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The antiapoptotic protein BCL2 plays critical roles in regulating lymphocyte development and immune responses, and has also been implicated in tumorigenesis and tumor survival. However, it is unknown whether BCL2 is critical for antitumor immune responses. We evaluated whether venetoclax, a selective small-molecule inhibitor of BCL2, would influence the antitumor activity of immune checkpoint inhibitors (ICI). We demonstrate in mouse syngeneic tumor models that venetoclax can augment the antitumor efficacy of ICIs accompanied by the increase of PD-1+ T effector memory cells. Venetoclax did not impair human T-cell function in response to antigen stimuli in vitro and did not antagonize T-cell activation induced by anti–PD-1. Furthermore, we demonstrate that the antiapoptotic family member BCL-XL provides a survival advantage in effector T cells following inhibition of BCL2. Taken together, these data provide evidence that venetoclax should be further explored in combination with ICIs for cancer therapy. Significance: The antiapoptotic oncoprotein BCL2 plays critical roles in tumorigenesis, tumor survival, lymphocyte development, and immune system regulation. Here we demonstrate that venetoclax, the first FDA/European Medicines Agency–approved BCL2 inhibitor, unexpectedly can be combined preclinically with immune checkpoint inhibitors to enhance anticancer immunotherapy, warranting clinical evaluation of these combinations. This article is highlighted in the In This Issue feature, p. 1

Previously, we have shown that sublethal exposure of blue light caused increased cell membrane permeability in Fusobacterium nucleatum. The aim of the present study was to test the effect of this exposure on the activity of Lavender, Sage, Echinacea and Mastic gum extracts against volatile sulfide compound (VSC) production by Fusobacterium nucleatum. Bacterial suspensions were pre-exposed to blue light (400-500 nm) bellow minimal inhibitory dosage (sub-MID). Exposed and nonexposed samples were inoculated into test tubes containing growth medium, filtered saliva with or without herbal extracts. Following incubation, test tubes were tested for malodor production (odor judge scores), VSC levels (OralChroma), salivary protein degradation (SDS-PAGE) and bacterial cell membrane damage (fluorescence microscopy). Results showed that sub-MID blue light exposure significantly increased the ability of Lavender and Echinacea to reduce VSC production by Fusobacterium nucleatum by more than 30%. These results suggest that sublethal blue light exposure may be useful to increase the efficacy of antimalodor agents.

Objectives The COVID-19 pandemic is a global health problem. We, as the EMERGE (EMErging RheumatoloGists and rEsearchers) group of PReS (Pediatric Rheumatology European Society) analyzed how the pandemic has affected pediatric rheumatology practice. Methods An online survey was developed to assess changes in pediatric rheumatology practice due to the pandemic. Results were analyzed using descriptive statistics. Results From 70 countries, 493 pediatric rheumatologists (80.3% in pediatric rheumatology practice for ≥5 years) responded to the survey. Around 70% disagreed that the pandemic led to reduced prescription of nonsteroidal anti-inflammatory drugs, conventional synthetic and biologic disease-modifying antirheumatic drugs. Almost half were more likely to taper corticosteroids faster. One-fifth hesitated to switch the major immunosuppressant during a flare. Patients encountering difficulties obtaining hydroxychloroquine and tocilizumab due to shortages were noted by 192 (38.9%) and 44 (8.9%), respectively. Twenty to 30% indicated that their patients had experienced a flare or delay in diagnosis/intervention due to postponed appointments.53% mentioned use of phone calls/smartphone applications while 47% shifted towards video consultations for patient care. Respondents indicated an increased number of patients with Kawasaki disease (30%), macrophage activation syndrome (15.6%), unusual vasculitic rashes (31.4%), and hyperinflammation (33.5%) during the pandemic. Conclusion This is the largest survey to date addressing changes in pediatric rheumatology practice due to the COVID-19 pandemic. Primary changes were due to delays in clinic appointments, increase in use of virtual technologies, and concerns about the use of immunosuppressive therapies. An increased number of patients with Kawasaki disease/hyperinflammation mentioned by the respondents is noteworthy., • COVID-19 pandemic threatens millions of lives worldwide. • As pediatric rheumatologists, we take care of a vulnerable population. • The main changes in pediatric rheumatology practice during the COVID-19 pandemic were due to delays in clinic appointments, increase in use of virtual technologies to decrease in-person visits, use of anti-rheumatic drugs treatment/prophylaxis, and concerns about using immunosuppressive therapies. • An increase in the number of patients with Kawasaki disease/hyperinflammation was also mentioned by the respondents of the survey. • Understanding the challenges imposed by the COVID-19 pandemic on the community of pediatric rheumatologists will help tailor future recommendations regarding the management of pediatric rheumatology patients during the pandemic according to the needs in daily clinical practice.

In the search for new therapeutic alternatives for Chagas disease, a series of six aryloxy -naphthoquinone derivatives were synthesized and evaluated in vitro against Trypanosoma cruzi epimastigotes of the Tulahuen 2, INC-5, and NINOA strains. The compounds 3d and 4a showed better or similar trypanosomicidal activity than the reference drug nifurtimox. In addition, 3d and 4a also elicited better trypanosomicidal activity than nifurtimox against T. cruzi bloodstream trypomastigotes. On the other hand, 3b showed the highest selective indexes (SI values between 44 and 500, in the three T. cruzi strains). Finally, molecular docking studies suggested that these compounds could be potential trypanothione reductase inhibitors. Therefore, based on these new results, we validated that the aryloxy-naphthoquinone scaffold is essential to obtain more selective cytotoxic and trypanosomicidal compounds.

Proteins of the bromodomain and extra-terminal (BET) domain family are epigenetic readers that bind acetylated histones through their bromodomains to regulate gene transcription. Dual-bromodomain BET inhibitors (DbBi) that bind with similar affinities to the first (BD1) and second (BD2) bromodomains of BRD2, BRD3, BRD4 and BRDt have displayed modest clinical activity in monotherapy cancer trials. A reduced number of thrombocytes in the blood (thrombocytopenia) as well as symptoms of gastrointestinal toxicity are dose-limiting adverse events for some types of DbBi1–5. Given that similar haematological and gastrointestinal defects were observed after genetic silencing of Brd4 in mice6, the platelet and gastrointestinal toxicities may represent on-target activities associated with BET inhibition. The two individual bromodomains in BET family proteins may have distinct functions7–9 and different cellular phenotypes after pharmacological inhibition of one or both bromodomains have been reported10,11, suggesting that selectively targeting one of the bromodomains may result in a different efficacy and tolerability profile compared with DbBi. Available compounds that are selective to individual domains lack sufficient potency and the pharmacokinetics properties that are required for in vivo efficacy and tolerability assessment10–13. Here we carried out a medicinal chemistry campaign that led to the discovery of ABBV-744, a highly potent and selective inhibitor of the BD2 domain of BET family proteins with drug-like properties. In contrast to the broad range of cell growth inhibition induced by DbBi, the antiproliferative activity of ABBV-744 was largely, but not exclusively, restricted to cell lines of acute myeloid leukaemia and prostate cancer that expressed the full-length androgen receptor (AR). ABBV-744 retained robust activity in prostate cancer xenografts, and showed fewer platelet and gastrointestinal toxicities than the DbBi ABBV-07514. Analyses of RNA expression and chromatin immunoprecipitation followed by sequencing revealed that ABBV-744 displaced BRD4 from AR-containing super-enhancers and inhibited AR-dependent transcription, with less impact on global transcription compared with ABBV-075. These results underscore the potential value of selectively targeting the BD2 domain of BET family proteins for cancer therapy. ABBV-744, a selective inhibitor of the BD2 domains of BET family proteins, is effective against prostate cancer in mouse xenograft models, with lower toxicities than the dual-bromodomain BET inhibitor ABBV-075.

