Early investigation of a novel SI306 theranostic prodrug for glioblastoma treatment.

Glioblastoma multiforme (GBM) is one of the most aggressive malignancies with a high recurrence rate and poor prognosis. Theranostic, combining therapeutic and diagnostic approaches, arises as a successful strategy to improve patient outcomes through personalized medicine. Src is a non-receptor tyrosine kinase (nRTK) whose involvement in GBM has been extensively demonstrated. Our previous research highlighted the effectiveness of the pyrazolo[3,4-d]pyrimidine SI306 and its more soluble prodrug CMP1 as Src inhibitors both in in vitro and in vivo GBM models. In this scenario, we decided to develop a theranostic prodrug of SI306, ProSI-DOTA(68 Ga) 1, which was designed to target GBM cells after hydrolysis and follow-up on the disease's progression and improve the therapy's outcome. First, the corresponding nonradioactive prodrug 2 was tested to evaluate its ADME profile and biological activity. It showed good metabolic stability, no inhibition of CYP3A4, suboptimal aqueous solubility, and slight gastrointestinal and blood-brain barrier passive permeability. Compound 2 exhibited a drastic reduction of cell vitality after 72 h on two different GBM cell lines (GL261 and U87MG). Then, 2 was subjected to complexation with the radionuclide Gallium-68 to give ProSI-DOTA(68 Ga) 1. The cellular uptake of 1 was evaluated on GBM cells, highlighting a slight but significant time-dependent uptake. The data obtained from our preliminary studies reflect the physiochemical properties of 1. The use of an alternative route of administration, such as the intranasal route, could overcome the physiochemical limitations and enhance the pharmacokinetic properties of 1, paving the way for its future development.

Photoprotective and Anti-Aging Properties of the Apical Frond Extracts from the Mediterranean Seaweed Ericaria amentacea.

There is a growing interest in using brown algal extracts thanks to the bioactive substances they produce for adaptation to the marine benthic environment. We evaluated the anti-aging and photoprotective properties of two types of extracts (50%-ethanol and DMSO) obtained from different portions, i.e., apices and thalli, of the brown seaweed, Ericaria amentacea. The apices of this alga, which grow and develop reproductive structures during summer when solar radiation is at its peak, were postulated to be rich in antioxidant compounds. We determined the chemical composition and pharmacological effects of their extracts and compared them to the thallus-derived extracts. All the extracts contained polyphenols, flavonoids and antioxidants and showed significant biological activities. The hydroalcoholic apices extracts demonstrated the highest pharmacological potential, likely due to the higher content of meroditerpene molecular species. They blocked toxicity in UV-exposed HaCaT keratinocytes and L929 fibroblasts and abated the oxidative stress and the production of pro-inflammatory cytokines, typically released after sunburns. Furthermore, the extracts showed anti-tyrosinase and anti-hydrolytic skin enzyme activity, counteracting the collagenase and hyaluronidase degrading activities and possibly slowing down the formation of uneven pigmentation and wrinkles in aging skin. In conclusion, the E. amentacea apices derivatives constitute ideal components for counteracting sunburn symptoms and for cosmetic anti-aging lotions.

Virtual Screening Combined with Enzymatic Assays to Guide the Discovery of Novel SIRT2 Inhibitors.

Sirtuin isoform 2 (SIRT2) is one of the seven sirtuin isoforms present in humans, being classified as class III histone deacetylases (HDACs). Based on the high sequence similarity among SIRTs, the identification of isoform selective modulators represents a challenging task, especially for the high conservation observed in the catalytic site. Efforts in rationalizing selectivity based on key residues belonging to the SIRT2 enzyme were accompanied in 2015 by the publication of the first X-ray crystallographic structure of the potent and selective SIRT2 inhibitor SirReal2. The subsequent studies led to different experimental data regarding this protein in complex with further different chemo-types as SIRT2 inhibitors. Herein, we reported preliminary Structure-Based Virtual Screening (SBVS) studies using a commercially available library of compounds to identify novel scaffolds for the design of new SIRT2 inhibitors. Biochemical assays involving five selected compounds allowed us to highlight the most effective chemical features supporting the observed SIRT2 inhibitory ability. This information guided the following in silico evaluation and in vitro testing of further compounds from in-house libraries of pyrazolo-pyrimidine derivatives towards novel SIRT2 inhibitors (1-5). The final results indicated the effectiveness of this scaffold for the design of promising and selective SIRT2 inhibitors, featuring the highest inhibition among the tested compounds, and validating the applied strategy.

