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Overexpression of CD70 has been documented in a variety of solid and hematological tumors, where it is thought to play a role in tumor proliferation and evasion of immune surveillance. Here, we describe ARGX-110, a defucosylated IgG1 monoclonal antibody (mAb) that selectively targets and neutralizes CD70, the ligand of CD27. ARGX-110 was generated by immunization of outbred llamas. The antibody was germlined to 95% human identity, and its anti-tumor efficacy was tested in several in vitro assays. ARGX-110 binds CD70 with picomolar affinity. In depletion studies, ARGX-110 lyses tumor cells with greater efficacy than its fucosylated version. In addition, ARGX-110 demonstrates strong complement-dependent cytotoxicity and antibody-dependent cellular phagocytosis activity. ARGX-110 inhibits signaling of CD27, which results in blocking of the activation and proliferation of Tregs. In a Raji xenograft model, administration of the fucosylated version of ARGX-110 resulted in a prolonged survival at doses of 0.1 mg/kg and above. The pharmacokinetics of ARGX-110 was tested in cynomolgus monkeys; the calculated half-life is 12 days.In conclusion, ARGX-110 is a potent blocking mAb with a dual mode of action against both CD70-bearing tumor cells and CD70-dependent Tregs. This antibody is now in a Phase 1 study in patients with advanced malignancies expressing CD70 (NCT01813539).

When activated by the sympathetic nervous system, brown adipose tissue (BAT) increases energy expenditure to produce heat. Augmenting BAT mass or increasing BAT activation could potentially be used to decrease obesity. Noninvasive methods to detect and monitor BAT mass are needed. Contrast ultrasound can estimate BAT blood flow and is able to measure the perfused volume of an organ and thus its mass. The objective of this study was to evaluate whether contrast ultrasound could characterize BAT mass in two mouse models of obesity: wild-type mice fed a high-fat diet and mutant db/dbmice.

SUMMARY Aromatase inhibitors (AIs) are increasingly being used to treat postmenopausal estrogen receptor-positive breast cancer. In the short term they enhance or induce menopausal symptoms that compromise AI adherence, while in the long-term they adversely affect bone and probably also cardiovascular health, although this is mainly when compared to tamoxifen, which is cardioprotective. The distinct decrease in estrogen levels following AI administration is believed to be the main reason for the emergence of these side effects, although exact mechanisms remain to be elucidated. As such, management strategies to meet these AI-induced toxicities are still mainly symptomatic. In this review, current treatment options and promising new modalities under research will be discussed.

The aim of this study was to assess the role of vitamin D in cancer development in postmenopausal osteoporotic women.

Nitric oxide mediates endothelium-dependent vasodilation, modulates cerebral blood flow, and determines stroke outcome. Nitric oxide signals in part by stimulating soluble guanylate cyclase (sGC) to synthesize cGMP. To study the role of sGC in stroke injury, we compared the outcome of cerebral ischemia and reperfusion in mice deficient in the 1subunit of sGC (sGC1−/−) with that in wild-type mice.

Sepsis-associated vasodilation and shock are centrally orchestrated by NO. Nevertheless, inhibition of NO synthesis may not be the target of choice for the treatment of septic shock because it increases morbidity and mortality. Potential other therapeutic targets include soluble guanulate cyclase (sGC) and K+channels. In this study, we investigated the protective effect of the sGC inhibitor methylene blue (MB) and various K+channel inhibitors on LPS- and TNF-induced mortality in mice. In TNF-induced shock, the importance of SK channels was underscored by the ability of a single treatment with the small-conductance calcium-activated SK channel inhibitor apamin to provide significant protection. The only other K+channel inhibitor that can add survival benefit to the apamin treatment was iberiotoxin, stressing the importance of large-conductance calcium-activated BK channels as well. Although MB can protect against TNF-induced shock and mortality, it cannot prevent LPS-induced mortality. Treatment with the nonspecific K+channel inhibitor tetraethylammonium or with inhibitors specific for adenosine triphosphate (ATP)-sensitive KATPchannels (glibenclamide), BK channels (iberiotoxin), or SK channels (apamin) could not protect either. However, when we combined MB treatment with a single dose of apamin and iberiotoxin, mice were completely protected against LPS-induced death for at least 2 days. In conclusion, the protective effect of MB in combination with apamin and iberiotoxin indicates an important role for SK and BK channels, rather than KATPchannels, during endotoxemia. Our results point to SK and BK channels as potential targets in septic shock treatment to be modulated preferentially together with sGC.

