Representing the major cause of morbidity and mortality for chronic lymphocytic leukemia (CLL) patients, immunosuppression is a common feature of the disease. Effectors of the innate and the adaptive ...immune response show marked dysfunction and skewing towards the generation of a tolerant environment that favors disease expansion. Major deregulations are found in the T lymphocyte compartment, with inhibition of CD8
cytotoxic and CD4
activated effector T cells, replaced by exhausted and more tolerogenic subsets. Likewise, differentiation of monocytes towards a suppressive M2-like phenotype is induced at the expense of pro-inflammatory sub-populations. Thanks to their B-regulatory phenotype, leukemic cells play a central role in driving immunosuppression, progressively inhibiting immune responses. A number of signaling cascades triggered by soluble mediators and cell-cell contacts contribute to immunomodulation in CLL, fostered also by local environmental conditions, such as hypoxia and derived metabolic acidosis. Specifically, molecular pathways modulating T-cell activity in CLL, spanning from the best known cytotoxic T lymphocyte antigen-4 (CTLA-4) and programmed cell death 1 (PD-1) to the emerging T cell immunoreceptor with immunoglobulin and immunoreceptor tyrosine-based inhibition motif domains (TIGIT)/CD155 axes, are attracting increasing research interest and therapeutic relevance also in the CLL field. On the other hand, in the microenvironment, the B cell receptor (BCR), which is undoubtedly the master regulator of leukemic cell behavior, plays an important role in orchestrating immune responses, as well. Lastly, local conditions of hypoxia, typical of the lymphoid niche, have major effects both on CLL cells and on non-leukemic immune cells, partly mediated through adenosine signaling, for which novel specific inhibitors are currently under development. In summary, this review will provide an overview of the molecular and microenvironmental mechanisms that modify innate and adaptive immune responses of CLL patients, focusing attention on those that may have therapeutic implications.
Damage-associated molecular patterns (DAMPs) are molecules that can be actively or passively released by injured tissues and that activate the immune system. Here we show that nicotinate ...phosphoribosyltransferase (NAPRT), detected by antibody-mediated assays and mass spectrometry, is an extracellular ligand for Toll-like receptor 4 (TLR4) and a critical mediator of inflammation, acting as a DAMP. Exposure of human and mouse macrophages to NAPRT activates the inflammasome and NF-κB for secretion of inflammatory cytokines. Furthermore, NAPRT enhances monocyte differentiation into macrophages by inducing macrophage colony-stimulating factor. These NAPRT-induced effects are independent of NAD-biosynthetic activity, but rely on NAPRT binding to TLR4. In line with our finding that NAPRT mediates endotoxin tolerance in vitro and in vivo, sera from patients with sepsis contain the highest levels of NAPRT, compared to patients with other chronic inflammatory conditions. Together, these data identify NAPRT as a endogenous ligand for TLR4 and a mediator of inflammation.
Proteasome inhibitors (PI) are extensively used for the therapy of multiple myeloma (MM) and mantle cell lymphoma. However, patients continuously relapse or are intrinsically resistant to this class ...of drugs. Here, to identify targets that synergize with PI, we carried out a functional screening in MM cell lines using a short hairpin RNA library against cancer driver genes. Isocitrate dehydrogenase 2 (IDH2) was identified as a top candidate, showing a synthetic lethal activity with the PI carfilzomib (CFZ). Combinations of US Food and Drug Administration–approved PI with a pharmacological IDH2 inhibitor (AGI-6780) triggered synergistic cytotoxicity in MM, mantle cell lymphoma, and Burkitt lymphoma cell lines. CFZ/AGI-6780 treatment increased death of primary CD138+ cells from MM patients and exhibited a favorable cytotoxicity profile toward peripheral blood mononuclear cells and bone marrow–derived stromal cells. Mechanistically, the CFZ/AGI-6780 combination significantly decreased tricarboxylic acid cycle activity and adenosine triphosphate levels as a consequence of enhanced IDH2 enzymatic inhibition. Specifically, CFZ treatment reduced the expression of nicotinamide phosphoribosyltransferase (NAMPT), thus limiting IDH2 activation through the NAD+-dependent deacetylase SIRT3. Consistently, combination of CFZ with either NAMPT or SIRT3 inhibitors impaired IDH2 activity and increased MM cell death. Finally, inducible IDH2 knockdown enhanced the therapeutic efficacy of CFZ in a subcutaneous xenograft model of MM, resulting in inhibition of tumor progression and extended survival. Taken together, these findings indicate that NAMPT/SIRT3/IDH2 pathway inhibition enhances the therapeutic efficacy of PI, thus providing compelling evidence for treatments with lower and less toxic doses and broadening the application of PI to other malignancies.
•IDH2 is a new synthetic lethal target to PI, efficacious in several hematological malignancies.•Inhibition of NAMPT/SIRT3/IDH2 pathway could enhance the therapeutic efficacy and overcome resistance to PI.
