Therapeutic effects of green tea involve an inhibitory function of its constituent polyphenol epigallocatechin gallate (EGCG) on cell signaling. The specificity and mechanism(s) by which EGCG ...inhibits cell signaling have remained unclear. Here, we demonstrate that green tea and EGCG induce suppressor of cytokine signaling 1 (SOCS1) gene expression, a negative regulator of specific cell signaling pathways. In mouse immune cells, EGCG induces SOCS1 expression via an oxidative (superoxide) pathway and activation of the signal transducer and activator of transcription 5 transcription factor. EGCG inhibited SOCS1-regulated cell signaling, but this inhibitory effect was abrogated in cells deficient in SOCS1. These findings identify a mechanism by which EGCG inhibits cell signaling with specificity, mediated by induction of the negative regulator SOCS1.
Abstract
Objectives : Previously, we demonstrated anti-cancer effect of our newly developed monoclonal antibody (mAb) targeting lipolysis-stimulated lipoprotein receptor (LSR) in LSR-positive ...epithelial ovarian cancer (EOC) in vitro and in vivo. In EOC treatment, platinum agents which induce apoptosis are the key drugs over the decades, therefore, we aimed to demonstrate the synergistic effect of anti-LSR mAb and platinum agent against LSR-positive EOC cells.
Methods : We administrated anti-LSR mAb and carboplatin to RMG-I cells (platinum resistant and LSR-positive EOC cell line) and evaluated cell proliferation in vitro and in vivo. We also investigated whether anti-LSR mAb reverse platinum resistance of RMG-I in vitro. In addition, we analyzed changes in protein expression related to MAPK/ERK and apoptosis using western blotting after treatment.
Results : In RMG-I, anti-LSR mAb inhibited phosphorylation of MAPK pathway-related proteins including MEK and ERK, and activated apoptosis-related proteins. Although RMG-I showed the refractory for carboplatin, anti-LSR mAb reversed its platinum resistance and decreased IC50 value of carboplatin. As a result, anti-LSR mAb induced the apoptotic effect of carboplatin in RMG-I. Finally, the combination therapy of anti-LSR mAb and carboplatin significantly inhibited the tumor growth of RMG-I in a synergistic manner in vitro and in vivo (p<0.05). The combination therapy caused strong apoptotic changes in tumor.
Conclusions : We confirmed the synergistic effect of the combination therapy of anti-LSR mAb and platinum agent against LSR-positive EOC cells via apoptotic pathway. Our findings demonstrated that this combination therapy might be a new treatment option of EOC.
Citation Format: Masashi Funauchi, Kosuke Hiramatsu, Satoshi Serada, Minoru Fujimoto, Tetsuji Naka. Anti-LSR mAb with carboplatin shows the synergistic effect in epithelial ovarian cancer abstract. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1211.
Leucine-rich α-2 glycoprotein (LRG), one of the acute phase proteins mainly produced by the liver, similar to C-reactive protein, has been recognized as an inflammatory biomarker for rheumatoid ...arthritis and inflammatory bowel diseases. We recently demonstrated that LRG was also increased in the sera of psoriasis patients and correlated well with disease activity with a sensitivity and specificity much higher than C-reactive protein; however, whether LRG mechanistically contributed to the pathogenesis of psoriasis remained unclear. In this study, we explored the role of LRG in psoriasiform inflammation using LRG-knockout (KO) mice in an imiquimod (IMQ)-mediated model. Following topical treatment with IMQ, serum levels of LRG and its expression in the liver were abruptly elevated. Similarly, an acute surge of proinflammatory cytokines was observed in the liver, including IL-1β, TNF-α, and IL-6, although LRG-KO mice showed delayed responses. LRG-KO mice showed less skin inflammation in the IMQ model than wild-type mice. K5.Stat3C mice developed psoriasis-like lesions following tape stripping, which also abruptly induced LRG expression in the liver. A deficiency of
mitigated tape stripping-induced lesions, similar to the IMQ model. These results indicate that LRG modulates both feed-forward and feedback loops of cytokines in the skin-liver axis involved with psoriasiform inflammation.
Recent evidence suggests that renal tubular injury plays a key role in deterioration of renal function in both chronic kidney disease (CKD) and acute kidney injury (AKI). Since commonly used ...biochemical indicators such as GFR, serum creatinine, blood urea nitrogen and creatinine clearance are inappropriate for detecting alteration in renal tubules, biomarkers reflecting renal tubular injury have been extensively explored.
