April 2021 – April 2025

FNR PRIDE DTU I2TRON program

Dr Ibrahim BOUSSAAD, Vera TSLAF

Feng HE (LIH), Markus OLLERT (DII – LIH)

It has been established that DJ-1 deficiency alters peripheral T cell compartment leading to an increase of naïve T cells and reduction of memory T lymphocytes and Tregs (Danileviciute et al., 2022; Zeng et al., 2022). Using human induced pluripotent stem cells (iPSC)-derived DJ-1 deficient neurons and isogenic control, we aim to investigate whether DJ-1 deficient neurons demonstrate an altered susceptibility to immune milieu. Immune system relevant markers and pathways were examined using RNA sequencing (RNA-seq) data obtained from the isogenic neuronal pair, and the differential expression of the identified markers is being currently validated in the in vitro cultures. Additionally, via in vitro co-culture model between the isogenic neuronal pair and primary human T lymphocytes, we aim to investigate whether DJ-1 deficient neurons show altered susceptibility to T cell immune response.

ongoing

Michael J Fox Foundation

 Dr. Jochen OHNMACHT

Center for Systems and Therapeutics and the Taube/Koret Center for Neurodegenerative Disease Research at Gladstone Institutes

Parkinson’s disease (PD) is the most frequent neurodegenerative movement disorder. While several genes causative for familial forms of PD as well as risks factors have already been identified, it is still unclear how patient’s genomes shape their predisposition to develop PD. For example, despite sharing the same mutation in the LRRK2 gene – the most common cause of late-onset familial PD – carriers display broad variation in severity of symptoms as well as the time of disease onset. To address this observed and unexplained variation, the LCSB takes part in an international effort, collaborating with partners in several European countries, to study families sharing both a history of PD and a G2019S mutation in the LRRK2 gene. By comparing clinical data and genetic profiles of patients a list of susceptibility factors has been generated (ongoing collaboration with Dr. Enrico Glaab and Dr. Patrick May at the LCSB). We are investigating the effects of these susceptibility factors on PD-related phenotypes using iPSC derived dopaminergic neuron cultures from PD patients and healthy controls .

January 2022 to August 2024

Fonds National de Recherche (FNR) – (CORE Junior grant to Giuseppe ARENA)

Dr. Giuseppe ARENA, Alexandre BARON

Enza Maria VALENTE (University of Pavia, Italy), Miriam CNOP (Université Libre de Bruxelles, Belgium), Axel METHNER (University of Mainz, Germany)

Epidemiological studies indicate that patients with T2D have an increased risk of developing Parkinson’s disease (PD). These two age-related chronic diseases share similar alterations in essential biological processes and molecular networks, suggesting common mechanisms underlying their pathogenesis. Mutations in the mitochondrial kinase PINK1 are the second most frequent cause of autosomal recessive PD. Based on the compelling evidence implicating PINK1 also in T2D, we decided to use complementary PINK1-deficient cellular models (PINK1-mutant iPSC-derived dopaminergic neurons and PINK1-silenced pancreatic β-cells) as prototype to decipher the cellular alterations leading to neurodegeneration in PD and β-cells failure in T2D, trying to clarify the functional interdependencies between the two diseases. Findings obtained from this project will allow the identification of novel potential therapeutic targets, thus contributing to patient-based biomedical research in both fields.

ongoing

Fonds National de Recherche (FNR) within the PEARL Excellence Program [FNR/P13/6682797], Jean Think Foundation Luxembourg, FNR [AFR PhD 12447024], Jump.

Dr. Ibrahim BOUSSAAD, François MASSART, Pauline MENCKE

Johannes MEISER

Familial forms of Parkinson’s disease (PD) offer the opportunity to generate human cell models based on induced pluripotent stem cell (iPSC) technology to study the pathophysiology of the disease. These models are used to identify molecular pathways involved in PD and to gain general insights not only into familial but also into idiopathic PD. One of the causes of familial PD is homozygous loss-of-function mutations of DJ-1. DJ-1, a protein encoded by the gene PARK7, has broad biological functions including effects on mitochondrial and lysosomal homeostasis (Krebiehl et al.). We have previously shown that fibroblasts obtained from PD patients carrying the homozygous mutation c.192G>C in the DJ-1 gene display a phenotype of impaired mitochondrial respiration, increased intra-mitochondrial reactive oxygen species, reduced basal autophagy and the accumulation of defective mitochondria.

In order to study the effect of DJ-1 loss of function on PD target cells, midbrain-specific dopaminergic (mDA) neurons, astrocytes and microglia, we have generated iPSC from these fibroblasts that are used in state-of-the-art differentiation protocols. Using pairs of disease-specific and isogenic control iPSC we’re identifying cellular phenotypes that can be used as read out for small chemical compound library screens. Furthermore, we are studying the metabolic implications for the cell caused by loss of DJ-1 in PD cell models and caused by DJ-1 upregulation in glioblastoma multiforme cell models.
Results have been published in Science Translational Medicine in September 2020.

