Intrinsic Oxidative Stress Research Articles

Stress response regulation of mRNA translation: Implications for antioxidant enzyme expression in cancer

From tumorigenesis to advanced metastatic stages, tumor cells encounter stress, ranging from limited nutrient and oxygen supply within the tumor microenvironment to extrinsic and intrinsic oxidative stress. Thus, tumor cells seize regulatory pathways to rapidly adapt to distinct physiologic conditions to promote cellular survival, including manipulation of mRNA translation. While it is now well established that metastatic tumor cells must up-regulate their antioxidant capacity to effectively spread and that regulation of antioxidant enzymes is imperative to disease progression, relatively few studies have assessed how translation and the hijacking of RNA systems contribute to antioxidant responses of tumors. Here, we review the major stress signaling pathways involved in translational regulation and discuss how these are affected by oxidative stress to promote prosurvival changes that manipulate antioxidant enzyme expression. We describe how tumors elicit these adaptive responses and detail how stress-induced translation can be regulated by kinases, RNA-binding proteins, RNA species, and RNA modification systems. We also highlight opportunities for further studies focused on the role of mRNA translation and RNA systems in the regulation of antioxidant enzyme expression, which may be of particular importance in the context of metastatic progression and therapeutic resistance.

Effect of dexpanthenol on cyclophosphamide-induced ovarian toxicity: a histological and molecular study in rats

Research questionDoes dexpanthenol work as an effective therapeutic agent against cyclophosphamide (CYC)-induced premature ovarian failure (POF) in rats? DesignA total of 28 female Wistar Albino rats were randomly divided into four groups (n = 7 per group). The POF and POF plus dexpanthenol groups were intraperitoneally administered CYC at an initial dose of 50 mg/kg, followed by 8 mg/kg for 14 days. The dexpanthenol and POF plus dexpanthenol groups were both intraperitoneally administered dexpanthenol at a dose of 500 mg/kg/day for 15 days. ResultsIn the group administered CYC, the following was observed: a decrease in the ovarian index; a decrease in the numbers of primordial, primary, secondary and antral follicles; an increase in the number of corpus luteum and atretic follicles; a decrease in proliferation cell nuclear antigen immunoreactivity; a significant reduction in anti-Müllerian hormone and oestradiol levels; and an increase in serum FSH levels compared with controls. Dexpanthenol, on the other hand, reversed these effects. Quantitative reverse transcription polymerase chain reaction analyses showed that dexpanthenol increased Bcl-2, Akt1, mTOR, Nrf2 and HO-1 in CYC-induced ovarian tissues, but decreased Bax, Cas3, Hsp27, Hsp70, and Hsp90. Dexpanthenol treatment has a potential for inhibiting the intrinsic apoptotic pathway and oxidative stress levels in ovarian tissues via the downregulation of the mRNA expression of heat shock proteins and the activation of Nrf2/HO-1 pathways. ConclusionsOur findings demonstrated that dexpanthenol is an effective agent against POF caused by CYC; however, further experimental and clinical data are needed to use it effectively.

Selective vulnerability of hippocampal CA1 and CA3 pyramidal cells: What are possible pathomechanisms and should more attention be paid to the CA3 region in future studies?

Transient ischemia and reperfusion selectively damage neurons in brain, with hippocampal pyramidal cells being particularly vulnerable. Even within hippocampus, heterogeneous susceptibility is evident, with higher vulnerability of CA1 versus CA3 neurons described for several decades. Therefore, numerous studies have focused exclusively on CA1. Pediatric cardiac surgery is increasingly focusing on studies of hippocampal structures, and a negative impact of cardiopulmonary bypass on the hippocampus cannot be denied. Recent studies show a shift in selective vulnerability from neurons of CA1 to CA3. This review shows that cell damage is increased in CA3, sometimes stronger than in CA1, depending on several factors (method, species, age, observation period). Despite a highly variable pattern, several markers illustrate greater damage to CA3 neurons than previously assumed. Nevertheless, the underlying cellular mechanisms have not been fully deciphered to date. The complexity is reflected in possible pathomechanisms discussed here, with numerous factors (NMDA, kainate and AMPA receptors, intrinsic oxidative stress potential and various radicals, AKT isoforms, differences in vascular architecture, ratio of pro- and anti-apoptotic Bcl-2 factors, vulnerability of interneurons, mitochondrial dysregulation) contributing to either enhanced CA1 or CA3 vulnerability. Furthermore, differences in expressed genome, proteome, metabolome, and transcriptome in CA1 and CA3 appear to influence differential behavior after damaging stimuli, thus metabolomics-, transcriptomics-, and proteomics-based analyses represent a viable option to identify pathways of selective vulnerability in hippocampal neurons. These results emphasize that future studies should focus on the CA3 field in addition to CA1, especially with regard to improving therapeutic strategies after ischemic/hypoxic brain injury.

