify the proteins that mediate the interactions of human microglia with synapses, and determine how these interactions are affected by AD pathology. To achieve this, we combine the complementary expertise of two labs: the Mancuso lab (VIB-UAntwerp), which specializes in microglial function and pioneered human microglia xenotransplantation in the mouse brain; and the De Wit lab (VIB-KU Leuven), which specializes in synaptic biology and advanced proteomic analysis of cell-cell interactions at synapses. We will address our hypothesis in two key aims: Aim 1) identify interacting proteins at human microglia-synapse contact sites, and determine how these interactions are altered by AD pathology; Aim 2) determine which of these proteins mediates excessive synaptic loss in a mouse model of AD. With this approach we aim to map the molecular nature of human microglia-synapse interactions in vivo, which has not been done before, and generate new insights in how AD pathology alters these interactions. Our goal is to uncover new potential treatment targets for AD. Furthermore, our findings on human microglia-synapse interactions may be relevant in a broader context, as microglia have been implicated in a range of neurodegenerative disease, as well as psychiatric disorders including schizophrenia or autism spectrum disorders.

Researcher(s)

• Promoter: Mancuso Renzo

Research team(s)

• VIB CMN - Microglia and Inflammation in Neurological Disorders (MIND) lab

Project type(s)

• Research Project

Abstract

In the recent meta-analysis of Alzheimer's disease (AD) genome-wide association studies by Sleegers Lab, 39 known and 44 novel variants were reported to have significant genome-wide signals. Pathway- and single cell enrichment analysis points to microglia as the major responsible for this genetic risk. However, none of these risk factors can explain alone the development of AD. I hypothesized that unknown combinations of these variants may have the potential to better stratify AD patients and controls. To test my hypothesis, I used multifactor dimensionality reduction and logistic regression over the AD-Belgian-Flemish Cohort to search for the putative combinations of 85 genetic risk factors. The preliminary analysis identified numerous high-risk statistically significant combinations, mainly composed by APOE ?4 and low-risk genetic factors. To validate my results, I am running the same pipeline in the largest European cohort of AD patients, the European AD Biobank consortium, where I will parallelly correlate several clinical parameters with the presence of the combos described. To functionally validate the in silico analysis, I will employ cutting-edge technology to A) recapitulate top 3 combos' effect in human microglia (MG) by multiplex gene editing and B) xenotransplant the edited MG into the brain of AD-like mice following the MIGRATE protocol recently published by the Mancuso Lab.

Researcher(s)

• Promoter: Mancuso Renzo
• Co-promoter: Sleegers Kristel
• Fellow: Leguía Fauró Gonzalo

Research team(s)

• VIB CMN - Microglia and Inflammation in Neurological Disorders (MIND) lab

Project type(s)

• Research Project

Abstract

Alzheimer's disease (AD) is a complex neurodegenerative disease and the most common form of dementia worldwide. The main pathological hallmark is the extensive accumulation of intracellular amyloid-? plaques and extracellular tau tangles, in which genetics play a fundamental role. Large-scale genetic studies place microglia dysfunction central to the etiology of AD, wherein genetic variants are classified in three major pathways: inflammation, lipid metabolism and endocytic trafficking. Yet, functional data validating the link between these variants, microglia pathways and AD pathology is still lacking. Thus, there is a critical need to understand the contribution of individual risk genes to microglia function. In this project, I aim to elucidate the role of the risk gene ATP-binding cassette transporter, subfamily A member 7 (ABCA7) in AD by uncovering the molecular mechanisms underlying two high-risk variants (odds ratio=2.8,4.5). Therefore, I will generate isogenic cell lines using patient iPSC-derived microglia and cutting-edge CRISPR technologies. I will use our novel humanized chimeric mice model and multi-OMICS approaches to reveal differences between diseased and healthy microglia. In addition, as ABCA7 is a major brain lipid distributor implicated in all three pathways, I will determine the impact of altered lipid homeostasis on said pathways. The outcome of this project will deliver invaluable insights concerning AD research and ultimately novel therapeutic targets.

