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Background Clear cell renal cell carcinoma (ccRCC) is the most common subtype of renal cancer. Large-scale metabolomic data have associated metabolic alterations with the pathogenesis and progression of renal carcinoma and have correlated mitochondrial activity with poor survival in a subset of patients. The aim of this study was to determine whether targeting mitochondria-lysosome interaction could be a novel therapeutic approach using patient-derived organoids as avatar for drug response. Methods RNAseq data analysis and immunohistochemistry were used to show overexpression of Purinergic receptor 4 (P2XR4) in clear cell carcinomas. Seahorse experiments, immunofluorescence and fluorescence cell sorting were used to demonstrate that P2XR4 regulates mitochondrial activity and the balance of radical oxygen species. Pharmacological inhibitors and genetic silencing promoted lysosomal damage, calcium overload in mitochondria and cell death via both necrosis and apoptosis. Finally, we established patient-derived organoids and murine xenograft models to investigate the antitumor effect of P2XR4 inhibition using imaging drug screening, viability assay and immunohistochemistry. Results Our data suggest that oxo-phosphorylation is the main source of tumor-derived ATP in a subset of ccRCC cells expressing P2XR4, which exerts a critical impact on tumor energy metabolism and mitochondrial activity. Prolonged mitochondrial failure induced by pharmacological inhibition or P2XR4 silencing was associated with increased oxygen radical species, changes in mitochondrial permeability (i.e., opening of the transition pore complex, dissipation of membrane potential, and calcium overload). Interestingly, higher mitochondrial activity in patient derived organoids was associated with greater sensitivity to P2XR4 inhibition and tumor reduction in a xenograft model. Conclusion Overall, our results suggest that the perturbed balance between lysosomal integrity and mitochondrial activity induced by P2XR4 inhibition may represent a new therapeutic strategy for a subset of patients with renal carcinoma and that individualized organoids may be help to predict drug efficacy.

The constant decline in fertility and older reproductive age is the major cause of low clinical pregnancy rates in industrialised countries. Epigenetic mechanisms impact on proper embryonic development in women undergoing in vitro fertilisation (IVF) protocols. Here, we describe the main epigenetic modifications that may influence female reproduction and could affect IVF success. Graphical Abstract

The original version of this Letter contains an error in Figure 5, concerning an experiment performed at University of Bologna, which shows the effect of the tested compounds on microglial activation. In the incorrect published Figure 5, the GAPDH band (housekeeping protein used as loading control) from a preliminary experiment for compound 1 was accidentally shown, and the correct image is used below. Furthermore, since we used the same LPS-treated control microglial cells for compounds 11 and 12, being that they were tested in the same experiment and plate, the right two panels have been modified for clarity. This is the correct Figure 5:.(Figure Presented).

Summary Global investigation of histone marks in acute myeloid leukemia (AML) remains limited. Analyses of 38 AML samples through integrated transcriptional and chromatin mark analysis exposes 2 major subtypes. One subtype is dominated by patients with NPM1 mutations or MLL-fusion genes, shows activation of the regulatory pathways involving HOX-family genes as targets, and displays high self-renewal capacity and stemness. The second subtype is enriched for RUNX1 or spliceosome mutations, suggesting potential interplay between the 2 aberrations, and mainly depends on IRF family regulators. Cellular consequences in prognosis predict a relatively worse outcome for the first subtype. Our integrated profiling establishes a rich resource to probe AML subtypes on the basis of expression and chromatin data.

Aristaless-related homeobox (ARX) is a crucial transcription factor involved in several interneuronopathies, including infantile spasms (ISSX1) and West Syndrome. Its polyAlanine motifs are hot spots for elongations, frequently found in kids affected by malignant Epilepsy with limited therapeutic options. Although in the last years several medical therapies have been tested to prevent epilepsy in West syndrome patients, an effective treatment is still needed. Noteworthy, the early onset of infantile spams followed by developmental delay and poor seizure control make these patients at high risk for suddenly unexpected death (SUDEP). The overall objectives of our project were to study Aristaless-related Homeobox Epilepsy models with the final goal of identifying druggable target molecules.

