Background Accumulation of extracellular matrix, recruitment of inflammatory macrophages and neutrophils secreting reactive oxygen species (ROS), and cytokines are characteristic of idiopathic pulmonary fibrosis (IPF) and renal fibrosis (RF). Signaling cascades triggered by TGF beta and involving cytokines and profibrotic factors are important therapeutic targets. The pro-inflammatory/pro-angiogenic chemokines ELR+CXCL and their receptors CXCR1/2 play a key role in promoting pulmonary fibrosis and nephropathy. In addition, VEGFC, a key driver of lymphangiogenesis produced by proinflammatory M2 macrophages, is critical for fibrosis via a cross-talk with TGF beta and ELR+CXCL stimulate lymphangiogenesis via VEGFC. To develop small therapies targeting these processes, the coordinator founded start-ups Roca therapeutics (RT) and Kekkan Biologics (KB). RT developed CXCR1/2 inhibitors and its drug candidate RCT001, and KB licensed the coordinator team's anti-VEGFC monoclonal antibodies. RCT001 inhibits the production of ROS, inflammation and fibrosis, is non-toxic to normal cells, stable, has good bioavailability and half-life. Anti-VEGFC Mab inhibits the proliferation of fibroblasts. Hypothesis Our hypothesis stipulates that ELR+CXCL/CXCR1/2 play a crucial role in fibrosis and that targeting CXCR1/2 can stop inflammation, mesenchymal cell activation, angiogenesis, and fibrosis. Interfering with the VEGFC pathway, which is also linked to CXCR1/2 signaling, may be effective in its own right or may further enhance the efficacy of RCT001. We believe that targeting two interconnected signaling pathways involved in chronic inflammation and abnormal lymphangiogenesis may represent innovative therapies. Methodology This translational project is based on the study of patient samples and the correlation between the presence of players in the ELR+CXCL/CXCR1/2 and VEGFC signaling pathways and the severity of IPF and/or RF. The effects of inhibiting both pathways individually or in combination on M2 macrophage polarisation and their ability to inhibit ROS formation will be tested in models of epithelial and endothelial cells and fibroblasts. The relevance of the two targetings will be evaluated functionally or biologically in models of IPF and RF in vivo. Expected results Pulmonary and renal fibrosis are currently at therapeutic impasses. Innovative therapies are urgently needed. Our patented molecules, backed by two start-ups, mean hope for patients. The two start-ups will licence the application patents for these inhibitors to carry out all the necessary steps to enter the clinical phase.

Constitutive heterochromatin (cHC), a key compartment of chromosomes essential for genome stability, is particularly difficult to replicate due to its dense organization and its enrichment in repetitive DNA. Recently a ground-breaking concept emerged from E. Gilson’s team (Partner 1) studies showing an unexpected role of a telomeric protein, TRF2, in the replication of the pericentromeric HC (PHC) regions. TRF2 is a member of the shelterin telomeric complex and protects chromosome ends from unwanted DNA damage response activation and illicit repair. It has also emerged as a multifunctional telomeric replication factor thanks to its capacity to recognize non-B DNA structures that can be formed on these repetitive sequences during replication (G4, chicken feet, DNA positive supercoils) and through the recruitment of several DNA targeting activities (helicases such as RTEL1 or WRN, nucleases such as APOLLO or MUS81-EME1). Our discovery of a key role of TRF2 in pericentromeric replication stems from the observation that TRF2 binds to PHC Satellite III DNA sequences during S phase, protects them from replicative DNA damage and is necessary for an efficient replication of PHC in terms of fork speed and origin firing (Mendez-Bermudez et al., Mol Cell 2018 and unpublished). TRF2 also protects other regions of the genome enriched in the H3K9me3 histone mark suggesting a much broader role of this protein in maintaining HC integrity and revealing the existence of a close functional relationship between telomeres and HC. Interestingly, both genomic regions are closely linked to cellular senescence and aging: one (telomeres) gradually shortening with age which ultimately leads to replicative senescence; the other (cHC) being subjected to profound structural changes at senescence. To decipher the mechanisms, the actors and the physiological consequences in cell senescence and aging of this telomere-cHC replication relationship, we designed a project named TELOCHROM that involves three teams, experts in telomere biology (Pr. E. Gilson, coordinator Dr. MJ Giraud-Panis) and in DNA replication (Pr. M. Debatisse and Dr. C. Chen). The aim of TELOCHROM is to Identify new pathways and telomere-associated actors of heterochromatin replication initiation and progression and their relationship with aging. TELOCHROM is organized in 3 work-packages aimed at identifying the mechanisms and actors involved in TRF2-dependent regulation of cHC replication (WP1), deciphering the effect of these different actors in replication origins firing and in replisome composition (WP2) and understanding the consequences of the identified pathways and actors in cell senescence (WP3). All teams are involved in each WP according to their technical expertise: Partner 1 will bring its essential know how in telomeres, pericentromeres biology and DNA topology; Partner 2 is an expert in DNA replication on hard to replicate regions such as fragile sites; Partner 3 has great expertise in top of the art Next Generation Sequencing and the analyses of the data generated. The originality and interest of the TELOCHROM project lies in: i) the innovative topic of a close functional link between telomeres and HC during replication; ii) the challenge of studying replication in highly repetitive, condensed and dynamic regions of the genome; iii) the top of the art technologies used (optical replication mapping, long read sequencing, Repli-seq, Topotools, Nascent Chromatin Capture); iv) the solid base of the project that stems from studies published recently or just submitted by the teams; v) the complementarity of the partners that have been successfully collaborating in recent years. Overall, TELOCHROM will undoubtedly bring new and important data on the basic mechanisms of HC replication, replicative senescence and on how telomeres and HC drive cell fate.

