Extrinsic Apoptosis Signaling Research Articles

Research and Development Progression of Oridonin for Hematological Malignancies Therapy.

Oridonin is a tetracyclic diterpenoid compound extracted from the medicinal herb Isodon and related species. Since 1976, studies have reported the significant anti-tumor activity of oridonin in vivo. Recently, an increasing number of studies have confirmed the anti-tumor effects of oridonin in various types of cancers, and its effect on hematological malignancies stands out. Herein, we have systematically reviewed the anti-- tumor effects of oridonin and its specific mechanisms in hematological malignancies, including the regulation of cancer proteins, activation of intrinsic and extrinsic apoptosis signaling pathways, accumulation of reactive oxygen species (ROS), modulation of chaperone proteins and miRNA expression, combination therapy with chemotherapeutic drugs, and the development of its derivatives. Taken together, oridonin exhibits multiple anti-tumor activities and serves as a multi-target agent, making it worthy of further investigation.

Unveiling nature's potential weapon: Magnolol's role in combating bladder cancer by upregulating the miR-124 and inactivating PKC-δ/ERK axis

BackgroundBladder cancer (BC) is a challenging disease to manage. Researchers have been investigating the potential of magnolol, a compound derived from Magnolia officinalis, as an anti-cancer agent. However, the exact regulatory mechanism of magnolol and its impact on the NF-κB signaling pathway in BC remain unclear. MaterialsTo comprehensively evaluate its therapeutic potential, the researchers conducted a series of experiments using BC cell lines (TSGH8301, T24, and MB49) and in vivo animal models. ResultsThe results of the study demonstrated that magnolol exhibits cytotoxic effects on BC cells by activating both the extrinsic and intrinsic apoptosis signaling pathways. Additionally, the expression of anti-apoptotic genes was downregulated by magnolol treatment. The researchers also uncovered the regulatory role of PKCδ/ERK and miR-124-3p in the NF-κB pathway, which may be influenced by magnolol. Treatment with magnolol led to the inactivation of PKCδ/ERK and an increase in miR-124-3p expression, effectively inhibiting NF-κB-mediated progression of BC. Importantly, the administration of magnolol did not result in significant toxicity in normal tissues, highlighting its potential as a safe adjunctive therapy with minimal adverse effects. ConclusionThese findings position magnolol as a promising therapeutic agent for the treatment of BC. By activating apoptosis signaling pathways and inhibiting NF-κB pathway through the upregulation of miR-124-3p and downregulation of PKCδ/ERK activation, magnolol holds promise for suppressing tumor progression and improving patient outcomes in BC. Further research and clinical trials are warranted to explore the full potential of magnolol in the future.

Caspase 3 Expressions in Children with Acute Lymphoblastic Leukemia During Induction Phase Chemotherapy

Introduction: Caspase-3 is a crucial mediator of the extrinsic apoptosis pathway. The role of caspase-3 for extrinsic apoptosis signalling is still a challenge and should be exploited in childhood ALL. This study aimed to compare the caspase-3 expression in the patient’s bone marrow before and after the induction phase of chemotherapy in childhood ALL. It will also to correlate the mean difference in caspase-3 expression between ALL standard-risk and ALL high-risk patients. Methods: Seventeen newly diagnosed ALL subjects were enrolled in this study. Caspase-3 expression in bone marrow was assessed using flow cytometry and monoclonal antibodies. A T-test and a paired T-test were used to compare between groups. The correlation coefficient between ALL groups was evaluated using Spearman’s test and linear regression with a significant p-value of 0.05. Results: The caspase-3 expression is higher after induction therapy. However, it showed an insignificant difference (16.56+12.91% vs 27.71+12.33%; p = 0.08, p > 0.05). The mean difference of caspase-3 in ALL high-risk groups was significantly higher than in ALL standard-risk groups with a positive correlation (p = 0.007, r = 0.756). Conclusion: The caspase-3 expression after induction phase chemotherapy was increased in all standard-risk and high-risk patients; other lymphoblast apoptosis markers need to be confirmed alongside caspase-3.

