Syk Signaling Pathway Research Articles

Metabolic degradation of polysaccharides from Lentinus edodes by Kupffer cells via the Dectin-1/Syk signaling pathway

It had been shown that lentinan (LNT) was mainly distributed in the liver after intravenous administration. The study aimed to investigate the integrated metabolic processes and mechanisms of LNT in the liver, as these have not been thoroughly explored. In current work, 5-([4,6-dichlorotriazin-2-yl] amino) fluorescein and cyanine 7 were used to label LNT for tracking its metabolic behavior and mechanisms. Near-infrared imaging demonstrated that LNT was captured mainly by the liver. Kupffer cell (KC) depletion reduced LNT liver localization and degradation in BALB/c mice. Moreover, experiments with Dectin-1 siRNA and Dectin-1/Syk signaling pathway inhibitors showed that LNT was mainly taken up by KCs via the Dectin-1/Syk pathway and promoted lysosomal maturation in KCs via this same pathway, which in turn promoted LNT degradation. These empirical findings offer novel insights into the metabolism of LNT in vivo and in vitro, which will facilitate the further application of LNT and other β-glucans.

Bone marrow-mesenchymal stem cells alleviate microglial Pyroptosis after intracerebral hemorrhage in rat by secreting C1q/tumor necrosis factor-related protein 3

BackgroundThere are no specific treatment methods for intracerebral hemorrhage (ICH). Neuroinflammation triggered by microglial pyroptosis plays an important role in ICH pathophysiology. Bone marrow mesenchymal stem cells (BMSCs) are widely used in the treatment of neurological diseases because of their paracrine function. In this study, we aimed to clarify whether BMSCs can alleviate microglial pyroptosis after ICH by secreting C1q/tumor necrosis factor-related protein 3 (CTRP3), a adiponectin paralog with established metabolic regulatory properties and neuroprotective effects. MethodsIn an in vitro study, microglia were stimulated with hemin for pyroptosis and then co-cultured with BMSCs, CTRP3, or CTRP3-small interfering RNA (siRNA)-BMSC; in an in vivo study, intracerebroventricular transplantation of BMSCs or siRNA-CTRP3-BMSCs was performed after ICH surgery. The expression of inflammation-related factors was detected by qRT-PCR and ELISA. Western blotting and immunofluorescence staining were performed to detect the expression of pyroptotic protein, and western blotting was used to detect the activation of phosphoinositide 3-kinase (PI3K), protein kinase B (AKT) and splenic tyrosine kinase (Syk). Behavioral changes were detected 7 days after transplantation. ResultsELISA and qRT-PCR results showed that the production of inflammatory cytokines in hemin-stimulated microglia was significantly downregulated following pretreatment with BMSCs or CTRP3. The Caspase-1 activity assay kit and western blotting results showed that BMSCs attenuated microglial pyroptosis by secreting CTRP3. Furthermore, the modulation functions of BMSCs or CTRP3 involve the promotion of PI3K/AKT and inhibition of Syk signaling pathway activation. Neurological deficits, edema, and disruption of tight junction protein were completely alleviated, while inflammation-related factors and microglial pyroptosis after ICH were significantly downregulated after BMSCs administration. ConclusionBMSCs can inhibit neuroinflammation by inhibiting microglial pyroptosis, thus alleviate ICH symptoms, likely by suppressing the Syk signaling pathway while promoting the PI3K/AKT signaling pathway activation through producing CTRP3.

Curcumin alleviates experimental colitis via a potential mechanism involving memory B cells and Bcl-6-Syk-BLNK signaling

