The inhibitor of apoptosis (IAP) family of proteins enhances cell survival through mechanisms that remain uncertain. In this report, we show that cIAP1 and cIAP2 promote cancer cell survival by ...functioning as E3 ubiquitin ligases that maintain constitutive ubiquitination of the RIP1 adaptor protein. We demonstrate that AEG40730, a compound modeled on BIR-binding tetrapeptides, binds to cIAP1 and cIAP2, facilitates their autoubiquitination and proteosomal degradation, and causes a dramatic reduction in RIP1 ubiquitination. We show that cIAP1 and cIAP2 directly ubiquitinate RIP1 and induce constitutive RIP1 ubiquitination in cancer cells and demonstrate that constitutively ubiquitinated RIP1 associates with the prosurvival kinase TAK1. When deubiquitinated by AEG40730 treatment, RIP1 binds caspase-8 and induces apoptosis. These findings provide insights into the function of the IAPs and provide new therapeutic opportunities in the treatment of cancer.
Moderate alcohol consumption is associated with increased plasma high‐density lipoprotein (HDL)‐cholesterol concentrations and reduced risk for cardiovascular disease. Plasma cholesteryl ester ...transfer activity (CETA) mediates the exchange of HDL‐cholesteryl ester (CE) for the triacylglycerol (TAG) of very‐low‐density lipoproteins. We compared the effects of oral challenges of Alcohol, saturated fat (SAT), and (Alcohol + SAT) on plasma CETA, cholesterol, nonesterified fatty acids (NEFA), and TAG among normo‐triglyceridemic (NTG) and mildly hypertriglyceridemic (HTG) volunteers having a range of plasma TAG concentrations. The major changes were (1) CETA increased more after ingestion of SAT and (Alcohol + SAT) in the HTG group versus the NTG group; (2) after all three challenges, elevation of plasma TAG concentration persisted longer in the HTG versus NTG group. Plasma cholesterol was not affected by the three dietary challenges, while Alcohol increased NEFA more in the HTG group than the NTG group. Plasma TAG best predicted plasma CETA, suggesting that intestinally derived lipoproteins are acceptors of HDL‐CE. Unexpectedly, ingestion of (Alcohol + SAT) reduced the strength of the correlation between plasma TAG and CETA, that is the effects of (SAT and Alcohol) on plasma CETA are not synergistic nor additive but rather mutually suppressive. The alcohol‐mediated inhibition of CE‐transfer to chylomicrons maintains a higher plasma HDL‐cholesterol concentration, which is athero‐protective, although the suppressive metabolite underlying this correlation could be acetate, the terminal alcohol metabolite, other factors, including CETA inhibitors, are also likely important.
Inhibition of carbonic anhydrase IX in glioblastoma multiforme Amiri, Abdolali; Le, Phuong Uyen; Moquin, Alexandre ...
European journal of pharmaceutics and biopharmaceutics,
December 2016, 2016-Dec, 2016-12-00, 20161201, Letnik:
109
Journal Article
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Carbonic anhydrase IX (CAIX) is a transmembrane enzyme upregulated in several types of tumors including glioblastoma multiforme (GBM). GBM is among the most aggressive tumors among ...gliomas. Temozolomide (TMZ) therapy combined with surgical or radiation approaches is the standard treatment but not effective in long term. In this study we tested the treatment with acetazolamide (ATZ), an inhibitor of CAIX, alone or combined with TMZ. The experiments were performed in 2D and 3D cultures (spheroids) using glioblastoma U251N and human brain tumor stem cells (BTSCs). Several proteins implicated in tumor cell death were also investigated. The key results from these studies suggest the following: (1) Cell death of human glioblastoma spheroids and BTSC is significantly increased with combined treatment after 7 days, and (2) the effectiveness of ATZ is significantly enhanced against BTSC and U251N when incorporated into nano-carriers. Collectively, these results point toward the usefulness of nano-delivery of CAIX inhibitors and their combination with chemotherapeutics for glioblastoma treatment.
Background:
Angiotensin-converting enzyme 2 (ACE2) hydrolyzes angiotensin (Ang) II to Ang-(1-7), promoting vasodilatation, and inhibiting oxidative stress and inflammation. Plasma membrane ACE2 is ...the receptor for all known SARS-CoV-2 (severe acute respiratory syndrome coronavirus 2) viral variants. In COVID-19 infection, soluble ACE2 variants may act as decoys to bind and neutralize the coronavirus, reducing its tissue infectivity. Furthermore, soluble ACE2 variants have been proposed as potential therapeutics for kidney disease and hypertensive disorders.
