Activation Of Antioxidant Pathway Research Articles

Increasing GSH-Px Activity and Activating Wnt Pathway Promote Fine Wool Growth in FGF5-Edited Sheep.

Fibroblast growth factor 5 (FGF5) plays key roles in promoting the transition from the anagen to catagen during the hair follicle cycle. The sheep serves as an excellent model for studying hair growth and is frequently utilized in various research processes related to human skin diseases. We used the CRISPR/Cas9 system to generate four FGF5-edited Dorper sheep and only low levels of FGF5 were detected in the edited sheep. The density of fine wool in GE sheep was markedly increased, and the proportion of fine wool with a diameter of 14.4-20.0 μm was significantly higher. The proliferation signal in the skin of gene-edited (GE) sheep was stronger than in wild-type (WT) sheep. FGF5 editing decreased cortisol concentration in the skin, further activated the activity of antioxidant enzymes such as Glutathione peroxidase (GSH-Px), and regulated the expression of Wnt signaling pathways containing Wnt agonists (Rspondins, Rspos) and antagonists (Notum) in hair regeneration. We suggest that FGF5 not only mediates the activation of antioxidant pathways by cortisol, which constitutes a highly coordinated microenvironment in hair follicle cells, but also influences key signals of the Wnt pathway to regulate secondary hair follicle (SHF) development. Overall, our findings here demonstrate that FGF5 plays a significant role in regulating SHF growth in sheep and potentially serves as a molecular marker of fine wool growth in sheep breeding.

Oxidative Stress in Spinocerebellar Ataxia Type 3 and Its Attenuation by Herbal Remedies in Traditional Chinese Medicine: A Systematic Review.

Spinocerebellar ataxia type 3 (SCA3) is an autosomal dominant neurodegenerative disorder that gives rise to motor incoordination and progressive functional disabilities. Although pharmacological interventions have revealed promising prospects in the management of SCA3, adverse effects may become unbearable. The use of herbal remedies in traditional Chinese medicine (TCM) may serve as potential alternative medicines to delay the progression of the disease. This systematic review is intended to identify, appraise, and summarize the findings of studies pertaining to the therapeutic roles of herbal remedies in TCM targeting oxidative stress in the management of SCA3. A literature search for relevant articles published from 1 January 2013 to 30 June 2023 in three databases, namely PubMed, Web of Science, and Scopus, was carried out according to the procedures of the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA). A total of ten preclinical studies met the inclusion criteria of the systematic review. We recognized the therapeutic potential of Brassica napus, Codonopsis pilosula, Curcuma sp., Gardenia jasminoides, Gastrodia elata, Ginkgo biloba, Glycyrrhiza inflata, Hericium erinaceus, Hyptis sp., Paeonia lactiflora, Panax ginseng, Poria cocos, Pueraria lobata, Rehmannia glutinosa, and Scrophularia ningpoensis. We identified the types of preclinical models expressing polyglutamine (polyQ) expanded mutant protein (mATXN3), inducers of oxidative stress that mimic the SCA3 pathogenesis, and effective doses of the herbal remedies. The modes of action contributing to the attenuation of oxidative stress are activation of antioxidant pathways, ubiquitin-proteasome system and autophagy, regulation of apoptosis, proinflammatory signaling pathway and chaperones, regulation of mitochondrial function and biogenesis, and restoration of neurotransmission and synaptic plasticity. In conclusion, herbal remedies in TCM may possibly delay the progression of SCA3, therefore providing justification for clinical trials.

TRIM21 Promotes Oxidative Stress and Ferroptosis through the SQSTM1-NRF2-KEAP1 Axis to Increase the Titers of H5N1 Highly Pathogenic Avian Influenza Virus.

Tripartite motif-containing protein 21 (TRIM21) is involved in signal transduction and antiviral responses through the ubiquitination of protein targets. TRIM21 was reported to be related to the imbalance of host cell homeostasis caused by viral infection. Our studies indicated that H5N1 highly pathogenic avian influenza virus (HPAIV) infection up-regulated TRIM21 expression in A549 cells. Western blot and qPCR results showed that knockdown of TRIM21 alleviated oxidative stress and ferroptosis induced by H5N1 HPAIV and promoted the activation of antioxidant pathways. Co-IP results showed that TRIM21 promoted oxidative stress and ferroptosis by regulating the SQSTM1-NRF2-KEAP1 axis by increasing SQSTM1 K63-linked polyubiquitination under the condition of HPAIV infection. In addition, TRIM21 attenuated the inhibitory effect of antioxidant NAC on HPAIV titers and enhanced the promoting effect of ferroptosis agonist Erastin on HPAIV titers. Our findings provide new insight into the role of TRIM21 in oxidative stress and ferroptosis induced by viral infection.