The Mechanism of Therapy Resistance By Lineage Plasticity in AML and How to Overcome It

Acute myeloid leukemia (AML) is an aggressive blood cancer driven predominantly by mutations in epigenetic regulators. This characteristic allows AML to thrive in a hostile and rapidly changing environment where it needs to adapt to stressors both extrinsic (chemotherapy) and intrinsic (replicative and oxidative stress) to survive. Our study is the first to demonstrate the mechanism of how AML uses epigenetic reprogramming to adjust to these stressors via lineage plasticity. Venetoclax and Azacitidine (Ven/Aza) co-treatment is increasingly the front-line therapy for AML, functioning through BH3 inhibition and DNA hypomethylation. However, most patients will relapse with a highly aggressive refractory AML. Some studies suggest that monocytic character of the AML is responsible for relapse, but more recent evidence demonstrates that Ven/Aza treatment itself can induce a lineage switch to monocytic character. A potent selection mechanism facilitated by epigenetic dysregulation generating plastic clones is likely driving resistance, though the mechanism remains unclear and represents a significant barrier for AML treatment. Using a combination of single cell transcriptomics with surface immunophenotype sequencing in multiple human xenograft and primary patient samples, we discovered that this plasticity is widespread and mediated by epigenetic reprogramming of the tumor in direct response to the therapy. Broadly, we found that AML responds to Ven/Aza by transitioning into a reversible state of cell cycle arrest with transcriptional character similar to hematopoietic stem cells. This transient state protects AML from Azacitidine through replication arrest and protects from Venetoclax by upregulation of anti-apoptotic transcriptional programs. These protections operate simultaneously with reprogramming events driven by a switch from MYC-driven transcriptional programs to a PU.1-driven program. Reprogramming is both facilitated by and dependent on repression of the Chromatin Assembly Factor 1 (CAF1) complex, a chromatin-bound master epigenetic regulator responsible for maintaining lineage fidelity through transcriptional regulation via its role as a nucleosome assembly factor. Repressing CAF1 facilitates reprogramming where the output is an altered cell population that is distinct from the initial tumor both transcriptionally and by ATACseq. AML can repeatedly return to this plastic state, regardless of the number of cycles of Ven/Aza, and introducing other cytotoxic compounds does not remove this plastic stem-like population but instead initiates the iterative process of reprogramming anew until a different resistant population emerges. To find therapeutic vulnerabilities, we generated several PDX models of Ven/Aza resistance so that we could track the stages of acquired resistance in real time. When single cell data from these experiments were projected onto a reference map of normal hematopoiesis, we found that Ven/Aza resistance stems from a plastic population of cells that actively samples the niche by generating short-lived multi-lineage hybrid differentiated products. While the predominating output of this process was monocytic, we found that the induced plastic stem-like population was durably retained even after Ven/Aza withdrawal. To undermine this process, we took advantage of the plastic state to drive differentiation through an external stimulus. We expected this strategy to be particularly effective because CAF1 was repressed (CAF1 repression facilitates differentiation in every metazoan model) and, based on our single cell data, these plastic populations of AML cells express pro-differentiation surface receptors. In particular, Ven/Aza induced the expression of CSF3R (the G-CSF receptor) which when stimulated, drives neutrophil maturation through the JAK/STAT pathway. While G-CSF as a single agent or in sequence with Ven/Aza was unable to achieve maturation, when used concurrently with Ven/Aza (or recapitulated by CAF1 depletion using shRNA) we were able to drive neutrophil differentiation in vitro and fully resolve primary mouse and PDX tumors in vivo, even when using Ven/Aza resistant tumors as starting material. Together, these data demonstrate the possibility that Ven/Aza resistance can be completely subverted by taking advantage of the plastic state induced by the therapy to stimulate differentiation and resolve the disease.

Identifying Gene-Treatment Interactions and Targetable Radiation Vulnerabilities in Glioblastoma through Coupling of In Vivo CRISPR Perturbation and Single Cell Transcriptomics