Aging is one of the major public health challenge. By impacting hematopoietic stem cells (HSCs) function, aging, exposure to irradiation, chemotherapy or chronic inflammation are highly associated with increased risk of developing cancers, and many other aging-related health disorders such as heart or neurocognitive diseases, through decline in the adaptive immunity and enhanced infections and inflammation. Epigenetic factors are key regulators of HSC function and epigenetic alterations have been observed in aged HSCs. Alteration of heterochromatin in HSCs during age is accompanied by overexpression of transposable elements (TEs) and increased inflammation. TEs are major contributors of gene regulatory networks. They can also be sensed as endogenous genomic parasites, eventually inducing an intrinsic sterile inflammation. Inflammation can in turn induce alterations in heterochromatin and TE expression and worsen the HSC phenotype in a vicious circle. We hypothesize here that heterochromatin alterations and TEs play a central role in the functional alterations of HSCs and in the production / function of myeloid cells during the aging process. Through the association of three partners with complementary expertise in epigenetics, transcription, biology, TEs, myelomonocyte differentiation and inflammatory response, we will explore whether: 1/ Alterations in heterochromatin and TE derepression are common events in age and stress 2/ TE derepression, via transcriptome alterations, act as the driving force behind HSC functional changes and their ability to produce monocytes with age 3/ TEs are involved in the inflammatory properties of aged HSCs and monocytes 4/ These pathways can be manipulated by reprogramming to rejuvenate HSC function This project should identify ways to restore lymphoid/myeloid HSC production and decrease chronic inflammation and immunosenescence by manipulating HSC heterochromatin and TE expression, thereby contributing to healthier aging.

Telomeres are the repeated sequences found at chromosome extremities. They are essential for genome integrity and control of cell proliferation. Telomere length (TL) at one chromosome end is a dynamic parameter subject to both shortening and elongation during cell divisions, thus resulting in a population-level distribution. Additionally, the TL distribution can be different at each chromosome end and the aggregated TL distribution over all ends in an isogenic population of cells is therefore a complex genomic property. How are the TL distributions at individual chromosome ends and aggregated over all ends regulated? What are the genetic determinants in trans and cis that control the natural diversity of TL and how do they interact with the environment? What are the fitness components associated with the natural TL diversity? To address these questions, our proposal DONATELO will leverage the natural diversity of Saccharomyces cerevisiae strains and the single molecule long-read sequencing technology that enable the measurement of TL distributions at each chromosome end. We recently established a large panel of high quality genome assemblies for 142 strains representing the phylogenetic and ecological diversity of the S. cerevisiae species, called the S. cerevisiae Reference Assembly Panel (ScRAP) (O’Donnell et al., 2023). DONATELO will (i) characterize the TL distributions and their stability across the genetically diverse ScRAP strains, as well as in an extended panel of strains, and various environmental conditions, (ii) uncover new genetic determinants of TL by tapping into the variants found in the ScRAP and the extended panel, and (iii) measure the phenotypes resulting from TL variations. In terms of approaches, our proposal will make the most of yeast genetics, genome engineering, Nanopore sequencing, bioinformatics and mathematical modelling. Overall, we anticipate that DONATELO will advance our understanding of telomere regulation and will address the fundamental question of whether and how this regulation is adaptive.

Clear cell Renal Cell Carcinoma (RCC) represents 85% of kidney cancers and 3% of adult cancers. The majority of RCCs present an inactivation of the VHL gene inducing the constitutive stabilization of HIF-1/2 transcription factors and leading to the overexpression the pro-inflammatory cytokines like ERL+CXCL exerting their actions via the CXCR1/2 receptors and of pro-angiogenic factors such as VEGFA. VEGFA promotes pathological angiogenesis, making these cancers among the most vascularized. This phenotype explains the use of anti-angiogenic drugs (sunitinib) as first-line treatment for metastatic RCCs over the past decade. Sunitinib increased progression-free survival without increasing overall survival, unlike immunotherapy. The first line treatment at now immunotherapies (nivolumab (anti-PD-1) + ipilimumab (anti-CTLA4)). Immunotherapy has resulted in long-term "remissions” but only 30% of patients are responsive to immunotherapy1. From these observations, the following objective arises: 1- Targeting all the players in the tumor niche to increase the effectiveness of immunotherapy: use of a home-made CXCR1/2 inhibitor 2- Non-invasive prediction of immunotherapy efficacy: towards precision medicine This project will allow: - To propose a new therapeutic strategy targeting all the actors of the tumor niche (RCC, endothelial cells, M2 macrophages and fibroblasts) and promoting the effectiveness of immunotherapy - To define a plasma signature at diagnosis to predict the resistance to immunotherapy. The ultimate goal of this project is to combine immunotherapy therapy with innovative compounds to prolong survival or induce very long-term remissions in patients with metastatic RCC.