Objective:
Soluble ACE2 variants conjugated to human Fc domains and selected for high-potency viral SARS-CoV-2 neutralization were prepared and evaluated for ACE2 activity in vitro. Lead candidates were then tested for systemic ACE2 activity, stability, and effects on blood pressure and albuminuria in mice with Ang II-induced hypertension.
Methods:
ACE2 activity of 10 soluble ACE2 variants was first assessed in cell-free conditions using a fluorogenic substrate, or by Ang II hydrolysis to Ang-(1-7). Hypertension was induced in male or female mice by implantation of osmotic minipumps containing Ang II. Two lead ACE2 variants were injected intravenously (i.v.) into hypertensive mice, followed by measurements of blood pressure (tail-cuff plethysmography), albuminuria, and tissue ACE2 activity and protein (immunoblots).
Results:
Soluble ACE2-Fc variants demonstrated significant ACE2 enzymatic activity, with kinetics comparable with human recombinant ACE2. In hypertensive mice, single dose i.v. injection of ACE2-Fc variant K (10 mg/kg) significantly decreased systolic blood pressure at 24 hours, with partial lowering sustained to 48 hours, and tendency to reduce albuminuria at 72 hours. By contrast, ACE2-Fc variant I had no effect on blood pressure or albuminuria in hypertensive mice; ACE2-Fc variant K was detected by immunoblotting in plasma, kidney, heart, lung, liver, and spleen lysates 72 hours after injection, associated with significantly increased ACE2 activity in all tissues except kidney and spleen. Angiotensin-converting enzyme 2-Fc variant I had no effect on plasma ACE2 activity.
Conclusions:
Soluble ACE2-Fc variant K reduces blood pressure and tends to lower albuminuria in hypertensive mice. Furthermore, soluble ACE2-Fc variant K has prolonged tissue retention, associated with increased tissue ACE2 activity. The results support further studies directed at the therapeutic potential of soluble ACE2-Fc variant K for cardiovascular and kidney protection.
Cancer cells can use X-linked inhibitor of apoptosis (XIAP) to evade apoptotic cues, including chemotherapy. The antitumor potential of AEG35156, a novel second-generation antisense oligonucleotide ...directed toward XIAP, was assessed in human cancer models when given as a single agent and in combination with clinically relevant chemotherapeutics.
AEG35156 was characterized for its ability to cause dose-dependent reductions of XIAP mRNA and protein in vitro and in vivo, to sensitize cancer cell lines to death stimuli, and to exhibit antitumor activity in multiple human cancer xenograft models as a single agent or in combination with chemotherapy.
AEG35156 reduced XIAP mRNA levels with an EC50 of 8 to 32 nmol/L and decreased XIAP protein levels by >80%. Loss of XIAP protein correlated with increased sensitization to tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-mediated apoptosis in Panc-1 pancreatic carcinoma cells. AEG35156 exhibited potent antitumor activity relative to control oligonucleotides in three human cancer xenograft models (prostate, colon, and lung) and was capable of inducing complete tumor regression when combined with taxanes. Antitumor effects of AEG35156 correlated with suppression of tumor XIAP levels.
AEG35156 reduces XIAP levels and sensitizes tumors to chemotherapy. AEG35156 is presently under clinical assessment in multiple phase I trials in cancer patients as a single agent and in combination with docetaxel in solid tumors or cytarabine/idarubicin in leukemia.
Deregulated expression of members of the IAP (inhibitor of apoptosis) family has been identified in a wide variety of neoplastic cells, and synthetic IAP antagonists represent a promising novel class ...of chemotherapeutic agents. Early work focused on the ability of these compounds to block the caspase-inhibitory function of XIAP (X-linked IAP). However, recent studies have shown that IAP antagonists, although primarily designed to target XIAP, trigger ubiquitin-mediated degradation of two related proteins, c-IAP (cellular IAP) 1 and c-IAP2, and through this process potentiates the death of tumour cells via autocrine cellular-signalling pathways. In this context, the relative contribution of XIAP as a target of this class of compounds is unclear. In the present study, we examine the involvement of XIAP using a recently described synthetic IAP antagonist, AEG40730, and through comparison of a human XIAP-depleted tumour cell line with its isogenic wild-type control line. Treatment with nanomolar concentrations of AEG40730 resulted in the loss of both XIAP and c-IAP1 proteins, albeit with different kinetics. Although XIAP-deficient HCT116 cells retained some sensitivity to external apoptotic stimuli, the results suggest that IAP antagonists, such as AEG40730, exert their apoptosis-enhancing effects through XIAP in addition to the c-IAPs. These results indicate that IAP antagonists can target multiple IAPs to augment distinct pro-apoptotic signalling pathways, thereby revealing the potential for these compounds in cancer therapy and underscoring the promise of IAP-targeted therapies.