In this paper, we propose an efficient method for collecting large amounts of malicious internet traffic. The key advantage of our method is that it does not need to maintain any state to emulate TCP services running on a large number of emulated end-systems. We implemented a prototype on the ASAX intrusion detection system and we provide several examples of the malicious activities that were collected on a campus network attached to the internet. We explain how we implemented various protocols in a stateless way. We also discuss how our method can be improved to make an accurate but still stateless emulation of stateful protocols.

Thin film InP/InGaAsP active opto-electronic devices were fabricated on top of Silicon-on-Insulator waveguide circuits. Device integration was done using adhesive die to wafer bonding. Both photodetectors and laser diodes were fabricated on and coupled to Silicon-on-Insulator waveguide circuits. Integration of laser diodes and photodetectors was done simultaneously using identical processing steps.

The bonding process of InP/InGaAsP dies to processed SOI using DVS-bis-BenzoCycloButene was developed. Surface preparation and the degree of planarization of bonding layers thinner than 300nm were optimized. Bonding strength and bonding quality was assessed, which shows to be sufficient for post-bonding processing. The heterogeneous integration of active photonic devices fabricated in this bonding layer on top of the SOI photonic layer is described.

Presents the author's experience of how his life was transformed with the evolution of agriculture. Challenges faced by the author and his family for earning their living; Benefits of studying professional agriculture on his family farming business; Contribution of the agricultural development on the author's farming business; Significance of the modernization on the agricultural sector.

Previous studies involving filter runs seeded with oocysts have not shown significantly poorer removals during ripening. However, these studies typically did not seed oocysts throughout the preceding filter run, which is necessary for representative numbers of oocysts to be present in the backwash‐remnant water. In this study, four trials consisting of three to five consecutive filter runs were continuously seeded with oocysts and/or microsphere surrogates so that they would appear in the backwash‐remnant water of the successive run. Compared with concentrations in the postripening period, the concentrations of oocysts and microspheres found in the effluent under the influence of backwash‐remnant particles ranged from no higher to more than an order of magnitude higher, depending on filter type. Significant carryover of different‐colored microspheres was not observed in consecutive filter runs following the filter‐ripening period. During this study, conventional and biological filters showed similar average postripening oocyst and microsphere removals (roughly 1.7 and 2.1 log, respectively), but the biological filters exhibited significantly lower concentrations of oocysts and microspheres in the initial effluent samples containing backwash‐remnant particles left in the filter at the conclusion of the backwashing procedure.

Presents the author's experience of fixing a nitrogen leakage in a pipe at his factory. Role of nitrogen in conveying under pressure, aluminum alkyl (AA) in engines and pipes; Ignition of AA in the presence of air and explosion in the presence of water; Distribution of nitrogen network everywhere in the factory; Safety measures taken by the author and his colleagues while fixing the nitrogen leakage.

The evaluation of environmental cleanliness of an electroplating facility, as compared to the best available practice, has been a challenge, particularly in small or mid-sized plants. This is mainly due to the fact that the detailed plant data necessary for evaluation is always difficult to obtain completely and precisely. To alleviate the data-scarce and lack-of-skill related problems in environmental performance evaluation for cleaner production, a fuzzy-logic-based decision analysis approach is introduced in this paper. The attractiveness of the approach is illustrated by the analysis of rinse system management. The approach is general and thus is suitable for any type of environmental cleanliness problems in the electroplating industry. [ABSTRACT FROM AUTHOR]