New Class of Benzodiazepinone Derivatives as Pro-Death Agents Targeting BIR Domains in Cancer Cells.

Inhibitor of Apoptosis Proteins (IAPs) are validated targets for cancer therapy, and the deregulation of their activities within the NF-κB pathway correlates with chemoresistance events, even after treatment with IAPs-antagonists in the clinic (Smac-mimetics). The molecule FC2 was identified as a NF-κB pathway modulator in MDA-MB-231 adenocarcinoma cancer cells after virtual screening of the Chembridge library against the Baculoviral IAP Repeat 1 (BIR1) domain of cIAP2 and XIAP. An improved cytotoxic effect is observed when FC2 is combined with Smac-mimetics or with the cytokine Tumor Necrosis Factor (TNF). Here, we propose a library of 22 derivatives of FC2, whose scaffold was rationally modified starting from the position identified as R1. The cytotoxic effect of FC2 derivatives was evaluated in MDA-MB-231 and binding to the cIAP2- and XIAP-BIR1 domains was assessed in fluorescence-based techniques and virtual docking. Among 22 derivatives, 4m and 4p display improved efficacy/potency in MDA-MB-231 cells and low micromolar binding affinity vs the target proteins. Two additional candidates (4b and 4u) display promising cytotoxic effects in combination with TNF, suggesting the connection between this class of molecules and the NF-κB pathway. These results provide the rationale for further FC2 modifications and the design of novel IAP-targeting candidates supporting known therapies.

Dysregulation in B-cell responses and T follicular helper cell function in ADA2 deficiency patients.

Adenosine deaminase 2 deficiency (DADA2) is an autoinflammatory disease characterized by inflammatory vasculopathy, early strokes associated often with hypogammaglobulinemia. Pure red cell aplasia, thrombocytopenia, and neutropenia have been reported. The defect is due to biallelic loss of function of ADA2 gene, coding for a protein known to regulate the catabolism of extracellular adenosine. We therefore investigated immune phenotype and B- and T-cell responses in 14 DADA2 patients to address if ADA2 mutation affects B- and T-cell function. Here, we show a significant decrease in memory B cells, in particular class switch memory, and an expansion of CD21low B cells in DADA2 patients. In vitro stimulated B lymphocytes were able to secrete nonfunctional ADA2 protein, suggesting a cell intrinsic defect resulting in an impairment of B-cell proliferation and differentiation. Moreover, CD4+ and CD8+ T cells were diminished; however, the frequency of circulating T follicular helper cells was significantly increased but they had an impairment in IL-21 production possibly contributing to an impaired B cell help. Our findings suggest that ADA2 mutation could lead to a B-cell intrinsic defect but also to a defective Tfh cell function, which could contribute to the immunodeficient phenotype reported in DADA2 patients.

Cannabis-like activity of Zornia latifolia Sm. detected in vitro on rat cortical neurons: major role of the flavone syzalterin.