Introduction: The World Health Organization establishes that 80% of the world’s population uses traditional medicine for their primary care, because they contain compounds responsible for their properties. Objective: To evaluate the antifungal effect of the essential oil of Citrus paradisi against C. albicans and the cytotoxic effect in three cell lines in vitro. Methods: The phytochemical characterization of the oil was carried out by chemical methods and Gas chromatography (GC-MS) and the antifungal effect against C. albicans (ATCC 90029) was evaluated by the Kirby-Bauer method, which evaluated concentrations of 0.75 - 20 μg/mL and compared with nystatin 100,000 Ul/mL as a positive control. The percentage of the relative inhibitory effect was calculated. The minimum inhibitory concentration (MIC) was determined at 24 hours. Moreover, the cytotoxic effect on C. albicans and cell lines was determined by the colorimetric MTT tetrazolium assay. Finally, the antifungal effect against Candida strains isolated from clinical samples was evaluated at a concentration of 20 μg/mL. Results: The essential oil showed an antifungal effect with a percentage of inhibition of 123%. The MIC was 2.5 μg/mL, and the cytotoxicity index was 5.44 μg/mL for C. albicans. The IC50 values were 21.060, 9.482 and 4.176 μg/mL for Vero E6, J774A.1 and MDBK cells respectively. Conclusion: These results show the use of C. paradisi essential oil as an alternative treatment in oral antifungal therapy, it is beneficial due to its antifungal effect and its low toxicity on cell cultures.

A previous study showed that sub-lethal exposure of blue light caused cell membrane damage in Fusobacterium nucleatum (Fn). The aim of the present study was to test the combined effect of blue light and silver nanoparticles against Fn. Bacterial suspensions were exposed to blue light (400–500 nm) with or without silver nanoparticles (10 nm). Exposed and non-exposed samples were studied for malodor production (Odor judge scores), VSC levels (Halimeter), reactive oxygen species (ROS) production (fluorimeter), and bacterial cell membrane damage (fluorescence microscopy). The results showed that combining blue light exposure and silver nanoparticles significantly reduced malodor and VSC production by Fn concomitant with increased ROS levels and bacterial cell membrane damage. These results suggest that silver nanoparticles may increase blue light phototoxicity against Fn.

The genus Zingiberaceae has been widely used for phytotherapeutic purposes in traditional medicine throughout the world for its anti-inflammatory activity. Experimental studies have established that inflammation caused by chronic infections represents a risk factor for different forms of cancer. The objective of this study was focused on determining the anti-inflammatory capacity and cytotoxic activity of aqueous extracts of Elettaria cardamomum (cardamom) and Curcuma Longa (turmeric). The extracts were obtained by maceration and, through GC-MS/MS, a total of 11 different chemical components were determined in the aqueous extract of cardamom and 7 in the extract of turmeric. The main compounds found in cardamom and turmeric were α-terpinyl acetate (54.46%) and β-turmerone (33.45%), respectively. RT-qPCR results showed significantly lower gene expression levels of innate inflammatory cytokines (IL-6 and TNF-α) compared to the control (LPS). Also, it was observed that the extracts do not possess cytotoxic activity against different cell lines, where E. cardamomum showed EC50 (µg/mL) of 473.84 (HeLa cells), 237.36 (J774A.1 cells), 257.51 (Vero E6 cells), and 431.16 (Balb/C peritoneal cells) and C. longa showed EC50 (µg/mL) of 351.17 (HeLa cells), 430.96 (J774A.1 cells), 396.24 (Vero E6 cells), and 362.86 (Balb/C peritoneal cells). The results of this research suggest that natural extracts of E. cardamomum and C. longa possess anti-inflammatory effects and no cytotoxic activity against HeLa, J774A.1, Vero E6, and Balb/C peritoneal cell lines. Finally, it was observed that the extracts also decreased nitric oxide (NO) production in peritoneal macrophages.

Endogenous heat shock cognate 71 kDa protein (HSC70) has a vital role in early embryonic development. This study assessed the effects of exogenous HSC70 on bovine embryo development and expression of genes associated with apoptosis. Expression analyses of HSPA1A, HSPA8, Bcl-2, and Bax genes were performed in bovine embryos in vivo on day 7 of development. Subsequently, expression of HSPA1A and HSPA8 were associated with apoptotic genes (Bcl-2 and Bax) in cultured bovine embryos in vitro that were supplemented with various concentrations (0 or control group, 50, and 100 ng) of HSC70. The results indicated that the control group (0 ng) in vitro embryos had higher expression of HSPA8, Bax, and Bcl-2 genes, compared with the vivo embryos (P

Oral malodour is considered to be caused mainly by the production of volatile sulfide compounds (VSC) by anaerobic gram-negative oral bacteria. Previous studies showed that these bacteria were susceptible to blue light phototoxicity mediated by the production of reactive oxygen species (ROS). In the present study, we tested the effect of blue light on the integrity Fusobacterium nucleatum's membrane, cellular proteins and DNA. Bacterial samples were exposed to high intensity blue light for 0, 70, 140 and 280 s (i.e. fluences of 0, 96, 192 and 384 J cm-2 , respectively). Following light exposure, bacterial samples were examined for membrane damage using fluorescence microscopy, intra-cellular protein analysis using electrophoresis (SDS-PAGE) and DNA fragmentation using ultra-filtration. Results showed that the increasing exposure of bacterial samples to blue light caused increased membrane permeability concomitant with a reduction in intra-cellular proteins and DNA fragments content. These results suggest that membrane damage is the main effect of high intensity blue light exposure on malodour producing bacteria.

The oral cavity is an ecosystem that provides ideal conditions for the growth of bacteria, the Streptococcus genus is important for the formation of biofilms that lead to the development of dental caries, which affects the population worldwide. The world health organization encourages the use of plants thanks to its various therapeutic actions. Origanum vulgare L. (oregano), is an aromatic plant with medicinal and culinary properties. The objective of this study was to investigate the in vitro antimicrobial and antibiofilm activity of the ethanolic extract of oregano, against the growth of Streptococcus mutans and Streptococcus sobrinus ATCC. Leaves of the plant were obtained and the ethanolic extract was made by maceration. Antimicrobial activity was evaluated using the Kirby-Bauer method and compared with 2% chlorhexidine, subsequently the extract was incorporated into a hydrogel and its effect on biofilm formation was assessed by fluorescence microscopy and the main compounds were identified. present in the extratco. The study revealed that the extract presented antimicrobial effect against both strains and at 2% it showed high antimicrobial action compared to chlorhexidine at the same concentration, with average inhibition halos of 26.3 mm and 19 mm for each microorganism analyzed, (p < 0.05). Likewise, the hydrogel prepared with 2% extract significantly eliminated the preformed Streptococcus biofilm, at 24 hours of exposure, due to the presence of a variety of chemical groups, such as sterols, triterpenes, flavonoids, flavanones, flavanonol s, lactones. sesquiterpenic, tannins and coumarins. The oregano extract presented high antimicrobial action for both species, with a greater effect towards Streptococcus mutans and an interesting antibiofilm action; These results show the importance of exploring treatment alternatives of plant origin, to be considered as interesting complementary aids in dental therapy.