Studying molecular convergence in neurodevelopmental disorders caused by mutations in specific disease-related genes permits us to define druggable molecular pathways. The purpose of our study is to assess the degree of damage associated with the ARX-KDM5C and to establish a correlation between similar phenotypes and same cellular functions. Mutations in Aristaless-related homeobox gene (ARX), a homeotic transcription factor with a key role in interneuron maturation, have been found in a spectrum of X-chromosome phenotypes including cortical malformations, chronic Epilepsy and X-Linked Intellectual Disabilities (XLID). About Lysine-specific demethylase 5C (KDM5C), its mutations have been reported as an important cause of XLID. Its protein is a histone demethylase acting as transcriptional repressor during brain development. Here we summarize functional analysis of two classes of ARX mutations : 1. PolyAlanine elongations affecting the first and the second PolyA tracts, frequently found in patients with Epilepsy or XLID; and 2. Missense mutations clustered in the paired-type homeodomain (HD) found in patients with Lissencephaly with abnormal genitalia (XLAG). Thus, we propose a "fault disease model" showing that the degree of spectrum of defects in transcription of ARX-target genes correlate with the severity of the neurophenotypes associated with ARX mutations.

The X-linked ARX gene encodes the Aristaless-related homeobox protein, ARX, an interneuron-specific transcription factor (TF) with a key role in GABAergic interneuron migration and maturation. Expansions of the first two polyalanine tracts (PA1 and PA2) cause refractory epilepsy (RE) with infantile spasms or X-linked Intellectual Disability (XLID). Here we report on an evolutionary conserved druggable ARX-dependent pathway damaged by the faulty functioning of the polyalanine elongated TFs, exerted through a poorly-understood pathogenic mechanism. We have combined the molecular validation of the conserved interaction of ARX with KDM5C from mammals to C.elegans with the analysis of relative neuronal outcomes in order to explore potential compounds able to correct molecular faults behind the ARX polyalanine dysfunction.

The X-linked ARX gene encodes the Aristaless-related homeobox protein, which belongs to a subset of morphogenetic transcription factors with a crucial role in cerebral development and patterning. Mutations in ARX cause a wide spectrum of X-linked neurodevelopmental disorders including lissencephaly, a severe cortical malformation and a catastrophic epileptic encephalopathy with recurrent and resistant seizures. The health problem to find efficient treatments for ARX-related diseases highlights the need for understanding the components and pathways that regulate brain development. Because the neuronal functioning is maintained by a complex regulatory network, the use of powerful genetic models like mouse and worm can complement studies on human genetics and physiology by offering new opportunities to dissect complicated and evolutionary conserved regulatory circuits. Here we describe for the first time the conservation of an ARX-dependent disease-pathway among human, mouse and worm establishing a gene-phenotype association from one organism to another.

Mistakes in histone methylation-demethylation rounds have been directly involved in several forms of Intellectual Disability (ID) with Epilepsy and/or Refractory Epilepsy (RE). Lysine-specific demethylase 5C (KDM5C) is an X-linked gene, which encodes a chromatin JmjC eraser with H3K4me2/3 demethylase activity. KDM5C is frequently mutated in a spectrum of X-linked ID (XLID) and/or RE. It functions as a transcriptional repressor that is critical for transition of neural progenitors to neurons. We identified a disease path, linking functionally KDM5C to another XLID/Epilepsy gene, encoding the homeotic transcription factor ARX, whose mutations impair severely KDM5C transcript regulation.Ongoing efforts will allow us to identify druggable hallmarks that could open up towards the exploitation of potential strategies to treat the growing group of ID and RE diseases caused by defects in chromatin and/or transcriptional regulators.

Mis-steps in histone methylation-demethylation rounds have been directly involved in several forms of Intellectual Disability (ID) with Epilepsy. Lysine-specific demethylase 5C (KDM5C) is an X-linked gene, which encodes a chromatin JmjC eraser with H3K4me2/3 demethylase activity. KDM5C is frequently mutated in a spectrum of XLID and/or malignant Epilepsy. It functions as a transcriptional repressor and interacts with REST/NSRF, a master epigenetic hub that is critical for transition of neural progenitors to neurons.Because chromatin modifications are reversible, it is possible that epigenetic drugs could compensate for the KDM5C-H3K4me3 deregulation. To achieve this aim, we screened a number of compounds targeting chromatin enzymes. We used, as cell disease model, neuronally-differentiated Arx KO/Kdm5C-depleted ES cells, which show GABAergic abnormalities in association with a global increase of H3K4me3 signal. Ongoing efforts in this direction allowed us to identify druggable hallmarks that could open up towards the exploitation of potential strategies to treat the growing group of ID and Epilepsy diseases that are caused by defects in chromatin and/or transcriptional regulators.

