Biology subjects

Baulande, S.

Publications and source records attributed to Baulande, S..

18 recordsLinked to original sources

bioRxiv2026

Oncogenic fusions induce an extensive cancer-restricted cryptic proteome in Ewing sarcoma

How tumors generate a cryptic "dark" proteome absent from healthy tissues, and whether it can be reversibly activated, remains unclear. In Ewing sarcoma, all tumors are driven by EWSR1::ETS fusions, making it a tractable model to study dark proteome activation. Integrating matched transcriptomes, translatomes and proteomes from 48 patient tumors with long-read RNA sequencing, single-cell Ribo-seq, proteomics and immunopeptidomics in cell line models, we show that EWSR1::FLI1 acts as a reversible switch recurrently inducing hundreds of cancer-specific neoproteins. Many arise from canonical coding regions via intragenic transcription start sites, generating truncated or out-of-frame neoproteins. We uncover a fusion-dependent increase in ribosomal readthrough into poly(A) tails, generating a stable 80-amino-acid TRPM4 neoprotein that, despite lacking a stop codon, is the most abundant tumor-specific microprotein across patients. Proteomic, immunopeptidomic, and immunofluorescence analyses validate neoproteins as tumor-restricted antigens, revealing a therapeutically actionable cancer dark proteome controlled by one oncogenic fusion.

cancer biology↗

bioRxiv2025

Myeloid reprogramming by poly(I:C) recruits progenitor-exhausted CD8+ T cells and sensitizes rhabdoid tumors to PD-1 blockade

Immune exclusion remains a major barrier to effective immunotherapy in solid tumors. Given the abundance and plasticity of tumor-associated macrophages (TAMs) in many tumors, including pediatric tumors, we investigated whether TLR3 activation could reprogram them to facilitate immune access. Single-cell and spatial profiling in a mouse model of rhabdoid tumors showed that they are dominated by TLR3-expressing TAMs, whose depletion delays tumor growth. Treatment with the TLR3 agonist poly(I:C) promotes immune cell infiltration, including progenitor-exhausted CD8+ T cells, by multiple mechanisms including the reduction and reprogramming of immunosuppressive TAMs, promoting nitric-oxide production by peritumoral macrophages, and inducing CXCL9/10 production. Combined poly(I:C) and PD-1 blockade elicited durable, complete tumor rejection. Human macrophages from tumor biopsies showed conserved TLR3 responsiveness, underscoring translational potential. These findings uncover a mechanism by which TLR3-driven myeloid reprogramming transforms immune-excluded tumors into checkpoint-responsive ones, revealing a therapeutic path to overcome resistance to PD-1 blockade.

immunology↗

bioRxiv2025

Dual PD-L1/TIGIT blockade induces PNAd+ HEV-like vessels and CD62L+ lymphocyte recruitment, driving rhabdoid tumor rejection

Rhabdoid tumors (RTs) are aggressive pediatric malignancies with poor prognosis and limited immunotherapy options. Here, we investigate the therapeutic potential of combined PD-L1 (Programmed cell death ligand 1) and TIGIT (T cell immunoreceptor with Ig and ITIM domains) immune checkpoint blockade in RTs using a preclinical murine model that recapitulates key features of human ATRT (Atypical teratoid rhabdoid tumors) subtypes. Transcriptomic analyses of human and murine RTs reveal co-expression of TIGIT and PD-1 (Programmed cell death 1) pathway components and their ligands, particularly in immune-infiltrated subtypes, supporting a rationale for dual blockade. Combination therapy induces complete tumor regression, prolongs survival, and reprograms the tumor immune microenvironment by enriching CD62L naive and central memory T cells and promoting selective T-cell clonal expansion. Notably, dual blockade initiates PNAd (Peripheral node addressin) high endothelial venule (HEV)-like structures, associated with focal lymphocyte clustering and enhanced immune cell recruitment. These findings reveal a mechanistic link between vascular remodeling and immune infiltration and support dual TIGIT and PD-L1 inhibition as a promising immunotherapeutic strategy for RTs.