We hypothesized that via exosomes, CLL cells turn endothelial cells into “CLL-supportive cells.” To test this hypothesis, we transfected vein-derived (HUVECs) and arterial-derived (HAOEC) endothelial cells with exosomes that we isolated from the peripheral blood of 45 treatment-naive patients. We found that endothelial cells take up CLL exosomes in a dose- and time-dependent manner. Since CLL cells are protected from apoptosis in an IL-6-rich environment, we wondered whether CLL exosomes turn endothelial cells into IL-6-producing cells. To test this, we exposed endothelial cells to CLL exosomes and found a 50% increase in IL-6 levels, suggesting that the endothelial cells exposed to CLL exosomes produced and secreted IL-6. Subsequently, we filtered out this growth medium and added CLL cells to this IL-6-enriched medium. After 15 minutes, STAT3 became phosphorylated, and there was a 40% decrease in the rate of apoptosis among these cells, indicating that IL-6 activated the STAT3-dependent anti-apoptotic pathway. Phosphor-proteomic analysis of endothelial cells that were loaded with CLL exosomes revealed 23 phosphor-proteins that were upregulated. Annotation analysis unraveled the central role of phosphor-β-catenin. To test whether β-catenin enhances the generation of IL-6 in these cells, we transfected HUVECs with a β-catenin-containing plasmid. We found, by ELISA, a 30% increase in the levels of IL-6 in the culture medium, and, by ChIP, the increased binding of 3 transcription factors (NF-κB, LEF/TCF, and C/EBP) to the IL-6 promoter. Taken together, we found that CLL cells communicate with endothelial cells through the exosomes that they release. Once these exosomes are taken up by endothelia, they turn them into IL-6-producing cells.

Dual bromodomain BET inhibitors that bind with similar affinities to the first and second bromodomains across BRD2, BRD3, BRD4, and BRDT have displayed modest activity as monotherapy in clinical trials. Thrombocytopenia, closely followed by symptoms characteristic of gastrointestinal toxicity, have presented as dose-limiting adverse events that may have prevented escalation to higher dose levels required for more robust efficacy. ABBV-744 is a highly selective inhibitor for the second bromodomain of the four BET family proteins. In contrast to the broad antiproliferative activities observed with dual bromodomain BET inhibitors, ABBV-744 displayed significant antiproliferative activities largely although not exclusively in cancer cell lines derived from acute myeloid leukemia and androgen receptor positive prostate cancer. Studies in acute myeloid leukemia xenograft models demonstrated antitumor efficacy for ABBV-744 that was comparable with the pan-BET inhibitor ABBV-075 but with an improved therapeutic index. Enhanced antitumor efficacy was also observed with the combination of ABBV-744 and the BCL-2 inhibitor, venetoclax compared with monotherapies of either agent alone. These results collectively support the clinical evaluation of ABBV-744 in AML (Clinical Trials.gov identifier: NCT03360006).

International audience; Over the past decades, membranotropic peptides such as positively charged cell-penetrating peptides (CPPs) or amphipathic antimicrobial peptides (AMPs) have received increasing interest in order to improve therapeutic agent cellular uptake.As far as we are concerned, we were interested in studying HCV fusion peptides as putative anchors. Two peptides, HCV6 and HCV7, were identified and conjugated to a fluorescent tag NBD and tested for their interaction with liposomes as model membranes. DSC and spectrofluorescence analyses demonstrate HCV7 propensity to insert or internalize in vesicles containing anionic lipids DMPG whereas no activity was observed with zwitterionic DMPC. This behavior could be explained by the peptide sequence containing a cationic arginine residue. On the contrary, HCV6 did not exhibit any membranotropic activity but was the only sequence able to induce liposomes' fusion or aggregation monitored by spectrofluorescence and DLS. This two peptides mild activity was related to their inefficient structuration in contact with membrane mimetics, which was demonstrated by CD and NMR experiments.Altogether, our data allowed us to identify two promising membrane-active peptides from E1 and E2 HCV viral proteins, one fusogenic (HCV6) and the other membranotropic (HCV7). The latter was also confirmed by fluorescence microscopy with CHO cells, indicating that HCV7 could cross the plasma membrane via an endocytosis process. Therefore, this study provides new evidences supporting the identification of HCV6 as the HCV fusion peptide as well as insights on a novel membranotropic peptide from the HCV-E2 viral protein.

Although the dynamics of telomeres during the life expectancy of normal cells have been extensively studied, there are still some unresolved issues regarding this research field. For example, the conditions required for telomere shortening leading to malignant transformations are not fully understood. In this work, we mass analyzed DNA of normal and cancer cells for comparing telomere isotopic compositions of white blood cells and cancer cells. We have found that the 1327 Da and 1672 Da characteristic telomere mass to charges cause differential mass distributions of about 1 Da for determining isotopic variations among normal cells relative to cancer cells. These isotopic differences are consistent with a prior theory that replacing primordial, common isotopes of 1H, 12C, 14N, 16O, 24Mg, 31P and/or 32S by nonprimordial, uncommon isotopes of 2D, 13C, 15N, 17O, 25Mg and/or 33S leads to altered enzymatic dynamics for modulating DNA and telomere codons towards transforming normal cells to cancer cells. The prior theory and current data are consistent also with a recently observed non-uniform methylation in DNA of cancer cells relative to more uniform methylation in DNA of normal cells. We observe further evidence of nonprimordial isotopic accelerations of acetylations, methylations, hydroxylations and aminations of nucleosides with alterations of phosphorylations of nucleotides for possibly explaining the induced mutations of DNA, RNA and proteins leading to cancer and more general alterations of DNA associated with aging. The different mass spectra of normal and cancer DNA may be reasoned by different functionalizations and isotopic enrichments as causing different motionally induced atomic and nucleotide orders by different nuclear magnetic moments (NMMs); many motionally induced oligonucleotides causing nanoscale disorder and chaos; and the many such motionally induced nanoscale chaoses of different genes causing order in macroscopic DNA for organelles organizations.

International audience; Epithelial ovarian cancers are insidious pathologies that give a poor prognosis due to their late discovery and the increasing emergence of chemoresistance. Development of small pharmacological anticancer molecules remains a major challenge. Ribavirin, usually used in the treatment of hepatitis C virus infections and more recently few cancers, has been a suggestion. However, Ribavirin has many side-effects, suggesting that the synthesis of analogs might be more appropriate. We have investigated the effect of a Ribavirin analog, SRO-91, on cancer cell behavioral characteristics considered as some of the hallmarks of cancer. Two human ovarian adenocarcinoma cell lines (SKOV3 and IGROV1) and normal cells (mesothelial and fibroblasts) have been used to compare the effects of SRO-91 with those of Ribavirin on cell behavior underlying tumor cell dissemination. SRO-91, like Ribavirin, inhibits proliferation, migration, clonogenicity and spheroids formation of cancer cells. Unlike Ribavirin, SRO-91 is preferentially toxic to cancer compared with normal cells. An in vitro physiologically relevant model showed that SRO-91, like Ribavirin or cisplatin, inhibits cancer cell implantation onto peritoneal mesothelium. In conclusion, SRO-91 analog effects on tumor dissemination and its safety regarding non-cancerous (normal) cells are encouraging findings a promising drug for the treatment of ovarian cancer.

This paper describes a simple, straightforward, and rapid method for producing microspheres from molten polymers by merely printing them in an inkjet-like manner onto a superoleophobic surface (microsphere printing, hence SPHRINT). Similar to 3D printing, a polymer melt is deposited onto a surface; however, in contrast to 2D or 3D printing, the surface is not wetted (i.e. exhibiting high contact angles with liquids, above 150°, due to its low surface energy), resulting in the formation of discrete spherical microspheres. In this study, microspheres were printed using polycaprolactone and poly(lactic-co-glycolic acid) loaded with a model active pharmaceutical ingredient-ibuprofen (IBU). The formation of microspheres was captured by high-speed imaging and was found to involve several physical phenomena characterized by non-dimensional numbers, including the thinning and breakup of highly viscous, weakly elastic filaments, which are first to be described in pure polymer melts. The resulting IBU-loaded microspheres had higher sphericity, reproducible sizes and shapes, and superior drug encapsulation efficiencies with a distinctly high process yield (>95%) as compared to the conservative solvent-based methods used presently. Furthermore, the microspheres showed sustained release profiles.

