To understand potential immune system alterations in newly diagnosed, untreated, pancreatic cancer patients and provide a foundation for immunotherapy, we profiled PBMC from pancreatic ductal adenocarcinoma (PDA) patients and age matched healthy controls using high dimensional CyTOF analysis. We developed two immune profiling panels: a broad profiling panel that includes 45 phenotypic markers that together permit the identification and enumeration of the main innate and adaptive immune cell subsets in humans, and a deep profiling panel that includes 45 features focusing on T cell phenotype and biology. We report a 2-fold increase in monocytes, and more plasmocytes in circulation in pancreatic cancer patients compared to age-matched controls. Using high dimensional approaches, we observe skewed T cell differentiation in pancreatic cancer patients, with CD8 T cells biased towards more CD45RA-positive CD27-positive CCR7-positive CD95-positive CD49d-positive stem cell memory cells (P=3x10-4), more CD45RA-negative CD27-positive CCR7-negative effector memory cells (P=0.002) and less CD45RA-positive CD27-negative CCR7-negative late effector memory cells (P=0.01) than age-matched controls. We further examinated alterations of T cell differentiation in CD8 T cell compartment in human spleens, and report increased proportions of late effector memory T cells in pancreatic cancer patients as compared to age-matched controls. Comparable observation of increased numbers of intra-splenic effector T cells was made in genetically engineered (KPC mice: LSL-KrasG12D/+;LSL-Trp53R172H/+;Pdx-1-Cre), as well as orthotopic mouse models of pancreatic cancer. In the former model, we observed an accumulation of adoptively transferred listeria-experienced effector T cells in the spleen. Together, these results demonstrate a trafficking defect of experienced T cells in the context of pancreatic cancer at baseline. We are now investigating the mechanisms underlying these observations, as well as their impact on T cell immunity of the patients. Our goal is to understand the nature of the skewing and how any changes in baseline immune health of the T cell compartment related to disease progression and/or response to therapy.  These studies should provide a foundation for improving therapy in pancreatic cancer patients. Additionally, they should provide new insights into the heterogeneity of T cell differentiation.

5 recent publications:

Chhatbar C, Detje CN, Grabski E, Borst K, Spanier J, Ghita L, Elliott DA, Jordão MJC, Mueller N, Sutton J, Prajeeth CK, Gudi V, Klein MA, Prinz M, Bradke F, Stangel M, Kalinke U. Type I Interferon Receptor Signaling of Neurons and Astrocytes Regulates Microglia Activation during Viral Encephalitis. Cell Rep. 2018 Oct 2;25(1):118-129.e4

Koestner W, Spanier J, Klause T, Tegtmeyer PK, Becker J, Herder V, Borst K, Todt D, Lienenklaus S, Gerhauser I, Detje CN, Geffers R, Langereis MA, Vondran FWR, Yuan Q, van Kuppeveld FJM, Ott M, Staeheli P, Steinmann E, Baumgärtner W, Wacker F, Kalinke U. Interferon-beta expression and type I interferon receptor signaling of hepatocytes prevent hepatic necrosis and virus dissemination in Coxsackievirus B3-infected mice. PLoS Pathog. 2018 Aug 3;14(8):e1007235.

Borst K, Frenz T, Spanier J, Tegtmeyer PK, Chhatbar C, Skerra J, Ghita L, Namineni S, Lienenklaus S, Köster M, Heikenwaelder M, Sutter G, Kalinke U. Type I interferon receptor signaling delays Kupffer cell replenishment during acute fulminant viral hepatitis. J Hepatol. 2017 Dec 21.

Hirche C, Frenz T, Haas SF, Döring M, Borst K, Tegtmeyer PK, Brizic I, Jordan S, Keyser K, Chhatbar C, Pronk E, Lin S, Messerle M, Jonjic S, Falk CS, Trumpp A, Essers MAG, Kalinke U. Systemic Virus Infections Differentially Modulate Cell Cycle State and Functionality of Long-Term Hematopoietic Stem Cells In Vivo. Cell Rep. 2017 Jun 13;19(11):2345-2356.

Frenz T, Grabski E, Buschjäger D, Vaas LA, Burgdorf N, Schmidt RE, Witte T, Kalinke U. CD4(+) T cells in patients with chronic inflammatory rheumatic disorders show distinct levels of exhaustion. J Allergy Clin Immunol. 2016 Aug;138(2):586-589.e10.