Zornia latifolia is a plant suspected to possess psychoactive properties and marketed as a marijuana substitute under the name 'maconha brava'. In this study, the effects of fractions obtained from a 2-propanol extract of aerial portions of the plant were determined by multielectrode array (MEA) analyses on cultured networks of rat cortical neurons. Lipophilic (ZL_lipo, mainly containing flavonoid aglycones), and hydrophilic (ZL_hydro, mainly containing flavonoid glycosides) fractions were initially obtained from the raw extract. ZL_lipo significantly inhibited mean firing rate (MFR) and mean bursting rate (MBR) of MEA recordings, while ZL_hydro induced no inhibition. Column chromatography separation of ZL_lipo yielded five fractions (ZL1-ZL5), among which ZL1 induced the strongest MFR and MBR inhibition. NMR and HPLC-MS analyses of ZL1 revealed the prevalence of the common flavonoids genistein (1) and apigenin (2) (in about a 1:1 ratio), and the presence of the rare flavone syzalterin (6,8-dimethylapigenin) (3) as a minor compound. Exposures of MEA to apigenin and genistein standards did not induce the MFR and MBR inhibition observed with ZL1, whereas exposure to syzalterin standard or to a 1:9 mixture syzalterin-genistein induced effects similar to ZL1. These inhibitory effects were comparable to that observed with high-THC hashish, possibly accounting for the plant psychoactive properties. Data indicate that Z. latifolia, currently marketed as a free herbal product, should be subjected to measures of control. In addition, syzalterin showed distinctive pharmacological properties, opening the way to its possible exploitation as a neuroactive drug.

Healthy Properties of a New Formulation of Pomegranate-Peel Extract in Mice Suffering from Experimental Autoimmune Encephalomyelitis.

A new formulation of a pomegranate-peel extract (PEm) obtained by PUAE (Pulsed Ultrasound-Assisted Extraction) and titrated in both ellagic acid (EA) and punicalagin is proposed, characterized and then analyzed for potential health properties in mice suffering from the experimental autoimmune encephalomyelitis (EAE). PEm effects were compared to those elicited by a formulation containing EA (EAm). Control and EAE mice were chronically administered EAm and Pem dissolved in the drinking water, starting from the day 10 post-immunization (d.p.i.), with a "therapeutic" protocol to deliver daily 50 mg/kg of EA. Treated EAE mice did not limit their daily access to the beverage, nor did they show changes in body weight, but they displayed a significant amelioration of "in vivo" clinical symptoms. "Ex vivo" histochemical analysis showed that spinal-cord demyelination and inflammation in PEm and EAm-treated EAE mice at 23 ± 1 d.p.i. were comparable to those in the untreated EAE animals, while microglia activation (measured as Ionized Calcium Binding Adaptor 1, Iba1 staining) and astrocytosis (quantified as glial fibrillar acid protein, GFAP immunopositivity) significantly recovered, particularly in the gray matter. EAm and PEm displayed comparable efficiencies in controlling the spinal pathological cellular hallmarks in EAE mice, and this would support their delivery as dietary supplementation in patients suffering from multiple sclerosis (MS).

First Evidence of Anti-Steatotic Action of Macrotympanain A1, an Amphibian Skin Peptide from Odorrana macrotympana.

Many different amphibian skin peptides have been characterized and proven to exert various biological actions, such as wound-healing, immunomodulatory, anti-oxidant, anti-inflammatory and anti-diabetic effects. In this work, the possible anti-steatotic effect of macrotympanain A1 (MA1) (FLPGLECVW), a skin peptide isolated from the Chinese odorous frog Odorrana macrotympana, was investigated. We used a well-established in vitro model of hepatic steatosis, consisting of lipid-loaded rat hepatoma FaO cells. In this model, a 24 h treatment with 10 µg/mL MA1 exerted a significant anti-steatotic action, being able to reduce intracellular triglyceride content. Accordingly, the number and diameter of cytosolic lipid droplets (LDs) were reduced by peptide treatment. The expression of key genes of hepatic lipid metabolism, such as PPARs and PLINs, was measured by real-time qPCR. MA1 counteracted the fatty acid-induced upregulation of PPARγ expression and increased PLIN3 expression, suggesting a role in promoting lipophagy. The present data demonstrate for the first time a direct anti-steatotic effect of a peptide from amphibian skin secretion and pave the way to further studies on the use of amphibian peptides for beneficial actions against metabolic diseases.

Expanding the Occurrence of Polysaccharides to the Viral World: The Case of Mimivirus.

