Gene therapy for management of lupus: Correction of Fas and Fas ligand-induced apoptosis in murine disease — therapeutic rationale and strategies

Background: Retinoic acid-inducible gene I (RIG-I)-like receptors (RLRs) play key roles in the antiviral response, but recent works show that RLR activation elicits anticancer activity as well, including apoptosis. Previously, we demonstrated that the anticancer activity of the RLR agonist Poly(I:C)-HMW/LyoVec™ [Poly(I:C)-HMW] against human lung cancer cells was enhanced by cotreatment with ionizing radiation (IR). In addition, cotreatment with Poly(I:C)-HMW and IR induced apoptosis in a Fas-independent manner, and increased Fas expression on the cell surface. Objective: The current study investigated the resultant hypothesis that Fas ligand (FasL) may enhance apoptosis in lung cancer cells cotreated with Poly(I:C)-HMW+IR. Methods: FasL was added into culture medium at 24 h following cotreatment with Poly(I:C)- HMW+IR, after upregulation of cell surface Fas expression on human lung cancer cells A549 and H1299 have already been discussed. Results: FasL enhanced the apoptosis of A549 and H1299 cells treated with Poly(I:C)-HMW+IR. Similarly, IR alone - and not Poly(I:C)-HMW - resulted in the upregulation of cell surface Fas expression followed by a high response to FasL-induced apoptosis, thus suggesting that the high sensitivity of cells treated with Poly(I:C)-HMW+IR to FasL-induced apoptosis resulted from the cellular response to IR. Finally, knockdown of Fas by siRNA confirmed that the high response of treated cells to FasL-induced apoptosis is dependent on Fas expression. Conclusion: In summary, the present study indicates that upregulated Fas expression following cotreatment with Poly(I:C)-HMW and IR is responsive to FasL-induced apoptosis, and a combination of RLR agonist, IR, and FasL could be a potential promising cancer therapy.

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Conformation-Specific Inhibitory Anti-MMP-7 Monoclonal Antibody Sensitizes Pancreatic Ductal Adenocarcinoma Cells to Chemotherapeutic Cell Kill

Cancers ◽

10.3390/cancers13071679 ◽

2021 ◽

Vol 13 (7) ◽

pp. 1679

Author(s):

Vishnu Mohan ◽

Jean P. Gaffney ◽

Inna Solomonov ◽

Maxim Levin ◽

Mordehay Klepfish ◽

...

Keyword(s):

Monoclonal Antibody ◽

Active Site ◽

Drug Targets ◽

Fas Ligand ◽

Specific Activity ◽

Matrix Metalloproteases ◽

Ductal Adenocarcinoma ◽

Enzyme Active Site ◽

Induced Apoptosis

Matrix metalloproteases (MMPs) undergo post-translational modifications including pro-domain shedding. The activated forms of these enzymes are effective drug targets, but generating potent biological inhibitors against them remains challenging. We report the generation of anti-MMP-7 inhibitory monoclonal antibody (GSM-192), using an alternating immunization strategy with an active site mimicry antigen and the activated enzyme. Our protocol yielded highly selective anti-MMP-7 monoclonal antibody, which specifically inhibits MMP-7′s enzyme activity with high affinity (IC50 = 132 ± 10 nM). The atomic model of the MMP-7-GSM-192 Fab complex exhibited antibody binding to unique epitopes at the rim of the enzyme active site, sterically preventing entry of substrates into the catalytic cleft. In human PDAC biopsies, tissue staining with GSM-192 showed characteristic spatial distribution of activated MMP-7. Treatment with GSM-192 in vitro induced apoptosis via stabilization of cell surface Fas ligand and retarded cell migration. Co-treatment with GSM-192 and chemotherapeutics, gemcitabine and oxaliplatin elicited a synergistic effect. Our data illustrate the advantage of precisely targeting catalytic MMP-7 mediated disease specific activity.