Druggable redox pathways against Mycobacterium abscessus in cystic fibrosis patient-derived airway organoids.

Mycobacterium abscessus (Mabs) drives life-shortening mortality in cystic fibrosis (CF) patients, primarily because of its resistance to chemotherapeutic agents. To date, our knowledge on the host and bacterial determinants driving Mabs pathology in CF patient lung remains rudimentary. Here, we used human airway organoids (AOs) microinjected with smooth (S) or rough (R-)Mabs to evaluate bacteria fitness, host responses to infection, and new treatment efficacy. We show that S Mabs formed biofilm, and R Mabs formed cord serpentines and displayed a higher virulence. While Mabs infection triggers enhanced oxidative stress, pharmacological activation of antioxidant pathways resulted in better control of Mabs growth and reduced virulence. Genetic and pharmacological inhibition of the CFTR is associated with better growth and higher virulence of S and R Mabs. Finally, pharmacological activation of antioxidant pathways inhibited Mabs growth, at least in part through the quinone oxidoreductase NQO1, and improved efficacy in combination with cefoxitin, a first line antibiotic. In conclusion, we have established AOs as a suitable human system to decipher mechanisms of CF-driven respiratory infection by Mabs and propose boosting of the NRF2-NQO1 axis as a potential host-directed strategy to improve Mabs infection control.

Plasma-derived extracellular vesicles released after endurance exercise exert cardioprotective activity through the activation of antioxidant pathways

Cardiovascular diseases (CVD) can cause various conditions, including an increase in reactive oxygen species (ROS) levels that can decrease nitric oxide (NO) availability and promote vasoconstriction, leading to arterial hypertension. Physical exercise (PE) has been found to be protective against CVD by helping to maintain redox homeostasis through a decrease in ROS levels, achieved by increased expression of antioxidant enzymes (AOEs) and modulation of heat shock proteins (HSPs). Extracellular vesicles (EVs) circulating in the body are a major source of regulatory signals, including proteins and nucleic acids. Interestingly, the cardioprotective role of EVs released after PE has not been fully described.The aim of this study was to investigate the role of circulating EVs, obtained through Size Exclusion Chromatography (SEC) of plasma samples from healthy young males (age: 26.95 ± 3.07; estimated maximum oxygen consumption rate (VO2max): 51.22 ± 4.85 (mL/kg/min)) at basal level (Pre_EVs) and immediately after a single bout of endurance exercise (30’ treadmill, 70% heart rate (HR) -Post_EVs). Gene ontology (GO) analysis of proteomic data from isolated EVs, revealed enrichment in proteins endowed with catalytic activity in Post_EVs, compare to Pre_EVs, with MAP2K1 being the most significantly upregulated protein. Enzymatic assays on EVs derived from Pre and Post samples showed increment in Glutathione Reductase (GR) and Catalase (CAT) activity in Post_EVs. At functional level, Post_EVs, but not Pre_EVs, enhanced the activity of antioxidant enzymes (AOEs) and reduced oxidative damage accumulation in treated human iPS-derived cardiomyocytes (hCM) at basal level and under stress conditions (Hydrogen Peroxide (H2O2) treatment), resulting in a global cardioprotective effect.In conclusion, our data demonstrated, for the first time, that a single 30-min endurance exercise is able to alter the cargo of circulating EVs, resulting in cardioprotective effect through antioxidant activity.