Immune dysfunction is the crucial cause in the pathogenesis of inflammatory bowel disease (IBD), which is mainly related to lymphocytes (T or B cells, incl-uding memory B cells), mast cells, activated neutrophils, and macrophages. As the precursor of B cells, the activation of memory B cells can trigger and differentiate B cells to produce a giant variety of inducible B cells and tolerant B cells, whose dysfunction can easily lead to autoimmune diseases, including IBD. To investigate whether or not curcumin (Cur) can alleviate experimental colitis by regulating memory B cells and Bcl-6-Syk-BLNK signaling. Colitis was induced in mice with a dextran sulphate sodium (DSS) solution in drinking water. Colitis mice were given Cur (100 mg/kg/d) orally for 14 con-secutive days. The colonic weight, colonic length, intestinal weight index, occult blood scores, and histological scores of mice were examined to evaluate the curative effect. The levels of memory B cells in peripheral blood of mice were measured by flow cytometry, and IL-1β, IL-6, IL-10, IL-7A, and TNF-α expression in colonic tissue homogenates were analyzed by enzyme-linked immunosorbent assay. Western blot was used to measure the expression of Bcl-6, BLNK, Syk, and other signaling pathway related proteins. After Cur treatment for 14 d, the body weight, colonic weight, colonic length, colonic weight index, and colonic pathological injury of mice with colitis were ameliorated. The secretion of IL-1β, IL-6, TNF-α, and IL-7A was statistically decreased, while the IL-35 and IL-10 levels were considerably increased. Activation of memory B cell subsets in colitis mice was confirmed by a remarkable reduction in the expression of IgM, IgG, IgA, FCRL5, CD103, FasL, PD-1, CD38, and CXCR3 on the surface of CD19+ CD27+ B cells, while the number of CD19+ CD27+ IL-10+ and CD19+ CD27+ Tim-3+ B cells increased significantly. In addition, Cur significantly inhibited the protein levels of Syk, p-Syk, Bcl-6, and CIN85, and increased BLNK and p-BLNK expression in colitis mice. Cur could effectively alleviate DSS-induced colitis in mice by regulating memory B cells and the Bcl-6-Syk-BLNK signaling pathway.

P461: MYELOID KINOME INHIBITOR HM43239 OVERCOMES ACQUIRED RESISTANCE IN ACUTE MYELOID LEUKEMIA MODELS

Background: Oral HM43239 is in development for the treatment of acute myeloid leukemia (AML) because of its capacity to potently inhibit kinases that drive myeloid malignancies, including diverse forms of the FLT3, SYK, JAK and c-KIT kinases. Wildtype FLT3 is overexpressed in most AML patients, and approximately 30% of newly diagnosed adult AML patients harbor internal tandem duplications (ITDs) or point mutations in the tyrosine kinase domain (TKD). These mutations drive aberrant activation of downstream proliferation pathways and are associated with a high risk of relapse. Likewise, the c-KIT alternative receptor kinase, as well as the SYK and JAK1/2 intracellular kinases, mediate oncogenic signaling in AML that can promote drug resistance to certain FLT3 inhibitors. HM43239 was developed to overcome shortcomings of other FLT3 inhibitors. It inhibits a broad set of mutant and wildtype forms of FLT3, while simultaneously disrupting downstream SYK, JAK/STAT5, ERK, and other rescue signaling pathways. This rationale supports the development of HM43239. Aims: Evaluate the activity of HM43239, an orally active drug, as a FLT3-focused myeloid kinome inhibitor in human AML models. Methods: Biochemical kinase assays were performed by RBC and Carna. The effects of HM43239 on cell proliferation (IC50), growth rate (GR50) and concentration at half-maximal effect (Growth Effective Concentration; GEC50) were determined using the MTS assay with vehicle controls. Cell-based inhibition of target phosphorylation was assessed by Western blot and flow cytometric analyses. The AML cell lines tested included MV-4-11, MOLM-13, MOLM-14 and BAF3/ITD. In vivo efficacy was assessed using the MOLM-14 FLT3-Mutated xenograft model. Results: HM43239 inhibited the enzymatic activities of FLT3-WT, -ITD, and -D835Y variants with IC50 values of 1.1, 1.8 and 1.0 nM, respectively. Binding Kd values for FLT3-WT, -ITD, -D835Y, -D835H, -ITD/D835V and -ITD/F691L were 0.58, 0.37, 0.29, 0.4, 0.48, and 1.3 nM, respectively. HM43239 killed AML cells that harbor the FLT3-ITD mutation (MV-4-11, MOLM-13 and MOLM 14) with IC50, GR50 and GEC50 values ranging from 4 to 10 nM. In cell lines expressing wild type FLT3 (KG1, HEL92.1, SKM-1, and THP-1) the values for these parameters ranged from 0.05 nM to 3 µM. Western blot and flow cytometric analyses of MV-4-11, MOLM-14 cells and Ba/F cells transfected with target proteins revealed that the phosphorylation of FLT3 and SYK was reduced by 50-90% at HM43239 concentrations of 10 – 100 nM, and that these concentrations produced >80% reduction in phosphorylation of ERK and JAK/STAT5 that function downstream of FLT3. HM43239 inhibited FcγR-induced SYK and JAK/STAT5 activation in KG-1a (FLT3-WT) cells that upregulate RAS signaling which is a mechanism of acquired resistance to gilteritinib. In murine xenograft studies, HM43239 was more effective than gilteritinib in both SC and orthotopic FLT3-F691L and FLT3-ITD/F691L mutant MOLM-14 models. Summary/Conclusion: HM43239 inhibits wild type and mutant forms of FLT3 at low nM concentrations and demonstrates in vivo efficacy at doses that are well tolerated. Its ability to also inhibit SYK and, by reducing the activity of these upstream kinases, to also impair the activity of EKR1/2 and JAK/STAT5 that participate in rescue pathways, makes this a particularly interesting molecule with the potential of offsetting the development of resistance that is common with other FLT3 inhibitors. A Phase 1/2 trial of HM43239 in patients with AML is open and accruing patients (NCT03850574).