Protein glycation is a non-enzymatic, irreversible modification of protein amino groups by reactive carbonyl species leading to the formation of advanced glycation end products (AGEs). Several proteins implicated in neurodegenerative diseases have been found to be glycated in vivo and the extent of glycation is related to the pathologies of the patients. Although it is now accepted that there is a direct correlation between AGEs formation and the development of neurodegenerative diseases related to protein misfolding and amyloid aggregation, several questions still remain unanswered: whether glycation is the triggering event or just an additional factor acting on the aggregation pathway. We have recently shown that glycation of the amyloidogenic W7FW14F apomyoglobin mutant significantly accelerates the amyloid fibrils formation providing evidence that glycation actively participates to the process. In the present study, to test if glycation can be considered also a triggering factor in amyloidosis, we evaluated the ability of different glycation agents to induce amyloid aggregation in the soluble wild type apomyoglobin. Our results show that glycation covalently modifies apomyoglobin and induces conformational changes that lead to the formation of oligomeric species that are not implicated in amyloid aggregation. Thus, AGEs formation does not trigger amyloid aggregation in the wild type apomyoglobin but only induce the formation of soluble oligomeric species able to affect cell viability. The molecular bases of cell toxicity induced by AGEs formed upon glycation of wild type apomyoglobin have been also investigated. This article is protected by copyright. All rights reserved.

Mis-steps in histone methylation-demethylation rounds have been directly involved in several forms of Intellectual Disability (ID) with Epilepsy. Lysine-specific demethylase 5C (KDM5C) is an X-linked gene which encodes a chromatin JmjC eraser with H3K4me2/3 demethylase activity. KDM5C is frequently mutated in a spectrum of XLID and/or malignant Epilepsy. Noteworthy, a defective KDM5C-H3K4me3 path has been found in association with a mis-regulation of XLID/Epilepsy effector genes. We report here on methods to correct KDM5C fault by testing epi-drugs to target KDM5C-H3K4me3 axis, specifically.In particular, we have undertaken an in vitro analysis of a number of compounds targeting chromatin enzymes with the aim of modifying the accessibility of the transcription machinery to KDM5C. A strong up-regulation of Kdm5C/KDM5C has been obtained in presence of an inhibitor of HDAC at different points of in vitro differentiation, both in WT and Kdm5C-defective cells. these methods allows to design new approaches to treat ID, malignant epilepsy diseases and many other neuropathologies with seizure linked to insufficient activity of chromatin and/or transcriptional regulators.

Understanding the links between genetic, epigenetic and non-genetic factors throughout the lifespan and across generations and their role in disease susceptibility and disease progression offer entirely new avenues and solutions to major problems in our society. To overcome the numerous challenges, we have come up with nine major conclusions to set the vision for future policies and research agendas at the European level. ispartof: BMC Genomics vol:15 issue:1 pages:487-487 ispartof: location:England status: published

Significance: Epigenetic inactivation of pivotal genes involved in cell growth is a hallmark of human pathologies, in particular cancer. Histone acetylation balance obtained through opposing actions of histone deacetylases (HDACs) and histone acetyltransferases is one epigenetic mechanism controlling gene expression and is, thus, associated with disease etiology and progression. Interfering pharmacologically with HDAC activity can correct abnormalities in cell proliferation, migration, vascularization, and death. Recent Advances: Histone deacetylase inhibitors (HDACi) represent a new class of cytostatic agents that interfere with the function of HDACs and are able to increase gene expression by indirectly inducing histone acetylation. Several HDACi, alone or in combination with DNA-demethylating agents, chemopreventive, or classical chemotherapeutic drugs, are currently being used in clinical trials for solid and hematological malignancies, and are, thus, promising candidates for cancer therapy. Critical Issues: (i) Non-specific (off-target) HDACi effects due to activities unassociated with HDAC inhibition. (ii) Advantages/disadvantages of non-selective or isoform-directed HDACi. (iii) Limited number of response-predictive biomarkers. (iv) Toxicity leading to dysfunction of critical biological processes. Future Directions: Selective HDACi could achieve enhanced clinical utility by reducing or eliminating the serious side effects associated with current first-generation non-selective HDACi. Isoform-selective and pan-HDACi candidates might benefit from the identification of biomarkers, enabling better patient stratification and prediction of response to treatment.

Epigenetic inactivation of pivotal genes involved in correct cell growth is a hallmark of human pathologies, in particular cancer. These epigenetic mechanisms, including crosstalk between DNA methylation, histone modifications and non-coding RNAs, affect gene expression and are associated with disease progression. In contrast to genetic mutations, epigenetic changes are potentially reversible. Re-expression of genes epigenetically inactivated can result in the suppression of disease state or sensitization to specific therapies. Small molecules that reverse epigenetic inactivation, so-called epi-drugs, are now undergoing clinical trials. Accordingly, the Food and Drug Administration (FDA) and the European Medicines Agency (EMA) for cancer treatment have approved some of these drugs. Here, we focus on the biological features of epigenetic molecules, analyzing the mechanism(s) of action and their current use in clinical practice.