ERC FUNDED PROJECTS

Acute Myeloid Leukemia (AML), the most common leukemia diagnosed in adults, represents the paradigm of resistance to front-line therapies in hematology. Indeed, AML is so genetically complex that only few targeted therapies are currently tested in this disease and chemotherapy remains the only standard treatment for AML since the past four decades. Despite an initial sustained remission achieved by chemotherapeutic agents, almost all patients relapse with a chemoresistant minimal residual disease (MRD). The goal of my proposal is to characterize the still poorly understood biological mechanisms underlying persistence and emergence of MRD. MRD is the consequence of the re-expansion of leukemia-initiating cells that are intrinsically more resistant to chemotherapy. This cell fraction may be stochastically more prone to survive front-line therapy regardless of their mutational status (the stochastic model), or genetically predetermined to resist by virtue of a collection of chemoprotective mutations (the mutational model). I have already generated in mice, by consecutive rounds of chemotherapy, a stochastic MLL-AF9-driven chemoresistance model that I examined by RNA-sequencing. I will pursue the comprehensive cell autonomous and cell non-autonomous characterization of this chemoresistant AML disease using whole-exome and ChIP-sequencing. To establish a mutationally-induced chemoresistant mouse model, I will conduct an innovative in vivo screen using pooled mutant open reading frame and shRNA libraries in order to predict which combinations of mutations, among those already known in AML, actively promote chemoresistance. Finally, by combining genomic profiling and in vivo shRNA screening experiments, I will decipher the molecular mechanisms and identify the functional effectors of these two modes of resistance. Ultimately, I will then be able to firmly establish the fundamental relevance of the stochastic and/or the mutational model of chemoresistance for MRD genesis.

1 500 000 €

Start date: 2018-03-01, End date: 2023-02-28

Blood and lymphatic vessels have been the subject of intense investigation due to their important role in cancer development and in cardiovascular diseases. The significant advance in the methods used to modify and analyse gene function have allowed us to obtain a much better understanding of the molecular mechanisms involved in the regulation of the biology of blood vessels. However, there are two key aspects that significantly diminish our capacity to understand the function of gene networks and their intersections in vivo. One is the long time that is usually required to generate a given double mutant vertebrate tissue, and the other is the lack of single-cell genetic and phenotypic resolution. We have recently performed an in vivo comparative transcriptome analysis of highly angiogenic endothelial cells experiencing different VEGF and Notch signalling levels. These are two of the most important molecular mechanisms required for the adequate differentiation, proliferation and sprouting of endothelial cells. Using the information generated from this analysis, the overall aim of the proposed project is to characterize the vascular function of some of the previously identified genes and determine how they functionally interact with these two signalling pathways. We propose to use novel inducible genetic tools that will allow us to generate a spatially and temporally regulated fluorescent cell mosaic matrix for quantitative analysis. This will enable us to analyse with unprecedented speed and resolution the function of several different genes simultaneously, during vascular development, homeostasis or associated diseases. Understanding the genetic epistatic interactions that control the differentiation and behaviour of endothelial cells, in different contexts, and with high cellular definition, has the potential to unveil new mechanisms with high biological and therapeutic relevance.

1 481 375 €

Start date: 2015-03-01, End date: 2020-02-29

Atherosclerosis is a chronic immune disease of arteries that causes vessel-narrowing atherosclerotic plaques. Its acute complications, myocardial infarction and stroke, are the leading causes of death worldwide. Atherosclerosis is accompanied by an inflammatory and autoimmune response with CD4+ T-helper cells that recognize self-antigens, including ApoB-100 (ApoB), the main protein in low-density lipoprotein (LDL) cholesterol. Although their existence has been inferred from indirect evidence, the existence and function of atherosclerosis-specific, self-reactive CD4+ T cells on a single-cell level remains elusive. In particular, it is unclear whether these are pro- or anti-inflammatory. Preliminary data suggest the existence of a natural pool of ApoB-reactive T-helper cells that share properties with atheroprotective T-regulatory cells but transform into pathogenic T-effector cells in the natural course of disease. This proposal aims to explore this loss of protective immunity on a cellular and function level. It employs novel tools to detect antigen-specific T cells in vivo by MHC-II multimers, mass cytometry (CyTOF), single cell RNA-sequencing (scRNA-seq), lineage-tracing mouse models, and live cell imaging. Based on the anticipated findings, this study will define a map of auto-reactive T-helper cell phenotypes in a temporal, spatial, and functional dimension. These insights will be used to identify novel immunomodulatory strategies to therapeutically stabilize the population of protective ApoB-specific T-helper cells, or to prevent their transformation into pathogenic T cell phenotypes by adoptive cells transfers, vaccination, or cytokine-blockade. In clinical association studies, a direct correlation of auto-immunity and clinical atherosclerosis will be tested. This proposal will decipher traits of protective immunity in atherosclerosis and help to build the conceptual framework to define novel therapeutic strategies for patients.

1 499 946 €

Start date: 2020-01-01, End date: 2024-12-31

The formation of a functional patterned vascular network is essential for development, tissue growth and organ physiology. Several human vascular disorders arise from the mis-patterning of blood vessels, such as arteriovenous malformations, aneurysms and diabetic retinopathy. Although blood flow is recognised as a stimulus for vascular patterning, very little is known about the molecular mechanisms that regulate endothelial cell behaviour in response to flow and promote vascular patterning. Recently, we uncovered that endothelial cells migrate extensively in the immature vascular network, and that endothelial cells polarise against the blood flow direction. Here, we put forward the hypothesis that vascular patterning is dependent on the polarisation and migration of endothelial cells against the flow direction, in a continuous flux of cells going from low-shear stress to high-shear stress regions. We will establish new reporter mouse lines to observe and manipulate endothelial polarity in vivo in order to investigate how polarisation and coordination of endothelial cells movements are orchestrated to generate vascular patterning. We will manipulate cell polarity using mouse models to understand the importance of cell polarisation in vascular patterning. Also, using a unique zebrafish line allowing analysis of endothelial cell polarity, we will perform a screen to identify novel regulators of vascular patterning. Finally, we will explore the hypothesis that defective flow-dependent endothelial polarisation underlies arteriovenous malformations using two genetic models. This integrative approach, based on high-resolution imaging and unique experimental models, will provide a unifying model defining the cellular and molecular principles involved in vascular patterning. Given the physiological relevance of vascular patterning in health and disease, this research plan will set the basis for the development of novel clinical therapies targeting vascular disorders.

1 618 750 €

Start date: 2016-09-01, End date: 2021-08-31