Heterogeneity and Plasticity in Metastatic Breast Cancer

Breast cancer is the most common cancer amongst women. As for most solid tumours, breast cancer-related deaths are the result of metastasis, a succession of complex steps leading to the dissemination of cancer cells and colonization of distant organs. The heterogeneity and plasticity of cancer cells facilitate the selection of clones that withstand the harsh conditions encountered during their journey as circulating tumour cells and invading a foreign microenvironment. While the main steps of the metastatic cascade have been described, the precise cellular and molecular mechanisms underlying each of its steps remain poorly understood. In my PhD studies, I aimed to identify metastasis drivers in breast cancer using in vivo models of metastasis and RNA-Seq. This overarching goal materialised into two different projects. The first part of my work leveraged bulk RNA-sequencing of matched primary tumours and metastases in different sites to identify molecular processes potentially involved in metastasis. Our initial findings identified glucocorticoid signalling as promoting lung metastasis. NR3C1, also known as the glucocorticoid receptor (GR), mediates the effect of stress hormones. We showed that upon stimulation, GR controls processes involved in steps of the metastatic cascade. Mechanistically this control translates into an up-regulation of the kinase ROR1, a WNT5A receptor, modulating the Wnt and Hippo pathways. We showed how GR and ROR1 ablation decrease metastasis. Our studies highlight the importance of glucocorticoid signalling in triple-negative breast cancer and call for caution when using activators of GR such as Dexamethasone to combat the side effects of chemotherapy. In the second part of my work, we compared matched primary tumours and lung metastases of different breast cancer PDX models using single-cell RNA-seq. This technique allowed us to assess potential mechanisms driving metastases at the level of single cells, probing biological processes occurring in specific subpopulations of cancer cells. Our data highlight the significant inter and intra-patient heterogeneity of breast cancer. We found that cell populations in primary tumours and lung macrometastases are transcriptionally similar, but that three main phenotypes are making up lesions in both sites. Each of these phenotypes appears to be engaged in the epithelial-to-mesenchymal transition (EMT), yet at varying levels, as indicated by canonical EMT and proliferation markers. We show that the top differentially expressed genes differentiating each of these phenotypes are controlled via partial EMT transcription factors (TFs). This finding argues for the importance of partial EMT in the metastatic process and sheds further light on the processes underlying the metastatic cascade

Glucocorticoids promote breast cancer metastasis

Diversity within or between tumours and metastases (known as intra-patient tumour heterogeneity) that develops during disease progression is a serious hurdle for therapy(1-3). Metastasis is the fatal hallmark of cancer and the mechanisms of colonization, the most complex step in the metastatic cascade(4), remain poorly defined. A clearer understanding of the cellular and molecular processes that underlie both intra-patient tumour heterogeneity and metastasis is crucial for the success of personalized cancer therapy. Here, using transcriptional profiling of tumours and matched metastases in patient-derived xenograft models in mice, we show cancer-site-specific phenotypes and increased glucocorticoid receptor activity in distant metastases. The glucocorticoid receptor mediates the effects of stress hormones, and of synthetic derivatives of these hormones that are used widely in the clinic as anti-inflammatory and immunosuppressive agents. We show that the increase in stress hormones during breast cancer progression results in the activation of the glucocorticoid receptor at distant metastatic sites, increased colonization and reduced survival. Our transcriptomics, proteomics and phospho-proteomics studies implicate the glucocorticoid receptor in the activation of multiple processes in metastasis and in the increased expression of kinase ROR1, both of which correlate with reduced survival. The ablation of ROR1 reduced metastatic outgrowth and prolonged survival in preclinical models. Our results indicate that the activation of the glucocorticoid receptor increases heterogeneity and metastasis, which suggests that caution is needed when using glucocorticoids to treat patients with breast cancer who have developed cancer-related complications.Peer reviewe

Bone marrow haematopoiesis in patients with COVID-19

AIMS Severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2) infection broadly affects organ homeostasis, including the haematopoietic system. Autopsy studies are a crucial tool for investigation of organ-specific pathologies. Here we perform an in-depth analysis of the impact of severe coronavirus disease 2019 (COVID-19) on bone marrow haematopoiesis in correlation with clinical and laboratory parameters. METHODS AND RESULTS Twenty-eight autopsy cases and five controls from two academic centres were included in the study. We performed a comprehensive analysis of bone marrow pathology and microenvironment features with clinical and laboratory parameters and assessed SARS-CoV-2 infection of the bone marrow by quantitative polymerase chain reaction (qPCR) analysis. In COVID-19 patients, bone marrow specimens showed a left-shifted myelopoiesis (19 of 28, 64%), increased myeloid-erythroid ratio (eight of 28, 28%), increased megakaryopoiesis (six of 28, 21%) and lymphocytosis (four of 28, 14%). Strikingly, a high proportion of COVID-19 specimens showed erythrophagocytosis (15 of 28, 54%) and the presence of siderophages (11 of 15, 73%) compared to control cases (none of five, 0%). Clinically, erythrophagocytosis correlated with lower haemoglobin levels and was more frequently observed in patients from the second wave. Analysis of the immune environment showed a strong increase in CD68+ macrophages (16 of 28, 57%) and a borderline lymphocytosis (five of 28, 18%). The stromal microenvironment showed oedema (two of 28, 7%) and severe capillary congestion (one of 28, 4%) in isolated cases. No stromal fibrosis or microvascular thrombosis was found. While all cases had confirmed positive testing of SARS-CoV-2 in the respiratory system, SARS-CoV-2 was not detected in the bone marrow by high-sensitivity PCR, suggesting that SARS-CoV-2 does not commonly replicate in the haematopoietic microenvironment. CONCLUSIONS SARS-CoV-2 infection indirectly impacts the haematological compartment and the bone marrow immune environment. Erythrophagocytosis is frequent and associated with lower haemoglobin levels in patients with severe COVID-19

PI3K inhibition circumvents resistance to SHP2 blockade in metastatic triple-negative breast cancer.

