Microbulles ultrasonores pour l'imagerie moléculaire et la délivrance de médicaments : détection de la Nétrine-1 dans le cancer du sein & modulation de la réponse immunitaire dans le carcinome hépatocellulaire

Ultrasound molecular imaging uses microbubbles as ultrasound contrast agents which are functionalized with targeting ligands. Upon intravenous injection, targeted microbubbles bind to molecular markers presented on the tumor endothelium and enable the non-invasive assessment cancer-related biomarkers. In the present thesis, ultrasound molecular imaging was developed for detection of netrin-1, which is upregulated in 70% of metastatic breast cancer and promotes cell survival. A newly developed netrin-1 interference therapy requires the identification of patients who overexpress the target protein and, could benefit from anti-netrin-1 therapy. In vivo imaging of netrin-1 showed a significantly increased imaging signal in netrin-1-positive breast tumors compared to netrin-1-negative breast tumors and normal mammary glands. The results suggest that ultrasound molecular imaging allows accurate detection of netrin-1 on the endothelium of netrin-1-positive tumors and has the potential to become a companion diagnostic for netrin-1 interference therapy in breast cancer patients.Ultrasound-targeted microbubble destruction triggers cavitation and sonoporation thereby permeabilizing the tissue and facilitating local drug delivery. Further, immune cell infiltration and tumor antigen release are induced and trigger anti-tumor immune responses. In the present thesis, ultrasound-targeted microbubble destruction-mediated delivery of anti-cancer microRNA-122 and anti-microRNA-21 is studied for immune response activation in hepatocellular carcinoma, in which the immune microenvironment is deregulated. Tumor lymph nodes showed pro-tumor cytokine downregulation and anti-tumor cytokine upregulation, suggesting an overall positive therapy response with regard to the tumor immunology. The results identified ultrasound-targeted microbubble destruction-mediated miRNA delivery as a potent immuno-modulatory therapeutic approachDans l'imagerie moléculaire par ultrasons, des microbulles sont fonctionnalisées avec des ligands. Après injection intraveineuse, ces microbulles ciblées s'accrochent aux marqueurs présents sur l'endothélium tumoral et permettent une détection non-invasive. Dans cette thèse, l'imagerie moléculaire par ultrasons a été développée pour la détection de la nétrine-1, qui est surexprimée dans 70% des cancers du sein et promeut la survie cellulaire. Une nouvelle thérapie moléculaire interférant avec la nétrine-1 a été développée et nécessite l'identification des patientes qui pourront bénéficier de ce traitement. Avec l'imagerie moléculaire de la nétrine-1, il a été possible de discriminer les tumeurs positives pour la nétrine-1 des tumeurs négatives. Par sa capacité à détecter de manière spécifique la nétrine-1 présentée sur l'endothélium des tumeurs, cette technique d'imagerie pourrait donc devenir un test d'accompagnement pour la thérapie d'interférence de la nétrine-1 chez les patientes atteintes de cancer du sein.La destruction ciblée des microbulles par ultrasons induit la cavitation et la sonoporation qui perméabilisent le tissu et facilite la délivrance locale de médicaments. De plus, cette destruction ciblée peut induire l'infiltration de cellules immunitaires et la libération d'antigènes tumoraux, déclenchant une réponse immunitaire anti-tumorale. Dans cette thèse, nous avons quantifié l'activation de la réponse immunitaire dans le carcinome hépatocellulaire, suivant la délivrance de nanoparticules chargés en microARN-122 et anti-microARN-21. Dans les nœuds lymphocytaires tumoraux, une baisse d'expression des cytokines pro-tumorales et une augmentation d'expression des cytokines anti-tumorales ont été observées, suggérant une réponse thérapeutique positive. L'approche thérapeutique de destruction ciblée des microbulles par ultrasons pour la délivrance de micro-ARN s'avère donc être un outil immuno-modulatoire puissan

Development of Targeted Microbubbles for Ultrasound Molecular Imaging of Breast Cancer

National audienceA subtype of metastatic breast cancer has recently been identified, in which netrin-1 is over-expressed. Netrin-1 binds to dependence receptors inducing survival, while its absence actively triggers apoptosis. An anti-netrin-1 therapy is under development , which disrupts ligand-receptor interaction and induces apoptosis. To identify potential responders for such a targeted therapy, netrin-1 expression has to be analyzed. Molecular imaging with ultrasound (US) is able to detect the expression of a specific protein when using targeted contrast agents. Thus, we aim to develop an US molecular imaging approach using microbubbles (MB) functionalized with an anti-netrin-1 antibody. Binding was tested on purified netrin-1 protein in static and dynamic conditions. Anti-netrin-1 MB were validated in in-vitro assays and will be further tested in pre-clinical studies using metastatic breast cancer animal models

Functionally distinct patterns of nucleosome remodeling at enhancers in glucocorticoid-treated acute lymphoblastic leukemia