Mice with Established Diabetes Show Increased Susceptibility to Renal Ischemia/Reperfusion Injury: Protection by Blockade of Jnk or Syk Signaling Pathways

Patients with diabetes are at an increased risk for acute kidney injury (AKI) after renal ischemia/reperfusion injury (IRI). However, there is a lack preclinical models of IRI in established diabetes. The current study characterized renal IRI in mice with established diabetes and investigated potential therapies. Diabetes was induced in C57BL/6J mice by low-dose streptozotocin injection. After 7 weeks of sustained diabetes, mice underwent 13 minutes of bilateral renal ischemia and were euthanized after 24 hours of reperfusion. Age-matched, nondiabetic controls underwent the same surgical procedure. Renal IRI induced two- and sevenfold increases in plasma creatinine level in nondiabetic and diabetic mice, respectively (P<0.001). Kidney damage, as indicated by histologic damage, tubular cell death, tubular damage markers, and inflammation, was more severe in the diabetic IRI group. The diabetic IRI group showed greater accumulation of spleen tyrosine kinase (Syk)-expressing cells, and increased c-Jun N-terminal kinase (Jnk) signaling in tubules compared to nondiabetic IRI. Prophylactic treatment with a Jnk or Syk inhibitor substantially reduced the severity of AKI in the diabetic IRI model, with differential effects on neutrophil infiltration and Jnk activation. In conclusion, established diabetes predisposed mice to renal IRI-induced AKI. Two distinct proinflammatory pathways, JNK and SYK, were identified as potential therapeutic targets for anticipated AKI in patients with diabetes.

Molecular structure, expression, and the emerging role of Siglec-15 in skeletal biology and cancer.

Siglec-15, a Siglec family member and type-1 transmembrane protein, is expressed mainly in human macrophages and dendritic cells. It is comprised of a lysine-containing transmembrane domain, two extracellular immunoglobulin (Ig)-like domains and a short cytoplasmic domain. Siglec-15 is highly conserved in vertebrates and acts as an immunoreceptor. It exerts diverse functions on osteoclast physiology as well as the tumor microenvironment. Siglec-15 interacts with adapter protein DAP12 - Syk signaling pathway to regulate the RANKL/RANK-mediated PI3K, AKT, and ERK signaling pathways during osteoclast formation in vitro. Consistently, the lack of the Siglec-15 gene in mice leads to impaired osteoclast activity and osteopetrosis in vivo. In addition, Siglec-15 is expressed by tumor-associated macrophages (TAMs) and regulates the tumor microenvironment by activating the SYK/MAPK signaling pathway. Interestingly, Siglec-15 shares sequence homology to programmed death-ligand 1 (PD-L1) and has a potential immune-regulatory role in cancer immunology. Thus, Siglec-15 might also represent an alternative target for the treatment of cancers that do not respond to anti-PD-L1/PD-1 immunotherapy. Understanding the role of Siglec-15 in osteoclastogenesis and the tumor microenvironment will help us to develop new treatments for bone disorders and cancer.

NLRP3 Inflammasome Contributes to Host Defense Against Talaromyces marneffei Infection.