October 2023 – October 2024

FNR (Mirisk-PD CORE grant to Rejko Krüger) and “King Baudouin Foundation United States” (to Rejko Krüger and Giuseppe Arena)

Dr. Giuseppe ARENA, Dr. Daniele FERRANTE, Alexandre BARON

Acurex Biosciences

Miro1 is a cytosolic calcium sensor anchored to the outer mitochondrial membrane (OMM) that plays a pivotal role in the regulation of mitochondrial dynamics and quality control (doi:10.3389/fneur.2020.00587). We identified and functionally characterized first PD-associated heterozygous mutations in the RHOT1 gene encoding Miro1: (i) c.815G>A à Miro1 R272Q; (ii) c.1348C>T à Miro1 R450C; (iii) c.1290A>G à Miro1 T351A; (iv) c.2067A>G à Miro1 T610A. Our findings in primary skin fibroblasts pointed to calcium dyshomeostasis as a major shared hallmark between the four PD-related mutations in patients with a positive family history of PD (doi:10.1089/ars.2018.7718; doi:10.3390/jcm8122226).

Moreover, induced pluripotent stem cells (iPSC)-derived neurons and 3D midbrain organoids from the Miro1 R272Q patient displayed impaired mitochondrial bioenergetics and higher α-synuclein levels compared to the respective isogenic, gene-corrected controls. This was accompanied by a significant loss of dopaminergic neurons both in Miro1 R272Q organoids and in the substantia nigra of 15-months old mice expressing the orthologous, Miro1 R285Q mutation, finally leading to impaired anterograde procedural memory (doi:10.1101/2023.12.19.571978).

We are now extending our analyses to iPSC-derived neurons from PD patients carrying distinct Miro1 mutations in different functional domains of the protein, aimed at exploring the potential interplay between Miro1 and α-synuclein, including the modulations of calcium channels.

February 2023 – ongonig

FNR NextImmune2 Doctoral Training Unit (PRIDE21/16749720/NEXTIMMUNE2), LCSB Tandem Funding Scheme

Ioanna BOUMPOUREKA, Dr Vyron GORGOGIETAS

Prof. Dr. Wim VANDENBERGHE (KU Leuven), Dr. Enrico GLAAB (LCSB)

Mutations in the vacuolar protein sorting 35 ortholog (VPS35) gene, encoding the core component of the retromer complex, have been identified as a cause of late-onset autosomal dominant Parkinson’s disease (PD). So far, a single missense mutation, Asp620Asn (D620N), has been unambiguously identified to cause PD in multiple individuals and families worldwide (Zimprich et al., 2011;  Vilariño-Güell et al., 2011). The retromer is responsible for the trafficking and localization of various proteins within the cell, via two key pathways: the endosome-to-Golgi retrograde transport pathway and the endosome-to-cell-membrane recycling pathway.

Various retromer cargos have been identified to date, pointing to the retromer’s indirect, yet important role in numerous cellular functions (Williams et al., 2017). The exact molecular mechanisms underlying VPS35-induced neurodegeneration are still under investigation. Our team has previously shown perturbed mitochondrial respiration, lysosomal dysfunction, and a-synuclein accumulation in induced pluripotent stem cell (iPSC)-derived neurons (Hanss et al., 2021). Recently, several studies have linked VPS35 deficiency to perturbed microglial activation and inflammatory phenotypes in immortalized cell lines (Lucin et al., 2013; Yin et al., 2016) and mouse models of Alzheimer’s disease and ischemic stroke (Ren et al., 2022; Ye et al., 2019).

However, no studies so far have assessed the roles of mutant VPS35 in microglial models of PD. With this project, we aim to uncover the novel role of the retromer in immune regulation in iPSC-derived microglial cells from patients with familial PD, and their respective isogenic controls, using wet lab techniques, integrated proteomics and metabolomics experiments, and bioinformatics tools.

Poster presentation ‘Uncovering the role of p.D620N VPS35 in immune regulation in a patient-derived microglial model of familial Parkinson’s disease’ in the EMBO Workshop ‘Microglia in Health and Disease’, Genoa, May 2024.