Abstract 393: Determinants of birinapant efficacy in triple negative breast cancer

Abstract Background: Triple-Negative Breast Cancer (TNBC) is an aggressive subtype of breast cancer with few targeted therapies, limiting patients to harsh systemic treatments and resulting in the lowest 5-year survival rate of all breast cancer subtypes. This survival rate plummets after metastasis to another site, making it crucial to identify new therapeutic options for TNBC patients to prevent recurrence and metastasis. The small molecule therapy birinapant, a secondary mitochondrial activator of caspases (SMAC) mimetic, represents a potential new avenue for TNBC treatment. Birinapant initiates apoptotic signaling through targeting of Inhibitor of Apoptosis (IAP) proteins in the Tumor Necrosis Factor Receptor (TNFR) extrinsic apoptosis signaling pathway, leading to the formation of a death complex, caspase activity, and apoptosis. We have found that approximately 25% of TNBC patient-derived xenograft (PDX) and xenograft-derived organoid (PDxO) models are exquisitely sensitive to birinapant treatment. However, the majority of our TNBC patient-derived lines remain resistant through unknown means. We seek to understand the parameters for birinapant response and the mechanism of inherent resistance in TNBC using our patient-derived TNBC models. Approach and Results: Preliminary Western blot experiments revealed that treatment of birinapant-sensitive and resistant PDX tumors in vivo and PDxOs in vitro with birinapant causes degradation of cIAP1 and cIAP2, but only causes apoptosis in birinapant-sensitive lines. Therefore, there may be a disruption of necessary downstream apoptotic signaling in the birinapant-resistant lines. Through follow-up Western blot experiments we determined that at baseline, birinapant-resistant PDxOs do not lack the TNFR extrinsic apoptosis proteins required for apoptosis signaling after IAP degradation, nor is there an abundance of these proteins in the birinapant-sensitive PDxOs. Furthermore, to determine if TNFR extrinsic apoptosis signaling is impaired in birinapant-resistant PDxOs, we treated birinapant-resistant and -sensitive PDxOs with the TNF-a cytokine, which is known to enhance extrinsic apoptosis signaling in the presence of birinapant. Treatment with TNF-a and birinapant did not induce apoptosis in the birinapant-resistant PDxOs, suggesting that the extrinsic apoptosis signaling pathway may be the culprit of resistance in these patient-derived lines. Discussion: Follow-up experiments will address whether an inability to execute extrinsic apoptosis through other death receptor pathways explains birinapant resistance in these models. This work will allow us to uncover what predisposes TNBC patients to be resistant or sensitive to birinapant, and determine how this may be harnessed therapeutically to improve outcomes for TNBC patients. This project was funded in part with federal funds under HHSN261201500003I and U54CA224076. Citation Format: Elisabeth A. Brown, Chieh-Hsiang Yang, Emilio Cortes-Sanchez, Zhengtao Chu, Jeevan Govindharajulu, Ralph Parchment, James Doroshow, Apurva Srivastava, Alana Welm. Determinants of birinapant efficacy in triple negative breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 393.

Caspase-8-driven apoptotic and pyroptotic crosstalk causes cell death and IL-1β release in X-linked inhibitor of apoptosis (XIAP) deficiency.

Genetic lesions in X-linked inhibitor of apoptosis (XIAP) pre-dispose humans to cell death-associated inflammatory diseases, although the underlying mechanisms remain unclear. Here, we report that two patients with XIAP deficiency-associated inflammatory bowel disease display increased inflammatory IL-1β maturation as well as cell death-associated caspase-8 and Gasdermin D (GSDMD) processing in diseased tissue, which is reduced upon patient treatment. Loss of XIAP leads to caspase-8-driven cell death and bioactive IL-1β release that is only abrogated by combined deletion of the apoptotic and pyroptotic cell death machinery. Namely, extrinsic apoptotic caspase-8 promotes pyroptotic GSDMD processing that kills macrophages lacking both inflammasome and apoptosis signalling components (caspase-1, -3, -7, -11 and BID), while caspase-8 can still cause cell death in the absence of both GSDMD and GSDME when caspase-3 and caspase-7 are present. Neither caspase-3 and caspase-7-mediated activation of the pannexin-1 channel, or GSDMD loss, prevented NLRP3 inflammasome assembly and consequent caspase-1 and IL-1β maturation downstream of XIAP inhibition and caspase-8 activation, even though the pannexin-1 channel was required for NLRP3 triggering upon mitochondrial apoptosis. These findings uncouple the mechanisms of cell death and NLRP3 activation resulting from extrinsic and intrinsic apoptosis signalling, revealhow XIAP loss can co-opt dual cell death programs, and uncover strategies for targeting the celldeath and inflammatory pathways that result from XIAP deficiency.

Oncogenic Role of ADAM32 in Hepatoblastoma: A Potential Molecular Target for Therapy.

