Apigenin Increases SHIP-1 Expression, Promotes Tumoricidal Macrophages and Anti-Tumor Immune Responses in Murine Pancreatic Cancer

Pancreatic cancer (PC) has an extremely poor prognosis due to the expansion of immunosuppressive myeloid-derived suppressor cells (MDSC) and tumor-associated macrophages (TAM) in the inflammatory tumor microenvironment (TME), which halts the recruitment of effector immune cells and renders immunotherapy ineffective. Thus, the identification of new molecular targets that can modulate the immunosuppressive TME is warranted for PC intervention. Src Homology-2 (SH2) domain-containing Inositol 5′-Phosphatase-1 (SHIP-1) is a lipid signaling protein and a regulator of myeloid cell development and function. Herein, we used the bioflavonoid apigenin (API) to reduce inflammation in different PC models. Wild type mice harboring heterotopic or orthotopic PC were treated with API, which induced SHIP-1 expression, reduced inflammatory tumor-derived factors (TDF), increased the proportion of tumoricidal macrophages and enhanced anti-tumor immune responses, resulting in a reduction in tumor burden compared to vehicle-treated PC mice. In contrast, SHIP-1-deficient mice exhibited an increased tumor burden and displayed augmented proportions of pro-tumor macrophages. These results provide further support for the importance of SHIP-1 expression in promoting pro-tumor macrophage development in the pancreatic TME. Our findings suggest that agents augmenting SHIP-1 expression may provide novel therapeutic options for the treatment of PC.

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Systemic but not MDSC-specific IRF4 deficiency promotes an immunosuppressed tumor microenvironment in a murine pancreatic cancer model

Cancer Immunology Immunotherapy ◽

10.1007/s00262-020-02605-9 ◽

2020 ◽

Vol 69 (10) ◽

pp. 2101-2112

Author(s):

Philipp Metzger ◽

Sabrina V. Kirchleitner ◽

Daniel F. R. Boehmer ◽

Christine Hörth ◽

Angelika Eisele ◽

...

Keyword(s):

Pancreatic Cancer ◽

Tumor Microenvironment ◽

Critical Role ◽

Suppressor Cells ◽

Myeloid Cell ◽

Ductal Adenocarcinoma ◽

Gm Csf ◽

Immunosuppressive Tumor Microenvironment ◽

And Function

Abstract Pancreatic ductal adenocarcinoma is characterized by a strong immunosuppressive network with a dense infiltration of myeloid cells including myeloid-derived suppressor cells (MDSC). Two distinct populations of MDSC have been defined: polymorphonuclear MDSC (PMN-MDSC) and monocytic MDSC (M-MDSC). Several factors influence the development and function of MDSC including the transcription factor interferon regulatory factor 4 (IRF4). Here, we show that IRF4 deficiency accelerates tumor growth and reduces survival, accompanied with a dense tumor infiltration with PMN-MDSC and reduced numbers of CD8+ T cells. As IRF4 has been described to modulate myeloid cell development and function, particularly of PMN-MDSC, we analyzed its role using MDSC-specific IRF4 knockout mice with the Ly6G or LysM knock-in allele expressing Cre recombinase and Irf4flox. In GM-CSF-driven bone marrow cultures, IRF4 deficiency increased the frequency of MDSC-like cells with a strong T cell suppressive capacity. Myeloid (LysM)-specific depletion of IRF4 led to increased tumor weight and a moderate splenic M-MDSC expansion in tumor-bearing mice. PMN cell (Ly6G)-specific depletion of IRF4, however, did not influence tumor progression or MDSC accumulation in vivo in accordance with our finding that IRF4 is not expressed in PMN-MDSC. This study demonstrates a critical role of IRF4 in the generation of an immunosuppressive tumor microenvironment in pancreatic cancer, which is independent of IRF4 expression in PMN-MDSC.

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Design Potential Selective Inhibitors for Treating Cancer by Targeting the Src Homology 2 (SH2) Domain-Containing Phosphatase 2 (Shp2) with Core Hopping Approach

Protein and Peptide Letters ◽

10.2174/0929866521666131223143913 ◽

2014 ◽

Vol 21 (6) ◽

pp. 556-563 ◽

Cited By ~ 10

Author(s):

Yu-Qing Duan ◽

Ying Ma ◽

Xue-Jiao Wang ◽

Yuan-Yuan Jin ◽

Run-Ling Wang ◽

...