Adipoquinas, envejecimiento y daño vascular

The main physiologic function of adipokines on the energy metabolism and the cardiovascular homeostasis, among others, are well known. In the last years, our laboratory is providing increasing evidence about the possible role of adipokines as mediators of vascular damage, by inducing different pro-inflammatory and pro-senescent mechanisms. The secretion of adipokines like visfatin or dipeptidylpeptidase-4 (DPP4) by adipose tissue, particularly by the visceral fat, is enhanced in obese and type 2 diabetic patients. Both adipokines are enzymes metabolically active that can induce the activation of specific receptors, namely type 4 “toll like receptors” (TLR4) for visfatin and “protease activated receptor 2” (PAR2) for DPP4. Stimulation of such receptors is triggering well known pro-inflammatory and pro-senescent mechanisms, like the nuclear transcription factor-κB (NF-κB) or the inflammasome NLRP3 (“nucleotide-binding, leucine-rich-repeat, pyrin domain containing 3”), a cell structure that transforms the immature forms of classic cytokines in their active derivates, interleukin-1β (IL-1β) or interleukin 18 (IL-18). It is worth to note that these classic cytokines are the final effectors for the harmful effects of the adipokines and its blockade can be a very relevant therapeutic approach. In our experiments, the specific antagonism of IL-1 receptors with anakinra prevents the inflammatory and senescent effects evoked by visfatin and DPP4. Moreover, this finding is in agreement with data from other researchers, as well as with the results of the CANTOS clinical trial, which demonstrate a very important cardioprotective cardiovascular effect mediated by the anti-inflammatory effect of the monoclonal antibody canakinumab. On the other hand, we have also provided experimental evidence about possible vasculopotective adipokines, such angiotensin-(1-7), which is able to induce, through activation of Mas receptors, the expression of klotho protein and the activation of antioxidant pathways, like the well known Nrf2-HO-1 (“nuclear factor erythroid-2 y hemoygenase-1”).

ATF4 Protects the Heart From Failure by Antagonizing Oxidative Stress.

Cellular redox control is maintained by generation of reactive oxygen/nitrogen species balanced by activation of antioxidative pathways. Disruption of redox balance leads to oxidative stress, a central causative event in numerous diseases including heart failure. Redox control in the heart exposed to hemodynamic stress, however, remains to be fully elucidated. Pressure overload was triggered by transverse aortic constriction in mice. Transcriptomic and metabolomic regulations were evaluated by RNA-sequencing and metabolomics, respectively. Stable isotope tracer labeling experiments were conducted to determine metabolic flux in vitro. Neonatal rat ventricular myocytes and H9c2 cells were used to examine molecular mechanisms. We show that production of cardiomyocyte NADPH, a key factor in redox regulation, is decreased in pressure overload-induced heart failure. As a consequence, the level of reduced glutathione is downregulated, a change associated with fibrosis and cardiomyopathy. We report that the pentose phosphate pathway and mitochondrial serine/glycine/folate metabolic signaling, 2 NADPH-generating pathways in the cytosol and mitochondria, respectively, are induced by transverse aortic constriction. We identify ATF4 (activating transcription factor 4) as an upstream transcription factor controlling the expression of multiple enzymes in these 2 pathways. Consistently, joint pathway analysis of transcriptomic and metabolomic data reveal that ATF4 preferably controls oxidative stress and redox-related pathways. Overexpression of ATF4 in neonatal rat ventricular myocytes increases NADPH-producing enzymes' whereas silencing of ATF4 decreases their expression. Further, stable isotope tracer experiments reveal that ATF4 overexpression augments metabolic flux within these 2 pathways. In vivo, cardiomyocyte-specific deletion of ATF4 exacerbates cardiomyopathy in the setting of transverse aortic constriction and accelerates heart failure development, attributable, at least in part, to an inability to increase the expression of NADPH-generating enzymes. Our findings reveal that ATF4 plays a critical role in the heart under conditions of hemodynamic stress by governing both cytosolic and mitochondrial production of NADPH.

Green Synthesis of Silymarin-Chitosan Nanoparticles as a New Nano Formulation with Enhanced Anti-Fibrotic Effects against Liver Fibrosis.

Background: Silymarin (SIL) has long been utilized to treat a variety of liver illnesses, but due to its poor water solubility and low membrane permeability, it has a low oral bioavailability, limiting its therapeutic potential. Aim: Design and evaluate hepatic-targeted delivery of safe biocompatible formulated SIL-loaded chitosan nanoparticles (SCNPs) to enhance SIL’s anti-fibrotic effectiveness in rats with CCl4-induced liver fibrosis. Methods: The SCNPs and chitosan nanoparticles (CNPs) were prepared by ionotropic gelation technique and are characterized by physicochemical parameters such as particle size, morphology, zeta potential, and in vitro release studies. The therapeutic efficacy of successfully formulated SCNPs and CNPs were subjected to in vivo evaluation studies. Rats were daily administered SIL, SCNPs, and CNPs orally for 30 days. Results: The in vivo study revealed that the synthesized SCNPs demonstrated a significant antifibrotic therapeutic action against CCl4-induced hepatic injury in rats when compared to treated groups of SIL and CNPs. SCNP-treated rats had a healthy body weight, with normal values for liver weight and liver index, as well as significant improvements in liver functions, inflammatory indicators, antioxidant pathway activation, and lipid peroxidation reduction. The antifibrotic activities of SCNPs were mediated by suppressing the expression of the main fibrosis mediators TGFβR1, COL3A1, and TGFβR2 by boosting the hepatic expression of protective miRNAs; miR-22, miR-29c, and miR-219a, respectively. The anti-fibrotic effects of SCNPs were supported by histopathology and immunohistochemistry (IHC) study. Conclusions: According to the above results, SCNPs might be the best suitable carrier to target liver cells in the treatment of liver fibrosis.

