Autosomal dominant polycystic kidney disease (ADPKD) is the most common hereditary kidney disease, with an estimated prevalence of ~1 in 1000 to 1 in 2500 individuals. Its course is characterized by the development of multiple kidney cysts, causing progressive loss of kidney function and frequently leading to end-stage kidney disease during or after the sixth decade. The most severe extra renal manifestation in ADPKD is the development of intracranial aneurysm (IA). IA prevalence is fivefold higher in ADPKD than in the general population, with estimates ranging from 9 to 12%. The main complication of IAs is subarachnoid hemorrhage following rupture, with a mean age of 41 years in ADPKD patients versus 51 years in the general population, and a high rate of mortality and morbidity. The only identified risk factor for IA in ADPKD is a familial history of unruptured or ruptured IA, with a prevalence of 22% in patients with a familial risk of IA, versus 6-8% in the absence of family history. Until now, despite strong familial clustering of IA cases in some ADPKD pedigrees, no genetic factor associated with the development of IA has been identified. Indication, timing, and frequency of IA screening in ADPKD patients are not well defined. In this translational research project, we aim to identify the genetic determinants of IA formation in ADPKD patients. The GENOVAS-PKD study will take advantage of international and local ongoing collaborations, allowing us to gather the largest cohort of ADPKD patients affected by IA worldwide. We will evaluate the respective influences of genetic and non-genetic risk factors, and revisit the genic and allelic influence of the ADPKD genes on IA risk. We will perform whole exome-sequencing in informative pedigrees and in a large cohort of unrelated ADPKD individuals with a past history of IA to identify candidate genes, and we will employ targeted next-generation sequencing to analyze these candidate genes in a large replication cohort. We will develop first-line functional assays to understand the pathogenic mechanisms resulting from the variants identified. Through the identification of genes influencing the development of IA in ADPKD patients, our hope is to improve the pre-symptomatic screening strategy, which will alleviate the mortality and morbidity burdens associated with this severe complication. Understanding the genetic determinants of IA formation will provide a better understanding of IA pathogenesis, hopefully translating into the discovery of pathways amenable to disease-modifying therapies.

Mutations in the gene encoding the CFTR protein affect mucus-producing epithelia, including the lungs and gastrointestinal tract. In the lungs, viscous mucus obstructs the bronchi, creating an environment conducive to chronic bacterial infection, which leads to the destruction of the lung parenchyma and the death of patients with Cystic Fibrosis. To date, ~65% of patients can be treated with drugs based on Ivacaftor (ex Kaftrio). Despite significant clinical benefits, in particular respiratory, have been reported, side effects may lead to the reduction and/or discontinuation of treatment due to the hepatic and pancreatic insufficiency inherent in the disease. Moreover, ~35% of patients with very rare mutated alleles are currently without a therapeutic solution. This may explain why gene therapy is a credible option to treat all patients regardless of their mutations. Since lung failure is responsible for mortality, gene therapy must be aerosolized to deliver the transgene. However, the high viscosity of CF mucus traps both viral and non-viral vectors and limits their clinical impact. In this context, extracellular vesicles (EVs) exhibiting a natural ability to diffuse into dense extracellular matrices, are relevant candidates to diffuse into mucus. The interest of ES derived from MSCs has been established in several pulmonary pathologies, mainly due to their natural antibacterial, anti-inflammatory and anti-fibrotic effects. Few studies have reported delivery of MSC-EV to the lungs after nebulization, demonstrating their safety but also showing heterogeneous results likely due to lack of stability during nebulization. Also, our project aims to nebulize the hMSC-EVs optimized to deliver an mRNA encoding CFTR and to evaluate them in the specific CF context.