Simple SummaryHepatoblastoma (HBL) is a rare hepatic malignancy occurring mainly in early childhood. Although recently developed treatment regimens have improved prognosis, many refractory cases still exist, and the anticancer drug cis-diamminedichloroplatinum—cisplatin—primarily used in HBL treatment often induces severe side effects. Therefore, more effective and safer therapies are needed. In this study, we demonstrate that the expression level of ADAM32 is particularly high in HBL tissue samples and is associated with poor prognosis in various cancer types through the regulation of cancer cell proliferation, stemness, migration, invasion, and acquired resistance to chemotherapy. Our results thus indicate that ADAM32 could be a promising candidate therapeutic target molecule, and provide new insights into the molecular mechanisms of HBL carcinogenesis.Outcomes of pediatric hepatoblastoma (HBL) have improved, but refractory cases still occur. More effective and safer drugs are needed that are based on molecular mechanisms. A disintegrin and metalloproteases (ADAMs) are expressed with high frequency in various human carcinomas and play an important role in cancer progression. In this study, we analyzed expression of ADAMs in HBL with a cDNA microarray dataset and found that the expression level of ADAM32 is particularly high. To investigate the role of ADAM32 in cancer, forced expression or knockdown experiments were conducted with HepG2 and HBL primary cells. Colony formation, cell migration and invasion, and cell viability were increased in HepG2 expressing ADAM32, whereas knockdown of ADAM32 induced a decrease in these cellular functions. Quantitative RT-PCR demonstrated an association between ADAM32 expression and the expression of genes related to cancer stem cells and epithelial–mesenchymal transition (EMT), suggesting a role of ADAM32 in cancer stemness and EMT. Furthermore, knockdown of ADAM32 increased cisplatin-induced apoptosis, and this effect was attenuated by a caspase-8 inhibitor, suggesting that ADAM32 plays a role in extrinsic apoptosis signaling. We conclude that ADAM32 plays a crucial role in progression of HBL, so it might be a promising molecular target in anticancer therapy.

Magnolol Induces Apoptosis Through Extrinsic/intrinsic Pathways and Attenuates NF-κB/STAT3 Signaling in Non-small-cell Lung Cancer Cells.

Non-small-cell lung cancer (NSCLC) is the most common type of lung cancer worldwide, and treatment outcomes are still poor. Magnolol, a hydroxylated biphenyl isolated from Magnolia officinalis, was found to be effective against hepatocellular carcinoma via inactivating nuclear-factor-kappa B (NF-B) signaling. However, whether magnolol targets not only NF-B but also other factors in NSCLC and may contribute to the suppression of tumor progression is unclear. Cell viability, flow cytometry, and western blotting assays were used to identify the mechanism of magnolol action in human lung adenocarcinoma cell lines A549 and CL1-5-F4. Our results indicated that magnolol induced cytotoxicity through extrinsic/intrinsic apoptosis signaling and suppressed phosphorylation of signal transducer and activator of transcription 3 (STAT3)/NF-B and expression of their downstream proteins. Magnolol not only induced extrinsic and intrinsic apoptosis signaling but also inactivated STAT3/NF-B and attenuated their signaling of epithelial-mesenchymal transition and metastasis-related protein expression in NSCLC.

Proinflammatory cytokines induce rapid, NO-independent apoptosis, expression of chemotactic mediators and interleukin-32 secretion in human pluripotent stem cell-derived beta cells

Aims/hypothesisThe aim of this study was to examine the effects of proinflammatory cytokines on cells of different developmental stages during the generation of stem cell-derived beta cells (SC-beta cells) from human pluripotent stem cells (hPSCs). We wanted to find out to what extent human SC-beta cells are suitable as an experimental cellular model and, with regard to a possible therapeutic use, whether SC-beta cells have a comparable vulnerability to cytokines as bona fide beta cells.MethodshPSCs were differentiated towards pancreatic organoids (SC-organoids) using a 3D production protocol. SC-beta cells and non-insulin-producing cells were separated by FACS and differential gene expression profiles of purified human SC-beta cells, progenitor stages and the human beta cell line EndoC-βH1, as a reference, were determined after 24 h incubation with the proinflammatory cytokines IL-1β, TNF-α and IFN-γ via a transcriptome microarray. Furthermore, we investigated apoptosis based on caspase cleavage, the generation of reactive oxygen species and activation of mitogen-activated protein-kinase (MAPK) stress-signalling pathways.ResultsA 24 h exposure of SC-beta cells to proinflammatory cytokines resulted in significant activation of caspase 3/7 and apoptosis via the extrinsic and intrinsic apoptosis signalling pathways. At this time point, SC-beta cells showed a markedly higher sensitivity towards proinflammatory cytokines than non-insulin-producing cells and EndoC-βH1 cells. Furthermore, we were able to demonstrate the generation of reactive oxygen species and rule out the involvement of NO-mediated stress. A transient activation of stress-signalling pathways p38 mitogen-activated protein kinases (p38) and c-Jun N-terminal kinase (JNK) was already observed after 10 min of cytokine exposure. The transcriptome analysis revealed that the cellular response to proinflammatory cytokines increased with the degree of differentiation of the cells. Cytokines induced the expression of multiple inflammatory mediators including IL-32, CXCL9 and CXCL10 in SC-beta cells and in non-insulin-producing cells.Conclusions/interpretationOur results indicate that human SC-beta cells respond to proinflammatory cytokines very similarly to human islets. Due to the fast and fulminant cellular response of SC-beta cells, we conclude that SC-beta cells represent a suitable model for diabetes research. In light of the immaturity of SC-beta cells, they may be an attractive model for developmentally young beta cells as they are, for example, present in patients with early-onset type 1 diabetes. The secretion of chemotactic signals may promote communication between SC-beta cells and immune cells, and non-insulin-producing cells possibly participate in the overall immune response and are thus capable of amplifying the immune response and further stimulating inflammation. We demonstrated that cytokine-treated SC-organoids secrete IL-32, which is considered a promising candidate for type 1 diabetes onset. This underlines the need to ensure the survival of SC-beta cells in an autoimmune environment such as that found in type 1 diabetes.Graphical abstract