Talaromyce marneffei is an important thermally dimorphic pathogen causing disseminated mycoses in immunocompromised individuals in southeast Asia. Previous studies have suggested that NLRP3 inflammasome plays a critical role in antifungal immunity. However, the mechanism underlying the role of NLRP3 inflammasome activation in host defense against T. marneffei remains unclear. We show that T. marneffei yeasts but not conidia induce potent IL-1β production. The IL-1β response to T. marneffei yeasts is differently regulated in different cell types; T. marneffei yeasts alone are able to induce IL-1β production in human PBMCs and monocytes, whereas LPS priming is essential for IL-1β response to yeasts. We also find that Dectin-1/Syk signaling pathway mediates pro-IL-1β production, and NLRP3-ASC-caspase-1 inflammasome is assembled to trigger the processing of pro-IL-1β into IL-1β. In vivo, mice deficient in NLRP3 or caspase-1 exhibit higher mortality rate and fungal load compared to wild-type mice after systemic T. marneffei infection, which correlates with the diminished recruitment of CD4 T cells into granulomas in knockout mice. Thus, our study first demonstrates that NLRP3 inflammasome contributes to host defense against T. marneffei infection.

BAY61‑3606 attenuates neuroinflammation and neurofunctional damage by inhibiting microglial Mincle/Syk signaling response after traumatic brain injury.

Neuroinflammatory processes mediated by microglial activation and subsequent neuronal damage are the hallmarks of traumatic brain injury (TBI). As an inhibitor of the macrophage-inducible C-type lectin (Mincle)/spleen tyrosine kinase (Syk) signaling pathway, BAY61-3606 (BAY) has previously demonstrated anti-inflammatory effects on some pathological processes, such as acute kidney injury, by suppressing the inflammatory macrophage response. In the present study, the potential effects of BAY on microglial phenotype and neuroinflammation after TBI were investigated. BAY (3 mg/kg) was first administered into mice by intraperitoneal injection after TBI induction in vivo and microglia were also treated with BAY (2 µM) in vitro. The levels of inflammatory factors in microglia were assessed using reverse transcription-quantitative PCR and ELISA. Cortical neuron, myelin sheath, astrocyte and cerebrovascular endothelial cell markers were detected using immunofluorescence. The levels of components of the Mincle/Syk/NF-κB signaling pathway [Mincle, phosphorylated (p)-Syk and NF-κB], in addition to proteins associated with inflammation (ASC, caspase-1, TNF-α, IL-1β and IL-6), apoptosis (Bax and Bim) and tight junctions (Claudin-5), were measured via western blotting and ELISA. Migration and chemotaxis of microglial cells were evaluated using Transwell and agarose spot assays. Neurological functions of the mice were determined in vivo using the modified neurological severity scoring system and a Morris water maze. The results of the present study revealed that the expression levels of proteins in the Mincle/Syk/NF-κB signaling pathway (including Mincle, p-Syk and p-NF-κB), inflammatory cytokines (TNF-α, IL-1β and IL-6), proteins involved in inflammation (ASC and caspase-1), apoptotic markers (Bax and Bim) and the tight junction protein Claudin-5 were significantly altered post-TBI. BAY treatment reversed these effects in both the cerebral cortex extract-induced cell model and the controlled cortical impact mouse model. BAY was also revealed to suppress activation of the microglial proinflammatory phenotype and microglial migration. In addition, BAY effectively attenuated TBI-induced neurovascular unit damage and neurological function deficits. Taken together, these findings provided evidence that BAY may inhibit the Mincle/Syk/NF-κB signaling pathway in microglia; this in turn could attenuate microglia-mediated neuroinflammation and improve neurological deficits following TBI.

Engagement of TLR and Dectin-1/Syk Signaling Is Required for Activation of Notch Targets in Dendritic Cells

Abstract Pattern-recognition receptors, such as toll-like receptors (TLRs), detect a wide range of microbial products and initiate innate immune responses leading to the production of inflammatory mediators. In addition, TLR signaling also activates expression of Notch target genes that play crucial roles in suppression of TLR-triggered inflammatory responses. However, whether TLR signaling pathways engaged by other classes of pattern-recognition receptors induce expression of Notch target genes remains unclear. Here we demonstrate that zymosan, a stimulus for TLR2 and dectin-1, strongly induces expression of multiple Notch target genes in both human and murine dendritic cells. Mechanistically, induction of Notch targets by zymosan is both TLR2- and Syk-dependent through activation of mitogen-activated protein kinases and the transcription factor c-Fos. Hence, our data reveals a novel mechanism that efficient induction of Notch target genes requires engagement of TLR and dectin-1/Syk signaling pathways.