Small, dense, electronegative low density lipoprotein LDL(−) is increased in patients with familial hypercholesterolemia and diabetes, populations at increased risk for coronary artery disease. It is ...present to a lesser extent in normolipidemic subjects. The mechanistic link between small, dense LDL(−) and atherogenesis is not known. To begin to address this, we studied the composition and dynamics of small, dense LDL(−) from normolipidemic subjects. NEFA levels, which correlate with triglyceride content, are quantitatively linked to LDL electronegativity. Oxidized LDL is not specific to small, dense LDL(−) or lipoprotein a (i.e., abnormal lipoprotein). Apolipoprotein C-III is excluded from the most abundant LDL (i.e., that of intermediate density: 1.034 < d < 1.050 g/ml) but associated with both small and large LDL(−). In contrast, lipoprotein-associated phospholipase A2 (LpPLA2) is highly enriched only in small, dense LDL(−). The association of LpPLA2 with LDL may occur through amphipathic helical domains that are displaced from the LDL surface by contraction of the neutral lipid core.
Targeting apoptosis control provides a novel therapeutic approach to the treatment of cancer and other proliferative disorders. We summarize the evidence for apoptosis deregulation in cancer and ...describe the pivotal role of XIAP, the X-linked Inhibitor-of-APoptosis. XIAP is the predominant inhibitor of caspases 3, 7 and 9 in cells, which suppresses the programmed cell death effector capability of these proteases. Evidence is presented validating XIAP as a cancer target. The inhibition or downregulation of XIAP in cancer cells lowers the apoptotic threshold, thereby inducing cell death and/or enhancing the cytotoxic action of chemotherapeutic agents. We describe the development of AEG35156 (also named GEM640), a second generation antisense compound targeting XIAP, from concept to in vivo preclinical proof-of-principle studies, through formal toxicology, and to a phase 1 clinical trial in cancer patients.
Small, dense, electronegative low density lipoprotein LDL(-) is increased in patients with familial hypercholesterolemia and diabetes, populations at increased risk for coronary artery disease. It is ...present to a lesser extent in normolipidemic subjects. The mechanistic link between small, dense LDL(-) and atherogenesis is not known. To begin to address this, we studied the composition and dynamics of small, dense LDL(-) from normolipidemic subjects. NEFA levels, which correlate with triglyceride content, are quantitatively linked to LDL electronegativity. Oxidized LDL is not specific to small, dense LDL(-) or lipoprotein a (i.e., abnormal lipoprotein). Apolipoprotein C-III is excluded from the most abundant LDL (i.e., that of intermediate density: 1.034 < d < 1.050 g/ml) but associated with both small and large LDL(-). In contrast, lipoprotein-associated phospholipase A₂ (LpPLA₂) is highly enriched only in small, dense LDL(-). The association of LpPLA₂ with LDL may occur through amphipathic helical domains that are displaced from the LDL surface by contraction of the neutral lipid core.
To understand how the balance between pro- and anti-apoptotic signals influences effector function in the immune system, we studied the X-linked inhibitor of apoptosis (XIAP), an endogenous regulator ...of cellular apoptosis. Real-time PCR showed increased XIAP expression in blood of mice with experimental autoimmune encephalomyelitis, correlating with disease severity. Daily administration (10 mg/kg/day i.p.) of a 19-mer antisense oligonucleotide specific for XIAP (ASO-XIAP) abolished disease-associated XIAP mRNA and protein expression, and given from day of onset, alleviated experimental autoimmune encephalomyelitis and prevented relapses. Prophylactic treatment also reduced XIAP expression and prevented disease. Random or 5-base mismatched ASO was not inhibitory, and ASO-XIAP did not affect T cell priming. In ASO-XIAP-treated animals, infiltrating cells and inflammatory foci were dramatically reduced within the CNS. Flow cytometry showed an 88-93% reduction in T cells. The proportion of TUNEL(+) apoptotic CD4(+) T cells in the CNS was increased from <1.6 to 26% in ASO-XIAP-treated mice, and the proportion of Annexin V-positive CD4(+) T cells in the CNS increased. Neurons and oligodendrocytes were not affected; neither did apoptosis increase in liver, where XIAP knockdown also occurred. ASO-XIAP increased susceptibility of T cells to activation-induced apoptosis in vitro. Our results identify XIAP as a critical controller of apoptotic susceptibility of effector T cell function.
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