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Abstract 028: CCN1 Enables Fas Ligand-Induced Apoptosis by Elevating Fas Expression in Primary Cardiomyocytes or by Increasing Cytoplasmic Smac in Cardiomyoblast H9c2 Cells

Circulation Research ◽

10.1161/res.113.suppl_1.a028 ◽

2013 ◽

Vol 113 (suppl_1) ◽

Author(s):

Bor-Chyuan Su ◽

Fan-E Mo

Keyword(s):

Cell Surface ◽

Fas Ligand ◽

Ischemia Reperfusion ◽

Surface Display ◽

Critical Role ◽

Mitogen Activated Protein Kinase ◽

Neonatal Rat ◽

Ros Generation ◽

H9c2 Cells ◽

Induced Apoptosis

Fas/Fas ligand (FasL) is implicated in cardiac ischemia/reperfusion injury. However, cardiomyocytes in culture are resistant to FasL-induced apoptosis, suggesting that additional factor(s) are required for FasL-induced apoptosis. Matricellular protein CCN1 has been demonstrated to promote cytotoxicity of FasL in human skin fibroblasts. CCN1 is induced in a variety of cardiac pathologies. We assessed the hypothesis that CCN1 may be involved in the regulation of FasL-induced apoptosis in cardiomyocytes. We found that either FasL or CCN1 did not induce cell death in neonatal rat ventricular cardiomyocytes (NRVM). Interestingly, the combination of FasL+CCN1 generated 2-fold induction of apoptosis (vs. control p<0.001). An integrin-α 6 β 1 -binding defective mutant CCN1, CCN1-DM failed to exert synergy with FasL to induce apoptosis, indicating a critical role of α 6 β 1 . The engagement between CCN1 and α 6 β 1 instigated the elevation of cellular reactive oxygen species (ROS), the activation of mitogen activated protein kinase p38, and followed by the induction of cell surface display of Fas, thereby sensitizing NRVM to FasL-induced apoptosis. Pretreatment of the p38 inhibitor SB202190 abolished the CCN1-induced cell-surface Fas expression and the apoptosis induced by FasL+CCN1. In addition, we tested the interaction between CCN1 and FasL on the cardiomyoblast H9c2 cells. We found that FasL or CCN1 alone did not cause apoptosis in H9c2, and required the combination of FasL+CCN1 to induced apoptosis (vs. control p<0.001) in H9c2 cells, reminiscent of the observation in NRVM. Mechanistically, CCN1 acted through binding to integrin α 6 β 1 , ROS generation, and p38 activation, however, did not increase the expression of cell surface Fas for its synergy with FasL in H9c2 cells. Instead, CCN1 induced Bax translocation to mitochondria, which in turn led to the release of Smac from mitochondria to cytosol. The cytosolic Smac functions to neutralize XIAP. Smac is critical for CCN1 action, because the knockdown of Smac blunted the apoptotic activities of CCN1. In conclusion, CCN1 may play a detrimental role in a stressed heart to both the differentiated cardiomyocytes and the proliferative cardioblasts through distinct signaling mechanisms.

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Induction of Fas (Apo-1, CD95)-Mediated Apoptosis of Activated Lymphocytes by Polyclonal Antithymocyte Globulins

Blood ◽

10.1182/blood.v91.7.2360.2360_2360_2368 ◽

1998 ◽

Vol 91 (7) ◽

pp. 2360-2368 ◽

Cited By ~ 2

Author(s):

Laurent Genestier ◽

Sylvie Fournel ◽

Monique Flacher ◽

Olga Assossou ◽

Jean-Pierre Revillard ◽

...

Keyword(s):

T Cells ◽

Peripheral Blood ◽

Fas Ligand ◽

Mononuclear Cells ◽

Resting Cells ◽

Activated T Cells ◽

Peripheral Blood Mononuclear ◽

Gene And Protein Expression ◽

Fas L ◽

Induced Apoptosis

Polyclonal horse antilymphocyte and rabbit antithymocyte globulins (ATGs) are currently used in severe aplastic anemia and for the treatment of organ allograft acute rejection and graft-versus-host disease. ATG treatment induces a major depletion of peripheral blood lymphocytes, which contributes to its overall immunosuppressive effects. Several mechanisms that may account for lymphocyte lysis were investigated in vitro. At high concentrations (.1 to 1 mg/mL) ATGs activate the human classic complement pathway and induce lysis of both resting and phytohemagglutinin (PHA)-activated peripheral blood mononuclear cells. At low, submitogenic, concentration ATGs induce antibody-dependent cell cytotoxicity of PHA-activated cells, but not resting cells. They also trigger surface Fas (Apo-1, CD95) expression in naive T cells and Fas-ligand gene and protein expression in both naive and primed T cells, resulting in Fas/Fas-L interaction-mediated cell death. ATG-induced apoptosis and Fas-L expression were not observed with an ATG preparation lacking CD2 and CD3 antibodies. Susceptibility to ATG-induced apoptosis was restricted to activated cells, dependent on IL-2, and prevented by Cyclosporin A, FK506, and rapamycin. The data suggest that low doses of ATGs could be clinically evaluated in treatments aiming at the selective deletion of in vivo activated T cells in order to avoid massive lymphocyte depletion and subsequent immunodeficiency.