The general perception of viruses is that they are small in terms of size and genome, and that they hijack the host machinery to glycosylate their capsid. Giant viruses subvert all these concepts: their particles are not small, and their genome is more complex than that of some bacteria. Regarding glycosylation, this concept has been already challenged by the finding that Chloroviruses have an autonomous glycosylation machinery that produces oligosaccharides similar in size to those of small viruses (6-12 units), albeit different in structure compared to the viral counterparts. We report herein that Mimivirus possesses a glycocalyx made of two different polysaccharides, now challenging the concept that all viruses coat their capsids with oligosaccharides of discrete size. This discovery contradicts the paradigm that such macromolecules are absent in viruses, blurring the boundaries between giant viruses and the cellular world and opening new avenues in the field of viral glycobiology.

The Remarkable Antioxidant and Anti-Inflammatory Potential of the Extracts of the Brown Alga Cystoseira amentacea var. stricta.

Inflammation and oxidative stress are part of the complex biological responses of body tissues to harmful stimuli. In recent years, due to the increased understanding that oxidative stress is implicated in several diseases, pharmaceutical industries have invested in the research and development of new antioxidant compounds, especially from marine environment sources. Marine seaweeds have shown the presence of many bioactive secondary metabolites, with great potentialities from both the nutraceutical and the biomedical point of view. In this study, 50%-ethanolic and DMSO extracts from the species C. amentacea var. stricta were obtained for the first time from seaweeds collected in the Ligurian Sea (north-western Mediterranean). The bioactive properties of these extracts were then investigated, in terms of quantification of specific antioxidant activities by relevant ROS scavenging spectrophotometric tests, and of anti-inflammatory properties in LPS-stimulated macrophages by evaluation of inhibition of inflammatory cytokines and mediators. The data obtained in this study demonstrate a strong anti-inflammatory effect of both C. amentacea extracts (DMSO and ethanolic). The extracts showed a very low grade of toxicity on RAW 264.7 macrophages and L929 fibroblasts and a plethora of antioxidant and anti-inflammatory effects that were for the first time thoroughly investigated. The two extracts were able to scavenge OH and NO radicals (OH EC50 between 392 and 454 µg/mL; NO EC50 between 546 and 1293 µg/mL), to partially rescue H2O2-induced RAW 264.7 macrophages cell death, to abate intracellular ROS production in H2O2-stimulated macrophages and fibroblasts and to strongly inhibit LPS-induced inflammatory mediators, such as NO production and IL-1α, IL-6, cyclooxygenase-2 and inducible NO synthase gene expression in RAW 264.7 macrophages. These results pave the way, for the future use of C. amentacea metabolites, as an example, as antioxidant food additives in antiaging formulations as well as in cosmetic lenitive lotions for inflamed and/or damaged skin.

Identification, Purification and Molecular Characterization of Chondrosin, a New Protein with Anti-tumoral Activity from the Marine Sponge Chondrosia Reniformis Nardo 1847.

: Chondrosia reniformis is a common marine demosponge showing many peculiarities, lacking silica spicules and with a body entirely formed by a dense collagenous matrix. In this paper, we have described the identification of a new cytotoxic protein (chondrosin) with selective activity against specific tumor cell lines, from C. reniformis, collected from the Liguria Sea. Chondrosin was extracted and purified using a salting out approach and molecular weight size exclusion chromatography. The cytotoxic fractions were then characterized by two-dimensional gel electrophoresis and mass spectrometry analysis and matched the results with C. reniformis transcriptome database. The procedure allowed for identifying a full-length cDNA encoding for a 199-amino acids (aa) polypeptide, with a signal peptide of 21 amino acids. The mature protein has a theoretical molecular weight of 19611.12 and an IP of 5.11. Cell toxicity assays showed a selective action against some tumor cell lines (RAW 264.7 murine leukemia cells in particular). Cell death was determined by extracellular calcium intake, followed by cytoplasmic reactive oxygen species overproduction. The in silico modelling of chondrosin showed a high structural homology with the N-terminal region of the ryanodine receptor/channel and a short identity with defensin. The results are discussed suggesting a possible specific interaction of chondrosin with the Cav 1.3 ion voltage calcium channel expressed on the target cell membranes.

Molecular Docking and QSAR Studies as Computational Tools Exploring the Rescue Ability of F508del CFTR Correctors.