Pulmonary Arterial Hypertension (PAH) is a rare, incurable and deadly disease of the pulmonary vessel. It is defined by an elevation of pulmonary arterial pressure, due to progressive and obstructive remodeling of small pulmonary arteries, leading to right heart failure. Existing treatments target vasoconstriction, are not curative and it remains an unmet need for anti-remodeling strategy. The only outcome is lung transplantation, with a survival of 50% at 5 years. A new player related to respiratory diseases, the pulmonary microbiota, is not yet taken into account in PAH. Asthma, COPD, idopathic fibrosis, cystic fibrosis are related to a pulmonary pathobiome with a decrease in diversity promoting progression of the disease, acute exacerbations and mortality, thus opening the way to new therapeutic avenues. LUMI aims to explore the pulmonary microbiota as a new actor directly impacting vascular remodeling and the progression of PAH, via its metabolites. The specific objectives are: a/ to determine the physiological impact of the microbiota on the architecture of the developing pulmonary vascular tree, b/ to translationally characterize the pulmonary microbiota in experimental and human PAH and c/ to evaluate the physiopathological and therapeutic consequences of this microbiota and its metabolites on vascular remodeling leading to PAH in a pre-clinical model. One of the challenges will be to demonstrate the link between pulmonary bacterial species, their metabolites and pulmonary vascular remodeling. The other challenge is how to ensure the translation of the initial observations on the pulmonary microbiome composition in PAH patients to pre-clinical models of PAH. Thus LUMI has emerged as a multidisciplinary consortium that brings together 3 complementary expert partners P1 (INSERM UMR_S 999, Paris Sud University/Paris Saclay University), P2 (MICALIS-INRA), and P3 (INSERM UMR_S 1078, UBO), respectively in the fields of Biology/Medicine (Pathophysiology of PAH and Therapeutic Innovation), Functional Metagenomics (METAFUN), and Lung Ecosystem (16S Metagenetics / Metatranscriptomics and MUCOBIOME Bioinformatics pipeline). LUMI has designed a research strategy focused directly on these metabolites, through both a targeted and comprehensive approach, to address these challenges with the unique opportunity of access to explanted lung tissue from PAH patients in relation to the National Reference Center hosted by P1. We believe that whatever the mechanisms leading to altered composition of the pulmonary microbiota – disruption of pulmonary homeostasis, bacterial translocation from intestine along the gut-lung axis, or migration of oropharyngeal bacteria – changes in the structure and diversity of the pulmonary microbiota, its composition and function may have direct effects on pulmonary vascular remodeling leading to PAH, through microbial metabolites produced in the pulmonary microenvironment. Our preliminary results indicate the role of certain targeted metabolites as negative or positive modulators of pulmonary vascular cell proliferation. The expected results of LUMI are: a) to contribute to new knowledge on the role of the microbiota in respiratory diseases, b) to open up and feed a new field of knowledge on pulmonary vascular development, vascular remodeling and pathophysology of PAH c) to lead to a breakthrough in our vision of the pathophysiology and management of PAH patients. LUMI will provide a first knowledge on the pathobiome diversity and the pulmonary microbiota signature of PAH, as a basis for identifying new biotherapeutic approaches, as well as PAH biomarkers based on identified circulating metabolites. The final products that could emerge from LUMI for further development could be based on bacteria or their metabolites. As new therapeutic agents, they could be used in add-on therapy to existing treatments, to restore lung bacterial homeostasis and reverse lung vascular remodeling.

Dormancy is an adaptive trait that is established during seed maturation and prevents seed germination on the parent plant or out of proper season after seed dispersal. It is also an important agronomic trait as germination before harvest (vivipary) is a major cause of crop yield losses. Abscisic acid (ABA) is the key phytohormone promoting dormancy whereas nitrate (NO3-) stimulates germination by triggering ABA catabolism. Partners 1 & 2 have previously identified a new whole MAPK (Mitogen-activated protein kinase) module which is activated by both ABA and NO3- in Arabidopsis plantlets. They have also recently shown that mutants impaired in this module produce seeds which are more dormant. Strikingly and coherently, mutations in homologous MAPK genes in wheat and barley were reported to reduce vivipary. Taken together, these preliminary results suggest that this MAPK module is a new player controlling seed dormancy conserved throughout angiosperms. The fact that this module is activated by both ABA and NO3- also suggests that it may have a pivotal role as integrator of signaling pathways controlling dormancy. This project aims to better characterize this module in the frame of seed germination using Arabidopsis as a model plant and to exploit the results to develop new strategies to manipulate crop germination in the field. To achieve these goals, the first WP will aim to functionally validate the module by unveiling where and when it is required to modulate seed dormancy and which are the kinases involved in this function, a MAPK module being composed of at least 3 kinases. Importantly, we will test how the MAPK signaling module is modulated by and/or modulates ABA and NO3- signaling by using a combination of biochemical and genetic approaches to study mutants impaired in these signaling cascades. Furthermore, the MAPK module presents unique features when compared to other plant and animal MAPK modules described so far. The second WP will thus be devoted to the characterization of these specificities and will particularly study the translational and post-translational regulations of the module as well as decipher the unknown function of protein domains in the central MAP2K. The third WP will focus on the downstream events that are regulated by the module. Firstly, we will identify substrates which are phosphorylated by MAPKs and are important to control seed dormancy. Secondly, we will unveil the cellular processes which are regulated by the module by performing transcriptomic and metabolomic studies of mutants impaired in the module. Finally, a fourth WP will aim to identify molecules targeting this MAPK module and use them as chemical probes to investigate to which extent, across the plant kingdom, this module is important for seed physiology and to modulate seed dormancy in crops. This project relies on the collaboration of 4 groups recognized as leaders in their respective fields, who will bring their expertise and skills to challenge the novel hypothesis that the recently discovered MAPK module integrates distinct environmental signaling pathways to trigger the downstream processes that determine whether seeds germinate or not. Their joint work will lead to a better understanding of the factors that control seed physiological traits and new essential knowledge to enhance resilience through advanced breeding programs and to provide guidelines for optimal seed production, treatment and storage. It will also use an original strategy based on chemical genetics in yeast aiming at the identification of small molecules that modify the activity of the MAPK module and modulate dormancy in model species and crops. Thus, the MAPKSEED project brings together multidisciplinary expertises to tackle an important issue for optimizing sustainable agriculture in a changing environment by novel and original basic research.