immunology↗

bioRxiv2025

Common origin for effector and regulatory Follicular and Tissue-Adapted CD4+ T cells in Non-Small Cell Lung Cancer

Tumor-invaded lymph nodes (LNs) serve as critical hubs for anti-tumor immunity, yet their role in orchestrating immune responses remains poorly understood. Using integrated single-cell RNA sequencing, T cell receptor sequencing, and chromatin accessibility profiling, we analyzed CD4+ T cells from matched blood, tumor-invaded LNs, and tumors of treatment-naive non-small cell lung cancer patients. We identified distinct immunological landscapes across these compartments. Compared to blood, tumor-invaded LNs and tumors were enriched for follicular regulatory T cells (Treg-Tfr), conventional T cell subsets with Tfh-like characteristics (Tconv-Tfh and Tconv-CXCL13), and tissue-resident memory Tregs (Treg-Trm). These populations share a BATF-dependent transcriptional program that governs T-cell activation and tissue adaptation, while simultaneously engaging distinct, subset-specific regulatory networks. Integrative TCR-RNA analysis revealed that tumor-reactive, neoantigen-specific T cell clones were enriched within these subsets and demonstrated extensive LN-tumor clonal sharing, indicating active recirculation between compartments. Through clonal coupling analysis and trajectory inference, we uncovered that Treg-Tfr cells function as multipotent progenitors that bifurcate into tissue-resident Treg-Trm or into ex-Tregs adopting a Tfh-like CXCL13+ ewector phenotype. Remarkably, follicular CD4+ T subsets from LNs and tumor were transcriptionally and epigenetically similar and localized to analogous germinal center niches. These findings establish tumor-invaded LNs as functional extensions of the tumor microenvironment that generate and maintain tumor-reactive CD4+ lineages. The identification of tissue-resident Treg-Tfr plasticity reveals a critical developmental checkpoint that could be therapeutically targeted to redirect immunosuppressive programs toward anti-tumor ewector responses.

immunology↗

bioRxiv2025

A recurrent pathogenic BRCA2 truncating variant reveals a role for BRCA2-PCAF complex in modulating NF-κB-driven transcription

Germline monoallelic truncating mutations in BRCA2, a critical mediator of homologous recombination (HR), predispose individuals to breast and ovarian cancer. While tumorigenesis is usually attributed to biallelic inactivation, emerging evidence suggests that haploinsufficiency may suffice in certain contexts. To investigate this, we recreated two BRCA2 pathogenic truncating variants in heterozygosis in non-tumorigenic breast epithelial cells. Cells carrying a truncating mutation that was not produced prompted sensitivity to PARP inhibitors (PARPi) and reduced the HR capacity indicating haploinsufficiency. Surprisingly, the other variant was expressed as a truncated product and prompted a transcriptional rewiring. Mechanistically, the truncated BRCA2 product formed abnormal oligomers with full-length BRCA2 and bound to the PCAF acetyltransferase, sequestering it. This led to reduced global histone H4 acetylation and decreased NF-{kappa}B transcriptional activity, ultimately impairing epithelial cell migration--a process also altered in tumors. Our findings uncover a previously unrecognized BRCA2-PCAF axis that modulates NF-{kappa}B-driven transcriptional program, a process that is co-opted by a recurrent BRCA2 pathogenic variant.

cancer biology↗

bioRxiv2025

Pathogenic Viruses, Genome Integrations, and Viral::Human Chimeric Transcripts Detected by VirusIntegrationFinder Across >30k Human Tumor and Normal Samples