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The structure, dynamics and interactions of nucleic acids, DNA in particular, sensitively respond to a broad range of environmental factors many of them being natively present in intracellular space.  In-cell NMR spectroscopy is a tailored adaptation of conventional high-resolution nuclear magnetic resonance spectroscopy that permits characterization of stereo-chemical features of nucleic acids directly in living cells. I will present applications of the method for characterization of DNA structure, stability and for monitoring of interactions between DNA and low molecular weight compounds (drug-like molecules) in vivo.  The limitations of this method as well as the general considerations, comprising of critical factors to be accounted for during the planning of in-cell NMR experiment as well as during the interpretation of in-cell NMR data, will be discussed.

Acknowledgment: The presented work has been supported by Czech Science Foundation (grant No. 16-10504S,17-12075S), European Regional Development Fund (CZ.02.1.01/0.0/0.0/15_003/-0000477) and Horizon 2020 Programme (iNEXT: grant No. 653706)

During the last decade, intravital optical microscopy has launched a new trend in the field of biology. By using this advanced imaging technique we have established a new system for visualizing in situ behavior of a diversity of living cells within intact tissues and organs. Among them, we succeeded in visualizing the various dynamic phenomena within bones and joints, where various kinds of immune cells are produced and functioning although poorly analyzed by conventional methodology such as histological analyses with decalcified sections. We have so far identified the real modes of migration, differentiation and function of bone-destroying osteoclasts, special kind of macrophages responsible for bone and joint erosions. In addition to the application to animal experimental models, we are currently trying to adapt this technique for evaluating disease status in local foci of human patients. In this presentation I will present the recent update on intravital imaging studies on immune and other systems for clarifying in vivo behaviors of cell and tissue dynamics.

Large-scale genomic studies have demonstrated that approximately 50% of high-grade serous ovarian cancers (HGSOCs) harbor genetic and epigenetic alterations in homologous recombination repair (HRR) pathway genes. The most commonly altered HRR genes are BRCA1 and BRCA2, followed by other Fanconi Anemia Genes.  Loss of HRR causes genomic instability, hyperdependence on alternative DNA repair mechanisms, and enhanced sensitivity to PARP-inhibitors (PARPi) through the mechanism of synthetic lethality. PARP inhibitor resistance has emerged as a vexing clinical problem for the treatment of BRCA1/2 deficient tumors. The most prevalent mechanism of PARPi resistance is secondary events that cancel the original HRR alteration and restore HRR proficiency. PARPi resistance also develops without restoration of HRR proficiency through enhanced replication fork (RF) stabilization. We have recently made the surprising observation that BRCA2-deficient tumor cells can stabilize their replication forks and become resistant to PARPi by downregulating the expression of various methyltransferases, such as MLL3/4 and EZH2. Other new mechanisms of PARPi resistance will be discussed. A molecular understanding of PARP inhibitor resistance mechanisms may allow the generation of a new class of drugs, or a re-purposing of existing drugs, which may reverse this resistance and extend the use of PARP inhibitors to more tumor types.

Extracellular vesicles (EVs), including exosomes, have emerged as novel mediators of cell/cell communication in multiple biological and pathological contexts. They are produced by all cell types, present in every body fluid and contain proteins and RNA (coding and non coding) able to change the phenotype of their receiving cell. Despite their increasing importance, their biogenesis, their dispersion and their function remain poorly understood. My work focuses on these aspects, using different animal models and favouring in vivo studies, intravital imaging and correlated light and electron microscopy. Over the past years, I used C. elegans to identify new genes involved in exosome biogenesis and secretion, and described the function of one of them, the RAL-1 GTPase (Hyenne et al. JCB 2015). We further show that mammalian RalA and RalB control EV levels, content and function in mouse mammary carcinoma, and that it could contribute to primary tumor growth and metastasis formation (unpublished data). In parallel, we developed the use of zebrafish as a novel animal model to track circulating EVs in a living organism at high temporal and spatial resolution (Hyenne et al. Dev. Cell 2019). We provided the first description of tumor EVs’ hemodynamic behavior and documented their intravascular arrest. We showed that circulating tumor EVs are rapidly taken up by endothelial cells and blood patrolling macrophages and subsequently stored in degradative compartments. Finally, we demonstrated that tumor EVs activate macrophages and promote metastatic outgrowth.

