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Nb?Si based alloys have drawn continuously increasing attention due to their excellent high-temperature mechanical properties. The addition of element Fe could improve their poor high-temperature oxidation resistance which largely restricts their application. With the aim to study the effect of Fe addition on the Nb?Si system and to design appropriate alloy composition, the Fe?Nb?Si ternary system was thermodynamically investigated using the CALPHAD (CALculation of PHAse Diagrams) approach aided with the formation enthalpies for ternary compounds at 0 K computed via ab initio calculations. A self-consistent thermodynamic description of the Fe?Nb?Si system was obtained in this work. Key isothermal sections and liquidus projection were presented, and the calculation results showed a good agreement with available experimental data.

Antibodies targeting CTLA-4 are emerging as an important class of cancer therapeutics. It is assumed that these antibodies cause tumor rejection by blocking negative signaling from the CTLA-4-B7 interactions to enhance the priming of naïve T cells in lymphoid organs. However, recent findings have shown that the effectiveness of CTLA-4 antibody critically depends on the Fc domain and the host Fc receptors. It remains unclear if the blocking function of CTLA-4 antibody is required for its anti-tumor activity. To address this, here we have selected a non-blocking anti-CTLA-4 antibody (D138) and assessed its binding property and antitumor activity in comparison with the therapeutic CTLA-4 antibody ipilimumab. Crystal structures of CTLA-4 complexed with these antibodies show that D138 binds to a distinctly different site to that of ipilimumab on the CTLA-4 surface. D138 binding did not block the association of cells expressing CTLA-4 and B7 whereas ipilimumab did. Subsequent antitumor assay revealed that D138 was similarly effective as ipilimumab in inhibiting tumor growth in mice. This antitumor activity required Fc function for efficacy and was correlated with selective reduction of intratumor regulatory T (Treg) cells, resulting in a significant increase in the ratio of CD8+ over Treg cells. Overall these data clearly demonstrate that blocking CTLA-4-B7 interaction is not required for CTLA-4 antibody mediated antitumor activity, opening prospects of developing non-blocking CTLA-4 antibodies or simple binders towards other Treg surface markers for Treg-targeted immunotherapy.

Microfluidics system enables high-throughput functional screening of antibodies for cancer immunotherapy., Currently, high-throughput approaches are lacking in the isolation of antibodies with functional readouts beyond simple binding. This situation has impeded the next generation of cancer immunotherapeutics, such as bispecific T cell engager (BiTE) antibodies or agonist antibodies against costimulatory receptors, from reaching their full potential. Here, we developed a highly efficient droplet-based microfluidic platform combining a lentivirus transduction system that enables functional screening of millions of antibodies to identify potential hits with desired functionalities. To showcase the capacity of this system, functional antibodies for CD40 agonism with low frequency (

In this paper, Lie symmetry analysis is performed for the equation derived from $(2+1)$-dimensional higher order Broer-Kaup equation. Meanwhile, the optimal system and similarity reductions based on the Lie group method are obtained. Furthermore, the conservation law is studied via the Ibragimov’s method.

Human immunoglobulin G (IgG) agonistic antibodies targeting costimulatory immunoreceptors represent promising cancer immunotherapies yet to be developed. Whether, and how, human IgG hinge and Fc impact on their agonistic functions have been disputed. Here, we show that different natural human IgGs confer divergent agonistic anti-CD40 immunostimulatory and antitumour activities in FcγR-humanized mice, including inactive IgG3 and superior IgG2. This divergence is primarily due to their CH1-hinges despite all human IgGs requiring Fc-FcγR binding for optimal agonistic activities. Unexpectedly, biophysical flexibility of these CH1-hinges inversely correlates with, and can modulate, their agonistic potency. Furthermore, IgG Fcs optimized for selective FcγR binding synergize with and still require IgG hinge, selected for rigidity, to confer improved anti-CD40 immunostimulatory and antitumour activities. These findings highlight the importance of both hinge rigidity and selective FcγR binding in antibody agonistic function, and the need for newer strategies to modulate antibody agonism for improved clinical application., Conserved regions of the antibody molecule impact its downstream biological effects. Here the authors show that a rigid hinge conformation increases the agonistic activity of CD40 and DR5 antibodies, distinctly from FcγR-binding, suggesting that the hinge and FcR binding regions can be separately modified to optimize therapies.

