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Mutations in TP53 gene play a pivotal role in tumorigenesis and cancer development. Here, we report that gain-of-function mutant p53 proteins inhibit the autophagic pathway favoring antiapoptotic effects as well as proliferation of pancreas and breast cancer cells. We found that mutant p53 significantly counteracts the formation of autophagic vesicles and their fusion with lysosomes throughout the repression of some key autophagy-related proteins and enzymes as BECN1 (and P-BECN1), DRAM1, ATG12, SESN1/2 and P-AMPK with the concomitant stimulation of mTOR signaling. As a paradigm of this mechanism, we show that atg12 gene repression was mediated by the recruitment of the p50 NF-κB/mutant p53 protein complex onto the atg12 promoter. Either mutant p53 or p50 NF-κB depletion downregulates atg12 gene expression. We further correlated the low expression levels of autophagic genes (atg12, becn1, sesn1, and dram1) with a reduced relapse free survival (RFS) and distant metastasis free survival (DMFS) of breast cancer patients carrying TP53 gene mutations conferring a prognostic value to this mutant p53-and autophagy-related signature. Interestingly, the mutant p53-driven mTOR stimulation sensitized cancer cells to the treatment with the mTOR inhibitor everolimus. All these results reveal a novel mechanism through which mutant p53 proteins promote cancer cell proliferation with the concomitant inhibition of autophagy. •GOF p53 mutant proteins inhibit the autophagic vesicle formation in cancer cells.•Mutant p53 proteins inhibit the expression of ATGs in cancer cells and patients.•Mutant p53/NF-κB p50 complex inhibits atg12 gene expression.•Mutant p53 proteins stimulate mTOR and repress AMPK signaling.•The expression of mutant p53 proteins sensitizes cancer cells to mTOR inhibition.

Essential oils (EOs) and their components have been reported to possess anticancer properties and to increase the sensitivity of cancer cells to chemotherapy. The aim of this work was to select EOs able to downregulate STAT3 signaling using Western blot and RT-PCR analyses. The molecular mechanism of anti-STAT3 activity was evaluated through spectrophotometric and fluorometric analyses, and the biological effect of STAT3 inhibition was analyzed by flow cytometry and wound healing assay. Herein, Pinus mugo EO (PMEO) is identified as an inhibitor of constitutive STAT3 phosphorylation in human prostate cancer cells, DU145. The down-modulation of the STAT3 signaling cascade decreased the expression of anti-proliferative as well as anti-apoptotic genes and proteins, leading to the inhibition of cell migration and apoptotic cell death. PMEO treatment induced a rapid drop in glutathione (GSH) levels and an increase in reactive oxygen species (ROS) concentration, resulting in mild oxidative stress. Pretreatment of cells with N-acetyl-cysteine (NAC), a cell-permeable ROS scavenger, reverted the inhibitory action of PMEO on STAT3 phosphorylation. Moreover, combination therapy revealed that PMEO treatment displayed synergism with cisplatin in inducing the cytotoxic effect. Overall, our data highlight the importance of STAT3 signaling in PMEO cytotoxic activity, as well as the possibility of developing adjuvant therapy or sensitizing cancer cells to conventional chemotherapy.

The imbalance of the intracellular redox state and, in particular, of the glutathione (GSH)/GSH disulfide couple homeostasis, is involved in the pathogenesis of a number of diseases. In many skin diseases, including psoriasis, oxidative stress plays an important role, as demonstrated by the observation that treatments leading to increase of the local levels of oxidant species ameliorate the disease. Recently, dehydrocostuslactone (DCE) and costunolide (CS), two terpenes naturally occurring in many plants, have been found to exert various anti-inflammatory and pro-apoptotic effects on different human cell types. These compounds decrease the level of the intracellular GSH by direct interaction with it, and, therefore, can alter cellular redox state. DCE and CS can trigger S-glutathionylation of various substrates, including the transcription factor STAT3 and JAK1/2 proteins. In the present study, we investigated on the potential role of DCE and CS in regulating inflammatory and proliferative responses of human keratinocytes to cytokines. We demonstrated that DCE and CS decreased intracellular GSH levels in human keratinocytes, as well as inhibited STAT3 and STAT1 phosphorylation and activation triggered by IL-22 or IFN-γ, respectively. Consequently, DCE and CS decreased the IL-22- and IFN-γ-induced expression of inflammatory and regulatory genes in keratinocytes, including CCL2, CXCL10, ICAM-1 and SOCS3. DCE and CS also inhibited proliferation and cell-cycle progression-related gene expression, as well as they promoted cell cycle arrest and apoptosis. In parallel, DCE and CS activated the anti-inflammatory EGFR and ERK1/2 molecules in keratinocytes, and, thus, wound healing in an in vitro injury model. In light of our findings, we can hypothesize that the employment of DCE and CS in psoriasis could efficiently counteract the pro-inflammatory effects of IFN-γ and IL-22 on keratinocytes, revert the apoptosis-resistant phenotype, as well as inhibit hyperproliferation in the psoriatic epidermis.