Cystic fibrosis (CF) is the autosomal recessive disorder most recurrent in Caucasian populations. Different mutations involving the cystic fibrosis transmembrane regulator protein (CFTR) gene, which encodes the CFTR channel, are involved in CF. A number of life-prolonging therapies have been conceived and deeply investigated to combat this disease. Among them, the administration of the so-called CFTR modulators, such as correctors and potentiators, have led to quite beneficial effects. Recently, based on QSAR (quantitative structure activity relationship) studies, we reported the rational design and synthesis of compound 2, an aminoarylthiazole-VX-809 hybrid derivative exhibiting promising F508del-CFTR corrector ability. Herein, we explored the docking mode of the prototype VX-809 as well as of the aforementioned correctors in order to derive useful guidelines for the rational design of further analogues. In addition, we refined our previous QSAR analysis taking into account our first series of in-house hybrids. This allowed us to optimize the QSAR model based on the chemical structure and the potency profile of hybrids as F508del-CFTR correctors, identifying novel molecular descriptors explaining the SAR of the dataset. This study is expected to speed up the discovery process of novel potent CFTR modulators.

A Bioactive Olive Pomace Extract Prevents the Death of Murine Cortical Neurons Triggered by NMDAR Over-Activation.

We have recently demonstrated that bioactive molecules, extracted by high pressure and temperature from olive pomace, counteract calcium-induced cell damage to different cell lines. Here, our aim was to study the effect of the same extract on murine cortical neurons, since the preservation of the intracellular Ca2+-homeostasis is essential for neuronal function and survival. Accordingly, we treated neurons with different stimuli in order to evoke cytotoxic glutamatergic activation. In these conditions, the high-pressure and temperature extract from olive pomace (HPTOPE) only abolished the effects of N-methyl-d-aspartate (NMDA). Particularly, we observed that HPTOPE was able to promote the neuron rescue from NMDA-induced cell death. Moreover, we demonstrated that HPTOPE is endowed with the ability to maintain the intracellular Ca2+-homeostasis following NMDA receptor overactivation, protecting neurons from Ca2+-induced adverse effects, including aberrant calpain proteolytic activity. Moreover, we highlight the importance of the extraction conditions used that, without producing toxic molecules, allow us to obtain protecting molecules belonging to proanthocyanidin derivatives like procyanidin B2. In conclusion, we can hypothesize that HPTOPE, due to its functional and nontoxic properties on neuronal primary culture, can be utilized for future therapeutic interventions for neurodegeneration.

It's Only a Part of the Story: Analytical Investigation of the Inks and Dyes Used in the Privilegium Maius.

The Privilegium maius is one of the most famous and spectacular forgeries in medieval Europe. It is a set of charters made in the 14th century upon commitment by Duke Rudolf IV, a member of the Habsburg family, to elevate the rank and the prestige of his family. These five charters, now kept at the Österreichisches Staatsarchiv in Vienna, have been subjected to a thorough interdisciplinary study in order to shed light on its controversial story. The charters are composed of pergamenaceous documents bound to wax seals with coloured textile threads. The present contribution concerns the characterisation of the inks used for writing and of the dyes used to colour to the threads: Are they compatible with the presumed age of the charters? Though showing only a part of the whole story of the charters, dyes analysis could contribute in assessing their complex history from manufacturing to nowadays. The dyes were characterised with non-invasive in situ measurements by means of fibre optic (FORS) and with micro-invasive measurements by means of Surface Enhanced Raman Spectroscopy (SERS) and High-Performance Liquid Chromatography with Mass Spectrometry (HPLC-MS) analysis. The results showed that the threads of four of the charters (three dyed with madder, one with orchil) were apparently coloured at different dyeing stages, then re-dyed in the 19-20th century.

The rare sugar N-acetylated viosamine is a major component of Mimivirus fibers.