Presence, function and targeting of G-quadruplexes in the human immunodeficiency virus-1 (HIV-1)

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Title & abstract TBD.

In the cell, many proteins begin folding co-translationally. To ensure proper function, co-translational folding must allow proteins to rapidly reach their native state while avoiding potentially harmful misfolded intermediates. Here we provide evidence that this process may be regulated in vivo by synonymous codon choice. Namely, we hypothesize that rare codons, which are known to be translated more slowly than their synonymous counterparts, may provide time for nascent polypeptide chains to begin folding. Using atomistic Monte Carlo simulations, we investigate the folding of various proteins whose sequences are evolutionarily enriched in rare codons and show that, indeed, these proteins can adopt stable native structure at translational intermediates corresponding to rare codon positions. Moreover, these translational intermediates fold more rapidly than their respective fully synthesized proteins, whose folding is drastically slowed by nonnative interactions. We then predict, using kinetic modeling, that these proteins’ proper co-translational folding relies on rare codons, and that synonymous substitution would dramatically reduce folding efficiency. This work sheds light on the factors that promote proper protein folding in the cellular environment, and on the role of crucial role of kinetics in biological self-assembly more broadly. 

- Amir Bitran, William Jacobs, Eugene Shakhnovich -

Title & abstract to come.

Rapid advances in DNA synthesis techniques have made it possible to engineer diverse genomic elements, pathways, and whole genomes, providing new insights into design and analysis of systems. The synthetic yeast genome project, Sc2.0 is well on its way with seven synthetic Saccharomyces cerevisiae chromosomes completed by a global team. The synthetic genome features several systemic modifications, including TAG/TAA stop-codon swaps, deletion of subtelomeric regions, introns, tRNA genes, transposons and silent mating loci. Strategically placed loxPsym sites enable genome restructuring using an inducible evolution system termed SCRaMbLE (Synthetic Chromosome Rearrangement and Modification by LoxP-mediated Evolution). SCRaMbLE can generate millions of derived variant genomes with predictable structures leading to complex genotypes and phenotypes.  The fully synthetic yeast genome provides a new kind of combinatorial genetics based on variations in gene content and copy number. Remarkably, the 3D structure of synthetic and native chromosomes are very similar despite the substantial changes introduced.

We recently completely engineered the yeast karyotype, by systematically fusing pairs of telomeres and deleting single centromeres, thus generating an isogenic series of yeast ranging from n=16 to n=2. These strains show reproductive isolation and a massively altered 3D genome structure, but are surprisingly “Normal” and show high fitness.

Finally, we have automated our big DNA synthesis pipeline (the GenomeFoundry@ISG), opening the door to parallelized big DNA assembly, including assembly of human genomic regions of 100 kb along with multiple designer synthetic variants thereof. We can precision deliver such segments to stem and cancer cells, and intend to use these methods to dissect genomic “dark matter”, perform transplants of specific human genomic regions to animal genomes, and endow human cells with new capabilities.

Dymond et al. 2011 Synthetic chromosome arms function in yeast and generate phenotypic diversity by design. Nature, 477:471-6.

Annaluru et al. 2014 Total synthesis of a functional designer eukaryotic chromosome. Science. 344:55-8.

Richardson et al. 2017 Design of a synthetic yeast genome, Sc2.0.  Science 355:1040-1044.

Mitchell et al. 2017 Synthesis, debugging and effects of synthetic chromosome consolidation: synVI and beyond. Science, 355 pii: eaaf4831.

Mercy et al. 2017 3D organization of synthetic and scrambled chromosomes. Science, 355 pii: eaaf4597.

Luo, Sun, Cormack and Boeke 2018 Karyotype engineering: chromosome fusion leads to reproductive isolation. Nature, 560:392-396.

Shen et al. 2018 Heterozygous diploid and interspecies SCRaMbLEing. Nat Commun. 9:1934.