Usnic acid-induced programmed cell death in ovarian cancer cells

Usnic acid (UA) is a secondary metabolite obtained by lichen species and exerts a significant anti-proliferative effect on different cancer cells. The current study aimed to determine the efficiency of UA on ovarian cancer and reveal the underlying molecular mechanisms of UA mediated anti-cancer activity. In this study, the cytotoxic effect of UA was evaluated by xCELLigence real-time online cell analysis in OVCAR-3, A2780 ovarian cancer cells and L929 non-cancerous cell compared with Carboplatin. After treatment with IC50 concentration of UA, apoptosis, cell cycle analysis, mRNA expression levels, invasion, wound healing and metastasis assays were performed to evaluate of the molecular mechanism of the apoptosis pathway. UA showed more anti-proliferative effect than carboplatin on OVCAR-3 which is the most aggressive cell among ovarian cancer cells. UA significantly decreased the cell proliferation of OVCAR-3 cells through apoptosis and G0/G1 arrest. Additionally, UA inhibited the invasion and particularly the migration of OVCAR-3 cells compared with control. Moreover, the expression level of 63 target genes from 87 apoptosis associated genes in the primary panel was determined after IC50 concentration of UA in OVCAR-3 cells. UA caused apoptotic cell death through the extrinsic pathway and over-expression of p53. Our findings demonstrated that UA-induced early and late apoptosis through G0/G1 arrest and extrinsic apoptosis signaling pathways and inhibited the migration and invasion of OVCAR-3 cells. Therefore, UA as a novel candidate molecule may be used for the treatment of ovarian cancer.Graphical abstractUsnic acid (UA) exerts anti-cancer effects and induces apoptosis through the extrinsic pathway and over-expression of p53. Additionally, UA potentially inhibits the migration ability of OVCAR-3 ovarian cancer cells. (Created with BioRender.com)

GBP5 promotes liver injury and inflammation by inducing hepatocyte apoptosis.

Liver injury is the first step in causing fibrosis, cirrhosis, and liver cancer, leading to mortality. However, the drivers of progressive liver injury are still incompletely defined. Here, we identify GBP5 as a major factor causing liver injury and inflammation. We show that the expression of GBP5 is abnormally elevated in the damaged liver, and its expression depends at least partially on the NF-κB-inducing kinase (NIK)/NF-κB2signaling pathway. Knockout of Gbp5 ameliorates D-galactosamine/lipopolysaccharide (GalN/LPS)-induced liver injury and inflammation. Conversely, liver-specific overexpression of GBP5 induces liver injury and inflammation. Mechanistically, GBP5 induces hepatocyte apoptosis through the activation of both calpain/caspase 12/caspase 3 and TNFα/caspase 8/caspase 3signaling pathways. Inhibition of either calpain activity or caspase 3 prevents GBP5-induced cell death. Our data demonstrate that GBP5 expression is induced by toxins or the NIK signaling pathway, which promotes both extrinsic and intrinsic apoptosis signaling pathways and further induces liver injury, providing a novel drug target for the treatment of liver injury and inflammation.

Suppression of EGFR/PKC-δ/NF-κB Signaling Associated With Imipramine-Inhibited Progression of Non-Small Cell Lung Cancer.

