Genoscience Pharma (France)

Abstract Description Autoimmune diseases, such as multiple sclerosis and type-1 diabetes (T1D), are mediated by autoantigen-specific CD4+ and CD8+ T cells. Treatment with antigen (Ag)-containing biodegradable poly(lactide-co-glycolide) (PLGA) nanoparticles, i.e. tolerogenic immune-modifying particles (TIMP, also denoted as CNPs), induces Ag-specific tolerance. While extensive work has shown the effect of Ag-specific TIMP treatment on CD4+ T cells, less is known on how Ag-specific TIMP treatment regulates CD8+ T cell activity. Our data show that treatment of NOD mice with TIMPs containing whole diabetogenic antigens results in an increased number of CD4+ and CD8+ T cells expressing regulatory markers, a decrease in disease incidence, and significantly decreases the number of T cells present within the pancreas. While these findings show that Ag-specific treatment can modulate CD8+ T cells, these effects may or may not require CD4+ Tregs. Therefore, we sought to determine if treatment of mice with TIMPs containing only a single CD4+ T cell peptide epitope within the same protein could result in modulated CD8+ T cell function. Utilizing the OTI CD8+ T cell transfer into B6.RIPmOVA mouse model of T1D, mice were treatment with unloaded TIMP vs TIMP-OVA323, i.e., a dominant CD4+ T cell epitope, when mice had an average blood sugar of > 225 mg/dL. The data show that TIMP-OVA323 treatment restored my to euglycemia for ∼9 weeks in a Treg-dependent manner, and anti-CD25 treatment block tolerance induction. Funding Sources This work was supported by grants to S.D.M. from Cour Pharmaceutical Development Company, NIH grant R01AI148076; and by gifts from the Johnnie Walkers MS Foundation, the Amy and David Fulton Foundation, the Crammer Family Foundation, the Thomas and Deige McLaughlin Foundation, and the Rottering Family Foundation. Topic Categories Immune Response Regulation: Cellular Mechanisms (IRC)

Palmitoyl-protein thioesterase-1 (PPT1) is a clinical stage druggable target for inhibiting autophagy in cancer. We aimed to determine the cellular and molecular activity of targeting PPT1 using ezurpimtrostat, in combination with an anti-PD-1 antibody. In this study we used a transgenic immunocompetent mouse model of hepatocellular carcinoma. Herein, we revealed that inhibition of PPT1 using ezurpimtrostat decreased the liver tumor burden in a mouse model of hepatocellular carcinoma by inducing the penetration of lymphocytes into tumors when combined with anti-programmed death-1 (PD-1). Inhibition of PPT1 potentiates the effects of anti-PD-1 immunotherapy by increasing the expression of major histocompatibility complex (MHC)-I at the surface of liver cancer cells and modulates immunity through recolonization and activation of cytotoxic CD8+ lymphocytes. Ezurpimtrostat turns cold tumors into hot tumors and, thus, could improve T cell-mediated immunotherapies in liver cancer.

In advanced cancer patients, tumor burden is calculated using the sum of the longest diameters (SLD) of the target lesions, a measure that lumps all lesions together and ignores intra-patient heterogeneity. Here, we used a rich dataset of 342 metastatic bladder cancer patients treated with a novel immunotherapy agent to develop a Bayesian multilevel joint model that can quantify heterogeneity in lesion dynamics and measure their impact on survival. Using a nonlinear model of tumor growth inhibition, we estimated that dynamics differed greatly among lesions, and inter-lesion variability accounted for 21% and 28% of the total variance in tumor shrinkage and treatment effect duration, respectively. Next, we investigated the impact of individual lesion dynamics on survival. Lesions located in the liver and in the bladder had twice as much impact on the instantaneous risk of death compared to those located in the lung or the lymph nodes. Finally, we evaluated the utility of individual lesion follow-up for dynamic predictions. Consistent with results at the population level, the individual lesion model outperformed a model relying only on SLD, especially at early landmark times and in patients with liver or bladder target lesions. Our results show that an individual lesion model can characterize the heterogeneity in tumor dynamics and its impact on survival in advanced cancerpatients.

Pulmonary-resident memory T cells (TRM) and B cells (BRM) orchestrate protective immunity to reinfection with respiratory pathogens. Developing methods for the in situ detection of these populations would benefit both research and clinical settings. To address this need, we developed a novel in situ immunolabelling approach combined with clinic-ready fibre-based optical endomicroscopy (OEM) to detect canonical markers of lymphocyte tissue residency in situ in human lungs undergoing ex vivo lung ventilation (EVLV). Initially, cells from human lung digests (confirmed to contain TRM/BRM populations using flow cytometry) were stained with CD69 and CD103/CD20 fluorescent antibodies and imaged in vitro using KronoScan, demonstrating it's ability to detect antibody labelled cells. We next instilled these pre-labelled cells into human lungs undergoing EVLV and confirmed they could still be visualised using both fluorescence intensity and lifetime imaging against background lung architecture. Finally, we instilled fluorescent CD69 and CD103/CD20 antibodies directly into the lung and were able to detect TRM/BRM following in situ labelling within seconds of direct intra-alveolar delivery of microdoses of fluorescently labelled antibodies. In situ, no wash, immunolabelling with intra-alveolar OEM imaging is a novel methodology with the potential to expand the experimental utility of EVLV and pre-clinical models.