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Nonylphenol and Octylphenol-Induced Apoptosis in Human Embryonic Stem Cells Is Related to Fas-Fas Ligand Pathway

Toxicological Sciences ◽

10.1093/toxsci/kfl114 ◽

2006 ◽

Vol 94 (2) ◽

pp. 310-321 ◽

Cited By ~ 42

Author(s):

S.-K. Kim ◽

B.-K. Kim ◽

J.-H. Shim ◽

J.-E. Gil ◽

Y.-D. Yoon ◽

...

Keyword(s):

Stem Cells ◽

Embryonic Stem Cells ◽

Human Embryonic Stem Cells ◽

Fas Ligand ◽

Embryonic Stem ◽

Induced Apoptosis

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CGP57148B (STI-571) induces differentiation and apoptosis and sensitizes Bcr-Abl–positive human leukemia cells to apoptosis due to antileukemic drugs

Blood ◽

10.1182/blood.v96.6.2246 ◽

2000 ◽

Vol 96 (6) ◽

pp. 2246-2253 ◽

Cited By ~ 192

Author(s):

Guofu Fang ◽

Caryn Naekyung Kim ◽

Charles L. Perkins ◽

Nimmanapalli Ramadevi ◽

Elliott Winton ◽

...

Keyword(s):

Myeloid Leukemia ◽

Fas Ligand ◽

Ectopic Expression ◽

K562 Cells ◽

Parp Cleavage ◽

Human Leukemia ◽

Sti 571 ◽

Tnf Α ◽

Cyt C ◽

Induced Apoptosis

Abstract The differentiation and apoptosis-sensitizing effects of the Bcr-Abl–specific tyrosine kinase inhibitor CGP57148B, also known as STI-571, were determined in human Bcr-Abl–positive HL-60/Bcr-Abl and K562 cells. First, the results demonstrate that the ectopic expression of the p185 Bcr-Abl fusion protein induced hemoglobin in the acute myeloid leukemia (AML) HL-60 cells. Exposure to low-dose cytosine arabinoside (Ara-C; 10 nmol/L) increased hemoglobin levels in HL-60/Bcr-Abl and in the chronic myeloid leukemia (CML) blast crisis K562 cells, which express the p210 Bcr-Abl protein. As compared with HL-60/neo, HL-60/Bcr-Abl and K562 cells were resistant to apoptosis induced by Ara-C, doxorubicin, or tumor necrosis factor-α (TNF-α), which was associated with reduced processing of caspase-8 and Bid protein and decreased cytosolic accumulation of cytochrome c (cyt c). Exposure to CGP57148B alone increased hemoglobin levels and CD11b expression and induced apoptosis of HL-60/Bcr-Abl and K562 cells. CGP57148B treatment down-regulated antiapoptotic XIAP, cIAP1, and Bcl-xL, without affecting Bcl-2, Bax, Apaf-1, Fas (CD95), Fas ligand, Abl, and Bcr-Abl levels. CGP57148B also inhibited constitutively active Akt kinase and NFκB in Bcr-Abl–positive cells. Attenuation of NFκB activity by ectopic expression of transdominant repressor of IκB sensitized HL-60/Bcr-Abl and K562 cells to TNF-α but not to apoptosis induced by Ara-C or doxorubicin. Importantly, cotreatment with CGP57148B significantly increased Ara-C– or doxorubicin-induced apoptosis of HL-60/Bcr-Abl and K562 cells. This was associated with greater cytosolic accumulation of cyt c and PARP cleavage activity of caspase-3. These in vitro data indicate that combinations of CGP57148B and antileukemic drugs such as Ara-C may have improved in vivo efficacy against Bcr-Abl–positive acute leukemia.

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