Discovering the Potential of Natural Antioxidants in Age-Related Macular Degeneration: A Review.

Age-related macular degeneration (AMD) is a multifactorial disease associated with anatomical changes in the inner retina. Despite tremendous advances in clinical care, there is currently no cure for AMD. This review aims to evaluate the published literature on the therapeutic roles of natural antioxidants in AMD. A literature search of PubMed, Web of Science and Google Scholar for peer-reviewed articles published between 1 January 2011 and 31 October 2021 was undertaken. A total of 82 preclinical and 18 clinical studies were eligible for inclusion in this review. We identified active compounds, carotenoids, extracts and polysaccharides, flavonoids, formulations, vitamins and whole foods with potential therapeutic roles in AMD. We evaluated the integral cellular signaling pathways including the activation of antioxidant pathways and angiogenesis pathways orchestrating their mode of action. In conclusion, we examined the therapeutic roles of natural antioxidants in AMD which warrant further study for application in clinical practice. Our current understanding is that natural antioxidants have the potential to improve or halt the progression of AMD, and tailoring therapeutics to the specific disease stages may be the key to preventing irreversible vision loss.

Abstract 14368: Valproic Acid Activates Irg1-Itaconate and Improve Donor Heart Function After Extended Storage Time

Background: Cold ischemic storage and subsequent reperfusion injury contributes to the primary graft dysfunction after heart transplantation. Valproic acid (VPA) has been shown to reduce myocardial infarction injury. However, the role of VPA in donor heart preservation is unknown. Methods: We investigate the role of VPA on donor heart preservation using both ex-vivo perfusion and heterotopic transplantation in murine models. Results: Our results demonstrate the administration of VPA to histidine-tryptophan-ketoglutarate (HTK) cardioplegia can significantly improve donor heart function after perfusion with extend storage time (16 hours, n=8). Specifically, VPA reduced the accumulation of succinate, a metabolite that involved in reperfusion injury, during cold storage. We further show that VPA increases the Irg1 expression. Irg1 is known to produce itaconate and suppress the succinate accumulation during ischemia. We found that the effect of VPA in donor heart function is significantly reduced in Irg1 knockout mice compared to wild type mice after 16 hours of cold storage. Furthermore, VPA reduced oxidative stress of donor heart via Nrf2 antioxidant pathway activation. However, Irg1 deletion decreased the induction of antioxidant genes expression by VPA. Administration of 4-octyl-itaconate rescued donor heart function after extended storage. Conclusion: Our results demonstrate that VPA could improve donor heart function and ischemic tolerance. This is in part through Irg1/itaconate mediated anti-oxidative activities. Importantly, VPA preserved cardiac function after perfusion with extended storage times. This strategy can extend the tolerable ischemic time for donor heart for heart transplant and may reduce the incidence of primary graft dysfunction.

Synthetic Triterpenoid RTA-408: Limits Radiation Damage to Normal Tissue.

To assess the efficacy and mechanism of action of a novel approach to mitigate acute and chronic radiation toxicity in a validated animal model. Randomized, prospective study using an in vivo rat model. Experimental animal study utilizing Sprague-Dawley rats divided into three cohorts: 1) radiation + dimethyl sulfoxide (DMSO) (inert vehicle); 2) radiation + RTA-408 (therapeutic drug); and 3) no radiation + DMSO. All animals in the radiation cohorts underwent 40 Gy of radiation with subsequent inferior epigastric axial rotational flap 30 days later in all cohorts with percentage of flap necrosis and vascular density calculated by blinded observers. In a second experiment, an additional three cohorts, underwent serial punch biopsies of the abdominal skin before, during, and after radiation and drug/vehicle control treatment. Transcriptome analysis utilizing gene set enrichment analysis and digital polymerase chain reaction were performed at various time points. The first experiment revealed average flap necrosis of 20% (95% confidence interval [CI] 16-45) in the radiation control group, 3% (95% CI 0-11) in the nonirradiated control, and 3% (95% CI 0.2-10) in the radiation group treated with RTA-408. Vascular density was preserved in the treatment group as compared to the radiated control. Nine rats were included in the second experiment, and transcriptome analyses in the treatment group revealed robust activation of antioxidant pathways with induced expression of genes associated with hypoxia and adipogenesis/angiogenesis. Administration of RTA-408 during radiation treatment in a rat model resulted in transcriptome changes which appear to mitigate the toxic effects of radiation, preserving capillary networks and improving flap survival and tissue healing after subsequent surgery. Foundational Evidence, Animal Research Laryngoscope, 132:1196-1204, 2022.