CLL is characterized by gradual accumulation of mature appearing long-lived lymphocytes that travel in blood and reside in lymph nodes, spleen and bone marrow. In these sites, pro inflammatory humoral factors support the survival and proliferation of the neoplastic cells. Previous studies showed that levels of the proinflammatory cytokine IL-6 are at least 10 folds higher in patients with CLL compared with healthy individuals. Yet, which cells produce and secrete IL-6 and what triggers this cellular activity in CLL is unknown. Secreted by all types of cells, exosomes are nano-scaled particles that travel in blood and carry a cargo that at least partially reflects the molecular makeup of its cell of origin. Exosomes, including those originating from neoplastic cells, function as stable intercellular transport vehicles that deliver their cargo to cells that engulf them. For example, CLL-derived exosomes are taken up by mesenchymal stromal cells, transforming them to cancer associated fibroblasts. Given the appropriate stimulation, endothelial cells produce IL-6 which provides CLL cells with a survival advantage. Therefore, we hypothesized that CLL-exosomes turn endothelial cells into "IL-6-secreting cells". To test this hypothesis, we transfected vein-derived (HUVECs) and arterial-derived (HAOEC) endothelial cells with exosomes that we isolated from the peripheral blood of 45 treatment naïve patients. We found that endothelial cells take-up CLL-exosomes in a dose- and time- dependent manner. Since CLL cells are protected from apoptosis in IL-6 rich environment, we wondered whether CLL-exosomes turn endothelial cells into IL-6-producing cells. To test this, we exposed endothelial cells to CLL-exosomes and found 50% increase in IL-6 levels, suggesting that the endothelial-exposed cells produced and secreted IL-6. Subsequently, we filtered out this growth medium and added CLL cells to this IL-6 enriched medium. After 15 minutes, STAT3 became phosphorylated and there was 40% decrease in the rate of apoptosis, indicating that IL-6 activated STAT3-dependent anti-apoptotic pathway. Phosphor-proteomics analysis of endothelial cells that were loaded with CLL-exosomes revealed 23 phosphor-proteins that were upregulated. Network analysis unraveled the central role of phosphor-b-catenin. To test whether b-catenin induces IL-6 in these cells, we transfected HUVECs with a b-catenin containing plasmid. We found by ELISA 30% increase in the levels of IL-6 in the culture medium and by chromatin immunoprecipitation assay an increased binding of 3 transcription factors (NFkB, LEF/TCF, and CEBP) to the IL-6 promoter. Taken together, we found that CLL cells communicate with endothelial cells through exosomes that they release. Once these exosomes are taken up by endothelial, they turn them into IL-6 producing cells, which in turn contributes to CLL cells' survival. Disclosures No relevant conflicts of interest to declare.

Background and Objectives: Nutritional deficiencies are one of the main triggers for the development of gastrointestinal diseases, such as ulcerative colitis (UC). Therefore, the objective of the present work consisted of determining the nutrients present in the bone broth (BB) and evaluating their anti-inflammatory properties in a murine model of UC, induced by intrarectal administration of 2, 4, 6-trinitrobenzene sulfonic acid (TNBS), and acetic acid (AcOH). The BB was prepared from the femur of bovine cattle and cooked in distilled water for 8 h at 100 ± 2 °C. Materials and Methods: The BB was administered ad libitum to BALB/c mice for 10 days before the induction of UC. Colon samples were collected for histological analysis and determination of cytokine expression levels by qPCR. Results: It was found that amino acids (AA) are the main nutritional contribution of BB, 54.56% of these correspond to essential AA. The prophylactic administration of BB in the murine model of UC reduced histological damage, decreased the expression of IL-1β (61.12%), IL-6 (94.70%), and TNF-α (68.88%), and increased the expression of INF-γ (177.06%), IL-4 (541.36%), and IL-10 (531.97%). Conclusions: This study shows that BB has anti-inflammatory properties, and its consumption can decrease the symptoms of UC.

The dynamic and reversible acetylation of proteins, catalysed by histone acetyltransferases (HATs) and histone deacetylases (HDACs), is a major epigenetic regulatory mechanism of gene transcription and is associated with multiple diseases. Histone deacetylase inhibitors are currently approved to treat certain cancers, but progress on the development of drug-like histone actyltransferase inhibitors has lagged behind. The histone acetyltransferase paralogues p300 and CREB-binding protein (CBP) are key transcriptional co-activators that are essential for a multitude of cellular processes, and have also been implicated in human pathological conditions (including cancer). Current inhibitors of the p300 and CBP histone acetyltransferase domains, including natural products, bi-substrate analogues and the widely used small molecule C646, lack potency or selectivity. Here, we describe A-485, a potent, selective and drug-like catalytic inhibitor of p300 and CBP. We present a high resolution (1.95 Å) co-crystal structure of a small molecule bound to the catalytic active site of p300 and demonstrate that A-485 competes with acetyl coenzyme A (acetyl-CoA). A-485 selectively inhibited proliferation in lineage-specific tumour types, including several haematological malignancies and androgen receptor-positive prostate cancer. A-485 inhibited the androgen receptor transcriptional program in both androgen-sensitive and castration-resistant prostate cancer and inhibited tumour growth in a castration-resistant xenograft model. These results demonstrate the feasibility of using small molecule inhibitors to selectively target the catalytic activity of histone acetyltransferases, which may provide effective treatments for transcriptional activator-driven malignancies and diseases.

ABBV-075 is a potent and selective BET family bromodomain inhibitor that recently entered phase I clinical trials. Comprehensive preclinical characterization of ABBV-075 demonstrated broad activity across cell lines and tumor models, representing a variety of hematologic malignancies and solid tumor indications. In most cancer cell lines derived from solid tumors, ABBV-075 triggers prominent G1 cell-cycle arrest without extensive apoptosis. In this study, we show that ABBV-075 efficiently triggers apoptosis in acute myeloid leukemia (AML), non-Hodgkin lymphoma, and multiple myeloma cells. Apoptosis induced by ABBV-075 was mediated in part by modulation of the intrinsic apoptotic pathway, exhibiting synergy with the BCL-2 inhibitor venetoclax in preclinical models of AML. In germinal center diffuse large B-cell lymphoma, BCL-2 levels or venetoclax sensitivity predicted the apoptotic response to ABBV-075 treatment. In vivo combination studies uncovered surprising benefits of low doses of ABBV-075 coupled with bortezomib and azacitidine treatment, despite the lack of in vitro synergy between ABBV-075 and these agents. The in vitro/in vivo activities of ABBV-075 described here may serve as a useful reference to guide the development of ABBV-075 and other BET family inhibitors for cancer therapy. Cancer Res; 77(11); 2976–89. ©2017 AACR.

Because of the emergence of new viruses, the need for new antiviral broad-spectrum compounds remains important. In this context, herein the synthesis of C-nucleosides, structurally close to ribavirin, a nucleoside presenting various biological activities and used until now particularly for its broad-spectrum antiviral properties, is reported. The compounds were designed in order to increase their stability and the number of hydrogen bond donor or acceptor in comparison to ribavirin, and to investigate the role of the carboxamide group on the biological activity. The efficient synthesis of 11 C-nucleosides is based on an indium-mediated alkynylglycosylation as the key step, followed by the construction of the triazole heterocycle. Amidation was performed with primary and secondary amines in yields up to 85%. An analogue nucleoside with a triazole without carboxamide group was also prepared in order to compare its activity. Finally, the carboxamide group was moved to the N-1 triazole position to mimic ribavirin.

