Biology subjects

Valente, S.

Publications and source records attributed to Valente, S..

10 recordsLinked to original sources

bioRxiv2026

Vorinostat (SAHA) alters the skeletal muscle differentiation program

Epigenetic regulation, particularly histone acetylation, plays a critical role in skeletal muscle differentiation by modulating gene expression programs without altering DNA sequence. Histone deacetylases (HDACs) tightly regulate myogenesis by controlling the timing of differentiation. Pharmacological inhibition of HDACs has shown context-dependent effects on muscle cells. We investigated the effects of 1 {micro}M SAHA (suberoylanilide hydroxamic acid) on C2C12 and L6 myoblasts during differentiation using morphological, immunofluorescence, transcriptomic, and proteomic analyses. SAHA delayed early differentiation, reducing myotube formation with partial recovery at later stages. Transcriptomic analysis revealed time-dependent changes in pathways related to cytoskeleton, cell cycle, and chromatin regulation. Proteomics showed increased mitochondrial metabolism and reduced cytoskeletal components in C2C12 cells, while L6 cells displayed alterations in muscle structural and extracellular matrix proteins. SAHA induces stage- and model-dependent reprogramming of myogenesis, highlighting the importance of timing and cellular context in HDAC-targeted therapies.

cell biology↗

bioRxiv2026

Neuroendocrine-like/EMT dedifferentiation mediates resistance to EGFR inhibitors via the NRG1/HER3 axis

Patients with non-small cell lung cancer (NSCLC) carrying activating EGFR mutations typically respond favorably to third-generation EGFR tyrosine kinase inhibitors (TKIs) such as osimertinib. Nevertheless, resistance almost inevitably emerges, ultimately limiting the durability of these treatments. We investigated non-genomic mechanisms enabling drug-tolerant persister cells to survive EGFR inhibition, likely co-opting compensatory HER3 activation, whose underlying mechanisms remain unclear. Using a combination of immortalized and patient-derived cellular models, together with single-cell RNA sequencing, we demonstrate that activation of EGFR/HER3 axis constitutes an early adaptive response to TKI exposure enriched in pulmonary alveolar type I and type II cancer cells. This response is driven by neuregulin-1 (NRG1), produced by stromal cells and by cancer cells undergoing NE-like/EMT dedifferentiation. Importantly, in vivo studies demonstrated that combining EGFR inhibition and NRG1 neutralization by monoclonal antibody successfully eradicated tumors. Together, these findings point to a therapeutic strategy to overcome TKI resistance in NSCLC through targeting HER3 signalling, its interplay with EGFR, and tumor microenvironment-derived cues.

cancer biology↗

bioRxiv2024

Quinoline-based compounds can inhibit diverse enzymes that act on DNA

DNA methylation, as exemplified by cytosine-C5 methylation in mammals and adenine-N6 methylation in bacteria, is a crucial epigenetic mechanism driving numerous vital biological processes. Developing non-nucleoside inhibitors to cause DNA hypomethylation is a high priority, in order to treat a variety of significant medical conditions without the toxicities associated with existing cytidine-based hypomethylating agents. In this study, we have characterized fifteen quinoline-based analogs. Notably, compounds with additions like a methylamine (9) or methylpiperazine (11) demonstrate similar low micromolar inhibitory potency against both human DNMT1 (which generates C5-methylcytosine) and Clostridioides difficile CamA (which generates N6-methyladenine). Structurally, compounds 9 and 11 specifically intercalate into CamA-bound DNA via the minor groove, adjacent to the target adenine, leading to a substantial conformational shift that moves the catalytic domain away from the DNA. This study adds to the limited examples of DNA methyltransferases being inhibited by non-nucleotide compounds through DNA intercalation, following the discovery of dicyanopyridine-based inhibitors for DNMT1. Furthermore, our study shows that some of these quinoline-based analogs inhibit other enzymes that act on DNA, such as polymerases and base excision repair glycosylases. Finally, in cancer cells compound 11 elicits DNA damage response via p53 activation. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=108 SRC="FIGDIR/small/587980v1_ufig1.gif" ALT="Figure 1"> View larger version (34K): org.highwire.dtl.DTLVardef@da2e94org.highwire.dtl.DTLVardef@27f909org.highwire.dtl.DTLVardef@4ec628org.highwire.dtl.DTLVardef@1b384df_HPS_FORMAT_FIGEXP M_FIG C_FIG HighlightsO_LISix of fifteen quinoline-based derivatives demonstrated comparable low micromolar inhibitory effects on human cytosine methyltransferase DNMT1, and the bacterial adenine methyltransferases Clostridioides difficile CamA and Caulobacter crescentus CcrM. C_LIO_LICompounds 9 and 11 were found to intercalate into a DNA substrate bound by CamA. C_LIO_LIThese quinoline-based derivatives also showed inhibitory activity against various base excision repair DNA glycosylases, and DNA and RNA polymerases. C_LIO_LICompound 11 provokes DNA damage response via p53 activation in cancer cells. C_LI