Excessive self-reactive and inadequate affinity-matured antigen-specific antibody responses have been reported to coexist in lupus, with elusive cellular and molecular mechanisms. Here, we report that the antigen-specific germinal center (GC) response―a process critical for antibody affinity maturation―is compromised in murine lupus models. Importantly, this defect can be triggered by excessive autoimmunity-relevant CD11c + Tbet + age-associated B cells (ABCs). In B cell-intrinsic Ship-deficient (ShipΔB) lupus mice, excessive CD11c + Tbet + ABCs induce deregulated follicular T-helper (T FH ) cell differentiation through their potent antigen-presenting function and consequently compromise affinity-based GC selection. Excessive CD11c + Tbet + ABCs and deregulated T FH cell are also present in other lupus models and patients. Further, over-activated Toll-like receptor signaling in Ship-deficient B cells is critical for CD11c + Tbet + ABC differentiation, and blocking CD11c + Tbet + ABC differentiation in ShipΔB mice by ablating MyD88 normalizes T FH cell differentiation and rescues antigen-specific GC responses, as well as prevents autoantibody production. Our study suggests that excessive CD11c + Tbet + ABCs not only contribute significantly to autoantibody production but also compromise antigen-specific GC B-cell responses and antibody-affinity maturation, providing a cellular link between the coexisting autoantibodies and inadequate affinity-matured antigen-specific antibodies in lupus models and a potential target for treating lupus.

The programmed cell death protein 1 (PD-1) ligands PD-L1 and PD-L2 on germinal center (GC) B cells deliver coinhibitory signals to follicular T cells. The PD-L1/L2–PD-1 axis modulates the quality and quantity of follicular T cells and has been shown to influence the GC responses. However, the transcriptional control of PD-1 ligands on GC B cells remains largely unknown. In this study, we report that the transcription factor BCL6 is a key negative regulator of the PD-1 ligands PD-L1 and PD-L2 in GC B cells. Acute deletion of Bcl6 in mature GC B cells resulted in marked upregulation of mRNA and protein abundance of PD-1 ligands. Moreover, the expression levels of BCL6 and PD-1 ligands were inversely correlated during GC B cell development and in human GC–derived lymphoma specimens. Mechanically, BCL6 directly bound to the promoter region of PD-L1 and intron 2 of PD-L2 to suppress their transcription. In addition, BCL6 indirectly inhibited the transcription of PD-1 ligands by repressing the expression of STAT1/STAT3 and IRF1. Moreover, BCL6 exerted these effects via its BTB domain. Finally, PD-1 blockade promoted cell survival to sustain the follicular T cell pool in the presence of Bcl6-deficinet GC B cells. In summary, B cell–specific expression of BCL6 dampens the PD-L1/L2–PD-1 signaling to maintain the size of follicular T cells during GC development.

Proper control of B cell growth and metabolism is crucial for B-cell-mediated immunity, but the underlying molecular mechanisms remain incompletely understood. In this study, Sin1, a key component of mTOR complex 2 (mTORC2), specifically regulates B cell growth and metabolism. Genetic ablation of Sin1 in B cells reduces the cell size at either the transitional stage or upon antigen stimulation and severely impairs metabolism. Sin1 deficiency also severely impairs B-cell proliferation, antibody responses, and anti-viral immunity. At the molecular level, Sin1 controls the expression and stability of the c-Myc protein and maintains the activity of mTORC1 through the Akt-dependent inactivation of GSK3 and TSC1/2, respectively. Therefore, our study reveals a novel and specific role for Sin1 in coordinating the activation of mTORC2 and mTORC1 to control B cell growth and metabolism.

IgG4 is the least potent human IgG subclass for the FcγR-mediated antibody effector function and is considered anti-inflammatory in the context of prolonged inflammation and allergic responses. Paradoxically, IgG4 is also the dominant IgG subclass of pathogenic autoantibodies in IgG4-mediated diseases. Here we show that the IgG subclass and Fc-FcγR interaction have a distinct impact on the pathogenic function of autoantibodies in different IgG4-mediated diseases. While IgG4 and its weak Fc-FcγR interaction have an ameliorative role in the pathogenicity of anti-ADAMTS13 autoantibodies isolated from thrombotic thrombocytopenic purpura (TTP) patients, they have an unexpected exacerbating effect on anti-Dsg1 autoantibody pathogenicity in pemphigus foliaceus (PF) models. Strikingly, a non-pathogenic anti-Dsg1 antibody variant optimized for FcγR-mediated effector function can attenuate the skin lesions induced by pathogenic anti-Dsg1 antibodies by promoting the clearance of dead keratinocytes. These studies suggest that IgG effector function contributes to the clearance of autoantibody-Ag complexes, which is harmful in TTP, but beneficial in PF and may provide new therapeutic opportunity.