Tannic acid protects aged brain against cerebral hypoperfusion via modulation of Nrf2 and inflammatory pathways

Current study purposed to investigate the neuroprotective effects of Tannic Acid (TA) on mild chronic cerebral hypoperfusion model in rats. Male Wistar rats were subjected to permanent Unilateral Common Carotid Artery Occlusion (UCCAO), followed by TA treatment (0.05% w/v) in drinking water for one month. Nuclear factor erythroid 2-related factor 2 (Nrf2), NAD(P)H: quinone oxidoreductase 1 (NQO-1), heme oxygenase-1 (HO-1), factor kappa-light-chain-enhancer of activated B cells (NF-κB), tumor necrosis factor-α (TNF-α), B-cell lymphoma 2 (Bcl-2), Bcl-2-associated X protein (Bax), caspase-3, blood triglyceride, blood glucose, and liver enzymes’ activity were detected after the experimental period. Also, behavioral tests, hematoxylin and eosin (H&E) staining, and PET scan were performed after treatment. Post-treatment of TA improved locomotion and memory function (P < 0.001), and reduced neural cell death (P < 0.001) in the treatment group compared to UCCAO rats. Furthermore, long-term TA treatment significantly increased the levels of Nrf2 (P < 0.001), NQO-1 (P < 0.001), and HO-1 (P < 0.001) in the hippocampus of the treatment group compared to the UCCAO group. TA consumption in the treatment group applied its anti-inflammatory effects via reducing the activity of NF-κB and TNF-α in comparison with the UCCAO group (P < 0.001 for both). Blood triglyceride, blood glucose, and liver enzymes did not change considerably in the groups (P > 0.05). The current results indicate that long-term post-treatment of TA exhibits protective effects against memory deficit and motor dysfunction. The cellular mechanism of TA in hypoperfused rats might be associated with the activation of antioxidant pathways, especially the Nrf2 pathway, and suppressing inflammatory factors like NF-κB and TNF-α.

Selenium Biofortification of Soybean Seeds Influences Physiological Responses of Seedlings to Osmotic Stress.

Climate change poses a serious threat to agricultural production. Water deficit in agricultural soils is one of the consequences of climate change that has a negative impact on crop growth and yield. Selenium (Se) is known to be involved in plant defense against biotic and abiotic stress through metabolic, structural, and physiological activity in higher plants. The aim of this study was to investigate the physiological response of Se-biofortified soybean (Glycine max (L.) Merrill) seedlings under osmotic stress. For this research, we used biofortified soybean grain obtained after foliar Se biofortification in 2020. The experiment was conducted in a growth chamber with two cultivars (Lucija and Sonja) grown on filter paper in three replicates. The experiment was carried out with two watering treatments: distilled water (PEG-0) and 2.5% polyethylene glycol 6000 (PEG-2.5) on Se-biofortified seeds (Se) and nonbiofortified seeds (wSe). Contents of lipid peroxidation product (LP), free proline (PRO), total phenolic content (TP), ferric reducing antioxidant power (FRAP), and ascorbic acid (AA) were analyzed in 7-days-old seedlings. Significant differences were detected in the Se content of soybean grains between the two cultivars. A milder reaction to PEG-2.5 was observed in cultivar Lucija in both Se and wSe treatments, which might represent the mitigating effects of Se on osmotic stress in this cultivar. Contrarily, in cultivar Sonja, Se adversely affected all analyzed traits in the PEG-2.5 treatment. Ultimately, Se is a pro-oxidant in Sonja, whereas it represents an anti-oxidant in Lucija. In conclusion, different soybean cultivars show contrasting physiological reactions to both osmotic stress and Se. However, the activation of antioxidant pathways in Sonja can also be interpreted as added value in soybean seedlings as a functional food.

Streptococcus thermophilus extends lifespan through activation of DAF-16-mediated antioxidant pathway in Caenorhabditis elegans.