Glioblastoma (GBM) is an incurable brain tumor comprised of dynamic malignant cell states and microenvironment components that underlie treatment resistance. Here we use genome-wide CRISPR/Cas9 functional genomics to define biological drivers and therapeutic vulnerabilities across human and mouse GBM models. To interrogate these mechanisms in the context of the tumor microenvironment and in vivo physiology, we established in vivo Perturb-seq intracranially, a technique coupling functional genomics with single cell transcriptomics, where each cell is an individual experiment. Orthotopic intracranial tumor models were established using human (GBM6, GBM43) or mouse (GL261, SB28) GBM cells stably expressing CRISPR interference (CRISPRi) machinery. Perturb-seq target selection for phenotyping of gene-treatment interactions was performed using genome-wide CRISPRi screens ± radiotherapy in cell cultures. Dual sgRNA lentivirus libraries were transduced either ex vivo prior to intracranial GBM cell transplantation or in vivo using intratumor convection enhanced delivery (CED). Transcriptional phenotyping was performed using single-cell RNA-seq with CRISPR direct capture following focal brain radiotherapy (2 Gy x 5) or mock treatment. GBM cell states were validated using single-nucleus RNA-seq data from 86 primary-recurrent patient-matched GBMs. Mechanistic and functional validation was performed using small molecule inhibitors, immunohistochemistry, clonogenic assays, and in vivo survival experiments. In vivo Perturb-seq ± radiotherapy of 48 genes underlying GBM radiotherapy responses, which were enriched for DNA damage response and metabolic pathways, was performed in > 425,000 single cells. Radiotherapy induced 16 distinct GBM cell states, and genetic perturbations reprogrammed these cell states in a treatment-dependent fashion. Quantitative modeling of gene/radiotherapy interactions using high dimensional manifolds revealed in vivo-specific genetic dependencies. We revealed a critical role for Prkdc, the catalytic subunit of DNA-dependent protein kinase (DNA-PK), as a radiotherapy sensitizer through regulation of cell intrinsic growth and oxidative stress pathways, and cell extrinsic interferon and signaling pathways that altered cell-cell interactions in vivo. These pathways were also disrupted in single-nucleus RNA-seq analysis of post-radiotherapy human GBM tumors. Inhibition of Prkdc using a Food and Drug Administration approved small molecule sensitized GBM cells to radiotherapy and extended survival in mice harboring human intracranial xenografts. We establish in vivo Perturb-seq in orthotopic GBM models as a platform for simultaneous functional genomic discovery and characterization of therapeutic targets, revealing an underappreciated role for Prkdc in GBM tumors in vivo that is targetable using small molecules. These tools are adaptable for a wide range of disease models and treatment modalities.

Cannabinoid Receptor Agonist WIN55, 212-2 Attenuates Injury in the Hippocampus of Rats after Deep Hypothermic Circulatory Arrest

Postoperative neurological deficits remain a challenge in cardiac surgery employing deep hypothermic circulatory arrest (DHCA). This study aimed to investigate the effect of WIN55, 212-2, a cannabinoid agonist, on brain injury in a rat model of DHCA. Twenty-four male Sprague Dawley rats were randomly divided into three groups: a control group (which underwent cardiopulmonary bypass (CPB) only), a DHCA group (CPB with DHCA), and a WIN group (WIN55, 212-2 pretreatment before CPB with DHCA). Histopathological changes in the brain were evaluated by hematoxylin-eosin staining. Plasma levels of superoxide dismutase (SOD) and proinflammatory cytokines including interleukin (IL)-1β, IL-6, and tumor necrosis factor-alpha (TNF-a) were determined using an enzyme-linked immunosorbent assay (ELISA). The expression of SOD in the hippocampus was detected by Western blot and immunofluorescence staining. Levels of apoptotic-related protein caspase-3 and type 1 cannabinoid receptor (CB1R) in the hippocampus were evaluated by Western blot. WIN55, 212-2 administration attenuated histopathological injury of the hippocampus in rats undergoing DHCA, associated with lowered levels of IL-1β, IL-6, and TNF-α (p < 0.05, p < 0.001, and p < 0.01, vs. DHCA, respectively) and an increased level of SOD (p < 0.05 vs. DHCA). WIN55, 212-2 treatment also increased the content of SOD in the hippocampus. The protein expression of caspase-3 was downregulated and the expression of CB1R was upregulated in the hippocampus by WIN55, 212-2. the administration of WIN55, 212-2 alleviates hippocampal injury induced by DHCA in rats by regulating intrinsic inflammatory and oxidative stress responses through a CB1R-dependent mechanism.

Targeted tumor killing by pomegranate polyphenols: Pro-oxidant role of a classical antioxidant