Richter’s syndrome (RS) is the transformation of chronic lymphocytic leukemia (CLL) into a high-grade B-cell malignancy. Molecular and functional studies have pointed out that CLL cells are close to the apoptotic threshold and dependent on BCL-2 for survival. However, it remains undefined how evasion from apoptosis evolves during disease transformation. Here, we employed functional and static approaches to compare the regulation of mitochondrial apoptosis in CLL and RS. BH3 profiling of 17 CLL and 9 RS samples demonstrated that RS cells had reduced apoptotic priming and lower BCL-2 dependence than CLL cells. While a subset of RS was dependent on alternative anti-apoptotic proteins and was sensitive to specific BH3 mimetics, other RS cases harbored no specific anti-apoptotic addiction. Transcriptomics of paired CLL/RS samples revealed downregulation of pro-apoptotic sensitizers during disease transformation. Albeit expressed, effector and activator members were less likely to colocalize with mitochondria in RS compared to CLL. Electron microscopy highlighted reduced cristae width in RS mitochondria, a condition further promoting apoptosis resistance. Collectively, our data suggest that RS cells evolve multiple mechanisms that lower the apoptotic priming and shift the anti-apoptotic dependencies away from BCL-2, making direct targeting of mitochondrial apoptosis more challenging after disease transformation.

The main purpose of the present study is to envisage the molecular mechanism of inhibitory action of dehydrocostuslactone (DCE) and costunolide (CS), two naturally occurring sesquiterpene lactones, towards the activation of signal transducer and activator of transcription 3 (STAT3). We report that, in human THP-1 cell line, they inhibit IL-6-elicited tyrosine phosphorylation of STAT3 and its DNA binding activity with EC(50) of 10 µM with concomitant down-regulation of the phosphorylation of the tyrosine Janus kinases JAK1, JAK2 and Tyk2. Furthermore, these compounds that contain an α-β-unsaturated carbonyl moiety and function as potent Michael reaction acceptor, induce a rapid drop in intracellular glutathione (GSH) concentration by direct interaction with it, thereby triggering S-glutathionylation of STAT3. Dehydrocostunolide (HCS), the reduced form of CS lacking only the α-β-unsaturated carbonyl group, fails to exert any inhibitory action. Finally, the glutathione ethylene ester (GEE), the cell permeable GSH form, reverts the inhibitory action of DCE and CS on STAT3 tyrosine phosphorylation. We conclude that these two sesquiterpene lactones are able to induce redox-dependent post-translational modification of cysteine residues of STAT3 protein in order to regulate its function.

The wild-type protein p53 plays a key role in preventing the formation of neoplasms by controlling cell growth. However, in more than a half of all cancers, the TP53 gene has missense mutations that appear during tumorigenesis. In most cases, the mutated gene encodes a full-length protein with the substitution of a single amino acid, resulting in structural and functional changes and acquiring an oncogenic role. This dual role of the wild-type protein and the mutated isoforms is also evident in the regulation of the redox state of the cell, with antioxidant and prooxidant functions, respectively. In this review, we introduce a new concept of the p53 protein by discussing its sensitivity to the cellular redox state. In particular, we focus on the discussion of structural and functional changes following post-translational modifications of redox-sensitive cysteine residues, which are also responsible for interacting with zinc ions for proper structural folding. We will also discuss therapeutic opportunities using small molecules targeting cysteines capable of modifying the structure and function of the p53 mutant isoforms in view of possible anticancer therapies for patients possessing the mutation in the TP53 gene.