I am a group leader and senior Lions Medical Research Foundation (LMRF) Fellow working at the University of Queensland Centre for Clinical Research (UQCCR). I am recognized as a national and international researcher working in the field of extracellular vesicles focusing on the reproductive biology (specifically in pregnancy and its complications) and I have been consistently invited as a speaker to both National and International meetings (e.g. Society of Reproductive investigation, American Diabetes Association, and International Society for Prenatal Diagnosis). I have 88 publications (42 as first or last author), which 68 are in the extracellular vesicles field. Within UQCCR, I have established and lead the EXOSOME BIOLOGY LABORATORY that conforms the ISO standards (ISO17025 and 13485), in which human exosomes can be isolated, characterised and their role elucidated to evaluate their clinical utility as biomarkers of disease and therapeutic interventions. Over the last 5 years, I have obtained exciting data suggesting that exosomes (i.e. membrane-bound nanovesicles) play a role throughout gestation, including, mediating a placental response to hyperglycaemia and insulin sensitivity. In addition, Ovarian cancer is the most lethal gynaecological cancer worldwide with the lack of early detection an important contributor to poor patient outcomes, thus, my group have been working on the potential role of exosomes in the diagnosis, drug delivery and real-time monitoring of ovarian cancer. 

March 12th 2019 will be a unique opportunity to discover original music compositions inspired by recent scientific discoveries !

This amazing work was created in the context of the “Aux Frontières des LabEx” PSL project ‘Muse-IC’, lead by Judith Miné-Hattab (Institut Curie researcher, LabEx DEEP). The project gathered six pairs of international composers and scientists working in the fields of biology, physics or biophysics (Institut Curie, MIT, Institut d’Astrophysique Spatiale), offering composers the opportunity to create original music scores inspired by recent scientific discoveries.

The concert is the culmination of this project. It is opened to all and free, so if you are interested, save this date and join us on March 12th, 20h30 Salle Cortot !

More information and registration on http://bit.ly/projet-muse-ic and https://www.weezevent.com/projet-muse-ic-musiques-inspirees-de-decouvertes-scientifiques

I will discuss recent insights into new therapeutic targets in cancer with an emphasis on lymphoma/leukemia. These will include mRNA translation, immune-receptors and unpublished data from genetic CRISPR/CAS9 screens for new and druggable mechanisms

Our lab uses biophysical methods, with particular emphasis on micromanipulation of single DNA molecules and single chromosomes, to study the internal structure of chromosomes in vivo, and to study chromosome-organizing proteins and DNA topoisomerases in vitro. We also develop mathematical models relevant to these types of experiments. Projects in progress involve combining fluorescence microscopy and force microscopy in experiments on DNA-protein complexes and whole chromosomes, and in-vivo studies of coupling of chromosome dynamics to gene expression.

Title & abstract to come.

Meeting co-sponsored by Gustave Roussy and Institut Curie that honors the memory of Professor Roger Monier, a prominent figure in French modern biology.

This year, we celebrate the 10th edition, a scientific day meeting focuses on a forefront issues in cancer biology: « Imaging: a revolution ».

For logistics reasons, please register with the following link http://minilien.curie.fr/aw75jj

Affiche du séminaire

Acute leukemia is the most common cancer of childhood and one of the most deadly due to relapsed disease. Predicting patients at high risk of relapse is an imperfect science and currently combines clinical features, somatic alterations and early response to therapy. Yet, despite this, for the 20% of children who will relapse, at least half will die of their leukemia. Using single-cell, high-parameter analysis of primary leukemia specimens from diagnosis, we demonstrate the ability to more accurately predict patients at high risk for relapse utilizing novel developmental classification and machine learning tools.  This approach also enables identification the cells that are responsible for relapse, thereby providing insight into potential therapeutic strategies.

Agenda to come.

more information to come 

Although cancer is governed by complex molecular systems, the composition and modular organization of these systems remains poorly understood. I will describe efforts by the Cancer Cell Map Initiative (CCMI) to generate large protein interaction maps of tumor cells, which we are integrating with existing molecular and structural data to assemble a comprehensive multiscale map of cancer cell biology. The current draft map contains a hierarchy of ~250 systems covering both known hallmarks and unexpected components. Integration with tumor mutation profiles suggests that the modules under strongest selective pressure in cancer are often not genes, but extend upwards in scale to include protein complexes, broad cellular processes, and organelles. This observation suggests a classification of cancer based on convergence of mutations at key bottlenecks in the hierarchy of cellular systems.

https://medschool.ucsd.edu/som/medicine/research/labs/ideker/Pages/default.aspx