Lyme disease is present worldwide but is most prevalent in the temperate regions of the hemisphere, where it is the primary vector-borne illness. Every year, numerous individuals in Europe and the United States receive a diagnosis and undergo treatment for Lyme disease. The transmission of tick-borne diseases is a complex process, and it is challenging to identify the factors that contribute to their spread. Epidemiology provides a means of analyzing the distribution of Lyme disease within a population and identifying risk factors

Treatment evaluation in advanced cancer mainly relies on overall survival and tumor size dynamics. Both markers and their association can be simultaneously analyzed by using joint models, and these approaches are supported by many softwares or packages. However, these approaches are essentially limited to linear models for the longitudinal part, which limit their biological interpretation. More biological models of tumor dynamics can be obtained by using nonlinear models, but they are limited by the fact that parameter identifiability require rich dataset. In that context Bayesian approaches are particularly suited to incorporate the biological knowledge and increase the information available, but they are limited by the high computing cost of Monte-Carlo by Markov Chains algorithms. Here, we aimed to assess the performances of the Hamiltonian Monte-Carlo (HMC) algorithm implemented in Stan for inference in a nonlinear joint model. The method was validated on simulated data where HMC provided proper posterior distributions and credibility intervals in a reasonable computational time. Then the association between tumor size dynamics and survival was assessed in patients with advanced or metastatic bladder cancer treated with atezolizumab, an immunotherapy agent. HMC confirmed limited sensitivity to prior distributions. A cross-validation approach was developed and identified the current slope of tumor size dynamics as the most relevant driver of survival. In summary, HMC is an efficient approach to perform nonlinear joint models in a Bayesian framework, and opens the way for the use of nonlinear models to characterize both the rapid dynamics and the intersubject variability observed during cancer immunotherapy treatment.

Among the acquired modifications in cancer cells, changes in lysosomal phenotype and functions are well described, making lysosomes a potential target for novel therapies. Some weak base lipophilic drugs have a particular affinity towards lysosomes, taking benefits from lysosomal trapping to exert anticancer activity. Here, we have developed a new lysosomotropic small molecule, GNS561, and assessed its activity in multiple in vitro intrahepatic cholangiocarcinoma models (HuCCT1 and RBE cell lines and patient-derived cells) and in a chicken chorioallantoic membrane xenograft model. GNS561 significantly reduced cell viability in two intrahepatic cholangiocarcinoma cell lines (IC50 of 1.5 ± 0.2μM in HuCCT1 and IC50 of 1.7 ± 0.1μM in RBE cells) and induced apoptosis as measured by caspases activation. We confirmed that GNS561-mediated cell death was related to its lysosomotropic properties. GNS561 induced lysosomal dysregulation as proven by inhibition of late-stage autophagy and induction of a dose-dependent build-up of enlarged lysosomes. In patient-derived cells, GNS561 was more potent than cisplatin and gemcitabine in 2/5 and 1/5 of the patient-derived cells models, respectively. Moreover, in these models, GNS561 was potent in models with low sensitivity to gemcitabine. GNS561 was also efficient in vivo against a human intrahepatic cholangiocarcinoma cell line in a chicken chorioallantoic membrane xenograft model, with a good tolerance at doses high enough to induce an antitumor effect in this model. In summary, GNS561 is a new lysosomotropic agent, with an anticancer activity against intrahepatic cholangiocarcinoma. Further investigations are currently ongoing to fully elucidate its mechanism of action.