Inhibitory activity of narirutin on RBL-2H3 cells degranulation

Context: It is an efficient strategy to apply inhibition of mast cell degranulation for evaluating anti-allergic effects of compounds. Previous works confirmed that narirutin had anti–allergic activity in OVA induced allergic asthma murine model. However, the mechanism is not clear. Objective: Here, inhibitory mechanism of narirutin on RBL-2H3 cells degranulation was investigated. Materials and methods: Cell viability was analyzed by CCK-8 kits, cell degranulation was analyzed by ELISA methods, morphology and ultrastructure of cells was observed by atomic force microscopy, intracellular Ca 2+ concentration was measured by fluorescence microscopre, mRNA expression were measured by PCR, and signaling pathways were measured by WB. Results: The results showed that narirutin have no direct effects on mRNA expression of FcεRI subunit. However, it inhibited Ca2+ influx by suppressing the phosphorylation of Syk, LAT and PLCγ1 signaling pathway transduction. Subsequently, the inhibition of Ca2+ influx directly leads to NF-κB signaling pathway transduction decreased. Narirutin can also suppress the phosphorylation of MAPK signaling pathways by decreasing the expression of P-p38, P-ERK and P-JNK, inhibit the synergistic effect for Ca2+ influx, and then reduce the release of IL-4, TNF-α, histamine and β-HEX. Conclusion: Our study suggested that the inhibitory mechanism of narirutin on RBL-2H3 cells degranulation could be related to regulate MAPK, NF-κB and Tyrosine kinase signaling pathway.

Caspofungin suppresses zymosan-induced cytokine and chemokine release in THP-1 cells: possible involvement of the spleen tyrosine kinase pathway

Systemic inflammatory response syndrome and sepsis are considered to contribute to hypercytokinemia in both patients with severe infection and immunocompromised condition. Past research has demonstrated that antibiotics and antifungals not only have antimicrobial efficacy but also affect the immune system. We previously examined whether immune cells were modulated by antibiotics such as tetracyclines or macrolides. The modulation of lipopolysaccharide-stimulated cells by those agents was elucidated. However, few reports about the modulation of the immune system by antifungal agents were found. In this study, the production of pro-inflammatory cytokines and chemokines and signaling pathways involved were investigated in zymosan-activated THP-1 cells. The effects were examined using antifungal agents such as echinocandin including caspofungin (CAS) and micafungin. Pro-inflammatory cytokine and chemokine levels were determined using enzyme-linked immunosorbent assay. Protein phosphorylation was evaluated by western blot analysis. CAS significantly decreased zymosan-induced pro-inflammatory cytokine and chemokine release in THP-1 cells. CAS (30 µg/mL) also downregulated tumor necrosis factor alpha levels, as shown by enzyme-linked immunosorbent assay. In western blot analysis, inhibitor of nuclear factor-kappa-B alpha, p38, c-Jun N-terminal kinase, extracellular signal-regulated kinase, and nuclear factor of activated T-cells phosphorylation and activation of caspase-1 and spleen tyrosine kinase (Syk) were downregulated. The major underlying mechanism of pro-inflammatory cytokine and chemokine suppression by CAS is to inhibit activation of Syk and its downstream signaling molecules. Based on the results, it can be concluded that CAS activity possibly involves Syk signaling pathways and has potential to prevent hypercytokinemia in fungal sepsis.

A Phase 2 Study of the Dual SYK/JAK Inhibitor Cerdulatinib Demonstrates Good Tolerability and Clinical Response in Relapsed/Refractory Peripheral T-Cell Lymphoma and Cutaneous T-Cell Lymphoma

A Phase 2 Study of the Dual SYK/JAK Inhibitor Cerdulatinib Demonstrates Good Tolerability and Clinical Response in Relapsed/Refractory Peripheral T-Cell Lymphoma and Cutaneous T-Cell Lymphoma

Ethanol Extract of Sesamum indicum Linn. Inhibits FcεRI-Mediated Allergic Reaction via Regulation of Lyn/Syk and Fyn Signaling Pathways in Rat Basophilic Leukemic RBL-2H3 Mast Cells.