2018 – 2024

FNR BRIDGES (MOTASYN), FNR INTERCORE (MAMASYN)

Dr. Vyron GORGOGIETAS, Dr. Ibrahim BOUSSAAD, Dr. Jochen OHNMACHT, Ioanna BOUMPOUREKA

Ksilink (France), Prof. Przedborski (Columbia University), Dr. Michel MITTELBRONN (LCSB/UL), Dr. Anne GRUNEWALD (LCSB/UL)

Following the discovery that SNCA encodes α-synuclein (Polymeropoulos et al., 1997) in an A53T autosomal dominant form of inheritance in a familial case of PD, a further five other point mutations have been identified in families with hereditary PD: A30P (Kruger et al., 1998), E46K (Zarranz et al., 2004), H50Q, (Appel-Cresswell et al., 2013), G51D (Lesage et al., 2013) and A53E (Pasanen et al., 2014). Furthermore, duplications and triplications of SNCA have been identified leading to the clinical manifestation of the disease, with the triplication of SNCA leading to an earlier disease onset and increased diseased severity. Several Genome-Wide Association Studies (GWAS) have also implicated the variability at the SNCA locus as a major risk factor in idiopathic PD (Simon-Sanchez et al., 2009). The oligomerization of the monomeric α-synuclein into toxic aggregates of amyloid-like fibrils is one of the main pathogenic features of α-synuclein, which is a major constituent of LB (Spillantini et al., 1998).

Our panel of patient-derived iPSC lines includes SNCA point mutations (A30P, A53T) or gene copy number alterations (duplication and triplication). We have generated a set of isogenic corrected cell lines and have introduced SNCA point mutations into age- and gender-matched healthy controls. Exploring PD-related phenotypes in dopaminergic neuron cultures, our particular focus is on aSyn roles at membranes, e.g. mitochondria/ER and exosomes.

Based on these phenotypes we are developing assays to screen in high-throughput small molecule libraries for early drug discovery and drug repurposing studies. In this context, using patient-derived SNCA triplication-derived neurons and their isogenic controls, we were able to identify phenotypic and molecular (mitochondrial)-based profiles and perform drug-repurposing studies, based on these readouts. We identified and validated the effect of promising compounds and we proposed the Mode-of-Action, respectively. Moreover, using the patient-derived SNCA mutations (A53T, A30P) neurons, we were able to validate-characterize these models and identify multiple neuronal phenotypes, that can be further used both for evaluation of the effect of α-synuclein in the ER-to-mitochondria axis and for drug repurposing studies. These projects were collaborations between our group and industrial partners and/or other international research institutes.

Considering the role of α-synuclein spreading in PD’s pathophysiology, we study the role of exosomes in PD. Scientists progressively study exosomes as they are a great, innovative, and promising tool for a better understanding of Parkinson’s disease (PD), but also a great tool for prognosis, diagnosis, and even drug treatment. Exosomal-based study of PD is promising, as it can reveal the spreading of PD pathophysiology in the brain. Based on our results, we are able to deeper understand the mechanisms participating in α-synuclein spreading and identify exosomal cargo alterations (exosomal “signatures”), that can possibly reveal putative PD-related biomarkers (including proteins, DNAs, RNAs, metabolites and lipids). Although we use PD-patient iPSC-derived neuronal models for our exosomal studies, our procedures can further be applied to clinical samples, as prognostic and diagnostic tools. Exosomal signatures can be correlated with specific PD cases, and response to treatment against PD symptoms, but can also be used as targets for new therapeutic approaches.

January 1st, 2023 (Scientific start date: February 1st), for 3 years

ERA PerMed project funded by INTER/FNR

Dr. Rejko Krüger, Dr. Yahya Salem, François Massart

Università degli Studi di Milano (Italy), IRCCS Istituti Clinici Scientifici Maugeri di Milano (Italy), Lund University (Sweden),Commissariat à l’Energie Atomique et aux énergies alternatives (France), University of Copenhagen (Denmark), The Danish Parkinson’s Association (Denmark), University of Zagreb (Croatia)

Evidence shows that individuals affected by idiopathic REM (Rapid Eye Movement) sleep behavior disorder (iRBD) have a high risk of conversion to Parkinson’s disease, dementia with Lewy bodies, or multiple system atrophy. Despite sharing a cellular pathological hallmark, the aggregation of alpha-synuclein, and some clinical features in the early stages, these conditions show different phenotypes in later stages with significant therapeutic and prognostic consequences for the patients.

The consortium DEEPEN-iRBD aims to develop a pathogenicity model for prediction of phenoconversion utilizing pre-clinical/clinical research and data analysis, taking into account a personalized medicine approach, based on the individual’s unique characteristics and optimization of strategies for the prevention, diagnosis and treatment of the individuals rather than the disease.

In this project, both existing and newly acquired data will be integrated, including advanced clinical assessments, physiological signal recordings, molecular markers derived from body fluids, skin biopsy, and iPSC-derived brain cells, in order to identify a specific profile at a very early stage, that is a prodromal phase, for each of the above conditions. This would ultimately allow to define a model for early risk stratification, diagnosis, treatment, and prognosis of patients with iRBD. 