More and more evidence supports the view that extracellular matrix (ECM) is essential for growth and patterning during development. However, the mechanisms underlying patterning and morphogenesis are unknown. In both adult and developing mammalians, we have characterized a new type of extracellular matrix structure that we have named fractones. Fractones are chemically similar to basement membranes but differ from the latter by their location. Fractones reside in the parenchyma of tissues and organs. They are are visualized as particles by light microscopy but often display a fractal ultrastructure. In contrast, basement membranes are located as sheaths at the connective tissue/parenchyma interface. We have shown that fractone‐associated heparan sulfates bind and activate growth factors to regulate stem cell proliferation in the neurogenic zone of the adult brain. Here, we used multiple fluorescence immunohistochemistry to visualize fractones, basement membranes and cell proliferation throughout mouse development, with a focus on brain structuration. Fractones preceded basement membranes and were associated with proliferating pluripotent stem cells in the inner cell mass of the blastocyst. During gastrulation, fractones were found in the three germinal tissues. During neurulation, fractones were visualized at the vicinity of proliferating neuroepithelial cells in the notoplate and in the ventricular zone. During corticalization, fractones gradually displaced through the neural tissue according to an in‐out gradient towards the basement membrane of superficial meninges. At this time, fractone distribution negatively correlated with neural cell proliferation. For white matter tract formation, axonal growth was accompanied by fractones along meningeal elongations that progressively patterned the forming brain. Then, brain fractones disappeared only to reappear soon after birth throughout the sub ventricular zone, a location that persisted into adulthood. Together, our results strongly suggest that fractones and basement membranes are essential for patterning and morphogenesis.Support or Funding InformationSupported by JSPS S09109This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Abstract Background: Hepatocellular carcinoma (HCC) remains a major health problem, often diagnosed at late stages with limited number of therapeutic options. New drugs with original mechanisms of action are urgently aimed to improve current armamentarium in HCC patients. Quinoline derivatives are novel class of oral small molecules inducing multiple cellular effects such as inhibition of autophagy, induction of apoptosis, and cell cycle modulation. The aim of these studies was to assess tolerance and efficacy of a new quinolone derivative GNS561. Material and methods: In vitro experiments were realized with viability, apoptosis and migration in tumor cells in HCC cell lines and primary tumor. Drug tolerance and plasma and liver pharmacokinetic were evaluated after single and repeated dosing in mice and rat. GNS561 and sorafenib efficacy in vivo were evaluated in a PDX orthotopic BALB/c-nu mouse model and in a diethylnitrosamine (DEN)-induced HCC cirrhotic rat model. AFP, cell proliferation and tumor weight and size were assessed in mice. In rat tumor progression was followed by MRI, pathological analysis (tumor size and number), immunohistochemistry and PCR analysis after 6weeks of treatment. Results: GNS 561 shows potent anti-proliferative activity when assayed against a panel of human tumor cell lines and notably against a panel of HCC patient primry tumors even in those with sorafenib resistance (Mean EC50 3µM vs 11µM for sorafenib). GNS561 is highly selectively trapped in the liver. Plasma and liver PK in mice and rats after single and repeated doses confirm this selectivity with good tolerance and oral bioavailability. In PDX mouse model, tumor growth was significantly reduced by GNS561 with a dose-response manner, this tumor regression was associated with AFP level decreases by 72% with GNS561 (p=0.002) and 54% with sorafenib (p=0.046) compared to control. In rat model, mean number of tumors was significantly lower in GNS561 at 15mg/kg group (n=50.6), in sorafanib at 10mg/kg (n=65.1) and in combination group (n=40.6), when compared to control (n=100.4; p=0.0024, p= 0.029 and p=0.0002). Tumor decrease measured by MRI was associated with a significantly reduced proliferation of tumor cells particularly in GNS561 group (70%) and combination (84%) compared to control, whereas the effect of sorafenib alone on proliferation was modest (30%). Conclusions: GNS561 is a liver selective drug with good tolerance and promising efficacy in different HCC animal models. GNS 561 was more efficient than sorafenib to control tumor growth in preclinical models. Based on its safe toxicity profile and potent activity in rodent models, GNS 561 is now aimed to further reach clinical development in patients with HCC in 2017. Citation Format: Firas BASSISSI, Zuzana Macek Jilkova, Sonia Brun, Jerome Courcambeck, Jennifer Tracz, Keerthi Kurma, Gaël S. Roth, Cindy Khaldi, Corinne Chaimbault, Benoit Quentin, Emilie Asseraf, Antoine Beret, Eric Raymond, Philippe Halfon, Thomas Decaens. GNS561 a new quinoline derivative inhibits the growth of hepatocellular carcinoma in a cirrhotic rat and human PDX orthotopic mouse models [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 5124. doi:10.1158/1538-7445.AM2017-5124

Water safety is a major concern for public health and for natural environment preservation. We propose to use bacteriophages to develop biosensor tools able to detect human and animal pathogens present in water. For this purpose, we take advantage of the highly discriminating properties of the bacteriophages, which specifically infect their bacterial hosts. The challenge is to use a fluorescent reporter protein that will be synthesized, and thus detected, only once the specific recognition step between a genetically modified temperate bacteriophage and its bacterial host has occurred. To ensure the accuracy and the execution speed of our system, we developed a test that does not require bacterial growth, since a simple 1-hour infection step is required. To ensure a high sensitivity of our tool and in order to detect up to a single bacterium, fluorescence is measured using a portable flow cytometer, also allowing on-site detection. In this study, we have constructed and characterized several "phagosensor" prototypes using the HK620 bacteriophage and its host Escherichia coli TD2158 and we successfully adapted this method to Salmonella detection. We show that the method is fast, robust and sensitive, allowing the detection of as few as 10 bacteria per ml with no concentration nor enrichment step. Moreover, the test is functional in sea water and allows the detection of alive bacteria. Further development will aim to develop phagosensors adapted on demand to the detection of any human or animal pathogen that may be present in water.