IntroductionMultiple sclerosis (MS), the most common neurological disease causing disability in young adults, is widely recognised as a major stress factor. Studies have shown that the first years after the diagnosis are distressing in terms of adjustment to the disease and that MS negatively affects patients’ psychological well-being, quality of life (QoL) and social functioning. However, the links between disease-specific variables at diagnosis, resilience and psychological adjustment of patients with MS remain largely unexplored, especially in adolescents and young adults. This observational study aims to fill the gap of knowledge on biopsychosocial characteristics and resilience of young adults with MS to evaluate the relationship among these variables and to develop a biopsychosocial model of resilience.Methods and analysisBiological and clinical characteristics of young adults newly diagnosed with MS will be investigated by collecting clinical information, performing neurological examinations, MRI and analysing cerebrospinal fluid and blood biomarkers (eg, measures of inflammation), body composition, gut microbiota and movement/perceptual markers. Psychosocial characteristics (eg, psychological distress, coping strategies), QoL, psychological well-being and resilience will be assessed by self-report questionnaires. Comparative statistics (ie, analysis of variance or unpaired samples t-test, correlation and regression analyses) will be applied to evaluate the relationship among biological, psychological and social factors. The results are expected to allow a comprehensive understanding of the determinants of resilience in young patients with MS and to inform resilience interventions, tailored to young patients’ specific needs, aiming to reduce the risk of maladaptive reactions to the disease and to improve psychological well-being and QoL.Ethics and disseminationThe study has been approved by the Verona University Hospital Ethics Committee (approval number: 2029CESC). The findings will be disseminated through scientific publications in peer-reviewed journals, conference presentations, social media and specific websites.Trial registration numberClinicalTrials.gov (NCT03825055).

STAT1 and STAT3 are nuclear transcription factors that regulate genes involved in cell cycle, cell survival and immune response. The cross-talk between these signaling pathways determines how cells integrate the environmental signals received ultimately translating them in transcriptional regulation of specific sets of genes. Despite being activated downstream of common cytokine and growth factors, STAT1 and STAT3 play essentially antagonistic roles and the disruption of their balance directs cells from survival to apoptotic cell death or from inflammatory to anti-inflammatory responses. Different mechanisms are proposed to explain this yin-yang relationship. Considering the redox aspect of STATs proteins, this review attempts to summarize the current knowledge of redox regulation of STAT1 and STAT3 signaling focusing the attention on the post-translational modifications that affect their activity.

The tumor microenvironment is a key factor in disease progression, local resistance, immune-escaping, and metastasis. The rapid proliferation of tumor cells and the aberrant structure of the blood vessels within tumors result in a marked heterogeneity in the perfusion of the tumor tissue with regions of hypoxia. Although most of the tumor cells die in these hypoxic conditions, a part of them can adapt and survive for many days or months in a dormant state. Dormant tumor cells are characterized by cell cycle arrest in G0/G1 phase as well as a low metabolism, and are refractive to common chemotherapy, giving rise to metastasis. Despite these features, the cells retain their ability to proliferate when conditions improve. An understanding of the regulatory machinery of tumor dormancy is essential for identifying early cancer biomarkers and could provide a rationale for the development of novel agents to target dormant tumor cell populations. In this review, we examine the current knowledge of the mechanisms allowing tumor dormancy and discuss the crucial role of the hypoxic microenvironment in this process.

BackgroundThe TP53 tumor suppressor gene is the most frequently altered gene in tumors and mutant p53 gain-of-function isoforms actively promote cancer malignancy.MethodsA panel of wild-type and mutant p53 cancer cell lines of different tissues, including pancreas, breast, skin, and lung were used, as well as chronic lymphocytic leukemia (CLL) patients with different TP53 gene status. The effects of mutant p53 were evaluated by confocal microscopy, reactive oxygen species production assay, immunoblotting, and quantitative reverse transcription polymerase chain reaction after cellular transfection.ResultsWe demonstrate that oncogenic mutant p53 isoforms are able to inhibit SESN1 expression and consequently the amount of SESN1/AMPK complex, resulting in the downregulation of the AMPK/PGC-1α/UCP2 axis and mitochondrial O2ˉ· production. We also show a correlation between the decrease of reduced thiols with a poorer clinical outcome of CLL patients bearing mutant TP53 gene. The restoration of the mitochondrial uncoupling protein 2 (UCP2) expression, as well as the addition of the radical scavenger N-acetyl-l-cysteine, reversed the oncogenic effects of mutant p53 as cellular hyper-proliferation, antiapoptotic effect, and resistance to drugs.ConclusionsThe inhibition of the SESN1/AMPK/PGC-1α/UCP2 axis contributes to the pro-oxidant and oncogenic effects of mutant p53, suggesting pro-oxidant drugs as a therapeutic approach for cancer patients bearing mutant TP53 gene.