Keyword(s):

Sh2 Domain ◽

Selective Inhibitors ◽

Src Homology 2 ◽

Src Homology

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PLC-γ directly binds activated c-Src, which is necessary for carbachol-mediated inhibition of NHE3 activity in Caco-2/BBe cells

AJP Cell Physiology ◽

10.1152/ajpcell.00277.2012 ◽

2013 ◽

Vol 305 (3) ◽

pp. C266-C275 ◽

Cited By ~ 5

Author(s):

Nicholas C. Zachos ◽

Luke J. Lee ◽

Olga Kovbasnjuk ◽

Xuhang Li ◽

Mark Donowitz

Keyword(s):

Sh2 Domain ◽

Signaling Proteins ◽

Multiprotein Complexes ◽

Sh2 Domains ◽

Src Homology 2 ◽

Src Inhibitor ◽

Intracellular Ca ◽

Direct Binding ◽

Src Homology

Elevated levels of intracellular Ca2+([Ca2+]i) inhibit Na+/H+exchanger 3 (NHE3) activity in the intact intestine. We previously demonstrated that PLC-γ directly binds NHE3, an interaction that is necessary for [Ca2+]iinhibition of NHE3 activity, and that PLC-γ Src homology 2 (SH2) domains may scaffold Ca2+signaling proteins necessary for regulation of NHE3 activity. [Ca2+]iregulation of NHE3 activity is also c-Src dependent; however, the mechanism by which c-Src is involved is undetermined. We hypothesized that the SH2 domains of PLC-γ might link c-Src to NHE3-containing complexes to mediate [Ca2+]iinhibition of NHE3 activity. In Caco-2/BBe cells, carbachol (CCh) decreased NHE3 activity by ∼40%, an effect abolished with the c-Src inhibitor PP2. CCh treatment increased the amount of active c-Src as early as 1 min through increased Y416phosphorylation. Coimmunoprecipitation demonstrated that c-Src associated with PLC-γ, but not NHE3, under basal conditions, an interaction that increased rapidly after CCh treatment and occurred before the dissociation of PLC-γ and NHE3 that occurred 10 min after CCh treatment. Finally, direct binding to c-Src only occurred through the PLC-γ SH2 domains, an interaction that was prevented by blocking the PLC-γ SH2 domain. This study demonstrated that c-Src 1) activity is necessary for [Ca2+]iinhibition of NHE3 activity, 2) activation occurs rapidly (∼1 min) after CCh treatment, 3) directly binds PLC-γ SH2 domains and associates dynamically with PLC-γ under elevated [Ca2+]iconditions, and 4) does not directly bind NHE3. Under elevated [Ca2+]iconditions, PLC-γ scaffolds c-Src into NHE3-containing multiprotein complexes before dissociation of PLC-γ from NHE3 and subsequent endocytosis of NHE3.

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Synthesis and in Vitro Evaluation of a Peptidomimetic Inhibitor Targeting the Src Homology 2 (SH2) Domain of STAT6

ACS Medicinal Chemistry Letters ◽

10.1021/ml4003919 ◽

2013 ◽

Vol 5 (1) ◽

pp. 69-72 ◽

Cited By ~ 15

Author(s):

Pietro Morlacchi ◽

Pijus K. Mandal ◽

John S. McMurray

Keyword(s):

Sh2 Domain ◽

In Vitro Evaluation ◽

Src Homology 2 ◽

Src Homology

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Enhanced Prediction of Src Homology 2 (SH2) Domain Binding Potentials Using a Fluorescence Polarization-derived c-Met, c-Kit, ErbB, and Androgen Receptor Interactome

Molecular & Cellular Proteomics ◽

10.1074/mcp.m113.034876 ◽

2014 ◽

Vol 13 (7) ◽

pp. 1705-1723 ◽

Cited By ~ 9

Author(s):

Kin K. Leung ◽

Ronald J. Hause ◽

John L. Barkinge ◽

Mark F. Ciaccio ◽

Chih-Pin Chuu ◽

...

Keyword(s):

Androgen Receptor ◽

Fluorescence Polarization ◽

Sh2 Domain ◽

Src Homology 2 ◽

Src Homology

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Evidence for a molecular complex consisting of Fyb/SLAP, SLP-76, Nck, VASP and WASP that links the actin cytoskeleton to Fcγ receptor signalling during phagocytosis

Journal of Cell Science ◽

10.1242/jcs.114.23.4307 ◽

2001 ◽

Vol 114 (23) ◽

pp. 4307-4318

Author(s):

Marc G. Coppolino ◽

Matthias Krause ◽

Petra Hagendorff ◽

David A. Monner ◽

William Trimble ◽

...