Streptococcus thermophilus bacteria, which are widely used as fermented starter for dairy production, exert various beneficial health effects. Nevertheless, even though pro-longevity effects of various probiotics have been reported, no report has described Streptococcus thermophilus effects on longevity. This study was conducted to evaluate Streptococcus thermophilus effects on lifespan extension and to elucidate the Streptococcus thermophilus-mediated longevity mechanism using Caenorhabditis elegans worms as a model animal. They were fed standard food (Escherichia coli OP50) or Streptococcus thermophilus from the young adult stage. Feeding with Streptococcus thermophilus, compared to Escherichia coli OP50, to Caenorhabditis elegans extend the lifespan, reduced lipofuscin accumulation, and maintain vigorous locomotion. Feeding with Streptococcus thermophilus did not alter the worm growth curve or the offspring number, indicating that the Streptococcus thermophilus-mediated lifespan extension is not attributable to caloric restriction. The qRT-PCR data showed that Streptococcus thermophilus increased the expression of daf-16 and some of its downstream antioxidant genes. Furthermore, the pro-longevity effects of Streptococcus thermophilus were decreased in loss-of-function mutant of daf-16. Results show that Streptococcus thermophilus extends the lifespan of Caenorhabditis elegans through DAF-16-mediated antioxidant pathway activation.

Abstract 2262: Comprehensive characterization of cancer drivers and therapeutic targets

Abstract Cancer is a family of diseases caused primarily by genetic “driver” mutations. Building upon a large-scale Pan-cancer study of ours, we expand our investigation to the proteogenomic level (including global proteome and phosphoproteome data) drawing &amp;gt;1,000 cases over 10 cancer types (BR, ccRCC, CO, EC, GBM, HNSCC, LSCC, LUAD, OV, PDAC) from the Clinical Proteomic Tumor Analysis Consortium (CPTAC). We concentrate on 6 aspects of drivers: (1) We identify genomic &amp; epigenomic drivers (mutations, SVs, CNVs, fusions, etc.), estimate their frequencies, and investigate their exclusivities &amp; co-occurrences. (2) We investigate the proteogenomic impact of drivers, especially recurrent ones, e.g. cis/trans effects, on miRNA, RNA, and protein expression. (3) We investigate their impact on Post Transcriptional Modifications (PTMs), comparing levels of protein and PTM, e.g. for one- and two-hit events. For example, we found upregulation of PTPN11 phosphorylation sites in EGFR-altered LUAD, GBM, and HNSCC samples, suggesting a pan-cancer effect, and observed high NRF2 phosphorylation and activation of antioxidant pathways in KEAP1 or NRF2-altered samples in lung cancer (LSCC and LUAD); (4) We compare tumors and normal adjacent tissues (NATs) to identify key protein level changes and regulations for oncogenic pathways and further examine cell type composition, proteomic variation, and association with driver mutations. For instance, we found VHL mutations often signify enrichment of endothelial cells. (5) We capture potential Significant Protein Drivers (SPRDs), independent of DNA level changes, to uncover hidden drivers in cause-ambiguous tumors. (6) We examine actionable driver alterations and expression changes across proteogenomic levels and associations with immune infiltration and immunogenic peptides to understand targeting, therapeutic implications and, influence on tumor micro-environment. We comment upon the clinical utility of comprehensive proteogenomics, especially in cases where a strictly genomic assessment is inconclusive. Proteomics can further prioritize gene targets with additional layers of evidence for differential expression among features such as cell-type composition, anatomical location, disease stage, and immune infiltration. Our approach has the potential for identifying cancer-type-specific and non-specific novel druggable targets, which would provide new biological and clinical insights into cancer. Citation Format: Yize Li, Oncogenic Drivers and Pathways GroupClinical Proteomic Tumor Analysis Consortium. Comprehensive characterization of cancer drivers and therapeutic targets [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 2262.

Inhibition of ROS-Induced Nrf2 Antioxidant Pathway Activation May Explain Synergy between Venetoclax and Hypomethylating Agents Against Acute Myeloid Leukemia