One of the key biochemical features that distinguish a cancer cell from normal cells is its persistent pro-oxidative state that leads to intrinsic oxidative stress. Malignant cells have evolved sophisticated adaptation systems that involve high dependency on antioxidant functions and upregulation of pro-survival molecules to counteract the deleterious effects of reactive species and to maintain dynamic redox balance. This situation renders them vulnerable to further oxidative challenges by exogenous agents. In the present study, we advocated that pomegranate polyphenols act as pro-oxidants and trigger ROS-mediated apoptosis in cancer cells. With the help of both in vitro and in vivo models, we have established that pomegranate fruit extract (PFE) can cause a significant reduction in tumor proliferation while leaving normal tissues and cells unharmed. Administration of PFE (0.2% v/v) in Erhlich's ascites carcinoma-bearing mice for 3 weeks, inhibited the nuclear factor (erythroid-derived 2)-like 2-antioxidant response element signaling cascade, increased intracellular reactive oxygen species content, altered glutathione cycle thereby activating reactive oxygen species-induced apoptotic pathway in Erhlich's ascites carcinoma cells. Moreover, PFE mitigated epithelial to mesenchymal transition and migration in triple negative breast cancer cells (MDA-MB 231 cells) by down-regulating nuclear factor kappa light-chain-enhancer of activated B cells. Pre-treatment of tumor cells with N-acetyl cysteine protected these cells from undergoing PFE-induced apoptosis while siRNA-mediated silencing of Nuclear factor (erythroid-derived 2)-like 2 and nuclear factor kappa light-chain-enhancer of activated B cells in tumor cells increased the cytotoxic potential and pro-oxidative activity of PFE, indicating a clear role of these transcription factors in orchestrating the anticancer/pro-oxidative properties of PFE. The seminal findings provided may be exploited to develop potential therapeutic targets for selective killing of malignant cells.

Tumor killing by a dietary curcumin mono-carbonyl analog that works as a selective ROS generator via TrxR inhibition

In comparison with normal cells, cancer cells feature intrinsic oxidative stress, thereby being more vulnerable to further production of reactive oxygen species (ROS) by pro-oxidative anticancer agents (PAAs). However, PAAs also inevitably generate ROS in normal cells, resulting in their narrow therapeutic window and toxic side effects that greatly limit their clinical application. To develop PAAs that generate ROS selectively in cancer cells over in normal cells, we rationally designed three series of 21 dietary curcumin 5-carbon mono-carbonyl analogs differentiated by either placement of the cyclohexanone, piperidone, and methylpiperidone linkers, or introduction of electron-withdrawing trifluoromethyl and electron-donating methoxyl groups on its two aromatic rings in the ortho, meta, or para position to the linkers. From the designed molecules, 2c, characterized of the presence of the meta-CF3-substituted mode and the piperidone linker, was identified as a potent selective ROS-generating agent, allowing its ability to kill selectively human non-small cell lung cancer NCI–H460 (IC50 = 0.44 μM) over human normal lung MRC-5 cells with a selectivity index of 32.0. Additionally, it was more potent and selective than the conventional chemotherapeutic agents (5-fluorouracil and camptothecin) did. Mechanistical investigation reveals that by means of its Michael acceptor unit and structure characteristics as described above, 2c could covalently modify the Sec-498 residue of intracellular thioredoxin reductase (TrxR) to generate ROS selectively, resulting in ROS-dependent apoptosis and ferroptosis of NCI–H460 cells. Noticeably, 2c inhibited significantly the growth of NCI–H460 cell xenograft tumor in nude mice without obvious toxicity to liver and kidney. Together, this work highlights a practical strategy of targeting TrxR overexpressed in cancer cells to develop PAAs capable of generating ROS selectively, as evidenced by the example of 2c.

Carbohydrate-derived bicyclic selenazolines as new dual inhibitors (cholinesterases/OGA) against Alzheimer’s disease

Concerned by the urgent need to explore new approaches for the treatment of Alzheimer’s disease, we herein describe the synthesis and evaluation of new multitarget molecules. In particular, we have focused our attention on modulating the activity of cholinesterases (AChE, BuChE) in order to restore the levels of the neurotransmitter acetylcholine, and of O-GlcNAcase (OGA), which is associated with hyperphosphorylation of tau protein, in turn related to the formation of neurofibrillary tangles in the brain.Specifically, we considered the possibility of using carbohydrate-fused 1,3-selenazolines, decorated with a 2-alkylamino or 2-alkoxy moieties. On the one hand, the presence of a selenium atom might be useful in modulating the intrinsic oxidative stress in AD. On the other hand, such bicyclic structure might behave as a transition state analogue of OGA hydrolysis. Moreover, upon protonation, it could mimic the ammonium cation of acetylcholine.The lead compound, bearing a propylamino moiety on C-2 position of the selenazoline motif, proved to be a good candidate against AD; it turned out to be a strong inhibitor of BuChE (IC50 = 0.46 µM), the most prevalent cholinesterase in advanced disease stages, with a roughly 4.8 selectivity index in connection to AChE (IC50 = 2.2 µM). This compound exhibited a roughly 12-fold increase in activity compared to galantamine, one of the currently marketed drugs against AD, and a selective AChE inhibitor, and virtually the same activity as rivastigmine, a selective BuChE inhibitor. Furthermore, it was also endowed with a strong inhibitory activity against human OGA, within the nanomolar range (IC50 = 0.053 µM for hOGA, >100 µM for hHexB), and, thus, with an outstanding selectivity (IC50(hHexB)/IC50(hOGA) > 1887). The title compounds also exhibited an excellent selectivity against a panel of glycosidases and a negligible cytotoxicity against tumor and non-tumor cell lines.Docking simulations performed on the three target enzymes (AChE, BuChE, and OGA) revealed the key interactions to rationalize the biological data.