The giant virus Mimivirus encodes an autonomous glycosylation system that is thought to be responsible for the formation of complex and unusual glycans composing the fibers surrounding its icosahedral capsid, including the dideoxyhexose viosamine. Previous studies have identified a gene cluster in the virus genome, encoding enzymes involved in nucleotide-sugar production and glycan formation, but the functional characterization of these enzymes and the full identification of the glycans found in viral fibers remain incomplete. Because viosamine is typically found in acylated forms, we suspected that one of the genes might encode an acyltransferase, providing directions to our functional annotations. Bioinformatic analyses indicated that the L142 protein contains an N-terminal acyltransferase domain and a predicted C-terminal glycosyltransferase. Sequence analysis of the structural model of the L142 N-terminal domain indicated significant homology with some characterized sugar acetyltransferases that modify the C-4 amino group in the bacillosamine or perosamine biosynthetic pathways. Using mass spectrometry and NMR analyses, we confirmed that the L142 N-terminal domain is a sugar acetyltransferase, catalyzing the transfer of an acetyl moiety from acetyl-CoA to the C-4 amino group of UDP-d-viosamine. The presence of acetylated viosamine in vivo has also been confirmed on the glycosylated viral fibers, using GC-MS and NMR. This study represents the first report of a virally encoded sugar acetyltransferase.

Elicited ROS Scavenging Activity, Photoprotective, and Wound-Healing Properties of Collagen-Derived Peptides from the Marine Sponge Chondrosia reniformis.

Recently, the bioactive properties of marine collagen and marine collagen hydrolysates have been demonstrated. Although there is some literature assessing the general chemical features and biocompatibility of collagen extracts from marine sponges, no data are available on the biological effects of sponge collagen hydrolysates for biomedical and/or cosmetic purposes. Here, we studied the in vitro toxicity, antioxidant, wound-healing, and photoprotective properties of four HPLC-purified fractions of trypsin-digested collagen extracts-marine collagen hydrolysates (MCHs)-from the marine sponge C. reniformis. The results showed that the four MCHs have no degree of toxicity on the cell lines analyzed; conversely, they were able to stimulate cell growth. They showed a significant antioxidant activity both in cell-free assays as well as in H2O2 or quartz-stimulated macrophages, going from 23% to 60% of reactive oxygen species (ROS) scavenging activity for the four MCHs. Finally, an in vitro wound-healing test was performed with fibroblasts and keratinocytes, and the survival of both cells was evaluated after UV radiation. In both experiments, MCHs showed significant results, increasing the proliferation speed and protecting from UV-induced cell death. Overall, these data open the way to the use of C. reniformis MCHs in drug and cosmetic formulations for damaged or photoaged skin repair.

Posidonia oceanica (L.) Delile Ethanolic Extract Modulates Cell Activities with Skin Health Applications.

Seagrasses are high plants sharing adaptive metabolic features with both terrestrial plants and marine algae, resulting in a phytocomplex possibly endowed with interesting biological properties. The aim of this study is to evaluate the in vitro activities on skin cells of an ethanolic extract obtained from the leaves of Posidonia oceanica (L.) Delile, family Potamogetonaceae, herein named Posidonia ethanolic extract (PEE). PEE showed high radical scavenging activity, high phenolic content, and resulted rich in chicoric acid, as determined through HPLC-MS analysis. The use of MTT assay on fibroblasts showed a PEE cytotoxicity threshold (IC05) of 50 µg/mL at 48 h, while a sub-toxic dose of 20 µg/mL induced a significant increase of fibroblast growth rate after 10 days. In addition, an ELISA assay revealed that PEE doses of 5 and 10 µg/mL induced collagen production in fibroblasts. PEE induced dose-dependent mushroom tyrosinase inhibition, up to about 45% inhibition at 1000 µg/mL, while 50% reduction of melanin was observed in melanoma cells exposed to 50 µg/mL PEE. Finally, PEE lipolytic activity was assessed by measuring glycerol release from adipocytes following triglyceride degradation. In conclusion, we have collected new data about the biological activities of the phytocomplex of P. oceanica seagrass on skin cells. Our findings indicate that PEE could be profitably used in the development of products for skin aging, undesired hyperpigmentation, and cellulite.

Effects on Energy Metabolism of Two Guanidine Molecules, (Boc)2 -Creatine and Metformin.