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NOTCH1 mutations in chronic lymphocytic leukemia (CLL) lead to accumulation of NOTCH1 intracellular domain (NICD) and prolong signaling. These mutations associate with a more aggressive disease ...compared to wild-type (WT) CLL. In this work we demonstrate a bidirectional functional relationship between NOTCH1 and the B cell receptor (BCR) pathways. By using highly homogeneous cohorts of primary CLL cells, activation of NOTCH1 is shown to increase expression of surface IgM, as well as LYN, BTK, and BLNK, ultimately enhancing BCR signaling responses, including global mRNA translation. Upon BCR cross-linking, NOTCH1 itself is actively translated and increased on cell surface. Furthermore, BCR ligation induces calcium mobilization that can facilitate ligand-independent NOTCH1 activation. These data suggest that the two pathways are functionally linked, providing a rationale for dual inhibition strategies. Consistently, addition of the γ-secretase inhibitor DAPT to ibrutinib significantly potentiates its effects, both in vitro and in a short-term patient-derived xenograft model. While this observation may find limited applications in the CLL field, it is more relevant for Richter's Syndrome (RS) management, where very few successful therapeutic options exist. Treatment of RS-patient-derived xenografts (RS-PDX) with the combination of ibrutinib and DAPT decreases disease burden and increases overall survival.
UVB radiation causes about 90% of non-melanoma skin cancers by damaging DNA either directly or indirectly by increasing levels of reactive oxygen species (ROS). Skin, chronically exposed to both ...endogenous and environmental pro-oxidant agents, contains a well-organised system of chemical and enzymatic antioxidants. However, increased or prolonged free radical action can overwhelm ROS defence mechanisms, contributing to the development of cutaneous diseases. Thus, new strategies for skin protection comprise the use of food antioxidants to counteract oxidative stress. Resveratrol, a phytoalexin from grape, has gained a great interest for its ability to influence several biological mechanisms like redox balance, cell proliferation, signal transduction pathways, immune and inflammatory response. Therefore, the potential of resveratrol to modify skin cell response to UVB exposure could turn out to be a useful option to protect skin from sunlight-induced degenerative diseases. To investigate into this matter, HaCaT cells, a largely used model for human skin keratinocytes, were treated with 25 or 100 µM resveratrol for 2 and 24 hours prior to UVB irradiation (10 to 100 mJ/cm(2)). Cell viability and molecular markers of proliferation, oxidative stress, apoptosis, and autophagy were analyzed. In HaCaT cells resveratrol pretreatment: reduces UVB-induced ROS formation, enhances the detrimental effect of UVB on HaCaT cell vitality, increases UVB-induced caspase 8, PARP cleavage, and induces autophagy. These findings suggest that resveratrol could exert photochemopreventive effects by enhancing UVB-induced apoptosis and by inducing autophagy, thus reducing the odds that damaged cells could escape programmed cell death and initiate malignant transformation.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Scaffolds populated with human cardiac progenitor cells (CPCs) represent a therapeutic opportunity for heart regeneration after myocardial infarction. In this work, square-grid scaffolds are prepared ...by melt-extrusion additive manufacturing from a polyurethane (PU), further subjected to plasma treatment for acrylic acid surface grafting/polymerization and finally grafted with laminin-1 (PU-LN1) or gelatin (PU-G) by carbodiimide chemistry. LN1 is a cardiac niche extracellular matrix component and plays a key role in heart formation during embryogenesis, while G is a low-cost cell-adhesion protein, here used as a control functionalizing molecule. X-ray photoelectron spectroscopy analysis shows nitrogen percentage increase after functionalization. O1s and C1s core-level spectra and static contact angle measurements show changes associated with successful functionalization. ELISA assay confirms LN1 surface grafting. PU-G and PU-LN1 scaffolds both improve CPC adhesion, but LN1 functionalization is superior in promoting proliferation, protection from apoptosis and expression of differentiation markers for cardiomyocytes, endothelial and smooth muscle cells. PU-LN1 and PU scaffolds are biodegraded into non-cytotoxic residues. Scaffolds subcutaneously implanted in mice evoke weak inflammation and integrate with the host tissue, evidencing a significant blood vessel density around the scaffolds. PU-LN1 scaffolds show their superiority in driving CPC behavior, evidencing their promising role in myocardial regenerative medicine.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Tumor immunosuppression is a major cause for treatment failure and disease relapse, both in solid tumors and leukemia. Local hypoxia is among the conditions that cause immunosuppression, acting at ...least in part through the upregulation of extracellular adenosine levels, which potently suppress T cell responses and skew macrophages towards an M2 phenotype. Hence, there is intense investigation to identify drugs that target this axis. By using the TCL1 adoptive transfer CLL mouse model, we show that adenosine production and signaling are upregulated in the hypoxic lymphoid niches, where intense colonization of leukemic cells occurs. This leads to a progressive remodeling of the immune system towards tolerance, with expansion of T regulatory cells (Tregs), loss of CD8+ T cell cytotoxicity and differentiation of murine macrophages towards the patrolling (M2-like) subset. In vivo administration of SCH58261, an inhibitor the A2A adenosine receptor, re-awakens T cell responses, while limiting Tregs expansion, and re-polarizes monocytes towards the inflammatory (M1-like) phenotype. These results show for the first time the in vivo contribution of adenosine signaling to immune tolerance in CLL, and the translational implication of drugs interrupting this pathway. Although the effects of SCH58261 on leukemic cells are limited, interfering with adenosine signaling may represent an appealing strategy for combination-based therapeutic approaches.