The selective Bcl-2 inhibitor venetoclax (ABT-199) has shown potent anti-leukemic activity in combination with hypomethylating agents (HMA) against Acute Myeloid Leukemia (AML). Induction of Reactive Oxygen Species (ROS) mediates the cytotoxicity of various AML therapies including HMA. Induction of ROS concurrently activates the Nrf2 antioxidant response pathway which in turn results in induction of antioxidant enzymes that in turn neutralize ROS. Nrf2 is critical to hematopoietic stem cell function and high Nrf2 expression has been implicated in chemoresistance of AML stem cells. In this study, we demonstrate that Nrf2 inhibition by venetoclax is an additional mechanism responsible for the marked anti-leukemic activity in AML seen with the combination of HMAs and venetoclax.Venetoclax increased cellular and mitochondrial ROS in AML cells. Venetoclax showed strong synergy with decitabine in inducing apoptosis and further augmented ROS induction by HMA in AML cell lines including MV-4-11, KG1A and Kasumi. For example, the percentage of apoptotic cells increased from 6% or 20.3% after decitabine or ABT-199 treatment alone, respectively, to 37.6 % after treatment with the combination in MV-4-11 cells. Cytotoxicity of HMA was inhibited by the ROS scavenger N-acetyl cysteine. Knock down of Nrf2 augmented the cytotoxicity of HMA against AML cells. Indeed, Nrf2 depletion by siRNA increases apoptotic ratio from 3.8% to 18.5% after treatment with 5-Azacitidine and from 5.4% to 32.8% after treatment with decitabine. Increased nuclear translocation of Nrf2 as well as induction of antioxidant enzyme response was observed after treatment of AML cells with decitabine. Treatment of AML cell lines as well as primary AML cells with venetoclax inhibited the increased nuclear translocation of Nrf2 and induction of downstream antioxidant enzymes including HO-1 and NQO1 seen in response to decitabine treatment alone (Figure 1, top panels). Using immunofluorescence and immunoprecipitation experiments we further show that Bcl-2 associates with Nrf2, and venetoclax treatment leads to dissociation of Bcl-2 from the Nrf2-Keap1 complex thereby targeting Nrf2 for ubiquitination and proteosomal degradation (Figure 1, bottom panels).Our results demonstrate an undiscovered mechanism namely Nrf2 inhibition that may underlie the synergistic effect of HMA and venetoclax in AML cells. Nrf2 inhibition by venetoclax has the potential to target leukemia stem cells and could explain the impressive results seen in early clinical studies of AML using combination therapy with HMA and venetoclax. We also show the novel role of Bcl-2 in regulating Nrf2 by its association with Nrf2 and Keap1 and further show increased ubiquitination of Nrf2 after venetoclax treatment. Dissociation of Bcl-2 binding to Nrf2/Keap1 complex leading to increased proteosomal degradation may explain the inhibition of nuclear Nrf 2 translocation seen with combination of venetoclax and HMA treatment in AML cells. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Toxicology and pharmacology of synthetic organoselenium compounds: an update.

Here, we addressed the pharmacology and toxicology of synthetic organoselenium compounds and some naturally occurring organoselenium amino acids. The use of selenium as a tool in organic synthesis and as a pharmacological agent goes back to the middle of the nineteenth and the beginning of the twentieth centuries. The rediscovery of ebselen and its investigation in clinical trials have motivated the search for new organoselenium molecules with pharmacological properties. Although ebselen and diselenides have some overlapping pharmacological properties, their molecular targets are not identical. However, they have similar anti-inflammatory and antioxidant activities, possibly, via activation of transcription factors, regulating the expression of antioxidant genes. In short, our knowledge about the pharmacological properties of simple organoselenium compounds is still elusive. However, contrary to our early expectations that they could imitate selenoproteins, organoselenium compounds seem to have non-specific modulatory activation of antioxidant pathways and specific inhibitory effects in some thiol-containing proteins. The thiol-oxidizing properties of organoselenium compounds are considered the molecular basis of their chronic toxicity; however, the acute use of organoselenium compounds as inhibitors of specific thiol-containing enzymes can be of therapeutic significance. In summary, the outcomes of the clinical trials of ebselen as a mimetic of lithium or as an inhibitor of SARS-CoV-2 proteases will be important to the field of organoselenium synthesis. The development of computational techniques that could predict rational modifications in the structure of organoselenium compounds to increase their specificity is required to construct a library of thiol-modifying agents with selectivity toward specific target proteins.

Anti‐Interleukin‐16 Neutralizing Antibody Treatment Alleviates Sepsis‐Induced Cardiac Injury and Dysfunction via the Nuclear Factor Erythroid‐2 Related Factor 2 Pathway in Mice