molecular biology↗

bioRxiv2023

Inhibition of the macrophage demethylase LSD1 reverses Leishmania amazonensis-induced transcriptomic changes and causes a decrease in parasite load

Intracellular pathogens exploit host cell functions to favor their own survival. In recent years, the subversion of epigenetic regulation has emerged as a key microbial strategy to modify host cell gene expression and evade antimicrobial immune responses. Using the protozoan parasite Leishmania as a model system, we have recently demonstrated that infection causes histone H3 hypomethylation, which is associated with the establishment of an anti-inflammatory phenotype, suggesting that host cell demethylases may play a role in the intracellular survival of these parasites. In this study, we combined pharmacological inhibition with RNA sequencing and quantitative immune-precipitation analysis to investigate the role of the macrophage lysine demethylase LSD1 (KDM1a) in Leishmania intracellular infection in vitro. Treatment of infected macrophages with validated, LSD1-specific inhibitors resulted in a significant reduction in parasite burden. We confirmed the impact of these inhibitors on LSD1 activity within macrophage nuclear extracts using an in vitro demethylase assay and established their LSD1 target engagement in situ by cellular thermal shift assay. RNA-seq analysis of infected and inhibitor-treated macrophages linked parasite killing to a partial reversion of infection-dependent expression changes, restoring the macrophage anti-microbial response and limiting cholesterol biosynthesis. While we ruled out any impact of Leishmania on LSD1 expression or localization, we uncovered significant alterations in LSD1 complex formation within infected macrophages, involving unique interactions with host cell regulatory proteins such as Rcor-1. Our study sheds important new light on the epigenetic mechanisms of macrophage immuno-metabolic subversion by intracellular Leishmania and identifies LSD1 as a potential candidate for host-directed, anti-leishmanial therapy.

cell biology↗

bioRxiv2021

Lipid droplets 3D full measurement by holographic in-flow tomography

The most recent discoveries in the biochemical field are highlighting the increasingly important role of lipid droplets (LDs) in several regulatory mechanisms in living cells. LDs are dynamic organelles and therefore their complete characterization in terms of number, size, spatial positioning and relative distribution in the cell volume can shed light on the roles played by LDs. Until now, fluorescence microscopy and transmission electron microscopy are assessed as the gold standard methods for identifying LDs due to their high sensitivity and specificity. However, such methods generally only provide 2D assays and partial measurements. Furthermore, both can be destructive and with low productivity, thus limiting analysis of large cell numbers in a sample. Here we demonstrate for the first time the capability of 3D visualization and the full LD characterization in high-throughput with a tomographic phase-contrast flow-cytometer, by using ovarian cancer cells and monocyte cell lines as models. A strategy for retrieving significant parameters on spatial correlations and LD 3D positioning inside each cell volume is reported. The information gathered by this new method could allow more in depth understanding and lead to new discoveries on how LDs are correlated to cellular functions.