A further important objective of the project deals with the ethical and social aspects of screening people in a prodromal stage of the diseases and of communicating the screening results.

Communication channels:

Twitter account-project: @DEEPEN_iRBD, https://twitter.com/DEEPEN_iRBD

LinkedIn account: DEEPEN iRBD, DEEPEN-iRBDLinkedIn

Main implications of the University of Luxembourg

In WP1 “Management and dissemination”, UniLu is a co-lead in the project management and a member of the project management committee responsible for establishing and providing documents and technical support. UniLu is a lead in data protection, will coordinate the establishment of the Data Management Plan and will support the consortium with the IT infrastructure at UNILU that will be used in sharing and distributing data across partners.

In WP2 “Molecular and neurophysiological profiling and clinical data integration”, UniLu is the lead of the WP and a co-lead in clinical data processing of data from individuals with polysomnography (PSG) validated iRBD participating in nation-wide REM-Sleep Behaviour Disorder Study. The iRBD cohort will be assessed on an annual basis (pre- and post-phenoconversion) with deep clinical (neurological and neuropsychological test battery) assessment and accompanied by biofluid collection and skin biopsy, thus patients be seen in this context. Data from iRBD patients with high risk of conversion will be complemented by longitudinal data and biosamples from more than 800 patients with PD, MSA and DLB from the Luxembourg Parkinson’s Study (NCER-PD) collected since 2015. UNILU will stratify iRBD cohort with NeuroChip (Illumina) and targeted sequencing of GBA gene (PacBio) to provide and cover the most frequent genetic risk factors in iRBD as well as in PD and DLB. Overnight PSG will be performed and interpreted according to standard guidelines at UNILU. Evaluation of the cardiovascular neural control in iRBD patients will be performed from the PSG recordings collected by UNILU.

In WP3 “Bio-samples processing”, UniLu will provide biological samples from patients whose pathology is characterized. In addition, UniLu provides and characterizes iPSC-derived brain cell types

generated from fibroblasts of RBD patients and healthy controls and will transfer the data obtained to Lund University for pathogenicity model implementation and prediction of phenoconversion from iRBD.

Press release:

ERA PerMed published a newsletter on the ERA PerMed website with the results of this call and public abstracts of the funded projects: erapermed/2023/Newsletter

EraPerMed-DEEPEN-iRBD

The tweet from ERA PerMed can be found here: twitter.com/ERANET_PerMed

Parkinson.lu/DEEPEN-iRBD

UNILI/DEEPEN-iRBD

February 2019 – February 2027

Fonds National de Recherche (FNR) (Mirisk-PD CORE to Rejko Krüger and AFR bilateral to Rejko Krüger and Anne Grünewald)

Dr. Giuseppe ARENA, Alexandre BARON, Iñigo Yoldi BERGUA, Vyron GORGOGIETAS

Christine KLEIN, Joanne TRINH & Philip Seibler (University of Lubeck), Bas BLOEM & Marcel VERBEECK (Radboud University Medical Centre, Nijmegen, The Netherlands), Thomas FOLTYNIE (Institute of Neurology, University College London), Oliver BANDMANN & Thomas PAYNE (Sheffield University), Manu SHARMA & Ashwin Ashok Kumar Sreelatha (University of Tuebingen), Nico DIEDERICH (Centre Hospitalier de Luxembourg), Ziv GAN-OR & Ted FON (McGill University, Montreal, Canada), Zied LANDOUSLI, Dajana GROSSMANN, Armelle VITALI, Sylvie DELCAMBRE, Paul ANTONY, Ibrahim Boussaad, Dheeraj Reddy BOBBILI, Lukas PAVELKA, Enrico GLAAB, Patrick MAY & Anne GRÜNEWALD (University of Luxembourg).

Given the large body of evidence pointing to mitochondrial dysfunction and lysosomal impairment as causative events in PD pathogenesis, these two organelles emerged early as logical targets for disease-modifying treatments in PD.  However, to date, all PD clinical trials applying drugs targeting mitochondrial (e.g., Coenzyme Q) or lysosomal (e.g., Ambroxol) pathways to unselected patient groups failed. In this light, genetic stratification of mitochondrial or lysosomal risk could help to define subgroups of idiopathic PD (iPD) patients, who may be more responsive to therapeutic compounds targeting mitochondrial or lysosomal defects, respectively. Here, we use computational approaches based on mitochondria- or lysosome-specific polygenic risk scores (mitoPRSs, lysoPRSs) to assess the synergistic effect of common variants in nuclear-encoded mitochondrial or lysosomal genes on PD risk. Importantly, we functionally validate PRS profiles in cellular models from iPD patients with high or low risk, paving the way for translating genetic prediction into more targeted therapeutic treatments.

Read the paper