Keyword(s):

Actin Cytoskeleton ◽

Molecular Complex ◽

Actin Filaments ◽

Sh2 Domain ◽

Fcγ Receptors ◽

Receptor Signalling ◽

Src Homology 2 ◽

Src Homology ◽

Particle Ingestion ◽

Syndrome Protein

Phagocytosis by macrophages and neutrophils involves the spatial and temporal reorganisation of the actin-based cytoskeleton at sites of particle ingestion. Local polymerisation of actin filaments supports the protrusion of pseudopodia that eventually engulf the particle. Here we have investigated in detail the cytoskeletal events initiated upon engagement of Fc receptors in macrophages. Ena/vasodilator-stimulated phosphoprotein (VASP) proteins were recruited to phagosomes forming around opsonised particles in both primary and immortalised macrophages. Not only did the localisation of Ena/VASP proteins coincide, spatially and temporally, with the phagocytosis-induced reorganisation of actin filaments, but their recruitment to the phagocytic cup was required for the remodelling of the actin cytoskeleton, extension of pseudopodia and efficient particle internalisation. We also report that SLP-76, Vav and profilin were recruited to forming phagosomes. Upon induction of phagocytosis, a large molecular complex, consisting in part of Ena/VASP proteins, the Fyn-binding/SLP-76-associated protein (Fyb/SLAP), Src-homology-2 (SH2)-domain-containing leukocyte protein of 76 kDa (SLP-76), Nck, and the Wiskott-Aldrich syndrome protein (WASP), was formed. Our findings suggest that activation of Fcγ receptors triggers two signalling events during phagocytosis: one through Fyb/SLAP that leads to recruitment of VASP and profilin; and another through Nck that promotes the recruitment of WASP. These converge to regulate actin polymerisation, controlling the assembly of actin structures that are essential for the process of phagocytosis.

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Src Homology 2 (SH2)-Containing 5′-Inositol Phosphatase Localizes to Podosomes, and the SH2 Domain Is Implicated in the Attenuation of Bone Resorption in Osteoclasts

Endocrinology ◽

10.1210/en.2005-1309 ◽

2006 ◽

Vol 147 (7) ◽

pp. 3307-3317 ◽

Cited By ~ 19

Author(s):

Keiichiro Yogo ◽

Megumi Mizutamari ◽

Kazuta Mishima ◽

Hiromi Takenouchi ◽

Norihiro Ishida-Kitagawa ◽

...

Keyword(s):

Bone Resorption ◽

Nuclear Factor Κb ◽

Sh2 Domain ◽

Terminal Region ◽

Spectrometric Analysis ◽

Inositol Phosphatase ◽

Amino Terminal ◽

Novel Proteins ◽

Src Homology 2 ◽

Src Homology

c-Src plays an important role in bone resorption by osteoclasts. Here, we show using wild-type and ship−/− osteoclasts that Src homology 2 (SH2)-containing 5′-inositol phosphatase (SHIP) appeared to negatively regulate bone resorption activated by c-Src. SHIP was found to localize to podosomes under the influence of c-Src, and the presence of either the amino-terminal region comprising the SH2 domain or the carboxyl-terminal region was sufficient for its localization. Although SHIP lacking a functional SH2 domain was still found in podosomes, it could not rescue the hyper-bone resorbing activity and hypersensitivity to receptor activator of nuclear factor-κB ligand in ship−/− osteoclasts, suggesting that the localization of SHIP to podosomes per se was not sufficient and the SH2 domain was indispensable for its function. Cas and c-Cbl, known to function in podosomes of osteoclasts, were identified as novel proteins binding to the SHIP SH2 domain by mass spectrometric analysis, and this interaction appeared to be dependent on the Src kinase activity. These results demonstrate that c-Src enhances the translocation of SHIP to podosomes and regulates its function there through the SH2 domain, leading to an attenuation of bone resorption.

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Triazolopyrimidinium salts: discovery of a new class of agents for cancer therapy

Future Medicinal Chemistry ◽

10.4155/fmc-2019-0317 ◽

2020 ◽

Vol 12 (5) ◽

pp. 387-402

Author(s):

Mariateresa Badolato ◽

Fabrizio Manetti ◽

Antonio Garofalo ◽

Francesca Aiello

Keyword(s):

Anticancer Agents ◽

Docking Studies ◽

In Vitro Models ◽

Sh2 Domain ◽

Activity Profile ◽

Cancer Cell Growth ◽

New Class ◽

Src Homology 2 ◽

Src Homology

Aim: The [1,2,4]triazolo[1,5- a]pyrimidine core is highly privileged in medicinal chemistry due to its versatile pharmacological activity profile. Recently, the search for novel anticancer agents has focused on [1,2,4]triazolo[1,5- a]pyrimidine derivatives. Results: Our hit functionalization has led to the discovery of new [1,2,4]triazolo[1,5- a]pyrimidinium salts with potential anticancer activity. Among a small library of molecules, compound 9 significantly inhibits cancer cell growth in a panel of in vitro models. Molecular docking studies and preliminary binding assay have displayed that 9 could directly bind the Src homology 2 (SH2) domain of STAT3 protein. Conclusion: Compound 9 is a novel promising lead compound that motivates additional evaluation of [1,2,4]triazolo[1,5- a]pyrimidinium salts as novel potential chemotherapeutics.

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