In IgHV-unmutated CLL (U-CLL), the proliferation rate is higher compared with IGHV-mutated CLL (M-CLL). Yet, why CLL cells differ in their mutational rates is unknown. Accumulating mutations in the IGHV gene lead to subsequent accumulation of DNA breaks. In rapidly dividing cells, DNA breaks are repaired by the efficient high-fidelity homology-directed DNA (HR) repair apparatus, whereas in slowly dividing cells, they are repaired by the inefficient low-fidelity nonhomologous end-joining (NHEJ) repair mechanism. Since HR is a high-fidelity DNA repair process, we expect that in cells that preferentially utilize HR, the mutations will be substituted by non-mutated (“germline”) nucleotides. We therefore postulate that proliferating U-CLLs utilize HR, which corrects the mutations accumulated during SHM. To test whether U-CLL cells preferentially use HR, we incubated CLL cells from two patients with U-CLL and two patients with M-CLL with antibodies that bind RPA and RAD51, key components in the HR repair pathway. By fluorescence microscopy, we detected these proteins in U-CLL, but not M-CLL, cells. To confirm these findings, we quantified the transcript levels of key players in the DNA repair process by qRT-PCR in four patients with M-CLL and two patients with U-CLL. Consistent with the immunohistochemistry data, the levels of FANCD2, RAD51, BRCA1 and BRCA2, which represent HR, were significantly higher in U-CLL compared with M-CLL. Levels of XRCC5, XRCC6, and PRKDC, which represent the NHEJ machinery, were low across all samples tested. Since HR is only operative in dividing cells, we hypothesized that inducing proliferation will turn on the HR enzymatic machinery. To induce proliferation, we incubated CLL cells from eight patients with CpG oligonucleotide. Using this agonist, proliferation increased by 110% (range: 42 to 250) in M-CLL cells but only by 25% (range 14 to 66) in U-CLL cells. The increase in proliferation rates correlated with upregulation of the HR machinery at the RNA and protein levels as measured by qRT-PCR and immunohistochemistry, respectively. Summary Here, we show that proliferating CLL cells utilize more often HR recombination repair machinery. Whether HR activity contributes to reduced mutational rates in the IGHV gene remains to be determined.

Aims: To use an agroindustrial waste (orange peels) as a source of polyphenols as a reducing medium for obtaining silver nanoparticles by greener method. Background: Several techniques have been employed for AgNPs synthesis, nevertheless, most of them involve the use of toxic chemicals in the process. The use of fungi, bacteria, and plant extracts as subtracts for green synthesis is an ecofriendly alternative, although hypothetic, route for AgNPs large scale synthesis. In the case of plant extracts, it is believed that polyphenols are the biomolecules responsible for the reduction and stabilization of the Ag+ ions into AgNPs, being a sustainable and ecological option; polyphenols could be obtained from plant waste and agroindustrial subproducts. Objective: To develop an efficient, greener, and low-cost method of AgNPs production using natural products. Methods: The basic principle of silver nanoparticles synthesis is the interaction in a mixture of silver nitrate (source of Ag+ ions) and the orange peel extract (reducing and stabilizing agent) under certain conditions. Five treatments were carried out, evaluating several parameters during AgNPs synthesis such as pH, orange peel extract-silver nitrate ratio, time and conditions of incubation, irradiation of UV light, irradiation of microwave, and temperature. Result: The synthesis of silver nanoparticles from an agroindustrial waste as the orange peel was successfully carried out and checked by visual evaluation, UV-Vis spectroscopy, and EDS analysis. The particle size was estimated between 42.82 nm to 151.75 nm, having a spherical and ovoid morphology. Discussion: Through the analysis of several synthesis conditions, it has become possible to establish a suitable treatment to increase antibacterial yield and evaluate morphology and size traits in order to acquire the best conditions for a future industrial scale synthesis. Conclusion: The orange peel aqueous extract resulted as a great source of polyphenols, allowing the successful synthesis of silver nanoparticles in mild conditions. Thus, obtained AgNPs revealed an increased antibacterial effect and potential against Gram-positive bacteria such as Staphyloccocus aureus.

Although the dynamics of telomeres during the life expectancy of normal cells have been extensively studied, there are still some unresolved issues regarding this research field. For example, the conditions required for telomere shortening leading to malignant transformation are not fully understood. In this work, we mass analyzed DNA of normal and cancer cells for comparing telomere isotopic compositions of white blood cells and cancer cells. We have found that the 1327 Da and 1672 Da characteristic telomere mass to charge cause differential mass distributions of about 1 Da for determining isotopic variations among normal cells relative to cancer cells. These isotopic differences are consistent with a prior theory that replacing primordial isotopes of 1H, 12C, 14N, 16O, 24Mg, 31P and/or 32S by nonprimordial, uncommon isotopes of 2D, 13C, 15N, 17O, 25Mg and/or 33S leads to altered enzymatic dynamics for modulating DNA and telomere codons towards transforming normal cells to cancer cells. The prior theory and current data are consistent also with a recently observed non-uniform methylation in DNA of cancer cells relative to more uniform methylation in DNA of normal cells.

Directed palladium-catalyzed C–H functionalisation of C2-amido glycals onto the anomeric position is described as a novel route to C-aryl/alkenylglycosides. An aminoquinoline-type directing group was used to successfully introduce diverse (hetero)aryl and alkenyl groups at position 1 of the sugar (20 examples). Its application to the synthesis of a dapagliflozin analogue is presented.

Background Common practice is to withhold vaccination with live-attenuated vaccines in patients with rheumatic diseases on high-dose DMARDs, glucocorticosteroids or biological agents, due to limited safety data, and the (theoretical) risk of introducing an infectious disease to the patient. Evidence for this approach is low. We collected data from pediatric rheumatologists who vaccinate these patients, to obtain additional safety data, which might update and revise this approach. Objectives To collect retrospective data in patients with JIA and other diseases who received live booster MMR or MMRV while on DMARDs, glucocorticosteroids or biological agents. Methods Data from 13 pediatric rheumatology centers in 10 countries were collected. Results 234 patients were reported; mean age 5± 2.7, 70% girls. 206 had JIA;. 46% oligoarticular, 36% polyarticular, 8% systemic, 5% SPA types, 5% JIA and uveitis. 48% of JIA patients were in remission on medication. Disease activity was low in 38%, high in 2%, moderate in 7%; 11 patients had juvenile dermatomyositis, 3 systemic and 2 localized scleroderma, 4 isolated idiopathic uveitis, 1 CINCA syndrome, 1 MKD, and 1 FMF. 110 patients had MMR/V booster while on MTX; 3 reported mild side-effects of local skin reaction and pain, none had disease flare. 76 had booster while on MTX+ anti-TNF; 7 reported mild and transient adverse events of local skin reaction, fever and URTI. 39 had booster while on anti-TNF alone; 1 reported fever. 3 had booster while on tocilizumab, 7 on anakinra, and 5 on canakinumab. There was no relation between disease activity, type or duration, sex, age and outcome of vaccinations. No vaccine infection related to measles, rubella, mumps and varicella were reported. Conclusion This large, retrospective data collection demonstrates that live-attenuated booster vaccine is probably safe in children with rheumatic diseases, on immunosuppressive therapies. This strengthens the new PRES recommendation: “Vaccination of live-attenuated vaccines in patients on high-dose DMARD, high-dose glucocorticosteroids or biological agents can be considered on a case-by-case basis, weighing the risk of infections against the hypothetical risk of inducing infection through vaccination.” These data provide the basis for a large, prospective data collection study that is planned by the PReS vaccination study group. It will increase the current level of evidence for the safety of vaccinations in our pediatric rheumatology population. Disclosure of Interests Yosef Uziel: None declared, Veronica Moshe Bergonzo: None declared, Beata Onozo: None declared, Andrea Kulcsar: None declared, Diana Trobert-Sipos : None declared, Jonathan Akikusa: None declared, Gecilmara Salviato Pileggi : None declared, Despoina Maritsi: None declared, Ozgur KASAPCOPUR: None declared, Roubini Smerla: None declared, Donato Rigante: None declared, Erato Atasali: None declared, Mariana Rodrigues: None declared, Balahan Makay Speakers bureau: Enzyvant, Novartis, Roche, Abbvie, Nico Wulffraat: None declared, Natasa Toplak: None declared