This study is aimed at determining whether Sesamum indicum Linn. beneficially influences FcεRI-mediated allergic reactions in RBL-2H3 mast cells; it is also aimed at further investigating Lyn/Fyn and Syk signaling pathways. To examine the antiallergic effect of Sesamum indicum Linn. extract (SIE), we treated antigen/immunoglobulin E- (IgE-) sensitized mast cells with extracts of various concentrations. We examined the degranulation release and concentrations of inflammatory mediators. Additionally, the expressions of genes involved in the FcεRI and arachidonate signaling pathways were examined. SIE inhibited the degranulation and secretion of inflammatory mediators in antigen/IgE-sensitized mast cells. SIE reduced the expressions of FcεRI signaling-related genes, such as Syk, Lyn, and Fyn, and the phosphorylation of extracellular signal-regulated kinase in antigen/IgE-sensitized mast cells. Additionally, in late allergic responses, SIE reduced PGD2 release and COX-2 and cPLA2 phosphorylation expression in FcεRI-mediated mast cell activation. Lastly, 250–500 mg/kg SIE significantly attenuated the Ag/IgE-induced passive cutaneous anaphylaxis (PCA) reaction in mice. The potent effect of SIE on RBL-2H3 mast cell activation indicates that the extract could potentially be used as a novel inhibitor against allergic reactions.

Classic and Novel Signaling Pathways Involved in Cancer: Targeting the NF-κB and Syk Signaling Pathways.

The nuclear factor kappa B (NF-κB) consists of a family of transcription factors involved in the regulation of a wide variety of biological responses. Growing evidence support that NF-κB plays a major role in oncogenesis as well as its well-known function in the regulation of immune responses and inflammation. Therefore, we made a review of the diverse molecular mechanisms by which the NF-κB pathway is constitutively activated in different types of human cancers and the potential role of various oncogenic genes regulated by this transcription factor in cancer development and progression. We also discussed various pharmacological approaches employed to target the deregulated NF-κB signaling pathway and their possible therapeutic potential in cancer therapy. Moreover, Syk (Spleen tyrosine kinase), non-receptor tyrosine kinase which mediates signal transduction downstream of a variety of transmembrane receptors including classical immune-receptors like the B-cell receptor (BCR), which can also activate the inflammasome and NF-κB-mediated transcription of chemokines and cytokines in the presence of pathogens would be discussed as well. The highlight of this review article is to summarize the classic and novel signaling pathways involved in NF-κB and Syk signaling and then raise some possibilities for cancer therapy.

Down-Regulation of Mir-107 Worsen Spatial Memory by Suppressing SYK Expression and Inactivating NF-ΚB Signaling Pathway.

Alzheimer's Disease (AD) is a chronic progressive neurodegenerative disorder in a central nervous system seen. We aimed to study the miR-107 in Alzheimer's Disease (AD) pathology through regulating SYK and NF-κB signaling pathway. Bioinformatics analysis was performed to screen NF-κB signaling pathway and differentially expressed genes. The target relationship between miR-107 and SYK was verified by dual luciferase assay. QRT-PCR and western blot analysis were used to verify the expression level of miR-107, SYK and NF- κB signaling pathway related proteins of hippocampus primary neurons. BAY61-3606 and BAY11-7082 were purchased for functional examination. Morris water maze tests were performed to access spatial memory of AD mice with SYK and NF-κB signaling pathway inhibition. Fluorescence microscope dyeing experiment investigated the neurons nuclear form and apoptosis. MiR-107 was lowly expressed while SYK was highly expressed in Tg19959 mouse model. Luciferase Assay confirmed the target relationship in miR-107 and SYK. With the inhibition of miR-107, SYK was up-regulated and the increase of p-p65 and the decrease of p-IκB-α suggested that NF-κB signaling pathway was activated in vitro. Morris water maze test indicated that the spatial memory of Tg19959 mice was increased with the treatment. The result of DAPI staining indicated that the inhibition of SYK or NF-κB signaling pathway reduced the apoptosis of Tg19959 mice neuron cell. MiR-107 exerts its effects through suppression of the NF-κB signaling pathway and SYK, the inhibition of SYK and NF-κB signaling pathway can improve spatial memory and suppress cell apoptosis.