Apoptotic cells are cleared by phagocytosis during development, homeostasis, and pathology. However, it is still unclear how necrotic cells are removed. We compared the phagocytic uptake by macrophages of variants of L929sA murine fibrosarcoma cells induced to die by tumor necrosis factor-induced necrosis or by Fas-mediated apoptosis. We show that apoptotic and necrotic cells are recognized and phagocytosed by macrophages, whereas living cells are not. In both cases, phagocytosis occurred through a phosphatidylserine-dependent mechanism, suggesting that externalization of phosphatidylserine is a general trigger for clearance by macrophages. However, uptake of apoptotic cells was more efficient both quantitatively and kinetically than phagocytosis of necrotic cells. Electron microscopy showed clear morphological differences in the mechanisms used by macrophages to engulf necrotic and apoptotic cells. Apoptotic cells were taken up as condensed membrane-bound particles of various sizes rather than as whole cells, whereas necrotic cells were internalized only as small cellular particles after loss of membrane integrity. Uptake of neither apoptotic nor necrotic L929 cells by macrophages modulated the expression of proinflammatory cytokines by the phagocytes.

Apoptotic cells are cleared by phagocytosis during development, homeostasis, and pathology. However, it is still unclear how necrotic cells are removed. We compared the phagocytic uptake by macrophages of variants of L929sA murine fibrosarcoma cells induced to die by tumor necrosis factor-induced necrosis or by Fas-mediated apoptosis. We show that apoptotic and necrotic cells are recognized and phagocytosed by macrophages, whereas living cells are not. In both cases, phagocytosis occurred through a phosphatidylserine-dependent mechanism, suggesting that externalization of phosphatidylserine is a general trigger for clearance by macrophages. However, uptake of apoptotic cells was more efficient both quantitatively and kinetically than phagocytosis of necrotic cells. Electron microscopy showed clear morphological differences in the mechanisms used by macrophages to engulf necrotic and apoptotic cells. Apoptotic cells were taken up as condensed membrane-bound particles of various sizes rather than as whole cells, whereas necrotic cells were internalized only as small cellular particles after loss of membrane integrity. Uptake of neither apoptotic nor necrotic L929 cells by macrophages modulated the expression of proinflammatory cytokines by the phagocytes.

In mice, tumor necrosis factor (TNF) displays a selective species specificity. In contrast to murine TNF (mTNF), human TNF (hTNF) only induces lethality at extremely high doses of about 500 μg/mouse, whereas it still has a powerful antitumor activity in combination with interferon-γ. The observation that hTNF does not interact with the p75 mTNF receptor seemed to provide a plausible explanation for these species-specific biological effects. Experiments in TNF receptor knockout mice and tests with hTNF muteins in baboons did not, however, support this hypothesis. We here show that an mTNF mutein selective for the p55 mTNF receptor induces lethality in a manner comparable to wild-type mTNF, and conclude that other differences between hTNF and mTNF must account for the reduced lethality of hTNF. Pharmacokinetics showed that hTNF is cleared much faster than mTNF or the mTNF mutein used. In contrast to the hardly lethal effect(s) of a bolus administration of hTNF, fractionated repetitive administration of the same total hTNF dose induced lethality. This suggests that prolonged exposure rather than peak levels determine the lethal effects of hTNF in mice. Experiments with receptor and ligand knockouts demonstrated that the difference in pharmacokinetics is independent of an interaction with (soluble) TNF receptor, TNF-induced effects or induction of endogenous TNF. These results show that manipulation of the clearance rate of TNF may broaden the therapeutic range of systemic treatments with TNF.