Cytoprotective effect of selenium polysaccharide from Pleurotus ostreatus against H2O2-induced oxidative stress and apoptosis in PC12 cells

One main fraction of Selenium-enriched Pleurotus ostreatus ( P. ostreatus ) polysaccharide (Se-POP-1) was extracted and purified by DEAE-52 and sephadex G-100. Se-POP-1 was an approximate homogenous polysaccharide with an average molecular weight of 1.62 × 10 4 Da, and mainly composed of glucose, mannose and galactose, with molar ratio of 5.30:1.55:2.14. The absorption peaks at 941 cm −1 and 1048 cm −1 in FT-IR analysis were ascribed as C-O-Se and Se=O bonds. Pr-treatment of Se-POP-1 (400 μg/mL) increased the cell survival of H 2 O 2 -stimulated PC12 cells and inhibited intrinsic apoptosis and oxidative stress in H 2 O 2 -stimulated PC12 cells via limiting DNA degradation and decreasing the reactive oxygen species (ROS) generation. In addition, up-regulation of anti-apoptotic protein Bcl-2, down- regulation of pro-apoptotic protein Bax, cleaved caspase 3, and cytochrome c were also observed. Se-POP-1 presented an obvious effect to alleviate oxidative damage and apoptosis in PC12 cells induced by H 2 O 2 . Therefore, Se-POP-1 possessed potent antioxidant and biological activities with the ability to prevent oxidation via scavenging ROS and free radicals in cells. It could be developed as organic selenium dietary supplement and functional food.

The Influence of Gut Microbiota on Neurogenesis: Evidence and Hopes.

Adult neurogenesis (i.e., the life-long generation of new neurons from undifferentiated neuronal precursors in the adult brain) may contribute to brain repair after damage, and participates in plasticity-related processes including memory, cognition, mood and sensory functions. Among the many intrinsic (oxidative stress, inflammation, and ageing), and extrinsic (environmental pollution, lifestyle, and diet) factors deemed to impact neurogenesis, significant attention has been recently attracted by the myriad of saprophytic microorganismal communities inhabiting the intestinal ecosystem and collectively referred to as the gut microbiota. A growing body of evidence, mainly from animal studies, reveal the influence of microbiota and its disease-associated imbalances on neural stem cell proliferative and differentiative activities in brain neurogenic niches. On the other hand, the long-claimed pro-neurogenic activity of natural dietary compounds endowed with antioxidants and anti-inflammatory properties (such as polyphenols, polyunsaturated fatty acids, or pro/prebiotics) may be mediated, at least in part, by their action on the intestinal microflora. The purpose of this review is to summarise the available information regarding the influence of the gut microbiota on neurogenesis, analyse the possible underlying mechanisms, and discuss the potential implications of this emerging knowledge for the fight against neurodegeneration and brain ageing.

The landscape of human tissue and cell type specific expression and co-regulation of senescence genes

BackgroundCellular senescence is a complex stress response that impacts cellular function and organismal health. Multiple developmental and environmental factors, such as intrinsic cellular cues, radiation, oxidative stress, oncogenes, and protein accumulation, activate genes and pathways that can lead to senescence. Enormous efforts have been made to identify and characterize senescence genes (SnGs) in stress and disease systems. However, the prevalence of senescent cells in healthy human tissues and the global SnG expression signature in different cell types are poorly understood.MethodsThis study performed an integrative gene network analysis of bulk and single-cell RNA-seq data in non-diseased human tissues to investigate SnG co-expression signatures and their cell-type specificity.ResultsThrough a comprehensive transcriptomic network analysis of 50 human tissues in the Genotype-Tissue Expression Project (GTEx) cohort, we identified SnG-enriched gene modules, characterized SnG co-expression patterns, and constructed aggregated SnG networks across primary tissues of the human body. Our network approaches identified 51 SnGs highly conserved across the human tissues, including CDKN1A (p21)-centered regulators that control cell cycle progression and the senescence-associated secretory phenotype (SASP). The SnG-enriched modules showed remarkable cell-type specificity, especially in fibroblasts, endothelial cells, and immune cells. Further analyses of single-cell RNA-seq and spatial transcriptomic data independently validated the cell-type specific SnG signatures predicted by the network analysis.ConclusionsThis study systematically revealed the co-regulated organizations and cell type specificity of SnGs in major human tissues, which can serve as a blueprint for future studies to map senescent cells and their cellular interactions in human tissues.