The Novel SYK/JAK Inhibitor Cerdulatinib Demonstrates Good Tolerability and Clinical Response in a Phase 2a Study in Relapsed/Refractory Peripheral T-Cell Lymphoma and Cutaneous T-Cell Lymphoma

The Novel SYK/JAK Inhibitor Cerdulatinib Demonstrates Good Tolerability and Clinical Response in a Phase 2a Study in Relapsed/Refractory Peripheral T-Cell Lymphoma and Cutaneous T-Cell Lymphoma

Syk and JNK signaling pathways are involved in inflammasome activation in macrophages infected with Streptococcus pneumoniae

Streptococcus pneumoniae is a pathogen of significant clinical importance worldwide that can cause severe invasive diseases, such as pneumonia, otitis media and meningitis. Inflammsomes has been reported to participate in host defense against S. pneumoniae infection. S. pneumoniae could induce the assembly of the nucleotide-binding oligomerization domain-like receptor family pyrin domain containing 3 (NLRP3)/absent in melanoma 2 (AIM2) inflammasome, which mediates the activation of caspase-1 and the subsequent maturation of Interleukin-1β (IL-1β). However, the precise signals that activate inflammasomes during pneumococcal infection remain to be fully elucidated. In the present study, primary mouse macrophages were selected as a cell model, and the effects of kinases on inflammasome activity induced by S. pneumoniae infection were examined by ELISA and western blotting after pretreatment with a kinase inhibitor. Here, we show that Syk and JNK signaling are required for S. pneumoniae-induced activation of the inflammasome. Inhibitors of Syk and JNK almost abolished the oligomerization of apoptosis-associated speck-like protein containing a caspase-activating and recruitment domain (ASC) and subsequent caspase-1 activation and IL-1β secretion. Moreover, pneumolysin (PLY) participated in this process and was critical for Syk/JNK activation. These results suggested that the Syk/JNK signaling pathway may play a vital role in the inflammasome activation and modulate host immune responses against S. pneumoniae.

Spinacetin Suppresses the Mast Cell Activation and Passive Cutaneous Anaphylaxis in Mouse Model.

We previously reported the anti-inflammatory and anti-asthmatic activities of the extract of the Inula japonica Thunb. Aiming for discovery of a novel anti-inflammatory compound, we isolated spinacetin from the extract and investigated its in vitro and in vivo anti-inflammatory effect and the related mechanism. Effect of spinacetin on the Syk signaling pathway was studied in bone marrow-derived mast cells (BMMCs), and that on the nuclear factor-κB (NF-κB) and mitogen-activated protein kinases (MAPKs) was investigated in Rat basophilic leukemia (RBL)-2H3 cells and human mast cell line (HMC-1). The in vivo anti-inflammatory activity was assessed with passive cutaneous anaphylaxis (PCA) reaction assay. Spinacetin significantly inhibited the release of histamine, and production of inflammatory mediators such as leukotriene C4 (LTC4) and interlukin-6 (IL-6) in IgE/Ag stimulated BMMCs. Analysis of the signaling pathways demonstrated that spinacetin inhibited activation of Syk, linker of activated T cells (LAT), phospholipase Cγ (PLCγ), cytosolic phospholipase A2 (cPLA2), MAPKs, Akt/NF-κB, and intracellular Ca2+ mobilization but with no effect on Fyn and Lyn. On the other hand, spinacetin suppressed IgE/Ag-induced activation of RBL-2H3 cells with inhibition against phosphorylation of extracellular signal regulated-protein kinase (ERK), c-Jun-NH2-terminal kinase (JNK), p38 MAPKs, PLCγ, translocation of cPLA2, and Akt/IκBα/NF-κB signal. However, spinacetin had no effect on PMA and A23187-induced activation of HMC-1. Furthermore, oral administration of spinacetin dose-dependently attenuated IgE/Ag-mediated PCA reaction in mouse model. Taken together, spinacetin showed the activities in preventing inflammatory processes, which might be at least partially attributed to the abolishment of Syk-dependent activation of IgE/Ag-mediated mast cells.