Researcher(s)

• Promoter: Mancuso Renzo
• Fellow: Premereur Jessie

Research team(s)

• VIB CMN - Microglia and Inflammation in Neurological Disorders (MIND) lab

Project type(s)

• Research Project

Abstract

Frontotemporal dementia (FTD) is the second most common form of dementia. It presents a heterogenous range of neuropathological hallmarks, including aberrant accumulation of TAU, FUS or TDP-43. FTD is also accompanied by inflammatory changes in the brain. Recent data place microglia, the immune cells of the brain, under the spotlight as central player in determining the susceptibility for developing FTD. In this proposal, we aim to define what is the role of one of the most relevant genetic causes of FTD, progranulin deficiency, in microglia using a combination of in vitro and in vivo approaches. We plan to generate independent series of genetically modified stem cells to partially or completely delete progranulin. Then, we will differentiate these cells into microglia to 1) study their basal alterations in in vitro systems, and 2) xenotransplant them in the mouse brain following our MIGRATE protocol. Using these cutting-edge in vivo systems, we expect to determine whether progranulin deficient microglia is sufficient to induce or can significantly modify FTD-like pathology. This project has major implications for our understanding of the cellular and molecular basis of FTD and will open new avenues for tailored treatments to tackle FTD progression.

Researcher(s)

• Promoter: Mancuso Renzo
• Fellow: van Hoek Maxim

Research team(s)

• VIB CMN - Microglia and Inflammation in Neurological Disorders (MIND) lab

Project type(s)

• Research Project

Abstract

Frontotemporal dementia (FTD) is a chronic neurodegenerative disease and is the second most common form of dementia worldwide. FTD is characterized by the build up of abnormal proteins inside nerve cells including TAU, FUS or TDP-43. FTD is accompanied by changes in the immune cells of the brain, the microglia. This inflammation in the brain is referred to as neuroinflammation, which may be a simple consequence of damage to nerve cells or may contribute to disease progression. FTD genetics has recently linked neuroinflammation and susceptibility to develop FTD, suggesting that inflammation might be a driver of the disease opposed to just a consequence. The current proposal is aimed at determining the role of one of the most important genetic causes of FTD, progranulin deficiency, by using novel models where human derived cells are injected into FTD mice, in order to expose those cells to the environment they would find in the pathological brain. By doing this with microglial cells, we will be able to dissect the contribution of progranulin in neuroinflammation and human microglia biology in FTD. These findings will have major implications for our understanding of the progression of human disease and would allow tailored treatments to slow down or arrest FTD progression. This approach addresses a critical component of the pathology of FTD and may yield novel drug targets with the potential to change the disease trajectory and patients' quality of life.

Researcher(s)

• Promoter: Mancuso Renzo

Research team(s)

• VIB CMN - Microglia and Inflammation in Neurological Disorders (MIND) lab

Project type(s)