BackgroundAnti-depressants have been reported to own anti-tumor potential types of cancers; however, the role of imipramine in non-small cell lung cancer (NSCLC) has not been elucidated. Epidermal growth factor receptor (EGFR) was known to be one of the key regulators that control NSCLC progression. Whether EGFR would be the target of imipramine for suppressing tumor signaling transduction and results in anti-tumor potential is remaining unclear.MethodsWe used CL-1-5-F4 cells and animal models to identify the underlying mechanism and therapeutic efficacy of imipramine. Cytotoxicity, apoptosis, invasion/migration, DNA damage, nuclear translocation of NF-κB, activation of NF-κB, phosphorylation of EGFR/PKC-δ/NF-κB was assayed by MTT, flow cytometry, transwell, wound healing assay, comet assay, immunofluorescence staining, NF-κB reporter gene assay and Western blotting, respectively. Tumor growth was validated by CL-1-5-F4/NF-κB-luc2 bearing animal model.ResultsImipramine effectively induces apoptosis of NSCLC cells via both intrinsic and extrinsic apoptosis signaling. DNA damage was increased, while, invasion and migration potential of NSCLC cells was suppressed by imipramine. The phosphorylation of EGFR/PKC-δ/NF-κB and their downstream proteins were all decreased by imipramine. Similar tumor growth inhibition was found in imipramine with standard therapy erlotinib (EGFR inhibitor). Non-obvious body weight loss and liver pathology change were found in imipramine treatment mice.ConclusionImipramine-triggered anti-NSCLC effects in both in vitro and in vivo model are at least partially attributed to its suppression of EGFR/PKC-δ/NF-κB pathway.

Abbv-621 Is a Novel and Potent TRAIL Receptor Agonist Fusion Protein That Induces Apoptosis Alone and in Combination with Navitoclax and Venetoclax in Hematological Tumors

Cell death can be initiated through activation of the extrinsic and intrinsic apoptotic signaling pathways. Tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL), a member of the TNF superfamily of cytokines, preferentially triggers the extrinsic apoptotic pathway by binding as a trimer to two closely related cell surface death receptors, TRAIL-R1 (DR4) and TRAIL-R2 (DR5). When these receptors undergo trimerization, it leads to the formation of the death-inducing signaling complex (DISC) to recruit and activate downstream caspases that ultimately leads to apoptotic cell death. Since TRAIL was found to induce apoptosis, several TRAIL receptor agonists have been developed for the treatment of cancer. ABBV-621 is a novel, second generation TRAIL receptor agonist that is currently being tested in Phase 1 clinical trials. It is an engineered fusion protein consisting as an IgG1-Fc linked to a single chain trimer of TRAIL subunits resulting in a total of six death receptor binding sites per molecule in order to maximize receptor clustering. ABBV-621 exhibited potent single agent activity in acute myeloid lymphoma (AML) and diffuse large B-cell lymphoma (DLBCL) cell lines following 24hrs treatment. This activity was rapid and mechanism-based since activation of downstream apoptotic signaling events (caspase activation, mitochondrial depolarization, phosphatidylserine exposure) were observed as early as 1 hour following the addition of ABBV-621, and cell death could be completely blocked with the pan caspase inhibitor z-VAD-FMK . In vivo, ABBV-621 demonstrated robust single agent anti-tumor activity in pre-clinical models of AML and DLBCL.The intrinsic apoptotic pathway is regulated by the BCL-2 protein family which controls the release of cytochrome c from the mitochondria resulting in the activation of downstream caspases. Because the two apoptotic pathways are linked, the intrinsic pathway can influence cell death mediated through the extrinsic pathway. The elevated expression of the anti-apoptotic proteins, BCL-2 and BCL-XL, can inhibit the activation of the intrinsic apoptotic pathway and enable resistance to anti-cancer therapeutics, including TRAIL. Venetoclax (ABT-199), a BCL-2 selective inhibitor, and navitoclax (ABT-263) a BCL-2/BCL-XL selective inhibitor, were tested in combination with ABBV-621 in a number of hematologic tumor cell lines in vitro and in vivo . The anti-tumor activity of ABBV-621 in combination with venetoclax or navitoclax exceeded that of either agent tested alone in pre-clinical and AML and DLBCL xenograft models.Collectively, these data highlight that targeting the extrinsic apoptosis signaling pathway with either ABBV-621 alone or in combination with venetoclax/navitoclax to harness the intrinsic apoptosis signaling pathway has potential therapeutic benefit in hematologic malignancies.Disclosures:All authors are employees of AbbVie. The design, study conduct, and financial support for this research were provided by AbbVie. AbbVie participated in the interpretation of data, review, and approval of the publication. DisclosuresTahir:Abbvie: Employment, Equity Ownership. Smith:Abbvie: Employment, Equity Ownership. Solomon:Abbvie: Employment, Equity Ownership. Zhang:Abbvie: Employment, Equity Ownership. Xue:Abbvie: Employment, Equity Ownership. Xiao:Abbvie: Employment, Equity Ownership. Cheng:Abbvie: Employment, Equity Ownership. Buchanan:Abbvie: Employment, Equity Ownership. Morgan-Lappe:Abbvie: Employment, Equity Ownership. Phillips:Abbvie: Employment, Equity Ownership.