Our research group identified leucine rich α-2 glycoprotein (LRG) as a novel serum biomarker for various inflammatory diseases such as rheumatoid arthritis and inflammatory bowel disease. In inflammatory diseases, LRG expression is up-regulated at the site of inflammation, in accordance with the induction of LRG in many cell types by various inflammatory cytokines. Recently, urinary LRG was reported as a possible biomarker for several renal diseases, but the mechanism of LRG excretion in urine is still unclear. In this study, by analyzing a mouse albumin (ALB) overload model that is commonly used to study proteinuria-induced renal tubular injury, we provided evidence that urinary LRG is produced in renal tubular epithelial cells by interleukin-1β (IL-1β) that is released during proteinuria-induced renal damage. In this model, urinary LRG became detectable after ALB overload. In kidney, mRNA expression of LRG together with that of NACHT LRR and PYD domains-containing protein 3 (NLRP3) and IL-1β was significantly up-regulated in ALB-overloaded mice, compared to PBS-treated mice. By pathological analysis of kidney, LRG was detected in the injured proximal tubules, distal tubules and collecting ducts in ALB-overloaded mice. Accordingly, in vitro stimulation of mouse renal cortical tubular epithelial cells with excessive ALB led to LRG mRNA up-regulation and its protein secretion, which was effectively blocked by IL-1 receptor antagonist. These results suggest that urinary LRG could be applied to a biomarker detecting renal tubular injury in various renal diseases.
•Urinary LRG, together with LRG mRNA expression in kidney is increased in albumin-overloaded mice.•LRG protein localizes to injured proximal tubules, distal tubules and collecting ducts in albumin-overloaded mice.•In mouse renal cortical tubular epithelial cells, albumin-induced IL-1β is critical for LRG production.•This study is helpful to develop new diagnostic agents for renal tubular injury.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UL, UM, UPCLJ, UPUK, ZRSKP
Lipolysis‐stimulated lipoprotein receptor (LSR) is known as a lipoprotein receptor. LSR is expressed in various solid tumors, including epithelial ovarian, gastric, and colon cancers. High LSR ...expression is significantly associated with poor prognosis, but its role in cancer has not been fully elucidated. LSR belongs to the Ig protein superfamily, which is conserved in B7 family. Here, we assessed LSR as a novel immune checkpoint molecule. We developed a novel anti‐LSR antibody (#27‐6 mF‐18) that defects antibody‐dependent cellular cytotoxicity and complement‐dependent cytotoxicity activity. The #27‐6 mF‐18 cross‐reacts with both human and mouse LSR. We found that LSR was expressed on 4T1 murine breast cancer cell line. The #27‐6 mF‐18 exhibited antitumor effects against the 4T1 syngeneic tumor model, a poor immunogenic model refractory to treatment with anti‐PD‐1 or anti‐CTLA‐4 antibodies. Compared with control antibody‐treated mice, mice treated with #27‐6 mF‐18 showed significantly increased numbers of CD8+ T cells and a ratio of activated CD8+ T cells infiltrated in the tumor tissue. This antitumor effect was abrogated by CD8+ T‐cell depletion through anti‐CD8 antibody treatment, indicating that LSR negatively regulates tumor immunity by repressing CD8+ T cells. These findings show that LSR negatively regulates T‐cell immune activity. LSR targeting could provide immune checkpoint inhibitors for cancer immunotherapy.
What's new?
Lipolysis‐stimulated lipoprotein receptor (LSR), a member of the Ig protein superfamily, is highly expressed in solid tumors. Nonetheless, its role in cancer remains largely uncharacterized. Here, the authors explored the relationship between LSR and tumor growth in an LSR‐positive 4T1 syngeneic tumor model, using anti‐LSR monoclonal antibodies. Antibody‐treated LSR‐positive 4T1 mice had increased CD8+ T‐cell levels and elevated CD8+ T‐cell infiltration in tumor tissue. Anti‐LSR antibody treatment significantly inhibited tumor growth, an effect that was abolished upon CD8+ T‐cell depletion. The findings suggest that anti‐LSR antibodies could be leveraged therapeutically to enhance tumor immunity in LSR‐positive tumors.