biophysics↗

bioRxiv2021

Identification of Potent Small Molecule Inhibitors of SARS-CoV-2 Entry

The severe acute respiratory syndrome coronavirus 2 responsible for COVID-19 remains a persistent threat to mankind, especially for the immunocompromised and elderly for which the vaccine may have limited effectiveness. Entry of SARS-CoV-2 requires a high affinity interaction of the viral spike protein with the cellular receptor angiotensin-converting enzyme 2. Novel mutations on the spike protein correlate with the high transmissibility of new variants of SARS-CoV-2, highlighting the need for small molecule inhibitors of virus entry into target cells. We report the identification of such inhibitors through a robust high-throughput screen testing 15,000 small molecules from unique libraries. Several leads were validated in a suite of mechanistic assays, including whole cell SARS-CoV-2 infectivity assays. The main lead compound, Calpeptin, was further characterized using SARS-CoV-1 and the novel SARS-CoV-2 variant entry assays, SARS-CoV-2 protease assays and molecular docking. This study reveals Calpeptin as a potent and specific inhibitor of SARS-CoV-2 and some variants.

microbiology↗

bioRxiv2021

Partial resistance to HDAC inhibitors in FAPs of dystrophic muscles at late stages of disease is associated to epigenetic and transcriptional features of cellular senescence.

Pharmacological treatment of Duchenne Muscular Dystrophy (DMD) with histone deacetylase inhibitors (HDACi) is currently being tested in clinical trials. Pre-clinical studies performed in mdx mice - the mouse model of DMD - have shown that HDACi promote compensatory muscle regeneration, while inhibiting fibro-adipogenic degeneration, by targeting fibro-adipogenic progenitors (FAPs); however, these beneficial effects are restricted to early stages of disease progression. We show here that FAPs from late stage mdx mice exhibit epigenetic and transcriptional features of senescence that could not be fully reversed by HDACi. In particular, genome-wide increase in H3K9/14 acetylation at gene promoters of Senescence Associated Secretory Phenotype (SASP) genes was associated with their upregulation in late stage mdx FAPs. Treatment with the HDACi Trichostatin A (TSA) could inhibit SASP gene activation in FAPs, by decreasing H3K9/14 acetylation. Conversely, combinatorial decrease of H3K27 and/or H3K9/14 acetylation at promoters of genes required for cycle activation and progression was associated with their downregulation in FAPs from late stage mdx mice. However, these epigenetic and transcriptional alterations could not be reversed by TSA, due to a general resistance exhibited by FAPs from late stage mdx mice to HDACi-induced H3K9/14 hyperacetylation. Overall, this data reveal that disease-associated features of senescence develop in FAPs of DMD muscle through epigenetically distinct and pharmacologically dissociable events, and suggests that HDACi might at least retain anti- fibrotic and inflammatory activity at late stages of DMD, by repressing FAP-derived SASP.

pharmacology and toxicology↗

bioRxiv2020

No evidence for prolactin's involvement in the post-ejaculatory refractory period

In many species, ejaculation is followed by a state of decreased sexual motivation, the post-ejaculatory refractory period. Several lines of evidence have suggested prolactin, a pituitary hormone released around the time of ejaculation in humans and other animals, to be a decisive player in the establishment of the refractory period. However, data supporting this hypothesis is controversial. We took advantage of two different strains of house mouse, a wild derived and a classical laboratory strain, that differ substantially in their sexual behavior, to investigate prolactins involvement in sexual motivation and the refractory period. First, we show that there is prolactin release during sexual behavior in male mice. Second, using a pharmacological approach, we show that acute manipulations of prolactin levels, either mimicking the natural release during sexual behavior or inhibiting its occurrence, do not affect sexual motivation or shorten the refractory period, respectively. Therefore, we show compelling evidence refuting the idea that prolactin released during copulation is involved in the establishment of the refractory period, a long-standing hypothesis in the field of behavioral endocrinology.