Molecular subtype-specific responses of colon cancer cells to the SMAC mimetic Birinapant

Colorectal cancer is a molecularly heterogeneous disease. Responses to genotoxic chemotherapy in the adjuvant or palliative setting vary greatly between patients, and colorectal cancer cells often resist chemotherapy by evading apoptosis. Antagonists of an inhibitor of apoptosis proteins (IAPs) can restore defective apoptosis signaling by degrading cIAP1 and cIAP2 proteins and by inhibition of XIAP. Due to the multiple molecular mechanisms-of-action of these targets, responses to IAP antagonist may differ between molecularly distinct colon cancer cells. In this study, responses to the IAP antagonist Birinapant and oxaliplatin/5-fluorouracil (5-FU) were investigated in 14 colon cancer cell lines, representing the consensus molecular subtypes (CMS). Treatment with Birinapant alone did not result in a substantial increase in apoptotic cells in this cell line panel. Annexin-V/PI assays quantified by flow cytometry and high-content screening showed that Birinapant increased responses of CMS1 and partially CMS3 cell lines to oxaliplatin/5-FU, whereas CMS2 cells were not effectively sensitized. FRET-based imaging of caspase-8 and -3 activation validated these differences at the single-cell level, with CMS1 cells displaying sustained activation of caspase-8-like activity during Birinapant and oxaliplatin/5-FU co-treatment, ultimately activating the intrinsic mitochondrial apoptosis pathway. In CMS2 cell lines, Birinapant exhibited synergistic effects in combination with TNFα, suggesting that Birinapant can restore extrinsic apoptosis signaling in the context of inflammatory signals in this subtype. To explore this further, we co-cultured CMS2 and CMS1 colon cancer cells with peripheral blood mononuclear cells. We observed increased cell death during Birinapant single treatment in these co-cultures, which was abrogated by anti-TNFα-neutralizing antibodies. Collectively, our study demonstrates that IAP inhibition is a promising modulator of response to oxaliplatin/5-FU in colorectal cancers of the CMS1 subtype, and may show promise as in the CMS2 subtype, suggesting that molecular subtyping may aid as a patient stratification tool for IAP antagonists in this disease.

Effect of nicotine on placental inflammation and apoptosis in preeclampsia-like model

AimsPlacental tissues from patients with preeclampsia (PE) and in the lipopolysaccharide (LPS)-induced PE-like model were used to investigate the implication of placental inflammation and apoptosis in PE. Whether the beneficial effects of nicotine are related to inhibition of placental inflammation and apoptosis in the PE-like model were investigated. Main methodsPlacental apoptosis was detected in PE patients and the PE-like rat model by TUNEL staining. Changes in the number of CD68+ macrophages in placental tissues from PE patients were detected by immunofluorescent staining. The mRNA expression of the pro-inflammatory cytokines tumor necrosis factor alpha (TNF-α), interleukin (IL-1β), MCP-1, and proteins involved in extrinsic or intrinsic apoptosis signaling in the PE-like model was determined by qRT-PCR; immunofluorescent staining was used to detect the expression of TNF-α receptor (TNFR1), MCP-1 and apoptosis-related proteins. Key findingsPlacental apoptosis was increased in both PE patients and the PE-like model, more macrophages infiltrated into placenta in PE patients. A significant upregulation in mRNA expression of TNF-α, IL-1β, MCP-1, and caspase 3, caspase 8, caspase 9 was found in the PE-like rats compared to the control animals, the immunoreactivity of placental MCP-1, TNFR1, and apoptosis-related proteins (caspase 3, caspase 8, caspase 9, Bax) was also enhanced; nicotine treatment significantly reversed those changes. SignificanceOur data suggests that the protective effects of nicotine are associated with inhibiting placenta inflammation and apoptosis, and nicotine might be a potentially therapeutic candidate for preventing preeclampsia.