Viruses are a leading cause of human morbidity and mortality. Certain viruses, including human papillomaviruses (HPVs), play a significant role in the etiology of cancer. Detection of viral DNA insertions in the human genome from next generation sequencing data defines viral associations with cancer and other diseases, identifies impacted organs and tissues, provides insights into disease mechanisms and has the potential to enhance clinical evaluations. In this study, we developed VirusIntegrationFinder (CTAT-VIF), a tool for surveying human genome insertions of various human viruses using both DNA and RNA sequencing data. We applied CTAT-VIF to analyze a dataset of over 30,000 tumor and normal samples, as well as more than 1,000 cancer cell lines. This effort resulted in the compilation of a catalog of over 30,0000 virus-human DNA or RNA junctions at more than 20,000 insertion loci and reassessed viral cancer-insertion hotspots across the human genome. Furthermore, we characterized the functional impacts of insertions with respect to human copy number alterations, effects on the expression of flanking human genes, and the identification of potentially oncogenic chimeric human and human/virus fusion transcripts at insertion loci. In addition to confirming known viral associations with specific tumor types, our study revealed both shared and virus-specific insertion hotspots in addition to variable functional impacts based on virus type. Besides some rare events of interest, we also found evidence for sequencing contamination, which underscores the need for vigilance when studying viral content or genome integrations.

cancer biology↗

bioRxiv2024

CCG interruptions are unstable and hypermethylated in DM1 patients

Myotonic dystrophy type 1 (DM1) exhibits highly heterogeneous clinical manifestations caused by an unstable CTG repeat expansion reaching up to 4,000 CTG. The dynamics of CTG repeats and clinical variability depends on the CTG repeat number, CNG repeat interruptions, DNA methylation, somatic mosaicism, but also gene modifiers. Around 10% of the DM1 population carries triplet repeat interruptions (CCG, CGG, CTC, CAG), which differ in number and nature between DM1 families. CCG interruptions have been associated with stabilization of the CTG repeat expansions and a milder phenotype in the DM1 population. However, the dynamics and precise role of interruptions in CTG repeat instability remain relatively underexplored due to the complexities of analyzing them. In this study, we showed that the number of CCG interruptions within the expansion varies within tissues and between generations using single-molecule real-time long-read sequencing. Although the interrupted expanded alleles showed global stabilization, the CCG interruptions themselves displayed variability across generations and within somatic tissues. Importantly, our findings reveal, for the first time, CCG hypermethylation within the CTG expansion, which is linked to downstream hypermethylation of the repeat. These results support the hypothesis that methylation of CCG interruptions within the expanded DMPK alleles may contribute to the stabilization of trinucleotide repeats.

molecular biology↗

bioRxiv2024

An extra-glycolytic function for hexokinase 2 as an RNA-binding protein regulating SOX10 mRNA translation in melanoma

Several studies have reported the importance of aerobic glycolysis in melanoma development. Although metabolic benefits of glycolysis have been extensively described in tumor cells, the extra-metabolic functions linked to this energetic pathway in melanoma growth and proliferation have not been clearly established yet. Recently, some key glycolytic enzymes, such as GAPDH and PKM2, were reported to regulate mRNA translation. Translational control of gene expression is considered as a critical effector in cancer biology, representing a highly promising area of research. Here, we report that Hexokinase 2 (HK2), a glucose kinase that catalyzes the first step of glycolysis, is an RNA binding protein (RBP) that regulates mRNA translation in melanoma. We show that siRNA-mediated HK2 depletion changes the translational landscape of melanoma cells. Polysome profiling experiments and RNA-Seq indicate that the translational regulation exerted by HK2 is partly independent of the metabolic status or the glycolytic pathway. We found that HK2 specifically regulates the translation of the mRNA encoding SOX10, a transcription factor implicated in the regulation of tumor initiation, maintenance and progression in melanoma. RNA-protein interaction assays, including crosslinking immunoprecipitation (CLIP), indicate that HK2 is an RBP whose interaction with RNA is independent of its hexokinase activity or subcellular localization. We also show that HK2 specifically associates with the 5 untranslated region (5UTR) of the SOX10 mRNA, and that several deletions in this region decreases both HK2-SOX10 mRNA association and SOX10 5 UTR-mediated translation. We further show that HK2-dependent SOX10 translational regulation is involved in melanoma cell proliferation and colony formation. Collectively, our data highlight a non-metabolic function of HK2, indicating that melanoma cells may enhance glycolysis for purposes beyond simple anabolism.

cancer biology↗

bioRxiv2024

TNFR2 blockade promotes anti-tumoral immune response in PDAC by targeting activated Treg and reducing T cell exhaustion