Background: Precise nucleosome positioning is an increasingly recognized feature of promoters and enhancers, reflecting complex contributions of DNA sequence, nucleosome positioning, histone modification and transcription factor binding to enhancer activity and regulation of gene expression. Changes in nucleosome position and occupancy, histone variants and modifications, and chromatin remodeling are also critical elements of dynamic transcriptional regulation, but poorly understood at enhancers. We investigated glucocorticoid receptor-associated (GR) nucleosome dynamics at enhancers in acute lymphoblastic leukemia. Results: For the first time, we demonstrate functionally distinct modes of nucleosome remodeling upon chromatin binding by GR, which we term central, non-central, phased, and minimal. Central and non-central remodeling reflect nucleosome eviction by GR and cofactors, respectively. Phased remodeling involves nucleosome repositioning and is associated with rapidly activated enhancers and induction of gene expression. Minimal remodeling sites initially have low levels of enhancer-associated histone modification, but the majority of these regions gain H3K4me2 or H3K27Ac to become de novo enhancers. Minimal remodeling regions are associated with gene ontologies specific to decreased B cell number and mTOR inhibition and may make unique contributions to glucocorticoid-induced leukemia cell death. Conclusions: Our findings form a novel framework for understanding the dynamic interplay between transcription factor binding, nucleosome remodeling, enhancer function, and gene expression in the leukemia response to glucocorticoids. Electronic supplementary material The online version of this article (doi:10.1186/s13072-015-0046-0) contains supplementary material, which is available to authorized users

MOESM1 of Functionally distinct patterns of nucleosome remodeling at enhancers in glucocorticoid-treated acute lymphoblastic leukemia

Additional file 1. GREAT output tables for the Gene Ontology (GO) and Mouse Phenotype ontologies (Excel format). GREAT analysis using all GR ChIP-Seq peaks (a), as well as subsets of GR peaks with central remodeling (b), minimal remodeling (c), non-central remodeling (d), or phased remodeling (e)

Ultrasound molecular imaging as a non-invasive companion diagnostic for netrin-1 interference therapy in breast cancer

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From netrin‐1‐targeted <scp>SPECT</scp> / <scp>CT</scp> to internal radiotherapy for management of advanced solid tumors

International audienceTargeted radionuclide therapy is a revolutionary tool for the treatment of highly spread metastatic cancers. Most current approaches rely on the use of vectors to deliver radionuclides to tumor cells, targeting membrane-bound cancer-specific moieties. Here, we report the embryonic navigation cue netrin-1 as an unanticipated target for vectorized radiotherapy. While netrin-1, known to be reexpressed in tumoral cells to promote cancer progression, is usually characterized as a diffusible ligand, we demonstrate here that netrin-1 is actually poorly diffusible and bound to the extracellular matrix. A therapeutic anti-netrin-1 monoclonal antibody (NP137) has been preclinically developed and was tested in various clinical trials showing an excellent safety profile. In order to provide a companion test detecting netrin-1 in solid tumors and allowing the selection of therapy-eligible patients, we used the clinical-grade NP137 agent and developed an indium-111-NODAGA-NP137 single photon emission computed tomography (SPECT) contrast agent. NP137-111 In provided specific detection of netrin-1-positive tumors with an excellent signal-to-noise ratio using SPECT/CT imaging in different mouse models. The high specificity and strong affinity of NP137 paved the way for the generation of lutetium-177-DOTA-NP137, a novel vectorized radiotherapy, which specifically accumulated in netrin-1-positive tumors. We demonstrate here, using tumor cell-engrafted mouse models and a genetically engineered mouse model, that a single systemic injection of NP137-177 Lu provides important antitumor effects and prolonged mouse survival. Together, these data support the view that NP137-111 In and NP137-177 Lu may represent original and unexplored imaging and therapeutic tools against advanced solid cancers

HIV modulates the expression of ligands important in triggering natural killer cell cytotoxic responses on infected primary T-cell blasts

The ability of natural killer (NK) cells to kill virus-infected cells depends on the presence of ligands for activation receptors on the target cells. We found the presence of few, if any, NKp30 and NK46 ligands on T cell blasts infected with HIV, although NKp44 ligands were found on infected cells. HIV does induce the NKG2D ligands ULBP-1, -2, and -3. These ligands are involved in triggering NK cells to kill autologous HIV-infected cells, because interfering with the interaction between NKG2D, but not NKp46, on NK cells and its ligands on HIV-infected cells drastically reduced the lysis of infected cells. Interfering with the binding of the NK-cell coreceptors NTB-A and 2B4 to their ligands also decreased destruction by NK cells. The coreceptor ligands, NTB-A and CD48, were also found to be down-regulated during the course of HIV infection. Thus, ligands for NK-cell receptors are modulated during the course of HIV infection, which may greatly alter NK cells' ability to kill the infected cells