• Research Project

Abstract

Frontotemporal lobar degeneration (FTLD) is an early-onset form of dementia representing 10-20% of all dementia cases. It is characterized by the loss of neurons in the frontal and temporal lobes resulting in a dramatic impact on core human qualities including personality, insight and verbal communication. Aggregates of the TAR DNA-binding protein 43 (TDP-43) are the hallmark of the most common pathological subtype of FTLD (FTLD-TDP). Importantly, over the past decade, the field of neurodegeneration has shifted from a proteinopathy-centric view towards the concept of a multicellular hypothesis underlying both initiation and perpetuation of disease. Genetic studies have helped significantly in deciphering the cellular substrate of neurodegeneration. In relation to FTLD, the Rademakers lab identified loss-of-function mutations in progranulin (GRN) as one of the major genetic causes of FTLD. They also described TMEM106B genetic variants as the first genetic risk factor for FTLD-TDP and found that TMEM106B protective variants can dramatically reduce the disease penetrance of GRN mutations. GRN and TMEM106B are enriched in separate cellular compartments (microglia versus neurons, respectively), depicting a scenario where different genetic factors interact from multiple cellular compartments. We hypothesise that the genetic risk shapes cellular responses and phenotypes promoting particular disease states in microglia which modify the vulnerability of neurons to degeneration. We propose to investigate this concept in the context of FTLD and age-related TDP-43 neuropathology using the established disease genes GRN and TMEM106B in microglia and neurons, respectively. However, the study of genotype-phenotype interactions in neurodegeneration is not trivial, as there is a limited homology between human and mouse in terms of expression of disease associated genes. To overcome this limitation, the Mancuso lab has developed a model of human iPSC-derived brain cells xenotransplantation to study human relevant genetic traits in a diseased mouse brain environment. In a joined effort from the Rademakers and Mancuso labs, we here propose to investigate the impact of GRN and TMEM106B genetic variants by iPSC microglia and neurons derivation, and xenotransplantation, in FTLD (Grn-/-) and wild type mice. We plan to 1) generate isogenic series of iPSC lines containing GRN and TMEM106B mutations; 2) determine the impact of GRN deficiency in human microglia and determine whether this deficiency is sufficient to induce neuropathology in the mouse brain; and 3) determine if changes in TMEM106B expression in human neurons lead to susceptibility or resilience against degeneration, in vivo. Our studies will provide critical knowledge on the diverse cellular processes underlying FTLD and we expect the novel insights gained from this research to be invaluable for the development of novel therapeutic strategies for FTLD patients and related disorders.  

Researcher(s)

• Promoter: Rademakers Rosa
• Co-promoter: Mancuso Renzo

Research team(s)

• VIB CMN - Applied and Translational Neurogenomics

Project type(s)

• Research Project

Abstract

Genetics shows that microglia are one of the central players in the pathogenesis of Alzheimer's disease (AD). The conceptual framework for the present proposal is that particular genetic risk profiles promote cellular disease states, particularly driving microglia into specific phenotypes that actively contribute to disease. However, the link between these genetic alterations and cellular function is yet to be elucidated. In silico analysis shows the majority of AD-risk genes appear to converge in a limited number of functional pathways, predominantly lipid metabolism. The ATP-binding cassette (ABC) transporter, subfamily A member 7 (ABCA7) is a major regulator of all lipid pathways, and has very a strong AD risk association in GWAS studies (compared to other relevant variants in SORL1 or TREM2). Changes in lipid composition, and likely alterations in ABCA7, may drastically affect important aspects of microglia function in AD – response to inflammatory stimuli and endolysosomal function. Therefore, we aim to decipher the link between ABCA7 genetics and microglial function in AD. There are fundamental differences between human and mouse microglia, especially in terms of expression of specific AD linked genes, which highlight the necessity of working in human systems. Additionally, many aspects of the biology of these cells cannot be easily studied in vitro – for example specific disease challenges (e.g amyloid-beta plaques) or complex interactions with other cell types. We will use a novel human microglia xenotransplantation model we pioneered, and elucidate the role of ABCA7 in human microglia biology in AD in vivo, and unravel the underlying mechanism of one of the highest GWAS-risk variants. We aim to investigate the rare PTC variant (p.Glu709fs; c.2126_2132del) that highly segregates with disease in a Belgian cohort as well as the high-risk pathogenic repeat expansion of a variable number of tandem repeats polymorphism (VNTR) in intron 18 by using patient-derived iPSCs. We plan to 1) Generate of an isogenic series of lines using CRISPR technology, 2) Determine the impact of ABCA7-associated AD-risk in human microglia, and 3) Determine the mechanism underlying the high-risk ABCA7 VNTR variant. The unique strategy we propose, will allow to explore human function in AD beyond current boundaries, and will lead to new therapeutic targets and specific biomarkers to diagnose, monitor and tackle AD. We expect to discover key phenotypic programs, as well as interactors and inflammatory soluble factors that contribute to neuronal dysfunction and degeneration.