BackgroundPancreatic ductal adenocarcinoma (PDAC) is one of the most aggressive cancers, highly resistant to standard chemotherapy and immunotherapy. Regulatory T cells (Tregs) expressing TNF receptor 2 (TNFR2) contribute to immunosuppression in PDAC. Treg infiltration correlates with poor survival and tumor progression in PDAC patients. We hypothesized that TNFR2 inhibition using a blocking monoclonal antibody (mAb) could shift the Treg-effector T cell balance in PDAC, thus enhancing anti-tumoral responses. MethodTo support this hypothesis, we first described TNFR2 expression in a cohort of 24 PDAC patients from publicly available single-cell analysis data. In orthotopic and immunocompetent mouse models of PDAC, we also described the immune environment of PDAC after immune cell sorting and single-cell analysis. The modifications of the immune environment before and after anti-TNFR2 mAb treatment were evaluated as well as effect on tumor progression. ResultsPDAC patients exhibited elevated TNFR2 expression in Treg, myeloid cells and endothelial cells but low level in ductal cells. By flow cytometry and single cell RNAseq analysis, we identified two Treg populations in orthotopic mouse models: resting and activated Tregs. The anti-TNFR2 mAb selectively targeted activated tumor-infiltrating Tregs, reducing T cell exhaustion markers in CD8+ T cells. However, anti-TNFR2 treatment alone had limited efficacy in activating CD8+ T cells and only slightly reduced the tumor growth. The combination of the anti-TNFR2 mAb with agonistic anti-CD40 mAbs promoted stronger T cell activation, tumor growth inhibition, and improved survival and immunological memory in PDAC-bearing mice. ConclusionOur data suggest that combining a CD40 agonist with a TNFR2 antagonist represents a promising therapeutic strategy for PDAC patients.

immunology↗

bioRxiv2023

The pelvic organs receive no parasympathetic innervation

The pelvic organs (bladder, rectum and sex organs) have been represented for a century as receiving autonomic innervation from two pathways -- lumbar sympathetic and sacral parasympathetic -- by way of a shared relay, the pelvic ganglion, conceived as an assemblage of sympathetic and parasympathetic neurons. Using single cell RNA sequencing, we find that the mouse pelvic ganglion is made of four classes of neurons, distinct from both sympathetic and parasympathetic ones, albeit with a kinship to the former, but not the latter, through a complex genetic signature. We also show that spinal lumbar preganglionic neurons synapse in the pelvic ganglion onto equal numbers of noradrenergic and cholinergic cells, both of which therefore serve as sympathetic relays. Thus, the pelvic viscera receive no innervation from parasympathetic or typical sympathetic neurons, but instead from a divergent tail end of the sympathetic chains, in charge of its idiosyncratic functions.

neuroscience↗

bioRxiv2022

Multi-omics comparison of malignant and normal uveal melanocytes reveals novel molecular features of uveal melanoma.

Uveal Melanoma (UM) is a rare cancer resulting from transformation of melanocytes residing in the uveal tract. Integrative analysis has identified four molecular and clinical subsets in UM. To improve our molecular understanding of UM we performed extensive multi-omics characterization comparing two aggressive UM patient-derived xenograft models with normal choroidal melanocytes, including DNA optical mapping, specific histone modifications, and DNA topology analysis by Hi-C. Our gene expression and cytogenetic analyses suggest that genomic instability is a hallmark of UM. We also identified a recurrent deletion in the BAP1 promoter resulting in loss of expression and associated with high risk of metastases in UM patients. Hi-C revealed chromatin topology changes associated with up-regulation of PRAME, an independent prognostic biomarker in UM and potential therapeutic target. Our findings illustrate how multi-omics approaches can improve the understanding of tumorigenesis and reveals two novel mechanisms of gene expression dysregulation in UM.