Sample-based modeling reveals bidirectional interplay between cell cycle progression and extrinsic apoptosis.

Apoptotic cell death can be initiated through the extrinsic and intrinsic signaling pathways. While cell cycle progression promotes the responsiveness to intrinsic apoptosis induced by genotoxic stress or spindle poisons, this has not yet been studied conclusively for extrinsic apoptosis. Here, we combined fluorescence-based time-lapse monitoring of cell cycle progression and cell death execution by long-term time-lapse microscopy with sampling-based mathematical modeling to study cell cycle dependency of TRAIL-induced extrinsic apoptosis in NCI-H460/geminin cells. In particular, we investigated the interaction of cell death timing and progression of cell cycle states. We not only found that TRAIL prolongs cycle progression, but in reverse also that cell cycle progression affects the kinetics of TRAIL-induced apoptosis: Cells exposed to TRAIL in G1 died significantly faster than cells stimulated in S/G2/M. The connection between cell cycle state and apoptosis progression was captured by developing a mathematical model, for which parameter estimation revealed that apoptosis progression decelerates in the second half of the cell cycle. Similar results were also obtained when studying HCT-116 cells. Our results therefore reject the null hypothesis of independence between cell cycle progression and extrinsic apoptosis and, supported by simulations and experiments of synchronized cell populations, suggest that unwanted escape from TRAIL-induced apoptosis can be reduced by enriching the fraction of cells in G1 phase. Besides novel insight into the interrelation of cell cycle progression and extrinsic apoptosis signaling kinetics, our findings are therefore also relevant for optimizing future TRAIL-based treatment strategies.

Promoted Viability and Differentiated Phenotype of Cultured Chondrocytes With Low Level Laser Irradiation Potentiate Efficacious Cells for Therapeutics.

Effective clinical treatments of cartilage lesions in affected joints require large numbers of viable chondrogenic cells generated through in vivo stimulation or ex vivo expansion of chondrocytes isolated from small biopsy specimens. Conventional passaging of chondrocytes in culture provides sufficient cells for treatments but these cells usually lose their differentiated phenotype. This leads to the formation of fibrocartilaginous tissue due to a malfunctioning repair process. Biostimulation of passaging chondrocytes with low level laser irradiation (LLLI) may theoretically produce more functional chondrocytes for cell-based repair of cartilage defects. Molecular and cellular analyses, cytochemistry, cell cultivation, and microscopy showed that LLLI treatments were found to (1) increase chondrocyte viability, (2) promote secretion of matrix proteins, (3) upregulate expression of chondrogenic genes, and (4) downregulate gene expression of cell destructive proteases and genes coding for mediators involved in the extrinsic apoptosis signaling pathway. Furthermore, LLLI attenuated induction of genes associated with cell death and matrix breakdown induced by IL-1β, some of which was seen at the protein level, with verification of effects on gene expression in the C28/I2 human chondrocyte line. LLLI treatments during culture generated larger numbers of viable chondrocytes compared to untreated cultures. Moreover, LLLI-treated chondrocytes in culture also rectified and simultaneously maintained their differentiated phenotype. Cultured chondrocytes treated with LLLI are a promising cell source for repairing cartilage lesions in vivo and restoration of articular function using tissue engineering strategies.

Fascinating Chemopreventive Story of Wogonin: A Chance to Hit on the Head in Cancer Treatment.

Cancer, global havoc, is a group of debilitating diseases that strikes family as well as society. Cancer cases are drastically increasing these days. Despite many therapies and surgical procedures available, cancer is still difficult to control due to limited effective therapies or targeted therapies. Natural products can produce lesser side effects to the normal cells, which are the major demerit of chemotherapies and radiation. Wogonin, a natural product extracted from the plant, Scutellaria baicalensis has been widely studied and found with a high caliber to tackle most of the cancers via several mechanisms that include intrinsic as well as extrinsic apoptosis signaling pathways, carcinogenesis diminution, telomerase activity inhibition, metastasis inhibition in the inflammatory microenvironment, anti-angiogenesis, cell growth inhibition and arrest of the cell cycle, increased generation of H2O2 and accumulation of Ca2+ and also as an adjuvant along with anticancer drugs. This article discusses the role of wogonin in various cancers, its synergism with various drugs, and the mechanism by which wogonin controls tumor growth.

Patient-specific logic models of signaling pathways from screenings on cancer biopsies to prioritize personalized combination therapies.