In many gastrointestinal tissues nitric oxide (NO) and vasoactive intestinal polypeptide (VIP) both play a role as inhibitory non‐adrenergic non‐cholinergic neurotransmitters. As the mode of interaction between NO and VIP remains controversial, the aim of this study was to investigate the interplay between NO and VIP in the mouse gastric fundus and to evaluate the nitric oxide synthase (NOS) isoform involved in VIP‐induced relaxation by using inducible NOS (iNOS), endothelial NOS (eNOS) and neuronal NOS (nNOS) knockout mice. The influence of NOS inhibitors on the relaxant effect of VIP was determined in isolated smooth muscle cells and smooth muscle strips of wild‐type and knockout mice. In isolated smooth muscle cells from wild‐type, eNOS knockout and nNOS knockout mice, the relaxation induced by VIP (10−9m) was inhibited by approximately 70–95 % by both the non‐selective NOS inhibitor NG‐nitro‐l‐arginine (l‐NA; 10−4m) and the selective inducible NOS inhibitor N‐(3‐(aminomethyl)‐benzyl)acetamidine (1400W; 10−6m). In cells isolated from iNOS knockout mice, VIP still induced full relaxation but it was not influenced by l‐NA or 1400W. In smooth muscle strips from wild‐type and knockout mice, the concentration‐dependent relaxation by VIP (10−9to 3 ×10−7m) was not influenced by l‐NA or 1400W. These results suggest that the experimental method determines the influence of NOS inhibitors on the relaxant effect of VIP. iNOS, probably induced by the isolation procedure, might be involved in the relaxant effect of VIP in isolated smooth muscle cells but not in classic smooth muscle strips.

Tumor necrosis factor (TNF) is a proinflammatory cytokine, which is centrally involved in several inflammatory disorders. Administration of TNF leads to a potentially lethal systemic inflammatory response syndrome (SIRS). We observed that (a) mice lacking functional genes for metallothionein 1 and 2 (MT-null) were protected compared with wild-type controls (P = 0.0078), and (b) mice overexpressing MT-1 (MT-TG) were more sensitized for the lethal effect of TNF than control mice (P = 0.0003), indicating a mediating role for MT in TNF induced SIRS. As MT is involved in the body zinc homeostasis, we tested whether zinc-deprivation or -supplementation alters the response to TNF. Although zinc-depletion strongly sensitized (P = 0.036), and pretreatment with zinc sulfate (ZnSO4) conferred protection against the deleterious effects of TNF (P < 0.0002), it was also found that the protection provided by zinc is independent of MT. Our observation that hsp70 is strongly induced in jejunum after ZnSO4 treatment, suggests a contribution of hsp70 in the protection against TNF. In addition, ZnSO4 cotreatment allowed complete regression of inoculated tumors with TNF and interferon γ, leading to a significantly better survival (P = 0.0045).

The bioactivity of tumor necrosis factor (TNF) is mediated by two TNF receptors (TNF-Rs), more particularly TNF-RI and TNF-RII. Although human TNF (hTNF) and murine TNF (mTNF) are very homologous, hTNF binds only to mTNF-RI. By measuring the binding of a panel of mTNF/hTNF chimeras to both mTNF-R, we pinpointed the TNF region that mediates the interaction with mTNF-RII. Using site-specific mutagenesis, we identified amino acids 71–73 and 89 as the main interacting residues. Mutein hTNF-S71D/T72Y/H73Δ/T89E interacts with both types of mTNF-R and is active in CT6 cell proliferation assays mediated by mTNF-RII. Mutein mTNF-D71S/Y72T/Δ73H/E89T binds to mTNF-RI only and is no longer active on CT6 cells. However, the L929s cytotoxicity of this mutein (an effect mediated by mTNF-RI triggering) was also 100-fold lower than that of wild-type mTNF due to enhanced dissociation during incubation at subnanomolar concentrations. The additional mutation of amino acid 102, resulting in the mutein mTNF-D71S/Y72T/Δ73H/E89T/P102Q, restored the trimer stability, which led to an enhanced specific activity on L929s cells. Hence the specific activity of a TNF species is governed not only by its receptor binding characteristics but also by its trimer stability after incubation at subnanomolar concentrations. In conclusion, the mutation of TNF amino acids 71–73, 89, and 102 is sufficient to obtain a mTNF mutein selective for mTNF-RI and a hTNF mutein that, unlike wild-type hTNF, also acts on mTNF-RII.