Several interleukin (IL) members have been reported to participate in sepsis. In this study, the effects of IL-16 on sepsis-induced cardiac injury and dysfunction were examined, and the related mechanisms were detected. IL-16 expression in septic mice was first measured, and the results showed that both cardiac and serum IL-16 expression levels were increased in mice with sepsis induced by LPS or cecal ligation and puncture (CLP) compared with control mice. Then, IL-16 was neutralized, and the effects on lipopolysaccharide- (LPS-) induced cardiac injury were detected. The results showed that an anti-IL-16 neutralizing antibody (nAb) significantly reduced mortality and increased serum lactate dehydrogenase (LDH), creatine kinase myocardial bound (CK-MB), and cardiac troponin T (cTnT) levels while improving cardiac function in mice with LPS-induced sepsis. Neutralization of IL-16 also increased the activation of antioxidant pathways and the expression of antioxidant factors in septic mice while decreasing the activation of prooxidant pathways and the expression of prooxidants. Treatment with the anti-IL-16 nAb increased mitochondrial apoptosis-inducing factor (AIF) expression, decreased nuclear AIF and cleaved poly-ADP-ribose polymerase (PARP) expression, and decreased TUNEL-positive cell percentages in LPS-treated mice. Additionally, treatment with CPUY192018, the nuclear factor erythroid-2 related factor 2 (Nrf2) pathway, significantly increased mortality and reversed the above effects in mice treated with LPS and the anti-IL-16 nAb. Our results showed that the anti-IL-16 nAb regulates oxidative stress through the Nrf2 pathway and participates in the regulation of cardiac injury in septic mice. Neutralization of IL-16 may be a beneficial strategy for the prevention of cardiac injury and dysfunction in sepsis patients.

Testosterone replacement therapy in blood donors modulates erythrocyte metabolism and susceptibility to hemolysis in cold storage.

Red blood cells (RBCs) derived from patients who receive testosterone replacement therapy (TRT) may be considered eligible for component production and transfusion. The aim of this study was to identify testosterone-dependent changes in RBC metabolism and to evaluate its impact on susceptibility to hemolysis during cold storage. We characterized stored RBCs from two cohorts of TRT patients who were matched with control donors (no TRT) based upon sex, age, and ethnicity. We further evaluated the impact of testosterone deficiency (orchiectomy) on RBC metabolism in FVB/NJ mice. RBC metabolites were quantified by ultra-high-pressure liquid chromatography-mass spectrometry. RBC storage stability was determined in RBC units from TRT and controls by quantifying storage, osmotic, and oxidative hemolysis. Orchiectomy in mice was associated with significant (P < 0.05) changes in RBC metabolism as compared with intact males including increased levels of acyl-carnitines, long-chain fatty acids (eg, docosapentaenoic acids), arginine, and dopamine. Stored RBCs from TRT patients exhibited higher levels of pentose phosphate pathway metabolites, glutathione, and oxidized purines (eg, hypoxanthine), suggestive of increased activation of antioxidant pathways in this group. Further analyses indicated significant changes in free fatty acids and acyl-carnitines in response to testosterone therapies. With regard to hemolysis, TRT was associated with enhanced susceptibility to osmotic hemolysis. Correlation analyses identified acyl-carnitines as significant modifiers of RBC predisposition to osmotic and oxidative hemolysis. These observations provide new insights into testosterone-mediated changes in RBC metabolome and biology that may impact the storage capacity and posttransfusion efficacy of RBCs from TRT donors.

Partial impairment of late-stage autophagic flux in murine splenocytes leads to sqstm1/p62 mediated nrf2-keap1 antioxidant pathway activation and induced proteasome-mediated degradation in malaria

Splenomegaly, a major symptom in Plasmodium infection, is extensively studied for its immunopathological role in mice malaria model infected with Plasmodium berghei ANKA. The status of autophagic regulation in hosts in malaria pathogenesis remains unreported till date. This study demonstrated the autophagy, proteasomal degradation and NRF2-KEAP1 antioxidant pathway status in the host during Plasmodium infection taking murine spleen as our organ of interest. Initial staining and autophagic gene expression indicate a possibility of autophagic pathway activation. Although the conversion of LC3A to LC3B and lysosome-autophagosome fusion increases, the final degradation step remains incomplete. Resultant upregulation of p62 and its altered phosphorylated status enhances its binding to keap1 causing NRF2 translocation to the nucleus. NRF2 act as transcription factor upregulating p62 level itself leading to an autoinduction loop of p62 expression. Interestingly, enhancement of P62 interaction with proteasome subunit RPT1 indicates a possible role in transporting ubiquitinated cargo to proteasome complex. Ubiquitination level increased with subsequent upregulation of all three modes of proteasomal degradation i.e trypsin-like, caspase-like and especially chymotrypsin-like. Sqstm1/p62 plays a critical central role in regulating autophagy, proteasomal degradation, and NRF2-KEAP1 pathway. The incomplete autophagic flux in the final step may be a key therapeutic target, as autophagic degradation and subsequent pathogenic peptide presentation is of utmost necessity for downstream immune response.