Researcher(s)

• Promoter: Mancuso Renzo

Research team(s)

• VIB CMN - Microglia and Inflammation in Neurological Disorders (MIND) lab

Project type(s)

• Research Project

Abstract

Alzheimer's disease (AD) is a complex neurodegenerative disease and the most common form of dementia worldwide. The main pathological hallmark is the extensive accumulation of intracellular amyloid-β plaques and extracellular tau tangles, in which genetics play a fundamental role. Large-scale genetic studies place microglia dysfunction central to the etiology of AD, wherein genetic variants are classified in three major pathways: inflammation, lipid metabolism and endocytic trafficking. Yet, functional data validating the link between these variants, microglia pathways and AD pathology is still lacking. Thus, there is a critical need to understand the contribution of individual risk genes to microglia function. In this project, I aim to elucidate the role the risk gene ATP-binding cassette transporter, subfamily A member 7 (ABCA7) in AD by uncovering the molecular mechanisms underlying two high-risk variants (odds ratio=2.8, 4.5). Therefore, I will generate isogenic cell lines using patient iPSC-derived microglia and cutting-edge CRISPR technologies. I will use our novel humanized chimeric mice models and multi-OMICS approaches to reveal differences between diseased and healthy microglia. In addition, as ABCA7 is a major brain lipid distributor implicated in all three pathways, I will determine the impact of altered lipid homeostasis on said pathways. The outcome of this project will deliver invaluable insights concerning AD research and ultimately novel therapeutic targets.

Researcher(s)

• Promoter: Mancuso Renzo
• Fellow: Bijnens Baukje

Research team(s)

• VIB CMN - Microglia and Inflammation in Neurological Disorders (MIND) lab

Project type(s)

• Research Project

Abstract

Gene therapy offers the promise of an efficient, single-dose therapeutic solution for many incurable diseases. In reality, despite a large effort of the scientific community and a strong industrial interest, bringing gene therapy to the market has proven challenging with only few products currently available. The major hurdles faced by gene therapy come in the form of its safety and efficacy with off-target effects caused by low specificity or inappropriate transgene expression levels. The use of well- designed synthetic cis-regulatory regions, called enhancers, could provide a solution to reach cell type specificity and high level of transgene expression. This SBO proposal aims at developing new computational and experimental tools and combining them in a pipeline aimed at identifying enhancers in complex tissues. This pipeline will exploit recent advances in single-cell multi-omics and deep learning. Next, we will utilize this pipeline to identify enhancers specific to three chosen brain cell types that are of high relevance for gene therapy. Particularly, our focus will be on excitatory neurons in the cortex, dopaminergic neurons, and microglia. For those three cell types we will: i) use mouse and human brain samples to generate a single-cell multi- omic profile; ii) select unique regulatory regions to each cell type and train enhancer models; iii) validate enhancer activity and specificity using in vivo massively parallel reporter assays. In parallel, we will use the AI-based enhancer models to create synthetic enhancer sequences with improved specificity and activity. Finally, we will focus on microglia as a case study for clinical translation in Alzheimer's disease (AD). We will xenotransplant human microglia progenitors to the mouse brain and use our optimized microglia enhancers to target APOE in normal and AD mouse brains. Our approach should demonstrate our ability to generate enhancers specific to any cell type and yield our first licensable enhancers. Roadmap for valorization: 1. An off-the-shelf library of proprietary, potent, compact, synthetic enhancers that control transgene expression in (1) excitatory cortical neurons; (2) dopaminergic neurons; or (3) microglia. Licensees will be given the option to i) test these enhancers in their proprietary systems, or ii) collaborate with selected consortium partners to finetune enhancers in order to meet their needs. 2. A fully integrated and validated EnhancerDesign platform, driven by an innovative combination of single cell analysis, massively parallel screening assays, and an improved AI-based bioinformatics toolbox. While the EnhancerDesign platform can be applied to all sorts of cells/tissues, we focus on neuronal cell types, for the following reasons: 1. these cell types are linked to neurodegenerative diseases with a high unmet medical need and huge market potential; 2. the CNS is the tissue with the highest cellular diversity (i.e. ambitious case to validate the EnhancerDesign pipeline); and 3. our timing is in line with the increasing interest of biotech and pharma companies in the CNS gene therapy field. A licensing strategy based on a non-exclusive access to the technology platform and resulting enhancers is being preferred. Where larger co- development deals might require exclusivity, an exclusive license within a field that is defined by (1) the therapeutic gene that gene therapy aims to deliver, in order to treat (2) a specified disease, might be an option.