Beneficial Role of Carica papaya Extracts and Phytochemicals on Oxidative Stress and Related Diseases: A Mini Review.

Simple SummaryThis review highlights the medicinal benefits of a natural remedy, the Carica papaya extracts and its phytochemicals. In this review, the potential of Carica papaya against various conditions, including cancer, inflammation, aging, healing of the skin, and lifelong diseases has been summarized and discussed. In short, more research and development should focus on this natural remedy that can potentially act as a prophylaxis against chronic diseases.Oxidative stress is a result of disruption in the balance between antioxidants and pro-oxidants in which subsequently impacting on redox signaling, causing cell and tissue damages. It leads to a range of medical conditions including inflammation, skin aging, impaired wound healing, chronic diseases and cancers but these conditions can be managed properly with the aid of antioxidants. This review features various studies to provide an overview on how Carica papaya help counteract oxidative stress via various mechanisms of action closely related to its antioxidant properties and eventually improving the management of various oxidative stress-related health conditions. Carica papaya is a topical plant species discovered to contain high amounts of natural antioxidants that can usually be found in their leaves, fruits and seeds. It contains various chemical compounds demonstrate significant antioxidant properties including caffeic acid, myricetin, rutin, quercetin, α-tocopherol, papain, benzyl isothiocyanate (BiTC), and kaempferol. Therefore, it can counteract pro-oxidants via a number of signaling pathways that either promote the expression of antioxidant enzymes or reduce ROS production. These signaling pathways activate the antioxidant defense mechanisms that protect the body against both intrinsic and extrinsic oxidative stress. To conclude, Carica papaya can be incorporated into medications or supplements to help manage the health conditions driven by oxidative stress and further studies are needed to investigate the potential of its chemical components to manage various chronic diseases.

Control of Oxidative Stress in Cancer Chemoresistance: Spotlight on Nrf2 Role.

Chemoresistance represents the main obstacle to cancer treatment with both conventional and targeted therapy. Beyond specific molecular alterations, which can lead to targeted therapy, metabolic remodeling, including the control of redox status, plays an important role in cancer cell survival following therapy. Although cancer cells generally have a high basal reactive oxygen species (ROS) level, which makes them more susceptible than normal cells to a further increase of ROS, chemoresistant cancer cells become highly adapted to intrinsic or drug-induced oxidative stress by upregulating their antioxidant systems. The antioxidant response is principally mediated by the transcription factor Nrf2, which has been considered the master regulator of antioxidant and cytoprotective genes. Nrf2 expression is often increased in several types of chemoresistant cancer cells, and its expression is mediated by diverse mechanisms. In addition to Nrf2, other transcription factors and transcriptional coactivators can participate to maintain the high antioxidant levels in chemo and radio-resistant cancer cells. The control of expression and function of these molecules has been recently deepened to identify which of these could be used as a new therapeutic target in the treatment of tumors resistant to conventional therapy. In this review, we report the more recent advances in the study of Nrf2 regulation in chemoresistant cancers and the role played by other transcription factors and transcriptional coactivators in the control of antioxidant responses in chemoresistant cancer cells.

DDRE-08. NRF2/GLUTATHIONE METABOLISM AS A NOVEL THERAPEUTIC TARGET FOR IDH1-MUTATED GLIOMA

BACKGROUND IDH1-mutated glioma is a recently defined disease entity with distinctive patterns of tumor cell biology, metabolism, and resistance to therapy. Although IDH1 mutations are highly prevalent in patients with WHO II/III glioma, curative molecular targeting approaches remain unavailable for this disease cluster.METHODSIn the present study, we investigated the glutathione de novo synthesis pathway through the TCGA patient cohort and patient-derived cell lines with IDH1 mutation. The biologic function of nuclear factor erythroid 2-related factor 2 (NRF2) was analyzed by biochemistry and cell biology assays. Finally, NRF2 inhibitors were evaluated in IDH1-mutated cell lines and preclinical models as an experimental therapy.RESULTS IDH1 mutant neomorphic activity depletes the cellular pools of enzyme cofactors such as nicotinamide adenine dinucleotide (NAD) and nicotinamide adenine dinucleotide phosphate (NADP). The limitation of NAD(P) not only affects the anabolic reactions, but also results in oxidative stress and damages on DNA and protein. Further, we showed that the reprogrammed redox landscape results in constitutive activation of NRF2-governed cytoprotective pathways through the decoupling of NRF2 from its E3 ligase Kelch-like ECH-associated protein 1. NRF2 mediated the transcriptional activation of GCLC, GCLM, and SLC7A11, which not only strengthens the glutathione de novo synthesis, but also relieves the metabolic burden in IDH1-mutated cells. The importance of the glutathione synthesis is further confirmed through COX regression analysis on lower-grade glioma. Blockade of the NRF2/glutathione metabolic pathway synergizes with the elevated intrinsic oxidative stress, which results in overwhelming oxidative damage, as well as a substantial reduction in tumor cell proliferation and xenograft expansion.CONCLUSIONWe report that the NRF2-guided cytoprotective pathways play pivotal roles in the disease progression of IDH1-mutated glioma. Targeting NRF2 and glutathione metabolism could be novel targeting strategies for IDH1-mutated glioma.