genomics↗

bioRxiv2021

Kronos scRT: a uniform framework for single-cell replication timing analysis

Mammalian genomes are replicated in a cell-type specific order and in coordination with transcription and chromatin organization. Although the field of replication is also entering the single-cell era, current studies require cell sorting, individual cell processing and have yielded a limited number (<100) of cells. Here, we have developed Kronos scRT (https://github.com/CL-CHEN-Lab/Kronos_scRT), a software for single-cell Replication Timing (scRT) analysis. Kronos scRT does not require a specific platform nor cell sorting, allowing the investigation of large datasets obtained from asynchronous cells. Analysis of published available data and droplet-based scWGS data generated in the current study, allows exploitation of scRT data from thousands of cells for different mouse and human cell lines. Our results demonstrate that, although most cells replicate within a close timing range for a given genomic region, replication can also occur stochastically throughout S phase. Altogether, Kronos scRT allows investigating the RT program at a single-cell resolution for both homogeneous and heterogeneous cell populations in a fast and comprehensive manner.

genomics↗

bioRxiv2021

Systematic multi-omics cell line profiling uncovers principles of Ewing sarcoma fusion oncogene-mediated gene regulation

Cell lines have been essential for major discoveries in cancer including Ewing sarcoma (EwS). EwS is a highly aggressive pediatric bone or soft-tissue cancer characterized by oncogenic EWSR1-ETS fusion transcription factors converting polymorphic GGAA-microsatellites (mSats) into neo-enhancers. However, further detailed mechanistic evaluation of gene regulation in EwS have been hindered by the limited number of well-characterized cell line models. Here, we present the Ewing Sarcoma Cell Line Atlas (ESCLA) comprising 18 EwS cell lines with inducible EWSR1-ETS knockdown that were profiled by whole-genome-sequencing, DNA methylation arrays, gene expression and splicing arrays, mass spectrometry-based proteomics, and ChIP-seq for EWSR1-ETS and histone marks. Systematic analysis of these multi-dimensional data illuminated hundreds of new potential EWSR1-ETS target genes, the nature of EWSR1-ETS-preferred GGAA-mSats, and potential indirect modes of EWSR1-ETS-mediated gene regulation. Moreover, we identified putative co-regulatory transcription factors and heterogeneously regulated EWSR1-ETS target genes that may have implications for the clinical heterogeneity of EwS. Collectively, our freely available ESCLA constitutes an extremely rich resource for EwS research and highlights the power of leveraging multidimensional and comprehensive datasets to unravel principles of heterogeneous gene regulation by dominant fusion oncogenes.

genomics↗

bioRxiv2021

Tissue-resident FOLR2+ macrophages associate with tumor-infiltrating CD8+ T cells and with increased survival of breast cancer patients.

Macrophage infiltration is a hallmark of solid cancers and overall macrophage infiltration is correlated with lower patient survival and resistance to therapy. However, tumor-associated macrophages are phenotypically and functionally heterogeneous. Specific tumor-associated macrophage subsets might be endowed with antagonistic role on cancer progression and on the development of anti-tumor immunity. For instance, monocyte-derived TREM2+ tumor-associated macrophages have pro-tumorigenic and immunosuppressive functions. Here, we identify a discrete population of FOLR2+ tumor-associated macrophages positively correlating with patient survival in breast cancer. FOLR2+ macrophages are evolutionarily conserved across species and populate human and murine healthy mammary gland. Moreover, FOLR2+ macrophages co-localize with lymphoid aggregates containing CD8+ T cells in breast cancer and across ten other types of cancers. This study highlights antagonistic roles for tumor-associated macrophage subsets and paves the way for subset-specific therapeutic interventions in macrophages-based cancer therapies.

immunology↗

bioRxiv2021

Interplay between intrinsic reprogramming potential and microenvironment controls neuroblastoma cell plasticity and identity

Two cell identities, noradrenergic and mesenchymal, have been characterized in neuroblastoma cell lines according to their epigenetic landscapes relying on specific circuitries of transcription factors. Yet, their relationship and relative contribution in patient tumors remain poorly defined. Here, we demonstrate that the knock-out of GATA3, but not of PHOX2A or PHOX2B, in noradrenergic cells induces a mesenchymal phenotype. Our results document spontaneous plasticity in several models between both identities and show that plasticity relies on epigenetic reprogramming. We demonstrate that an in vivo microenvironment provides a powerful pressure towards a noradrenergic identity for these models. Consistently, tumor cells with a mesenchymal identity are not detected in a series of PDX models. Further study of the intra-tumor noradrenergic heterogeneity reveals two distinct cell populations exhibiting features of chromaffin-like or sympathoblast-like cells. This work emphasizes that both external cues of the environment and intrinsic factors control plasticity and cell identity in neuroblastoma.