Researcher(s)

• Promoter: Mancuso Renzo

Research team(s)

• VIB CMN - Microglia and Inflammation in Neurological Disorders (MIND) lab

Project type(s)

• Research Project

Abstract

Over the last decade, organoids emerged as an attractive middle ground between 2D cell cultures, which do not fully recapitulate the 3D environment and animal models, which pose technical and ethical limits. In particular, cerebral organoids are emerging as the next step in patient-derived in vitro models for both neurodevelopmental as well as neurodegenerative diseases. However, cerebral organoids have mostly been based on neuronal cells alone, while evidence increases that the role of non-neuronal types (microglia, astrocytes, endothelial cells) is critical in these conditions. Integration of these cell types will more closely mimic the in vivo cellular environment in health and disease and constitutes the major challenge of this project. The established neuroimmune organoid technology will find a wide range of applications as models for studying fundamental mechanisms underlying cellular biology and genetic pathophysiology as well as for efficient drug screening.

Researcher(s)

• Promoter: Sleegers Kristel
• Co-promoter: Jordanova Albena
• Co-promoter: Malysheva Valeriya
• Co-promoter: Mancuso Renzo
• Co-promoter: Weckhuysen Sarah

Research team(s)

• VIB CMN - Genetics of Alzheimer’s Disease

Project type(s)

• Research Project

Abstract

The key to explaining neurodegenerative disease may lie in our understanding of microglial interactions with other CNS cells, which I plan to unravel using a combined human xenotransplantation model and barcoded rabies virus approach. Microglia (MG) are the resident immune cells of the brain, constantly extending and retracting their processes to monitor the surrounding environment. However they are not mere responders to disease or damage. Involved in neurogenesis, regulation of neuronal activity, and synapse formation and pruning, MG play important roles in shaping the brain and its normal functioning. However, we still do not know the cellular and molecular pathways responsible for maintaining a homeostatic profile in microglia, and how these change in disease. Nor do we know how communication with other cell types helps to regulate the microglial response in the development of disease such as Alzheimer's (AD). AD is the most common form of dementia, making up 60% of all cases, and extensive studies to understand the etiology has revealed MG as key players in AD, which is characterized by multiple neuropathologies. Using a novel human MG xenotransplantation model from my host group and cutting-edge techniques to label cells that have directly interacted with MG, my project focuses on the interplay between hMG and other cells of the brain. I will identify molecular and transcriptional networks involved in MG-centric interactions and how these contribute to AD development.

Researcher(s)

• Promoter: Mancuso Renzo
• Fellow: Mohebiany Alma Nazlie

Research team(s)

• VIB CMN - Microglia and Inflammation in Neurological Disorders (MIND) lab

Project type(s)

• Research Project

Abstract

Alzheimer's disease (AD) is a complex neurodegenerative disease and the most common form of dementia worldwide. The main pathological hallmark is the extensive accumulation of intracellular amyloid-? plaques and extracellular tau tangles, in which genetics play a fundamental role. Large-scale genetic studies place microglia dysfunction central to the etiology of AD, wherein genetic variants are classified in three major pathways: inflammation, lipid metabolism and endocytic trafficking. Yet, functional data validating the link between these variants, microglia pathways and AD pathology is still lacking. Thus, there is a critical need to understand the contribution of individual risk genes to microglia function. In this project, I aim to elucidate the role of the risk gene ATP-binding cassette transporter, subfamily A member 7 (ABCA7) in AD by uncovering the molecular mechanisms underlying two high-risk variants (odds ratio=2.8,4.5). Therefore, I will generate isogenic cell lines using patient iPSC-derived microglia and cutting-edge CRISPR technologies. I will use our novel humanized chimeric mice model and multi-OMICS approaches to reveal differences between diseased and healthy microglia. In addition, as ABCA7 is a major brain lipid distributor implicated in all three pathways, I will determine the impact of altered lipid homeostasis on said pathways. The outcome of this project will deliver invaluable insights concerning AD research and ultimately novel therapeutic targets.