The importance of telomerase, the enzyme that maintains telomere length, has been reported in many malignancies in general and in multiple myeloma (MM) in particular. Proteasome inhibitors are clinically used to combat effectively MM. Since the mechanism of action of proteasome inhibitors has not been fully described we sought to clarify its potential effect on telomerase activity (TA) in MM cells. Previously we showed that the first generation proteasome inhibitor bortezomib (Brt) inhibits TA in MM cells by both transcriptional and post-translational mechanisms and has a potential clinical significance. In the current study we focused around the anti- telomerase activity of the new generation of proteasome inhibitors, epoxomicin (EP) and MG-132 in order to clarify whether telomerase inhibition represents a class effect. We have exposed MM cell lines, ARP-1, CAG, RPMI 8226 and U266 to EP or MG and the following parameters were assessed: viability, TA, hTERT expression, the binding of hTERT (human telomerase reverse transcriptase) transcription factors and post-translational modifications. Epoxomicin and MG-132 differentially downregulated the proliferation and TA in all MM cell lines. The downregulation of TA and the expression of hTERT were faster in CAG than in ARP-1 cells. Epoxomicin was more potent than MG-132 and therefore further mechanistic studies were performed using this compound. The inhibition of TA was mainly transcriptionally regulated. The binding of three positive regulator transcription factors: SP1, c-Myc and NF-&kappa, B to the hTERT promoter was decreased by EP in CAG cells as well as their total cellular expression. In ARP-1 cells the SP1 and c-MYC binding and protein levels were similarly affected by EP while NF-&kappa, B was not affected. Interestingly, the transcription factor WT-1 (Wilms&rsquo, tumor-1) exhibited an increased binding to the hTERT promoter while its total cellular amount remained unchanged. Our results combined with our previous study of bortezomib define telomerase as a general target for proteasome inhibitors. The inhibitory effect of TA is exerted by several regulatory levels, transcriptional and post translational. SP1, C-Myc and NF-&kappa, B were involved in mediating these effects. A novel finding of this study is the role of WT-1 in the regulation of telomerase which appears as a negative regulator of hTERT expression. The results of this study may contribute to future development of telomerase inhibition as a therapeutic modality in MM.

The design of molecules that mimic biologically relevant glycans is a significant goal for understanding important biological processes and may lead to new therapeutic and diagnostic agents. In this study we focused our attention on the trisaccharide human natural killer cell-1 (HNK-1), considered the antigenic determinant of myelin-associated glycoprotein and the target of clinically relevant auto-antibodies in autoimmune neurological disorders such as IgM monoclonal gammopathy and demyelinating polyneuropathy. We describe a structure-activity relationship study based on surface plasmon resonance binding affinities aimed at the optimization of a peptide that mimics the HNK-1 minimal epitope. We developed a cyclic heptapeptide that shows an affinity of 1.09 x 10(-7) m for a commercial anti-HNK1 mouse monoclonal antibody. Detailed conformational analysis gave possible explanations for the good affinity displayed by this novel analogue, which was subsequently used as an immunological probe. However, preliminary screening indicates that patients' sera do not specifically recognize this peptide, showing that murine monoclonal antibodies cannot be used as a guide to select immunological probes for the detection of clinically relevant human auto-antibodies.

BACKGROUND: Primary glioblastoma multiforme (GBM) have a high proportion (~80%) of TERT-expressing tumors, indicating that this is the primary mechanism of telomerase (hTERT) activation. Exosomes are nano-sized secreted vesicles containing nucleic acids and proteins, which reflect the content of the mother cells. We have previously shown that hTERT transcripts in serum exosomes may serve as a “pan-cancer” diagnostic method, reflecting the load of hTERT in systemic cancer cells. The goal of the current study was to evaluate whether exosomal hTERT may serve as a circulating biomarker for GBM. METHODS: hTERT mRNA levels were determined in serum derived exosomes obtained from 20 GBM patients and 45 healthy controls. The level of exosomal hTERT mRNA was measured prior to surgery in all GBM patients. In 10 patients additional longitudinal evaluation was performed in blood samples obtained prior to and after concurrent radio/chemotherapy and again during adjuvant temozolomide treatment. RESULTS: Circulating hTERT transcripts were absent in controls and were variably detected in 40% of GBM patients with significantly elevated mean level at diagnosis (p=0.049). These transcripts were gradually downregulated on longitudinal evaluations and during treatment period with significant reduction observed after 3 months of adjuvant treatment when compared to initial sampling (p=0.026). In 10 patients an analysis of hTERT promoter mutation was performed and in 8/10 one of the two common mutations (C228T and C250T) was detected on tumor samples. CONCLUSIONS: hTERT mRNA levels may reflect the tumor burden and the clinical status of patients with GBM. In systemic tumors exosomal hTERT mRNA expression is detected in about 60% of cases while in GBM the percentage is probably lower. In patients with detectable levels, this assay may serve as a serum biomarker. These results warrant further confirmation and an update on an extended cohort of patients will be presented at the meeting.

Encephalitozoon cuniculi is an obligate macrophage parasite of vertebrates that commonly infects rodents, monkeys, dogs, birds, and humans. In the present study, we aimed to assess the phagocytosis and intracellular survival of E. cuniculi spores using untreated and lipopolysaccharide (LPS)-activated J774A.1 murine macrophages and assess the macrophage viability. The experimental groups comprised untreated spores, spores killed by heat treatment at 90 °C, and spores killed by treatment with 10% formalin. LPS-activated macrophages significantly increased the phagocytosis of spores and reduced their intracellular growth after 24 and 48 h (P

Due to the cadmium toxicity, a new research line has surged up on the past years, solar cells cadmium free, and so, the request of materials with similar properties than CdS in order to replace it as buffer and window layer on solar cells. In this way, In 2 S 3 has shown adequate physical properties to be applied on solar cells. In this work, In 2 S 3 thin films were deposited by chemical bath deposition, the samples were thermally treated under different atmospheres. In 2 S 3 thermally treated on air presented the best conditions to be applied on CdTe solar cells.

The synthesis of four C -pyrimidyl nucleosides is described by condensation of small nitrogen molecules (amidines and ureas) onto alkynyl riboside derivatives. These last compounds were obtained by indium mediated stereoselective alkynylation of suitably protected ribose derivatives and the condensation reaction conditions were studied in order to favor the N -attack of the nitrogen molecules leading to the pyrimidine ring formation.

Progress in understanding tumor stromal biology has been constrained in part because cancer-associated fibroblasts (CAF) are a heterogeneous population with limited cell-type–specific protein markers. Using RNA expression profiling, we identified the membrane protein leucine-rich repeat containing 15 (LRRC15) as highly expressed in multiple solid tumor indications with limited normal tissue expression. LRRC15 was expressed on stromal fibroblasts in many solid tumors (e.g., breast, head and neck, lung, pancreatic) as well as directly on a subset of cancer cells of mesenchymal origin (e.g., sarcoma, melanoma, glioblastoma). LRRC15 expression was induced by TGFβ on activated fibroblasts (αSMA+) and on mesenchymal stem cells. These collective findings suggested LRRC15 as a novel CAF and mesenchymal marker with utility as a therapeutic target for the treatment of cancers with LRRC15-positive stromal desmoplasia or cancers of mesenchymal origin. ABBV-085 is a monomethyl auristatin E (MMAE)-containing antibody–drug conjugate (ADC) directed against LRRC15, and it demonstrated robust preclinical efficacy against LRRC15 stromal-positive/cancer-negative, and LRRC15 cancer-positive models as a monotherapy, or in combination with standard-of-care therapies. ABBV-085′s unique mechanism of action relied upon the cell-permeable properties of MMAE to preferentially kill cancer cells over LRRC15-positive CAF while also increasing immune infiltrate (e.g., F4/80+ macrophages) in the tumor microenvironment. In summary, these findings validate LRRC15 as a novel therapeutic target in multiple solid tumor indications and support the ongoing clinical development of the LRRC15-targeted ADC ABBV-085. Significance: These findings identify LRRC15 as a new marker of cancer-associated fibroblasts and cancers of mesenchymal origin and provide preclinical evidence for the efficacy of an antibody-drug conjugate targeting the tumor stroma. Cancer Res; 78(14); 4059–72. ©2018 AACR.