Mechanistic modeling of signaling pathways mediating patient‐specific response to therapy can help to unveil resistance mechanisms and improve therapeutic strategies. Yet, creating such models for patients, in particular for solid malignancies, is challenging. A major hurdle to build these models is the limited material available that precludes the generation of large‐scale perturbation data. Here, we present an approach that couples ex vivo high‐throughput screenings of cancer biopsies using microfluidics with logic‐based modeling to generate patient‐specific dynamic models of extrinsic and intrinsic apoptosis signaling pathways. We used the resulting models to investigate heterogeneity in pancreatic cancer patients, showing dissimilarities especially in the PI3K‐Akt pathway. Variation in model parameters reflected well the different tumor stages. Finally, we used our dynamic models to efficaciously predict new personalized combinatorial treatments. Our results suggest that our combination of microfluidic experiments and mathematical model can be a novel tool toward cancer precision medicine.

Next-generation hypomethylating agent SGI-110 primes acute myeloid leukemia cells to IAP antagonist by activating extrinsic and intrinsic apoptosis pathways.

Therapeutic efficacy of first-generation hypomethylating agents (HMAs) is limited in elderly acute myeloid leukemia (AML) patients. Therefore, combination strategies with targeted therapies are urgently needed. Here, we discover that priming with SGI-110 (guadecitabine), a next-generation HMA, sensitizes AML cells to ASTX660, a novel antagonist of cellular inhibitor of apoptosis protein 1 and 2 (cIAP1/2) and X-linked IAP (XIAP). Importantly, SGI-110 and ASTX660 synergistically induced cell death in a panel of AML cell lines as well as in primary AML samples while largely sparing normal CD34+ human progenitor cells, underlining the translational relevance of this combination. Unbiased transcriptome analysis revealed that SGI-110 alone or in combination with ASTX660 upregulated the expression of key regulators of both extrinsic and intrinsic apoptosis signaling pathways such as TNFRSF10B (DR5), FAS, and BAX. Individual knockdown of the death receptors TNFR1, DR5, and FAS significantly reduced SGI-110/ASTX660-mediated cell death, whereas blocking antibodies for tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) or FAS ligand (FASLG) failed to provide protection. Also, TNFα-blocking antibody Enbrel had little protective effect on SGI-110/ASTX660-induced cell death. Further, SGI-110 and ASTX660 acted in concert to promote cleavage of caspase-8 and BID, thereby providing a link between extrinsic and intrinsic apoptotic pathways. Consistently, sequential treatment with SGI-110 and ASTX660-triggered loss of mitochondrial membrane potential (MMP) and BAX activation which contributes to cell death, as BAX silencing significantly protected from SGI-110/ASTX660-mediated apoptosis. Together, these events culminated in the activation of caspases-3/-7, nuclear fragmentation, and cell death. In conclusion, SGI-110 and ASTX660 cooperatively induced apoptosis in AML cells by engaging extrinsic and intrinsic apoptosis pathways, highlighting the therapeutic potential of this combination for AML.

Apoptosis induction and ERK/NF-κB inactivation are associated with magnolol-inhibited tumor progression in hepatocellular carcinoma in vivo.

Although hepatitis B and/or hepatitis C virus were recognized as major risk factor for the development of hepatocellular carcinoma (HCC), certain occupational, environmental, and lifestyle factors also play key roles in HCC tumorigenesis. Moreover, in molecular signaling route, extracellular signal-regulated kinase (ERK)/nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) signaling was found to be overexpressed and linked to poor prognosis in HCC. Thus, to identify possible nature compound that can suppress ERK/NF-κB may be benefit to HCC patient. Magnolol, a natural compound derived from herbal plant Magnolia officinalis, has been recognized as a liver protection and antitumor reagent. However, whether magnolol-inhibited HCC progression correlates with disruption of ERK/NF-κB signaling is remained unclear. In this studies, we performed SK-Hep1/luc2 HCC bearing animal model to investigate the anticancer efficacy and mechanism of magnolol on tumor progression. Tumor size and tumor growth rate were dramatically suppressed after treatment of magnolol. In addition, expression of phospho-ERK (p-ERK), NF-κB p65 (Ser536), and tumor progression-associated proteins, such as matrix metallopeptidase 9 (MMP-9), vascular endothelial growth factor (VEGF), X-linked inhibitor of apoptosis protein (XIAP), and CyclinD1 were all significantly decreased by magnolol. Most important, major extrinsic and intrinsic apoptosis signaling factors, including active caspase-8 and caspase-9 were both enhanced by magnolol. This study indicated that apoptosis induction through extrinsic/intrinsic pathways and blockage of ERK/NF-κB activation were associated with magnolol-inhibited tumor progression in HCC in vivo.