Researcher(s)

• Promoter: Mancuso Renzo
• Fellow: Premereur Jessie

Research team(s)

• VIB CMN - Microglia and Inflammation in Neurological Disorders (MIND) lab

Project type(s)

• Research Project

Abstract

Alzheimer's disease (AD) is a chronic neurodegenerative disease and is the most common form of dementia worldwide. AD is characterized by the build up of abnormal protein, in the form of amyloid beta plaques and tau tangles. AD is accompanied by changes in the immune cells of the brain, the microglia. This inflammation in the brain is referred to as neuroinflammation, which may be a simple consequence of damage to nerve cells or may contribute to disease progression. Recent genetic studies reveal a link between neuroinflammation and susceptibility to develop AD, suggesting that inflammation might be a driver of the disease opposed to just a consequence. The current proposal has two complementary approaches aimed at determining the role of neuroinflammation in human AD by generating novel models where human derived cells are injected into AD mice, in order to expose those cells to the environment they would find in the pathological brain. By doing this both with neural and microglial cells, we will be able to dissect the contribution of neuroinflammation in the AD brain in two crucial human cell types. These findings would have major implications for our understanding of the progression of human AD and would allow tailored treatments to slow down or arrest AD progression. This approach addresses a critical component of the pathology in the AD brain and might yield novel drug targets with the potential to change the disease trajectory and patients' quality of life

Researcher(s)

• Promoter: Mancuso Renzo

Research team(s)

• VIB CMN - Microglia and Inflammation in Neurological Disorders (MIND) lab

Project type(s)

• Research Project

Abstract

Alzheimer's disease (AD) is the most common form of dementia worldwide. AD is characterized by the build up of abnormal protein, in the form of amyloid beta plaques and tau tangles. AD is accompanied by changes in the immune cells of the brain, the microglia. This inflammation in the brain is referred to as neuroinflammation, which may be a simple consequence of damage to nerve cells or may contribute to disease progression. Genetic studies reveal a link between neuroinflammation and susceptibility for AD, suggesting that inflammation might be a driver of the disease opposed to just a consequence. All these genes seem to be involved in the metabolism of lipids and cholesterol, pointing to a link between microglial function and lipid metabolism. We aimed at determining the link between AD genetic risk, microglia and lipid metabolism by combining novel models where human derived cells are injected into AD mice, and cutting edge single cell RNA sequencing for in depth analysis of microglial function. By doing this, we will be able to dissect the contribution of microglia and lipid metabolism in the AD brain in a crucial human system. These findings would have major implications for our understanding of the progression of human AD and would allow tailored treatments to slow down or arrest AD. We plan to addresses critical components of the pathology and might yield novel drug targets with the potential to change the disease trajectory and patients' quality of life.

Researcher(s)

• Promoter: Mancuso Renzo

Research team(s)

• VIB CMN - Microglia and Inflammation in Neurological Disorders (MIND) lab

Project type(s)

• Research Project

Abstract

Early Career Researcher Award by the French Alzheimer's Research Foundation for my contributions to the field or microglia biology and neuroinflammation in Alzheimer's disease, with special focus on the role of APOE in human microglia, in vivo.

Researcher(s)

• Promoter: Mancuso Renzo

Research team(s)

• VIB CMN - Microglia and Inflammation in Neurological Disorders (MIND) lab

Project type(s)

• Research Project