The protein tyrosine phosphatase SHP2 activates oncogenic pathways downstream of most receptor tyrosine kinases (RTK) and has been implicated in various cancer types, including the highly aggressive subtype of triple-negative breast cancer (TNBC). Although allosteric inhibitors of SHP2 have been developed and are currently being evaluated in clinical trials, neither the mechanisms of the resistance to these agents, nor the means to circumvent such resistance have been clearly defined. The PI3K signaling pathway is also hyperactivated in breast cancer and contributes to resistance to anticancer therapies. When PI3K is inhibited, resistance also develops for example via activation of RTKs. We therefore assessed the effect of targeting PI3K and SHP2 alone or in combination in preclinical models of metastatic TNBC. In addition to the beneficial inhibitory effects of SHP2 alone, dual PI3K/SHP2 treatment decreased primary tumor growth synergistically, blocked the formation of lung metastases, and increased survival in preclinical models. Mechanistically, transcriptome and phospho-proteome analyses revealed that resistance to SHP2 inhibition is mediated by PDGFRβ-evoked activation of PI3K signaling. Altogether, our data provide a rationale for co-targeting of SHP2 and PI3K in metastatic TNBC

Journée de formation Polar

En lien avec le rectorat de Lyon, le réseau des bibliothèques municipales de Lyon et la DRAC Auvergne-Rhône-Alpes, cette journée a pour but de sensibiliser les enseignants et bibliothécaires au polar et de leur donner des pistes pour amorcer un travail sur le genre avec les adultes et les enfants. Le programme de cette journée s’articule autour d’interventions d’éditeurs, auteurs, libraires et autres professionnels du livre qui viendront notamment présenter des nouveautés polar 2018-2019, apporter des éclairages sur les raisons du succès du polar nordique et aborderont les spécificités du roman policier jeunesse

Masterclass de l’éditrice Viviane Hamy à l’occasion des 25 ans de la collection Chemins nocturnes

Interventions de la cadre de la journée de formation au polar

Unbiased screen for pathogens in human paraffin-embedded tissue samples by whole genome sequencing and metagenomics.

Identification of bacterial pathogens in formalin fixed, paraffin embedded (FFPE) tissue samples is limited to targeted and resource-intensive methods such as sequential PCR analyses. To enable unbiased screening for pathogens in FFPE tissue samples, we established a whole genome sequencing (WGS) method that combines shotgun sequencing and metagenomics for taxonomic identification of bacterial pathogens after subtraction of human genomic reads. To validate the assay, we analyzed more than 100 samples of known composition as well as FFPE lung autopsy tissues with and without histological signs of infections. Metagenomics analysis confirmed the pathogenic species that were previously identified by species-specific PCR in 62% of samples, showing that metagenomics is less sensitive than species-specific PCR. On the other hand, metagenomics analysis identified pathogens in samples, which had been tested negative for multiple common microorganisms and showed histological signs of infection. This highlights the ability of this assay to screen for unknown pathogens and detect multi-microbial infections which is not possible by histomorphology and species-specific PCR alone

Mechanism of MRX inhibition by Rif2 at telomeres

International audienceSpecific proteins present at telomeres ensure chromosome end stability, in large part through unknown mechanisms. In this work, we address how the Saccharomyces cerevisiae ORC-related Rif2 protein protects telomere. We show that the small N-terminal Rif2 BAT motif (Blocks Addition of Telomeres) previously known to limit telomere elongation and Tel1 activity is also sufficient to block NHEJ and 5' end resection. The BAT motif inhibits the ability of the Mre11-Rad50-Xrs2 complex (MRX) to capture DNA ends. It acts through a direct contact with Rad50 ATP-binding Head domains. Through genetic approaches guided by structural predictions, we identify residues at the surface of Rad50 that are essential for the interaction with Rif2 and its inhibition. Finally, a docking model predicts how BAT binding could specifically destabilise the DNA-bound state of the MRX complex. From these results, we propose that when an MRX complex approaches a telomere, the Rif2 BAT motif binds MRX Head in its ATP-bound resting state. This antagonises MRX transition to its DNA-bound state, and favours a rapid return to the ATP-bound state. Unable to stably capture the telomere end, the MRX complex cannot proceed with the subsequent steps of NHEJ, Tel1-activation and 5' resection

Table_1_Unbiased screen for pathogens in human paraffin-embedded tissue samples by whole genome sequencing and metagenomics.xlsx

Identification of bacterial pathogens in formalin fixed, paraffin embedded (FFPE) tissue samples is limited to targeted and resource-intensive methods such as sequential PCR analyses. To enable unbiased screening for pathogens in FFPE tissue samples, we established a whole genome sequencing (WGS) method that combines shotgun sequencing and metagenomics for taxonomic identification of bacterial pathogens after subtraction of human genomic reads. To validate the assay, we analyzed more than 100 samples of known composition as well as FFPE lung autopsy tissues with and without histological signs of infections. Metagenomics analysis confirmed the pathogenic species that were previously identified by species-specific PCR in 62% of samples, showing that metagenomics is less sensitive than species-specific PCR. On the other hand, metagenomics analysis identified pathogens in samples, which had been tested negative for multiple common microorganisms and showed histological signs of infection. This highlights the ability of this assay to screen for unknown pathogens and detect multi-microbial infections which is not possible by histomorphology and species-specific PCR alone.</p