Incubation of murine tumor necrosis factor (mTNF) at subnanomolar concentrations results in partial dissociation of the trimers, coinciding with a decrease in bioactivity. Using size-exclusion chromatography, we observed that the conversion of labeled mTNF to monomers is not only prevented by coincubation with an excess of unlabeled mTNF but also with unlabeled human TNF (hTNF). Moreover, after coincubation of mTNF and hTNF four different TNF complexes were revealed by native polyacrylamide gel electrophoresis, viz. homotrimeric mTNF and hTNF, as well as two complexes with an intermediate migration pattern. Analytical gel filtration in combination with native polyacrylamide gel electrophoresis and Western blot immunodetection indicated that these new complexes consisted of heterotrimeric TNF molecules. We conclude that an exchange of monomers takes place during coincubation of two different species of TNF, which results in homotrimeric and heterotrimeric TNF. To assess receptor interaction in vitro, TNF heterotrimeric molecules were used as obtained after incubation of mTNF with labeled hTNF (which only binds to mTNF receptor I) or with labeled mutein mTNF75 (specific for mTNF receptor II). These heterotrimers were retained by both mTNF receptors, which means that the mTNF subunits incorporated in heterotrimeric complexes still can bind to both types of TNF receptor. In addition, the gradual decrease in mTNF bioactivity during preincubation at subnanomolar concentrations was prevented by the presence of mutein mTNF75, which is inactive in an L929 cytotoxicity assay, indicating that heterotrimerization can influence the overall bioactivity.

The proinflammatory cytokine tumor necrosis factor alpha (TNF-alpha) induces lethal hepatitis when injected into D-(+)-galactosamine-sensitized mice on the one hand or systemic inflammatory response syndrome (SIRS) in normal mice on the other hand. We studied whether serum amyloid P component (SAP), the major acute-phase protein in mice, plays a protective role in both lethal models. For this purpose, we used SAP(0/0) mice generated by gene targeting. We studied the lethal response of SAP(0/0) or SAP(+/+) mice to both lethal triggers but found no differences in the sensitivity of both types of mice. We also investigated whether SAP is involved in establishing two types of endogenous protection: one using a single injection of interleukin-1beta (IL-1beta) for desensitization and clearly involving a liver protein, the other by tolerizing mice for 5 days using small doses of human TNF-alpha. Although after IL-1beta or after tolerization the SAP levels in the serum had risen fourfold in the control mice and not in the SAP(0/0) mice, the same extents of desensitization and tolerization were achieved. Finally, we observed that the induction of hemorrhagic necrosis in the skin of mice by two consecutive local injections with TNF-alpha was not altered in SAP(0/0) mice. We conclude that the presence or absence of SAP has no influence on the sensitivity of mice to TNF-alpha-induced hepatitis, SIRS, and hemorrhagic necrosis or on the endogenous protective mechanisms of desensitization or tolerization.

Tumor necrosis factor (TNF) induces hepatitis when injected in human beings or in rodents. The molecular mechanism by which TNF induces hepatic distress remains largely unknown, although induction of apoptosis of hepatocytes appears to be an essential step. In order to increase the therapeutic value of TNF, we have studied the protective activity of several molecules and found that four chemically totally different substances confer significant protection in the model of TNF‐induced lethal hepatitis in mice sensitized with D‐(+)‐galactosamine (GalN), but not in mice sensitized with actinomycin‐D(ActD) or against anti‐Fas‐induced lethal hepatitis. Verapamil, a calcium‐channel blocker, tannic acid, picotamide, a thromboxane A2receptor antagonist, and K76COOH, an inhibitor, amongst others, of complement, protected significantly against induction of lethality, release of the liver‐specific enzyme alanine aminotransferase (ALT) and induction of apoptosis in the liver after TNF/GalN, except for K76COOH, which paradoxically increased ALT values after challenge, and which also protected against TNF/GalN in complement‐deficient mice. The data suggest that activation of platelets and neutrophils, as well as induction of inflammation occur in the TNF/GalN model, but not in the TNF/ActD or anti‐Fas models, in which direct induction of apoptosis of hepatocytes may be more relevant. The protective activity of the drugs may lead to an increase in therapeutic value of TNF.