cancer biology↗

bioRxiv2021

H3K27me3 is a determinant of chemotolerance in triple-negative breast cancer

Triple-negative breast cancer is associated with the worst prognosis and the highest risk of recurrence among all breast cancer subtypes1. Residual disease, formed by cancer cells persistent to chemotherapy, remains one of the major clinical challenges towards full cure2,3. There is now consensus that non-genetic processes contribute to chemoresistance in various tumor types, notably through the initial emergence of a reversible chemotolerant state4-6. Understanding non-genetic tumor evolution stands now as a prerequisite for the design of relevant combinatorial approaches to delay recurrence. Here we show that the repressive histone mark H3K27me3 is a determinant of cell fate under chemotherapy exposure, monitoring epigenomes, transcriptomes and lineage with single-cell resolution. We identify a reservoir of persister basal cells with EMT markers and activated TGF-{beta} pathway leading to multiple chemoresistance phenotypes. We demonstrate that, in unchallenged cells, H3K27 methylation is a lock to the expression program of persister cells. Promoters are primed with both H3K4me3 and H3K27me3, and removing H3K27me3 is sufficient for their transcriptional activation. Leveraging lineage barcoding, we show that depleting H3K27me3 alters tumor cell fate under chemotherapy insult - a wider variety of tumor cells tolerate chemotherapy. Our results highlight how chromatin landscapes shape the potential of unchallenged cancer cells to respond to therapeutic stress.

cancer biology↗

bioRxiv2020

Contribution of resident and circulating precursors to tumor-infiltrating CD8+ T cell populations in non-small cell lung cancer patients

Tumor-infiltrating lymphocytes (TILs) in general, and CD8+ TILs in particular, represent a favorable prognostic factor in non-small cell lung cancer (NSCLC). The tissue origin, regenerative capacities, and differentiation pathways of TIL subpopulations, however, remain poorly understood. Using a combination of single cell RNA and T cell receptor (TCR) sequencing, we investigate the functional organization of TIL populations in primary NSCLC. We identify two CD8+ TIL subpopulations expressing memory-like gene modules: one is also present in blood (circulating precursors) and the other one in juxta-tumor tissue (tissue resident precursors). In tumors, these two precursor populations converge through a unique transitional state into terminally differentiated cells, often referred to as dysfunctional or exhausted. Differentiation is associated with TCR expansion, and transition from precursor to late differentiated states correlates with intratumor T cell cycling. These results provide a coherent working model for TIL origin, filiation and functional organization in primary NSCLC.

immunology↗

bioRxiv2019

Transcriptional programs define intratumoral heterogeneity of Ewing sarcoma at single cell resolution

EWSR1-FLI1, the chimeric oncogene specific for Ewing sarcoma (EwS), induces a cascade of signaling events leading to cell transformation. However, it remains elusive how genetically homogeneous EwS cells can drive heterogeneity of transcriptional programs. Here, we combined independent component analysis of single cell RNA-sequencing data from diverse cell types and model systems with time-resolved mapping of EWSR1-FLI1 binding sites and of open chromatin regions to characterize dynamic cellular processes associated with EWSR1-FLI1 activity. We thus defined an exquisitely specific and direct, super-enhancer-driven EWSR1-FLI1 program. In EwS tumors, cell proliferation was associated with a well-defined range of EWSR1-FLI1 activity; moreover, cells with a high EWSR1-FLI1 activity presented a strong oxidative phosphorylation metabolism. In contrast, a subpopulation of cells from below and above optimal EWSR1-FLI1 activity was characterized by increased hypoxia. Overall, our study reveals sources of intratumoral heterogeneity within Ewing tumors.

cancer biology↗