Abstract 2371: Entospletinib, a potent SYK inhibitor, blocks constitutive and FCgRI activated signaling in FLT3-ITD cell lines

Abstract Spleen tyrosine kinase (SYK), a non-receptor protein tyrosine kinase, is an important mediator of immune receptor signaling in B cells, mast cells, neutrophils, and macrophages. SYK is expressed in 90% of AML blasts and has been reported to be is constitutively activated (Hahn et al., Cancer Cell 2009). High expression of phosphorylated SYK (pSYK) is associated with unfavorable outcomes, independent of other AML prognostic predictors such as age, cytogenetics, and white blood cell count (Boros et al., Oncotarget 2015). Entospletinib (ENTO) is a potent and selective inhibitor of SYK currently in clinical trials for the treatment of AML as monotherapy and in combination with standard-of-care therapies. We investigated the effects of ENTO on SYK phosphorylation and signaling in FLT3-ITD mutant AML cells. We found that ENTO blocked constitutive phosphorylation of pSYKY352, pSTAT5Y694, and MYC protein expression in FLT3-ITD-driven AML cell lines, MV4-11 and MOLM14. The concentration of ENTO required to inhibit pSYKY352 and pSTAT5Y694 correlated with inhibition of cell viability. Phosphorylation at these sites was inhibited by the selective FLT3 inhibitor, quizartinib; but not inhibited by Src family kinase inhibitor (dasatinib) or JAK inhibitors (ruxolitinib or tofacitinib). Further, a second-generation, highly selective SYK inhibitor, GS-9876, failed to inhibit the constitutive phosphorylation of these sites. Comparison of the biochemical and cellular kinase activities of the SYK and FLT3 inhibitors suggests that the observed inhibition of constitutive phosphorylation of pSYKY352 and pSTAT5Y694 and MYC expression was mediated by the direct activity of ENTO against FLT3-ITD, rather than SYK. Activation of FcγRI has been previously shown to activate SYK (Oellerich et al., Blood 2013). We demonstrate SYK activation at pSYKY348 and pSYKY323 after stimulation of the FCγRI with human IgG in the FLT3-ITD mutant cell line, MV4-11. Downstream signaling molecules pERK1/2T202/Y204, pVAV1Y160, and pLYNY507 were also activated by FCγR stimulation. ENTO inhibited phosphorylation of pSYKY348, pSYKY323 and pVAV1 Y160. The second-generation SYK inhibitor, GS-9876, also inhibited FCγRI-mediated activation of pSYKY348,pSYKY323 and pVAV1 Y160, suggesting that these phosphorylation events were mediated by SYK activity. In conclusion, ENTO inhibited constitutive and FCγRI-activated SYK phosphorylation and signaling pathways in FLT3-ITD mutant cell lines, resulting in inhibition of cell viability. Inhibition of constitutive SYK phosphorylation is mediated by direct activity of ENTO against FLT3-ITD; however, inhibition of FCγRI activated SYK results from direct inhibition of SYK. Citation Format: Stacey L. Tannheimer, Adam Kashishian, Rick Sorensen, Kathleen S. Keegan. Entospletinib, a potent SYK inhibitor, blocks constitutive and FCgRI activated signaling in FLT3-ITD cell lines [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2371.

Role of Hepatic Deposited Immunoglobulin G in the Pathogenesis of Liver Damage in Systemic Lupus Erythematosus.

The onset of hepatic disorders in patients with systemic lupus erythematosus (SLE) is frequent; however, the etiology and liver pathogenesis of SLE remain unknown. In the present study, the role of hepatic deposited immunoglobulin G (IgG) in SLE-derived liver damage was investigated. From a retrospective analysis of the medical records of 404 patients with lupus and from experimental studies on mice models, we found that liver dysfunction is common in SLE and liver damage with IgG deposition spontaneously develops in lupus-prone mice. Liver injury was recreated in mice by injecting IgG from lupus serum intrahepatically. The inflammation intensity in the liver decreased with IgG depletion and the lupus IgG-induced liver inflammation in FcγRIII-deficient mice was comparatively low; while, inflammation was increased in FcγRIIb-deficient mice. Macrophages, Kupffer cells, natural killer cells, and their products, but not lymphocytes, are required for the initiation of SLE-associated liver inflammation. Blocking IgG signaling using a spleen tyrosine kinase (Syk) inhibitor suppressed the liver damage. Our findings provided evidence of spontaneously established liver damage in SLE. They also suggested that hepatic-deposited lupus IgG is an important pathological factor in the development of liver injury and that hepatic inflammation is regulated by the Syk signaling pathway. Thus, Syk inhibition might promote the development of a therapeutic strategy to control liver damage in patients with SLE.