The functional unit within the ovary is the ovarian follicle, which is also a morphological unit composed of three basic cell types: the oocyte, granulosa, and theca cells. Similar to human ovarian follicles, mouse follicles can be isolated from their ovarian environment and cultured in vitro to study folliculogenesis, or follicle development for days or weeks. Over the course of the last decade, follicle culture in a three-dimensional (3D) environment exponentially improved the outcomes of in vitro folliculogenesis. Follicle culture in 3D environments preserves follicle architecture and promotes the cross talk between cells in the follicle. Hydrogels, such as polyethylene glycol (PEG), have been used for various physiological systems for regenerative purposes because they provide a 3D environment similar to soft tissues, allow diffusion of nutrients, and can be readily modified to present biological signals, including cell adhesion ligands and proteolytic degradation facilitated by enzymes secreted by the encapsulated cells. This chapter outlines the application of PEG hydrogels to the follicle culture, including the procedures to isolate, encapsulate, and culture mouse ovarian follicles. The tunable properties of PEG hydrogels support co-encapsulation of ovarian follicles with somatic cells, which further promote follicle survival and growth in vitro through paracrine and juxtacrine interactions.

Th-T-Al system, where T- 3d transition metals, was studied at ThT 2 Al 20 stoichiometry to establish the influence of T on the structural stability of ternary aluminide formed. Different alloys were prepared, varying T in the row from Ti to Fe. Using electron microscopy and X-ray diffraction methods it was found that ThT 2 Al 20 phase adopts CeCr 2 Al 20 structure type when T = Ti, V, and Cr. Starting from Mn, the symmetry of the stable Al-rich phase, which forms in the alloys with the same composition, decreases from cubic to orthorhombic. The results of Density Functional Theory (DFT) calculations coincide with experiments. Concepts of the Theory of Coordination Compounds and Jahn–Teller effect were used to explain the observed abrupt change of the symmetry. These considerations were supported by DFT calculations.

The blanching of easel paintings can affect the varnish layer and also the paint layer with a blurring effect. The understanding of the physicochemical and optical phenomena involved in the whitening process remains an important challenge for the painting conservation. A set of ca. 50 microsamples from French, Flemish, and Italian blanched oil paintings, from sixteenth to nineteenth century, have been collected for in deep investigations. In parallel, the reproduction of the alteration was achieved by preparing some paint layers according to historical treatises and altering them in a climatic chamber in a humid environment or directly by immersing in ultrapure water. The observation of raw samples with a field-emission gun scanning electron microscope revealed for the first time that the altered layers have an unexpected highly porous structure with a pore size ranging from ca. 40 nm to 2 μm. The formation mechanism of these pores should mostly be physical as the supplementary analyses (Fourier transform infrared spectroscopy, gas chromatography–mass spectrometry) do not reveal any noticeable molecular modification. Considering the tiny size of the pores, the alteration can be explained by the Rayleigh or Mie light scattering.

The expression of tumor necrosis factor (TNF) -α, interleukin (IL) -4 and IL-10, as well as apoptosis and nitric oxide (NO) levels were measured in the brain and kidneys of immunocompetent and immunosuppressed New Zealand White rabbits infected with Encephalitozoon cuniculi. All of the animals had clinical signs histopathological lesions compatible with encephalitozoonosis and were E. cuniculi-positive by using a carbon immunoassay test. Encephalitozoon cuniculi infection promoted the expression of TNF-α and NO production in the kidneys of infected rabbits, and a synergic effect was observed in animal treated with dexamethasone. The IL-4 expression was similar in the brain and kidneys of infected rabbits, regardless of their immunologic status. The IL-10 mRNA expression in the brain of infected immunosuppressed rabbits was elevated when compared with positive controls. Apoptosis of granuloma mononuclear-like cells was detected in immunocompetent E. cuniculi-infected rabbits, but it was more evident in infected-immunosuppressed animals. Nitric oxide levels were elevated both in immunocompetent and immunosuppressed infected animals, but it was more apparent in the kidneys. These data suggest that modulation of the immune response by E. cuniculi could contribute to the survival of the parasite within phagocytic cells in granulomas via an as yet undetermined mechanism.

An efficient synthetic pathway leading to two carbonated analogues of ribavirin is described. The key-steps in the synthesis of these ribosyltriazoles bearing a quaternary carbon atom in the 2’-position are an indium-mediated alkynylation and a 1,3-dipolar cyclization.

International audience; An efficient synthetic pathway leading to two carbonated analogues of ribavirin is described. The key-steps in the synthesis of these ribosyltriazoles bearing a quaternary carbon atom in the 2'-position are an indium-mediated alkynylation and a 1,3-dipolar cyclization.

International audience; The azide-alkyne cycloaddition reaction was investigated under catalytic conditions involving a copper (II) loaded silica based mesoporous material. Cu(II) SBA-15 demonstrated a high catalytic effect in 1,4-triazoles synthesis in organic. No additives such as a base or a reductant are required. Quantitative yields were obtained and a mere filtration of the mesoporous material which retains copper (II) allows the recovery of the catalyst. In addition, up to 5 times recycling of the catalyst was achieved without loss of the activity affording 1,4-triazoles in a yield up to 98%.