Currently high throughput approaches are lagged for isolation of antibodies whose function goes beyond simple binding, which have prevented the next generation cancer immunotherapeutics, such as bispecific T cell engager antibodies or agonist antibody of costimulatory receptor, from reaching their full potential. Here we developed a highly efficient droplet-based microfluidics platform combining with lentivirus transduction system that enables functional screening of millions of antibodies. To showcase the capacity of the system, functional antibodies for CD40 agonism with low frequency (

Summary Differentiation and homeostasis of Foxp3+ regulatory T cells (Tregs) are tightly controlled by the interleukin-2 receptor (IL-2R) signaling, yet the mechanisms governing these processes are incompletely understood. Here, we report that transcription factor Bach2 attenuates IL-2R signaling to coordinate Treg differentiation and homeostasis. Bach2 is required for the quiescence, survival, and maintenance of resting Treg cells (rTregs). Unexpectedly, Bach2 directly represses CD25 (IL-2Rα) and subsequently attenuates IL-2R signaling in Tregs. Upregulated CD25/IL-2R signaling in Bach2-deficient rTregs acts as a parallel pathway to partially counteract their poor survival and maintenance. Furthermore, Bach2 suppresses CD25/IL-2R signaling in T follicular regulatory (Tfr) cells. Bach2 deficiency in Tregs prevents the formation of highly differentiated Tfr cells, associated with aberrant GC response. Finally, a mild and late onset of autoimmune disease is observed in mice with Bach2-deficient Tregs. Thus, Bach2 balances IL-2R signaling to orchestrate development and homeostasis of various Treg subsets.

Chronic subdural hematoma (CSDH) is an inflammatory angiogenic disease. It is believed that vascular endothelial growth factor (VEGF) plays an important role in pathological CSDH angiogenesis.In this study, magnetic resonance imaging (MRI) results were used to assign 115 primary CSDH patients to four MRI types. The four MRI types are described as follows: type 1 (T1-weighted low, T2-weighted low), type 2 (T1-weighted high, T2-weighted low), type 3 (T1-weighted mixed, T2-weighted mixed), and type 4 (T1-weighted low/high, T2-weighted high). The four MRI types were then correlated with CSDH stage and patient hematoma fluid and serum VEGH concentrations that were measured using an enzyme-linked immunosorbent assay (ELISA). Neurological status was assessed by Markwalder scoring at admission and six-month follow-up.The mean VEGF concentration was significantly higher in CDSH hematoma fluid samples than in patient sera (p0.01). In unilateral CSDH hematoma fluid samples, VEGF concentration was highest in type 1 (21,613.5±1473.3pg/ml), next highest in type 2 (18,071.8±1737.1pg/ml), lower in type 3, and lowest in type 4 patients (13,153.7±3854.4pg/ml, 7265.7±726.2pg/ml, respectively). High VEGF concentrations strongly correlated with MRI type (unilateral CSDH group r=0.838, bilateral CSDH group r=0.851, p0.01). Moreover, higher hematoma fluid VEGF concentrations correlated with markedly higher recurrence in type 1 (3/19, 15.8%) vs. type 4 unilateral CSDH patients (1/27, 3.7%).The present study reports a significant correlation between CSDH hematoma fluid VEGF concentration and MRI results. Therefore, MRI results could be used to predict hematoma fluid VEGF concentrations in CSDH patients.