Genomic, Transcriptomic, and Functional Alterations in DNA Damage Response Pathways as Putative Biomarkers of Chemotherapy Response in Ovarian Cancer.

Simple SummarySeveral chemotherapy drugs are approved for ovarian cancer treatment in the neo-adjuvant/adjuvant setting as well as following relapse. These include carboplatin, paclitaxel, doxorubicin, topotecan, PARP inhibitors (PARPi), and gemcitabine. However, except for PAPRi, there are no predictive biomarkers to guide the choice of drug. The majority of chemotherapeutic drugs function by inducing DNA damage or inhibiting its repair. However, the association of DNA damage repair (DDR) pathway alterations with therapy response remain unclear. In this study, using a panel of 14 ovarian cancer cell lines, 10 patient ascites-derived primary cultures and bioinformatic analysis of The Cancer Genome Atlas (TCGA) ovarian cancer dataset, we identified the role of genomic/transcriptomic and/or functional alterations in DDR pathways as determinants of therapy response.Defective DNA damage response (DDR) pathways are enabling characteristics of cancers that not only can be exploited to specifically target cancer cells but also can predict chemotherapy response. Defective Homologous Recombination Repair (HRR) function, e.g., due to BRCA1/2 loss, is a determinant of response to platinum agents and PARP inhibitors in ovarian cancers. Most chemotherapies function by either inducing DNA damage or impacting on its repair but are generally used in the clinic unselectively. The significance of HRR and other DDR pathways in determining response to several other chemotherapy drugs is not well understood. In this study, the genomic, transcriptomic and functional analysis of DDR pathways in a panel of 14 ovarian cancer cell lines identified that defects in DDR pathways could determine response to several chemotherapy drugs. Carboplatin, rucaparib, and topotecan sensitivity were associated with functional loss of HRR (validated in 10 patient-derived primary cultures) and mismatch repair. Two DDR gene expression clusters correlating with treatment response were identified, with PARP10 identified as a novel marker of platinum response, which was confirmed in The Cancer Genome Atlas (TCGA) ovarian cancer cohort. Reduced non-homologous end-joining function correlated with increased sensitivity to doxorubicin, while cells with high intrinsic oxidative stress showed sensitivity to gemcitabine. In this era of personalised medicine, molecular/functional characterisation of DDR pathways could guide chemotherapy choices in the clinic allowing specific targeting of ovarian cancers.

Bcl-xL as a Modulator of Senescence and Aging.

Many features of aging result from the incapacity of cells to adapt to stress conditions. When cells are overwhelmed by stress, they can undergo senescence to avoid unrestricted growth of damaged cells. Recent findings have proven that cellular senescence is more than that. A specific grade of senescence promotes embryo development, tissue remodeling and wound healing. However, constant stresses and a weakening immune system can lead to senescence chronicity with aging. The accumulation of senescent cells is directly related to tissue dysfunction and age-related pathologies. Centenarians, the most aged individuals, should accumulate senescent cells and suffer from their deleterious effects, however, they enjoy a compression of morbidity. We have shown that they overexpress B-cell lymphoma-extra large (Bcl-xL). Bcl-xL could avoid an excessive burden of senescent cells through the regulation of intrinsic apoptosis, mitochondrial bioenergetics and oxidative stress. On the other hand, Bcl-xL maintains a fully functional immune system that ensures an efficient clearance of senescent cells. Moreover, there is a paradox, as inhibitors of Bcl-xL have been employed as senolytic agents, which have been shown to protect from aging in animal models. In this review, we aim to discuss how Bcl-xL could modulate senescence-associated harmful effects in centenarians, protecting them from the burden of accumulation of senescent cells.

Mitochondrial Bioenergetics at the Onset of Drug Resistance in Hematological Malignancies: An Overview.