animal behavior and cognition↗

bioRxiv2018

Histological evidences of diabetic kidney disease precede clinical diagnosis

In the absence of a histhological diagnosis, persistent albuminuria is globally accepted as the main diagnostic criteria for diabetic kidney disease.\n\nIn the present retrospective study, we evaluated data from an Italian cohort of 42 diabetic deceased donors (mainly type 2 diabetes). Using kidney biopsies obtained at time of donation to evaluate single or double allocation based on Karpinski score, we determined the prevalence of histological lesions attributable to diabetes.\n\nAll 42 donors presented with normoalbuminuria and an estimated Glomerular Filtration Rate (CKD-EPI) > 60 ml/min/1.73m2. Of the 36 patients with available kidney biopsy, one was non interpretable and 32 had histopathological lesions consistent with diabetic kidney disease and encompassing all histological classes.\n\nThus, we found a relatively high proportion of histologically proven diabetic kidney disease that was clinically undiagnosed as none of the patient had significant albuminuria and eGFR <60 cc/min/1.73 m2. Our data support the need to implement routine kidney biopsies in normoalbuminuric diabetic subjects in early stages of Chronic Kidney Disease. As others have shown that histological lesions associated with diabetic kidney disease predict disease progression, such strategy may help to risk stratify patients and may guide therapeutic decisions at early disease stages.

scientific communication and education↗

bioRxiv2018

Stepping into a dangerous quagmire: environmental determinants of human-lancehead pit vipers (Bothrops genus) contact resulting in injuries, Brazilian Amazon

Despite significant and successful efforts in Brazil regarding snakebites in the areas of research, antivenom manufacture and quality control, training of health professionals in the diagnosis and clinical management of bites, little is known about determinants of snakebites incidence in order to further plan interventions to reduce the impact of this medical condition. Understanding the complexity of ecological interactions in a geographical region is important for prediction, prevention and control measures of snakebites. The aim of this investigation is to describe spatial distribution and identify environmental determinants of human- lancehead pit vipers (Bothrops genus) contact resulting in injuries, in the Brazilian Amazon. Aggregated data by municipality was used to analyze the spatial distribution of Bothrops bites cases and its relationship with geographic and environmental factors. Eight geo-environmental factors were included in the analysis as independent variables: (1) tree canopy loss increase; (2) area with vegetation cover; (3) area covered by water bodies; (4) altitude; (5) precipitation; (6) air relative humidity; (7) soil moisture; and (8) air temperature. Human- lancehead pit vipers (Bothrops genus) contact resulting in envenomings in the Amazon region is more incident in lowlands [-0.0006827 (IC95%: -0.0007705; - 0.0005949), p<0.0001], with high preserved original vegetation cover [0.0065439 (IC95%: 0.0070757; 0.0060121), p<0.0001], with heaviest rainfall [0.0000976 (IC95%: 0.0000925; 0.0001026), p<0.0001] and higher air relative humidity [- 0.0081773 (IC95%: -0.0107681; -0.0055865), p<0.0001]. This association is interpreted as the result of the higher forest productivity and abundance of pit vipers in such landscapes.\n\nAuthor summaryDespite successful efforts in Brazil regarding snakebites in the areas of research, antivenom manufacture and quality control and training of health professionals, little is known about determinants of snakebites incidence in order to further plan interventions to reduce the impact of this medical condition. Understanding the complexity of ecological interactions in a geographical region is important for prediction, prevention and control measures of snakebites. The aim of this study is to describe spatial distribution and identify environmental determinants of human- lancehead pit vipers (Bothrops genus) contact resulting in injuries, in the Brazilian Amazon. An increase in the forest productivity with a higher availability of some types of prey, such as frogs and amphibians, anurans and lizards, was suggested as a cause for the higher snake abundance in the rainy season. Probably due to the higher forest productivity and abundance of pit vipers in such landscapes, human-lancehead pit vipers contact resulting in envenomings in the Amazon region is more incident in lowlands, with high preserved original vegetation cover, with heaviest rainfall and higher air relative humidity.

animal behavior and cognition↗