BACKGROUND Glioblastoma multiforme (GBM) is the most common malignant and lethal type of primary central nervous system tumor in humans. In spite of its high lethality, a small percentage of patients have a relatively good prognosis, with median survival times of 36 months or longer. The identification of clinical subsets of GBM associated with distinct molecular genetic profiles has made it possible to design therapies tailored to treat individual patients. METHODS We compared microarray data sets from long-term survivors (LTSs) and short-term survivors (STSs) to screen for prognostic biomarkers in GBM patients using the WebArrayDB platform. We focused on FBLN4, IGFBP-2, and CHI3L1, all members of a group of 10 of the most promising, differentially regulated gene candidates. Using formalin-fixed paraffin-embedded GBM samples, we corroborated the relationship between these genes and patient outcomes using methylation-specific polymerase chain reaction (PCR) for MGMT methylation status and quantitative reverse transcription PCR for expression of these genes. RESULTS Expression levels of the mRNAs of these 3 genes were higher in the GBM samples than in normal brain samples and these 3 genes were significantly upregulated in STSs compared to the levels in LTS samples (P

Breast cancer is the most commonly diagnosed cancer and the leading cause of cancer death among women in the worldwide. Triple-negative breast cancer (TNBC) has a poor clinical outcome. The antitumor efficacy of Ilamycins, natural products with anti-tuberculosis activity isolated from deep sea-derived Streptomyces atratus, in TNBC has not been investigated, and the mechanisms remain elusive. Here, we demonstrated that Ilamycin-E, but not -F, decreases cell viability, inhibits G1/S cell cycle progression, and promotes apoptosis in the TNBC cell lines HCC1937 and MDA-MB-468. Ilamycin E promotes apoptosis via activation of endoplasmic reticulum (ER) stress, increasing the expression of CHOP, and down-regulating the expression of anti-apoptotic protein Bcl-2. Depletion of CHOP or overexpression of Bcl2 significantly rescued Ilamycin E-induced apoptosis. These findings indicate that Ilamycin E has anti-cancer activity in TNBC.

While engagement of the inhibitory Fcγ-receptor (FcγR) IIB is an absolute requirement for in vivo antitumor activity of agonistic mouse anti-CD40 monoclonal antibodies (mAbs), a similar requirement for human mAbs has been disputed. By using a mouse model humanized for its FcγRs and CD40, we revealed that FcγRIIB engagement is essential for the activity of human CD40 mAbs, while engagement of the activating FcγRIIA inhibits this activity. By engineering Fc variants with selective enhanced binding to FcγRIIB, but not to FcγRIIA, significantly improved antitumor immunity was observed. These findings highlight the necessity of optimizing the Fc domain for this class of therapeutic antibodies by using appropriate preclinical models that accurately reflect the unique affinities and cellular expression of human FcγR.

Triterpenoids extracted from Cimicifuga foetida have been reported to inhibit cancer by inducing cell cycle arrest and apoptosis. In this study, KHF16 (24-acetylisodahurinol-3-O-β-D-xylopyranoside), a cycloartane triterpenoid isolated from the rhizomes of C. foetida, showed potent anti-cancer activity in multiple ERα/PR/HER2 triple-negative breast cancer (TNBC) cell lines. KHF16 significantly induces cell cycle G2/M phase arrest and apoptosis in both MDA-MB-468 and SW527 TNBC cell lines. KHF16 reduces the expression levels of XIAP, Mcl-1, Survivin and Cyclin B1/D1 proteins. Importantly, KHF16 inhibits TNFα-induced IKKα/β phosphorylation, IKBα phosphorylation, p65 nuclear translocation and NF-κB downstream target gene expression, including XIAP, Mcl-1 and Survivin, in TNBC cells. These results suggest that KHF16 may inhibit TNBC by blocking the NF-κB signaling pathway in part.

Fungal infection stimulates the canonical C-type lectin receptor (CLR) signaling pathway via activation of the tyrosine kinase Syk. Here we identify a crucial role for the tyrosine phosphatase SHP-2 in mediating CLR-induced activation of Syk. Ablation of the gene encoding SHP-2 (Ptpn11; called 'Shp-2' here) in dendritic cells (DCs) and macrophages impaired Syk-mediated signaling and abrogated the expression of genes encoding pro-inflammatory molecules following fungal stimulation. Mechanistically, SHP-2 operated as a scaffold, facilitating the recruitment of Syk to the CLR dectin-1 or the adaptor FcRγ, through its N-SH2 domain and a previously unrecognized carboxy-terminal immunoreceptor tyrosine-based activation motif (ITAM). We found that DC-derived SHP-2 was crucial for the induction of interleukin 1β (IL-1β), IL-6 and IL-23 and anti-fungal responses of the TH17 subset of helper T cells in controlling infection with Candida albicans. Together our data reveal a mechanism by which SHP-2 mediates the activation of Syk in response to fungal infection.