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BFBNIB, FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SBCE, SBMB, UL, UM, UPUK
Abstract
Cholangiocarcinoma (CCA) is one of highly malignant cancers. Systemic chemotherapy is used in many patients, however, there are few chemotherapy options, and their prognosis are poor. ...Therefore, it is necessary to develop new treatments. We have identified glypican-1 (GPC1) as a novel cancer antigen and reported that enhanced expression of GPC1 in esophageal squamous cell carcinoma and pancreatic cancer is significantly associated with poorer prognosis (Hara, Naka, et al. Br J Cancer. 2016, Nishigaki, Naka, et al. Br J Cancer. 2020). GPC1 is a heparan sulfate proteoglycan that is linked to the cell surface by a glycosylphosphatidylinositol anchor and promotes tumor growth, metastasis, and invasion by acting as a coreceptor, enhancing various signaling pathways. We recently identified that the expression of GPC1 was increased in cholangiocarcinoma. The present study aimed to develop a new therapy for cholangiocarcinoma using an antibody-drug conjugate (ADC) targeting glypican-1 (GPC1). Expression of GPC1 was evaluated in resected cholangiocarcinoma specimens and cell lines. GPC1 knockout cell was established from GPC1-positive cholangiocarcinoma cell line. The antitumor effect of monomethyl auristatin F conjugated GPC1-ADC was investigated in vitro and in vivo. GPC1 was highly expressed in cholangiocarcinoma cells and tissues. Immunohistochemical analysis of 49 extrahepatic cholangiocarcinoma patients revealed that 23 patients (47%) were high expression of GPC1, 24 patients (49%) were low expression of GPC1 and 2 patients (4%) were GPC1 negative. High-expression group demonstrated significantly poorer prognosis compared with the low-expression group in terms of disease-free survival and overall survival (p < 0.05). GPC1 was also expressed in tumor vessels of cholangiocarcinoma, but not on the vessels of nontumor tissues. Monomethyl auristatin F-conjugated GPC1-ADC significantly inhibited tumor growth against GPC1-positive cholangiocarcinoma cells in vitro and in vivo. In a GPC1 knockout xenograft model, GPC1-ADC partially inhibited tumor growth. Vascular endothelial cells in tumor tissues of GPC1-negative xenograft mice expressed GPC1 and were arrested in the G2/M phase of cell cycle by GPC1-ADC. In the present study, GPC1-ADC inhibits tumor growth and tumor angiogenesis for glypican-1 positive cholangiocarcinoma. Our preclinical data demonstrated GPC1-ADC as a promising strategy for GPC1-positive cholangiocarcinoma.
Citation Format: Keiichiro Yokota, Satoshi Serada, Shigehiro Tsujii, Ichiro Murakami, Kazuhiro Hanazaki, Tetsuji Naka. Antibody-drug conjugate targeting glypican-1 inhibits tumor growth and tumor angiogenesis for glypican-1 positive cholangiocarcinoma abstract. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1315.
Background:
Reliable biomarkers for monitoring disease activity have not been clinically established in ulcerative colitis (UC). This study aimed to investigate whether levels of serum leucine‐rich ...alpha‐2 glycoprotein (LRG), identified recently as a potential disease activity marker in Crohn's disease and rheumatoid arthritis, correlate with disease activity in UC.
Methods:
Serum LRG concentrations were determined by enzyme‐linked immunosorbent assay (ELISA) in patients with UC and healthy controls (HC) and were evaluated for correlation with disease activity. Expression of LRG in inflamed colonic tissues from patients with UC was analyzed by western blotting and immunohistochemistry. Interleukin (IL)‐6‐independent induction of LRG was investigated using IL‐6‐deficient mice by lipopolysaccharide (LPS)‐mediated acute inflammation and dextran sodium sulfate (DSS)‐induced colitis.
Results:
Serum LRG concentrations were significantly elevated in active UC patients compared with patients in remission (P < 0.0001) and HC (P < 0.0001) and were correlated with disease activity in UC better than C‐reactive protein (CRP). Expression of LRG was increased in inflamed colonic tissues in UC. Tumor necrosis factor alpha (TNF‐α), IL‐6, and IL‐22, serum levels of which were elevated in patients with active UC, could induce LRG expression in COLO205 cells. Serum LRG levels were increased in IL‐6‐deficient mice with LPS‐mediated acute inflammation and DSS‐induced colitis.
Conclusions:
Serum LRG concentrations correlate well with disease activity in UC. LRG induction is robust in inflamed colons and is likely to involve an IL‐6‐independent pathway. Serum LRG is thus a novel serum biomarker for monitoring disease activity in UC and is a promising surrogate for CRP. (Inflamm Bowel Dis 2012;)
Background
Constitutive activation of STAT3 promotes oncogenesis and growth of oral squamous cell carcinoma (OSCC). We investigated the mechanism of action of suppressor of cytokine signaling 1 ...(SOCS1), an endogenous inhibitor of JAK, as gene therapy for OSCC.
Methods
Antitumor effect of SOCS1 was compared to JAK inhibitor I by cell proliferation assay, cell cycle analysis, and apoptosis analysis in vitro. In addition, antitumor effect was evaluated using xenograft mouse models in vivo.