Despite advances in understanding of the biology of acute myeloid leukemia (AML), cure remains elusive for the majority of patients. ABT-199 (Venetoclax) is a small-molecule BH3 mimetic that selectively inhibits BCL-2 causing cell death. First generation BET inhibitor ABBV-075 and Venetoclax were recently shown to be synergistic in AML cell lines (Bui MH,Cancer Res 2017). ABBV-744 is a highly selective inhibitor for the BDII of BET family proteins, exhibiting greater than 300-fold more potent binding affinity to the BDII bromodomain of BRD4 relative to BDI (Warren Kati AACR 2018; Xiaoyu Lin AACR 2018). In this study, we evaluated the anti-leukemia efficacy of the concomitant BCL-2 blockade by venetoclax and of BDII inhibition with ABBV-744 in primary AML samples. Anti-leukemia activity of venetoclax and ABBV-744 was examined in 21 primary AML samples with diverse genomic alterations. The combination significantly enhanced cell death (57.0 ± 6.3%) compared to the single agent treatment (43.9 ± 5.7% in ABT-199 10 nM group, p In two AML primary samples tested, combination treatment of ABBV-744 and ABT-199 induced apoptosis with caspase-3 activation and PARP cleavage through regulation of the key proteins regulating survival and proliferation pathways (e.g. (BCL-2, BCL-XL, MCL-1, c-Myc)). To identify biomarkers of response to therapy, we performed the baseline transcriptome analysis of AML cells used for in vitro response assessment (n=25) by RNA-sequencing (RNA-seq), and correlated baseline gene expression levels with in vitro response to therapy. Based on response to venetoclax or combination, samples were divided into 3 groups: sensitive to venetoclax (n=10), samples with no response to single agent or combination ("low apoptosis", n=7) and samples resistant to venetoclax as a single agent but responsive to venetoclax/ ABBV-744 combination ("synergy", n=4). AML samples sensitive to venetoclax and venetoclax/ABBV-744 combination were characterized by high level of BCL2 and lower levels of MCL1 and BCL2L1 transcripts, consistent with known inability of venetoclax to inhibit MCL-1 and BCL2L1 (Fig.1C).The resistant samples additionally expressed higher levels of anti-apoptotic genes such as GADD45, BCL2L10, PMAIP1. AML cells that showed synergy between venetoclax/ABBV-744 expressed low levels of AR, IL1R1 genes and had high CCND1 expression. The gene expression analysis indicated that the genes differentially expressed in the synergy vs. low apoptosis samples overlap with the genes inhibited by dual BCL-2/BCL-XL inhibitor ABT-737. To test the efficacy of this regimen in vivo, we established a patient-derived xenograft (PDX) from an AML patient with FLT3-ITD, DNMT3A, EGFR, IDH1, NPM1, TET2 mutations in NSG mice. Upon engraftment, mice were randomized to receive vehicle; single agent venetoclax at 50 mg/kg; ABBV-744 at 9.4 mg/kg; or venetoclax plus ABBV-744 for 21 days. After 21 days of therapy, flow cytometry data demonstrated significantly reduced leukemia burden in venetoclax treated group (9.5% ± 1.7%) but not in ABBV-744 group (22.3% ± 5.8%) compared to controls (30.8% ± 3.9%), with lowest tumor burden in the combination group (5.0% ± 0.8%, p In summary, combinatorial blockade of BDII bromodomain and of BCL-2 anti-apoptotic pathway facilitates apoptotic cell death, suppresses proliferation in the majority of primary AML cells and produces anti-AML activity in AML PDX models in vivo at tolerable doses of both agents. This combination is currently undergoing testing in a Phase I clinical trial in AML (NCT03360006). Disclosures Kuruvilla: The University of Texas M.D.Anderson Cancer Center: Employment. Lin:AbbVie: Employment. Uziel:AbbVie: Employment, Other: stock or other options. Lu:AbbVie: Employment. Zhang:AbbVie: Employment. Huang:AbbVie: Employment. Zhang:The University of Texas M.D.Anderson Cancer Center: Employment. Shen:AbbVie: Employment. Konopleva:Astra Zeneca: Research Funding; Ablynx: Research Funding; Eli Lilly: Research Funding; Kisoji: Consultancy, Honoraria; Ascentage: Research Funding; Agios: Research Funding; Reata Pharmaceuticals: Equity Ownership, Patents & Royalties; Amgen: Consultancy, Honoraria; AbbVie: Consultancy, Honoraria, Research Funding; Cellectis: Research Funding; Genentech: Honoraria, Research Funding; Stemline Therapeutics: Consultancy, Honoraria, Research Funding; Calithera: Research Funding; F. Hoffman La-Roche: Consultancy, Honoraria, Research Funding; Forty-Seven: Consultancy, Honoraria.

Background: The BCL-2 family of anti-apoptotic proteins (e.g., BCL-2, BCL-XL, MCL-1) are important regulators of lymphocyte development and are therapeutic targets for hematologic malignancies, including multiple myeloma (MM). Venetoclax (Ven) is a highly selective, potent, oral BCL-2 inhibitor that induces apoptosis in MM cells and has shown synergistic activity with bortezomib (Velcade; V) and dexamethasone (Dex or d). Combination of the CD38 monoclonal antibody (mAb) daratumumab (D) with Ven is hypothesized to further increase anti-myeloma activity based upon dual mechanisms of pro-apoptotic effects on tumor cells as well as enhanced immune stimulation. Pre-clinically and in healthy human subjects, Ven treatment leads to enrichment of CD8+ T effector memory cells, while resulting in loss of CD4+ and CD8+ naïve T-cells, however the potential immunomodulatory effect of Ven in MM is unknown. Results presented herein describe the pharmacodynamic changes observed in immune cell subsets in relapsed/refractory (R/R) MM patients (pts) treated with VenDex, VenDd, and VenDVd. Methods: Peripheral blood samples were collected at day 1 of cycles 1-5 to characterize pharmacodynamic changes in B- and T-cell sub-populations by multicolor flow cytometry in Ven MM clinical trials M13-367 (NCT01794520) and M15-654 (NCT03314181). M13-367 is a phase 1/2 study of VenDex in t(11;14) R/R MM, and M15-654 is a phase 1/2 study of VenDd in t(11;14) R/R MM (Part 1) and VenDVd in R/R MM (Part 2). As of 12 June 2019, 15 out of 31 pts treated with VenDex in phase 2 of M13-367, 19 out of 24 pts treated in Part 1 (VenDd), and 19 out of 24 pts treated in Part 2 (VenDVd) of M15-654 had baseline and post-treatment specimens available for analysis. Results: Consistent with previous findings that B-cells are highly dependent upon BCL-2 for cell survival, each Ven-containing regimen (VenDex, VenDd, and VenDVd) resulted in rapid and sustained reduction (~90% decrease from baseline by end of cycle 1) in peripheral B-cell (CD19+/CD5-) counts (Figure). In subgroup analyses, naïve B-cells (CD27-/IgM+) were significantly reduced in pts treated with each regimen, however regulatory B-cells (CD27+/CD24+) remained largely unaffected. In addition, a decrease in plasmablasts (CD27+CD38+CD20-) were only observed in pts treated with D-containing regimens (VenDd and VenDVd), which is consistent with expression of CD38 in this subgroup. In contrast to B-cells, CD3+ T-cells were minimally reduced (~20-30% reduction from baseline through end of cycle 4) with VenDex and VenDd treatment. A more prominent reduction was observed in pts treated with VenDVd (~50-60% reduction from baseline through end of cycle 4). CD4+ T-cells were more sensitive than CD8+ T-cells to VenDex and VenDd, while both CD4+ and CD8+ T-cells were reduced in VenDVd treated pts (Figure). T-cell subgroup analyses were also performed, including naïve T-cells (CD45RA+/CD197+), effector (CD45RA+/CD197-), effector memory (CD45RA-/CD197-), and central memory (CD45RA-/CD197+). Overall, there were no significant changes in the composition of the T-cell pool (Tnaive, TCM, TEM, TEMRA) or in Th1 (CD4+/CD183+/CD196-), Th2 (CD4+/CD183-/CD196-), and Th17 (CD4+/CD183-/CD196+) cells in pts treated with VenDex, VenDd, or VenDVd. Finally, a decrease in Tregs (CD4+/CD25+/CD127low/CD194+) was observed in VenDd and VenDVd treated pts, but not in VenDex treated pts (Figure). While no reduction in overall Tregs was observed with VenDex, a decrease in the relative proportion of activated Tregs (HLA-DR+/CD45RO+) to naïve (HLA-DR-/CD45RO-) and memory (HLA-DR-/CD45RO+) Tregs was observed. Conclusions: Comprehensive immunophenotyping studies were conducted to characterize the potential effects of Ven in B- and T-cell subsets when used in combination with standard of care anti-myeloma agents of various mechanisms of action (i.e. glucocorticoid, CD38 mAb and/or proteasome inhibitor). While each Ven combination regimen resulted in significant reductions in circulating B-cells, T-cell effects were largely dependent upon the combination regimen. Minimal reduction in total T-cell counts were observed with VenDex and VenDd, which were primarily due to reduction in CD4+ T-cells. However, the extent of T-cell depletion (CD4+ and CD8+ T-cells) was greatest in pts treated with the bortezomib-containing regimen VenDVd. Correlative studies between immune profiles and pt outcomes are ongoing. Disclosures Vishwamitra: AbbVie: Employment, Other: stock or other options. Mantis:AbbVie Inc: Employment, Other: Stock/stock options. Haribhai:AbbVie: Employment, Other: stock or other options. Uziel:AbbVie: Employment, Other: stock or other options. Leverson:AbbVie Inc: Employment, Other: Stock or options. Pauff:AbbVie Inc: Employment, Other: may own stock or stock options. Bueno:AbbVie: Employment, Other: Stock/stock options. Maciag:AbbVie: Employment, Other: Stock/stock options. Ross:AbbVie: Employment, Other: Stock/stock options.