It is well known that the mechanical properties of hypoeutectic Al–Si alloys are influenced by the size, morphology and distribution of primary α-Al and eutectic Si crystals. In the present study, the effect of in-situ γ-Al 2 O 3 particles and heat treatment on the size and morphology of the primary α-Al dendrites and eutectic Si phases in A356 alloy has been investigated. The as-cast and solution treatment samples were characterized by scanning electron microscopy (SEM). Microstructural analyses demonstrated that primary α-Al was significantly refined by in-situ γ-Al 2 O 3 particles. The secondary dendrite arm space (SDAS) of unrefined α-Al is about 25 μm. The SDAS decreases to 11 μm when the content of in-situ γ-Al 2 O 3 particles is 0.6%. In addition, the eutectic Si structure was modified from coarse needle-like/platelet-like structure to short plate-like structure and some discrete particles. The mechanical properties were investigated by tensile test with various concentrations of γ-Al 2 O 3 particles. It was found that the ultimate tensile strength (UTS) and elongation (El) increased by 41% and 137%, respectively, due to decreasing SDAS of primary α-Al dendrites and modifying eutectic Si crystals. Moreover, the change of mechanical properties corresponds to the evolution of microstructure.

Chronic subdural hematoma (CSDH) is a common neurosurgical condition. Currently, surgery is the most effective medical intervention for treatment of this disorder. Because CSDH is an inflammatory angiogenic disease involving multifactorial mechanisms, a better understanding of CSDH pathogenesis should facilitate clinical management. Therefore, the purpose of this review is to describe recent progress in elucidation of molecular mechanisms causing CSDH and to summarize the body of knowledge gained from past drug treatment studies. Because hematoma fluid and outer membrane characteristics may be linked to pathology, they could serve as disease biomarkers. Moreover, past drug treatment studies have shown that such biomarkers may mutually synergize to initiate and promote CSDH progression. These findings suggest that modulation of biomarker expression or function using drug therapy may benefit CSDH patients.

The inhibitory FcγR FcγRIIB is widely expressed on B cells, dendritic cells (DCs), and myeloid effector cells and modulates a variety of Ab-driven in vivo functions. Although it has been established that FcγRIIB plays an important role in the maintenance of peripheral tolerance, the responsible cell-specific FcγRIIB expression remains to be determined. In this study, we generated mice with selective deletion of FcγRIIB in B cells, DCs, and myeloid effector cells and evaluated these novel strains in models of tolerance and autoimmune diseases. Our results demonstrate that mice with selective deletion of FcγRIIB expression in B cells and DCs have increased Ab and T cell responses, respectively, and display enhanced susceptibility to disease in distinct models, suggesting that FcγRIIB expression in distinct cellular populations contributes to the maintenance of peripheral tolerance through different mechanisms.

Significance Agonistic antibodies targeting key TNF receptor (TNFR) molecules involved in antitumor responses have been demonstrated as potent antitumor therapies in preclinical studies. However, no effective agonistic anti-TNFR therapies have been successfully developed to date. In contrast, cytotoxic antitumor antibodies targeting tumor antigens or antagonistic antibodies blocking key inhibitory checkpoints are widely used in clinical settings. One explanation for this discrepancy has been recently provided by the finding that agonistic anti-TNFR antibodies have a previously unappreciated requirement for inhibitory Fcγ receptor FcγRIIB coengagement. Understanding the differential FcγRIIB coengagement requirement by different antibodies and the in vivo cross-linking function of FcγRIIB, as defined by this study, not only has implications for the development of potent agonistic anti-TNFR therapies but also for the understanding of TNFR activation mechanisms.