Results
JAK inhibitor I inhibited the proliferation of KOSC2 cl3‐43 or T3M‐1 clone2 OSCC cell lines in vitro. While JAK inhibitor I arrested both cell lines at the G2/M phase, induction of apoptosis was observed in T3M‐1 clone2 cells, but not KOSC2‐cl3‐43 cells. An adenoviral vector expressing SOCS1 (AdSOCS1) significantly decreased the proliferation of both OSCC cell lines and induced G2/M phase cell cycle arrest and apoptosis, suggesting that induction of apoptosis of KOSC2 cl3‐43 cells by AdSOCS1 is regulated by the JAK/STAT independent pathway. Overexpression of SOCS1 inhibited activation of the JAK/STAT and p44/42 MAPK pathways, while JAK inhibitor I inhibited activation of the JAK/STAT pathway only. Consistently, expression of Mcl‐1 was decreased by overexpression of SOCS1, but not JAK inhibitor I. Additionally, KOSC2 cl3‐43 or T3M‐1 clone2 OSCC cells were subcutaneously implanted in the flanks of two xenograft mouse models. As compared to a control adenovirus vector (AdLacZ), intratumor injection of AdSOCS1 significantly decreased the tumor volume and induced apoptosis in vivo.
Conclusion
SOCS1 gene therapy may be a beneficial approach for the treatment of OSCC.
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CMK, FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SBCE, SBMB, UL, UM, UPUK
Plasmodium falciparum histidine-rich protein 2 (PfHRP2) has been suggested to be an initiator of the polymerization of heme, which is produced as by-product on the digestion of hemoglobin, and a ...promoter of the H2O2-induced degradation of heme in food vacuoles of the malarial parasite. In this work, we have designed PfHRP2 model peptides, R18 and R27 (18 and 27 residues, respectively), and used them for optical and electron spin resonance spectroscopic measurements to confirm that the axial ligands of the heme-PfHRP2 complex are the nitrogenous donors derived from the imidazole moieties of histidine residues of PfHRP2. In addition, we revealed that the affinities of R18 and R27 for heme (Kd = 2.21 × 10–6 M and 0.71 × 10–6 M, respectively) might be as high as that of PfHRP2 (Kd = 0.94 × 10–6 M). The R27 peptide can remove heme from membrane-intercalated heme and inhibit heme-induced hemolysis. Therefore, we suggest another function of PfHRP2: it may play an important role in the neutralization of toxic heme in the parasite cytoplasm and infected erythrocytes by removing heme from heme-bound membranes or reducing heme-induced hemolysis.
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IZUM, KILJ, NUK, PILJ, PNG, SAZU, UL, UM, UPUK
Abstract
Introduction: Advanced endometrial cancer (EC) still has poor prognosis and requires the development of new therapeutic agents. In ovarian and gastric cancer, we revealed that high ...expression of lipolysis-stimulated lipoprotein receptor (LSR), a cell-surface membrane protein, was associated with poor prognosis and LSR could be a new therapeutic target. However, in EC, the clinical and cell biological role of LSR is still unclear. We aimed to investigate the functions of LSR in EC.
Methods: We evaluated LSR expression by immunohistochemistry and analyzed overall survival (OS) and clinicopathological factors in 228 EC patients. To investigate the mechanism by which LSR affects the prognosis of EC patients, the pathway enrichment analysis and gene ontology analysis were conducted using the publicly available proteogenomic dataset of EC. Cell proliferation was analyzed by WST-8 assay using two LSR-knockdown cell lines (HEC1 and HEC116) developed by transfection of LSR-siRNA, and the activity of several signaling pathways were examined by Western blotting.
Results: Patients were divided into two groups based on LSR expression; High (darkly stained cells in ≥25% of the area, n=153) and Low (darkly stained cells in <25% of the area, n=75) groups. 5-year OS rate in High group was significantly lower than Low group (hazard ratio: 3.53, 95% confidence interval: 1.35 - 9.24, p=0.01). Stage III-IV, deep myometrial invasion (≥75% of uterine myometrium), tumor involvement of adnexa or serosa, and distant metastasis were more frequently observed in High group than in Low group (p<0.05, respectively). The pathway analysis demonstrated that genes correlated with high LSR expression were enriched in MAPK signaling pathway. In gene ontology analysis, these genes were enriched in regulation of ERK1/2 and MAPK cascade. In vitro, LSR-knockdown suppressed cell proliferation (p<0.01) and phosphorylation of MEK1/2, ERK1/2, and p90RSK, suggesting that LSR promoted cell proliferation via MEK/ERK pathway. However, LSR did not regulate the other signaling pathways (SAPK/JNK, p38-MAPK, JAK/STAT3, and PI3K/AKT/mTOR pathway).
Conclusion: LSR regulated cell proliferation via MEK/ERK pathway, and contributed poor prognosis in EC. LSR may be a new therapeutic target of advanced EC.
Citation Format: Yoshikazu Nagase, Kosuke Hiramatsu, Satoshi Nakagawa, Shinya Matsuzaki, Toshihiro Kimura, Satoshi Serada, Yutaka Ueda, Tetsuji Naka, Tadashi Kimura. LSR is a novel prognostic factor by regulating cell proliferation via MEK/ERK pathway in endometrial cancer:Analysis of signal transduction, bioinformatics, and in vitro/in vivo study abstract. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1989.
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