Antimicrobial resistant bacteria are emerging biological contaminants of the environment. In aquatic ecosystems, they originate mainly from hospitals, livestock manure and private households sewage ...water, which could contain antimicrobial agents and resistant microorganisms. Aeromonas spp. occur ubiquitously in aquatic environments and they cause disease in fish. Motile aeromonads are also associated with human gastrointestinal and wound infections and fish can act as a transmission route of antimicrobial resistance (AMR) aeromonads to humans. The environmental ubiquity, the natural susceptibility to antimicrobials and the zoonotic potential of Aeromonas spp. make them optimal candidates for studying the AMR in aquatic ecosystems.
The AMR patterns of 95 motile aeromonads isolated from freshwater fish during 2013 and 2016 were analyzed. All samples from fish came from farms and natural water bodies located in northern Italy, which is an area characterized by high anthropic impact on the environment. The isolates were biochemically identified as Aeromonas hydrophila, Aeromonas sobria or Aeromonas caviae and AMR was determined by the standard disk diffusion method. All isolates were resistant to cloxacillin, spiramycin and tilmicosin. High AMR frequencies (> 95%) were detected for tylosin, penicillin and sulfadiazine. AMR to danofloxacin, enrofloxacin, flumequine, ceftiofur, aminosidine, colistin, doxycycline, gentamicin, marbocyl and florfenicol was observed at low levels (< 10%). No AMR to cefquinome was found. Logistic regression showed several differences in antimicrobial activity between complexes. According to the source of aeromonads, only few differences in AMR between isolates from farmed and wild fish were observed.
Our data revealed an increasing trend of AMR to neomycin and apramycin among Aeromonas isolates during the study period, while resistance to erythromycin, tetracycline and thiamphenicol decreased. All isolates were multidrug resistance (MDR), but A. caviae showed the highest number of MDR per isolate. In most isolates, various degrees of MDR were detected to macrolides, quinolones, fluoroquinolones, polymyxins and cephalosporins (third and fourth generations), which are listed, by the World Health Organisation, to be among the highest priority and critically important antimicrobials in human medicine. Our findings underlined that freshwater fish can act as potential source of MDR motile aeromonads. Due to their zoonotic potential, this can pose serious threat to human health.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Gene expression is controlled by epigenetic deregulation, a hallmark of cancer. The DNA methylome of canine diffuse large B-cell lymphoma (cDLBCL), the most frequent malignancy of B-lymphocytes in ...dog, has recently been investigated, suggesting that aberrant hypermethylation of CpG loci is associated with gene silencing. Here, we used a multi-omics approach (DNA methylome, transcriptome and copy number variations) combined with functional in vitro assays, to identify putative tumour suppressor genes subjected to DNA methylation in cDLBCL. Using four cDLBCL primary cell cultures and CLBL-1 cells, we found that
,
and
, which were significantly suppressed in DLBCL samples, were hypermethylated and also responsive (at the DNA, mRNA and protein level) to pharmacological unmasking with hypomethylating drugs and histone deacetylase inhibitors. The regulatory mechanism underneath the methylation-dependent inhibition of those target genes expression was then investigated through luciferase and in vitro methylation assays. In the most responsive CpG-rich regions, an in silico analysis allowed the prediction of putative transcription factor binding sites influenced by DNA methylation. Interestingly, regulatory elements for
and
were commonly identified in all three genes. This study provides a foundation for characterisation and experimental validation of novel epigenetically-dysregulated pathways in cDLBCL.
Pleural mesothelioma (PM) is an aggressive cancer with poor prognosis and no effective therapies, mainly caused by exposure to asbestos. Antagonists of growth hormone-releasing hormone (GHRH) display ...strong antitumor effects in many experimental cancers, including lung cancer and mesothelioma. Here, we aimed to determine whether GHRH antagonist MIA-690 potentiates the antitumor effect of cisplatin and pemetrexed in PM. In vitro, MIA-690, in combination with cisplatin and pemetrexed, synergistically reduced cell viability, restrained cell proliferation and enhanced apoptosis, compared with drugs alone. In vivo, the same combination resulted in a strong growth inhibition of MSTO-211H xenografts, decreased tumor cell proliferation and increased apoptosis. Mechanistically, MIA-690, particularly with chemotherapeutic drugs, inhibited proliferative and oncogenic pathways, such as MAPK ERK1/2 and cMyc, and downregulated cyclin D1 and B1 mRNAs. Inflammatory pathways such as NF-kB and STAT3 were also reduced, as well as oxidative, angiogenic and tumorigenic markers (iNOS, COX-2, MMP2, MMP9 and HMGB1) and growth factors (VEGF and IGF-1). Overall, these findings strongly suggest that GHRH antagonists of MIA class, such as MIA-690, could increase the efficacy of standard therapy in PM.
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