Anti–CTLA-4 antibody induces selective depletion of T reg cells within tumor lesions in a manner that is dependent on the presence of Fc gamma receptor-expressing macrophages within the tumor microenvironment., Treatment with monoclonal antibody specific for cytotoxic T lymphocyte–associated antigen 4 (CTLA-4), an inhibitory receptor expressed by T lymphocytes, has emerged as an effective therapy for the treatment of metastatic melanoma. Although subject to debate, current models favor a mechanism of activity involving blockade of the inhibitory activity of CTLA-4 on both effector (T eff) and regulatory (T reg) T cells, resulting in enhanced antitumor effector T cell activity capable of inducing tumor regression. We demonstrate, however, that the activity of anti–CTLA-4 antibody on the T reg cell compartment is mediated via selective depletion of T reg cells within tumor lesions. Importantly, T reg cell depletion is dependent on the presence of Fcγ receptor–expressing macrophages within the tumor microenvironment, indicating that T reg cells are depleted in trans in a context-dependent manner. Our results reveal further mechanistic insight into the activity of anti-CTLA-4–based cancer immunotherapy, and illustrate the importance of specific features of the local tumor environment on the final outcome of antibody-based immunomodulatory therapies.

By virtue of their ability to induce apoptosis and regulate growth, differentiation, and cytokine responses, the tumor necrosis factor receptor (TNFR) superfamily members have emerged as attractive targets for anticancer therapeutics. Agonistic antibodies to apoptosis-inducing TNFRs, such as death receptor 5 (DR5), although displaying impressive activities against a variety of tumors in preclinical models, appear to be less active in clinical trials. We report that the in vivo apoptotic and antitumor activities of these antibodies have an absolute requirement for the coengagement of an inhibitory Fcγ receptor, FcγRIIB. Anti-DR5 antibodies of the type currently in clinical trials have weak FcγRIIB binding and thus are compromised in their proapoptotic and antitumor activities in both colon and breast carcinoma models. Enhancing FcγRIIB engagement increases apoptotic and antitumor potency. Our results demonstrate that Fc domain interactions are critical to the therapeutic activity of anti-DR5 antibodies and, together with previous reports on agonistic anti-CD40 antibodies, establish a common requirement for FcγRIIB coengagement for optimal biological effects of agonistic anti-TNFR antibodies.

The in vivo biological activities of IgG antibodies result from their bifunctional nature, in which antigen recognition by the Fab is coupled to the effector and immunomodulatory diversity found in the Fc domain. This diversity, resulting from both amino acid and glycan heterogeneity, is translated into cellular responses through Fcγ receptors (FcγRs), a structurally and functionally diverse family of cell surface receptors found throughout the immune system. Although many of the overall features of this system are maintained throughout mammalian evolution, species diversity has precluded direct analysis of human antibodies in animal species, and, thus, detailed investigations into the unique features of the human IgG antibodies and their FcγRs have been limited. We now report the development of a mouse model in which all murine FcγRs have been deleted and human FcγRs, encoded as transgenes, have been inserted into the mouse genome resulting in recapitulation of the unique profile of human FcγR expression. These human FcγRs are shown to function to mediate the immunomodulatory, inflammatory, and cytotoxic activities of human IgG antibodies and Fc engineered variants and provide a platform for the detailed mechanistic analysis of therapeutic and pathogenic IgG antibodies.

Somatic hypermutation (SHM), coupled with Ag selection, provides a mechanism for generating Abs with high affinity for invading pathogens. Class-switch recombination (CSR) ensures that these Abs attain pathogen-appropriate effector functions. Although the enzyme critical to both processes, activation-induced cytidine deaminase, has been identified, it remains unclear which cis-elements within the Ig loci are responsible for recruiting activation-induced cytidine deaminase and promoting its activity. Studies showed that Ig gene-transcription levels are positively correlated with the frequency of SHM and CSR, making the intronic, transcriptional enhancer Eμ a likely contributor to both processes. Tests of this hypothesis yielded mixed results arising, in part, from the difficulty in studying B cell function in mice devoid of Eμ. In Eμ’s absence, VH gene assembly is dramatically impaired, arresting B cell development. The current study circumvented this problem by modifying the murine Igh locus through simultaneous insertion of a fully assembled VH gene and deletion of Eμ. The behavior of this allele was compared with that of a matched allele carrying the same VH gene but with Eμ intact. Although IgH transcription was as great or greater on the Eμ-deficient allele, CSR and SHM were consistently, but modestly, reduced relative to the allele in which Eμ remained intact. We conclude that Eμ contributes to, but is not essential for, these complex processes and that its contribution is not as a transcriptional enhancer but, rather, is at the level of recruitment and/or activation of the SHM/CSR machinery.