Several enzymes are involved in the energy production, becoming a possible target for new anti-cancer drugs. In this paper, we used biochemical and in silico studies to evaluate the effects of two guanidine molecules, (Boc)2 -creatine and metformin, on creatine kinase, an enzyme involved in the regulation of intracellular energy levels. Our results show that both drugs inhibit creatine kinase activity; however, (Boc)2 -creatine displays a competitive inhibition, while metformin acts with a non-competitive mechanism. Moreover, (Boc)2 -creatine is able to inhibit the activity of hexokinase with a non-competitive mechanism. Considering that creatine kinase and hexokinase are involved in energy metabolism, we evaluated the effects of (Boc)2 -creatine and metformin on the ATP/AMP ratio and on cellular proliferation in healthy fibroblasts, human breast cancer cells (MDA-MB-468), a human neuroblastoma cell line (SH-SY5Y), a human Hodgkin lymphoma cell line (KMH2). We found that healthy fibroblasts were only partially affected by (Boc)2 -creatine, while both ATP/AMP ratio and viability of the three cancer cell lines were significantly decreased. By inhibiting both creatine kinase and hexokinase, (Boc)2 -creatine appears as a promising new agent in anticancer treatment. Further research is needed to understand what types of cancer cells are most suitable to treatment by this new compound. J. Cell. Biochem. 118: 2700-2711, 2017. © 2017 Wiley Periodicals, Inc.

ADA2 deficiency (DADA2) as an unrecognised cause of early onset polyarteritis nodosa and stroke: a multicentre national study.

OBJECTIVES: To analyse the prevalence of CECR1 mutations in patients diagnosed with early onset livedo reticularis and/or haemorrhagic/ischaemic strokes in the context of inflammation or polyarteritis nodosa (PAN). Forty-eight patients from 43 families were included in the study. METHODS: Direct sequencing of CECR1 was performed by Sanger analysis. Adenosine deaminase 2 (ADA2) enzymatic activity was analysed in monocyte isolated from patients and healthy controls incubated with adenosine and with or without an ADA1 inhibitor. RESULTS: Biallelic homozygous or compound heterozygous CECR1 mutations were detected in 15/48 patients. A heterozygous disease-associated mutation (p.G47V) was observed in two affected brothers. The mean age of onset of the genetically positive patients was 24 months (6 months to 7 years). Ten patients displayed one or more cerebral strokes during their disease course. Low immunoglobulin levels were detected in six patients. Thalidomide and anti-TNF (tumour necrosis factor) blockers were the most effective drugs. Patients without CECR1 mutations had a later age at disease onset, a lower prevalence of neurological and skin manifestations; one of these patients displayed all the clinical features of adenosine deaminase 2deficiency (DADA2) and a defective enzymatic activity suggesting the presence of a missed mutation or a synthesis defect. CONCLUSIONS: DADA2 accounts for paediatric patients diagnosed with PAN-like disease and strokes and might explain an unrecognised condition in patients followed by adult rheumatologist. Timely diagnosis and treatment with anti-TNF agents are crucial for the prevention of severe complications of the disease. Functional assay to measure ADA2 activity should complement genetic testing in patients with non-confirming genotypes.

SIRT6 inhibitors with salicylate-like structure show immunosuppressive and chemosensitizing effects.

The NAD+-dependent deacetylase SIRT6 is an emerging cancer drug target, whose inhibition sensitizes cancer cells to chemo-radiotherapy and has pro-differentiating effects. Here we report on the identification of novel SIRT6 inhibitors with a salicylate-based structure. The new SIRT6 inhibitors show improved potency and specificity compared to the hit inhibitor identified in an in silico compound screen. As predicted based on SIRT6 biological roles, the new leads increase histone 3 lysine 9 acetylation and glucose uptake in cultured cells, while blocking TNF-α production and T lymphocyte proliferation. Notably, the new SIRT6 inhibitors effectively sensitize pancreatic cancer cells to gemcitabine. Finally, studies of compound fingerprinting and pharmacokinetics defined the drug-like properties of one of the new SIRT6 inhibitors, potentially allowing for subsequent in vivo proof-of-concept studies. In conclusion, new SIRT6 inhibitors with a salicylate-like structure were identified, which are active in cells and could potentially find applications in disease conditions, including cancer and immune-mediated disorders.