The combined derangements in mitochondria network, function and dynamics can affect metabolism and ATP production, redox homeostasis and apoptosis triggering, contributing to cancer development in many different complex ways. In hematological malignancies, there is a strong relationship between cellular metabolism, mitochondrial bioenergetics, interconnections with supportive microenvironment and drug resistance. Lymphoma and chronic lymphocytic leukemia cells, e.g., adapt to intrinsic oxidative stress by increasing mitochondrial biogenesis. In other hematological disorders such as myeloma, on the contrary, bioenergetics changes, associated to increased mitochondrial fitness, derive from the adaptive response to drug-induced stress. In the bone marrow niche, a reverse Warburg effect has been recently described, consisting in metabolic changes occurring in stromal cells in the attempt to metabolically support adjacent cancer cells. Moreover, a physiological dynamic, based on mitochondria transfer, between tumor cells and their supporting stromal microenvironment has been described to sustain oxidative stress associated to proteostasis maintenance in multiple myeloma and leukemia. Increased mitochondrial biogenesis of tumor cells associated to acquisition of new mitochondria transferred by mesenchymal stromal cells results in augmented ATP production through increased oxidative phosphorylation (OX-PHOS), higher drug resistance, and resurgence after treatment. Accordingly, targeting mitochondrial biogenesis, electron transfer, mitochondrial DNA replication, or mitochondrial fatty acid transport increases therapy efficacy. In this review, we summarize selected examples of the mitochondrial derangements in hematological malignancies, which provide metabolic adaptation and apoptosis resistance, also supported by the crosstalk with tumor microenvironment. This field promises a rational design to improve target-therapy including the metabolic phenotype.

Anti-Survival and Pro-Apoptotic Effects of 6-Shogaol on SW872 Human Liposarcoma Cells via Control of the Intrinsic Caspase Pathway, STAT-3, AMPK, and ER Stress.

Notably, 6-Shogaol, a bioactive natural substance, has anticancer effects on many types of tumors. Up to date, the anticancer effect and mode of action of 6-Shogaol on liposarcoma are not known. In this study, we investigated whether 6-Shogaol inhibits the growth of SW872 and 93T449 cells, two different human liposarcoma cell lines. Of note, 6-Shogaol inhibited the growth of SW872 and 93T449 cells without affecting that of normal 3T3-L1 preadipocytes. Specifically, 6-Shogaol further induced the apoptosis of SW872 cells, as evidenced by nuclear DNA fragmentation, increased sub G1 population, activation of the intrinsic caspase pathway, and PARP cleavage. However, pretreatment with either z-VAD-fmk, a pan-caspase inhibitor, or N-acetylcysteine, an antioxidant, attenuated the 6-Shogaol’s growth-suppressive and apoptosis-inducing effects on SW872 cells. Moreover, 6-Shogaol activated AMPK while inhibited STAT-3 in SW872 cells, and siRNA-based genetic silencing of AMPK or STAT-3 considerably blocked the growth-suppressive and apoptotic response of 6-Shogaol to SW872 cells. Moreover, 6-Shogaol also upregulated the expression and phosphorylation of GRP-78, eIF-2α, ATF4, and CHOP, known ER stress markers, in SW872 cells, illustrating the induction of ER stress. These findings collectively demonstrate that 6-Shogaol has strong antigrowth and proapoptotic effects on SW872 cells through regulation of the intrinsic caspase pathway, oxidative stress, STAT-3, AMPK, and ER stress.

Selenium modulates inorganic mercury induced cytotoxicity and intrinsic apoptosis in PC12 cells

Mercury (Hg) in its all forms, including inorganic Hg (iHg) is an environmental contaminant due to toxicity and diseases in human. However, a little is known about the underlying mechanisms responsible for iHg toxicity. Selenium (Se) is an essential trace element, recognized as an antioxidant and protective agent against metal toxicities. The purpose of this research was to investigate ameliorations of Se counter to iHg-mediated toxicity in PC12 cells. Cytotoxic assays have been shown that iHg (5 μM) caused oxidative stress and intrinsic apoptosis via ROS generation, oxidizing glutathione, damaging DNA, degrading cell membrane integrity, down-regulating mTOR, p-mTOR, akt and ERK1, and up-regulating cleaved caspase 3 and cytochrome c release in PC12 cells 48 h after incubation. Co-treatment of Se (5 μM) inhibited intrinsic apoptosis and oxidative stress induced by iHg (5 μM) via inhibiting ROS formation, boosting GPx contents, increasing reduced glutathione, limiting DNA degradation, improving cell membrane integrity, up-regulating mTOR, p-mTOR, akt, ERK1 and caspase 3, and down-regulating cleaved caspase 3 and cytochrome c leakage in PC12 cells. In conclusion, these results recommended that excessive ROS generation acts a critical role in iHg-influenced oxidative stress and co-treatment of Se attenuates iHg-cytotoxicity through its antioxidant properties.