Publications by authors named "Mattia Mori"

111 Publications

Hepatic miR-144 Drives Fumarase Activity Preventing NRF2 Activation During Obesity.

Gastroenterology 2021 Aug 21. Epub 2021 Aug 21.

Center for Infectious Medicine (CIM), Department of Medicine, Karolinska Institutet, Huddinge, Sweden. Electronic address:

Background And Aims: Oxidative stress plays a key role in the development of metabolic complications associated with obesity, including insulin resistance and the most common chronic liver disease worldwide, nonalcoholic fatty liver disease. We have recently discovered that the microRNA miR-144 regulates protein levels of the master mediator of the antioxidant response, nuclear factor erythroid 2-related factor 2 (NRF2). On miR-144 silencing, the expression of NRF2 target genes was significantly upregulated, suggesting that miR-144 controls NRF2 at the level of both protein expression and activity. Here we explored a mechanism whereby hepatic miR-144 inhibited NRF2 activity upon obesity via the regulation of the tricarboxylic acid (TCA) metabolite, fumarate, a potent activator of NRF2.

Methods: We performed transcriptomic analysis in liver macrophages (LMs) of obese mice and identified the immuno-responsive gene 1 (Irg1) as a target of miR-144. IRG1 catalyzes the production of a TCA derivative, itaconate, an inhibitor of succinate dehydrogenase (SDH). TCA enzyme activities and kinetics were analyzed after miR-144 silencing in obese mice and human liver organoids using single-cell activity assays in situ and molecular dynamic simulations.

Results: Increased levels of miR-144 in obesity were associated with reduced expression of Irg1, which was restored on miR-144 silencing in vitro and in vivo. Furthermore, miR-144 overexpression reduces Irg1 expression and the production of itaconate in vitro. In alignment with the reduction in IRG1 levels and itaconate production, we observed an upregulation of SDH activity during obesity. Surprisingly, however, fumarate hydratase (FH) activity was also upregulated in obese livers, leading to the depletion of its substrate fumarate. miR-144 silencing selectively reduced the activities of both SDH and FH resulting in the accumulation of their related substrates succinate and fumarate. Moreover, molecular dynamics analyses revealed the potential role of itaconate as a competitive inhibitor of not only SDH but also FH. Combined, these results demonstrate that silencing of miR-144 inhibits the activity of NRF2 through decreased fumarate production in obesity.

Conclusions: Herein we unravel a novel mechanism whereby miR-144 inhibits NRF2 activity through the consumption of fumarate by activation of FH. Our study demonstrates that hepatic miR-144 triggers a hyperactive FH in the TCA cycle leading to an impaired antioxidant response in obesity.
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http://dx.doi.org/10.1053/j.gastro.2021.08.030DOI Listing
August 2021

Active Components from Prevent HIV-1 Entry by Distinct Mechanisms of Action.

Int J Mol Sci 2021 May 10;22(9). Epub 2021 May 10.

Department of Infection and Immunity, Luxembourg Institute of Health, L-4354 Esch-sur-Alzette, Luxembourg.

is widely used in Sub-Saharan Africa for treating many diseases, including HIV-1 infection. We have recently described the chemical structures of 28 compounds isolated from an alcoholic crude extract of barks and roots of , and showed that six bioactive compounds inhibit HIV-1 infection. In the present study, we demonstrate that the six compounds block HIV-1 entry into cells: oleanolic acid, palmitic acid, taxifolin, piceatannol, guibourtinidol-(4α→8)-epiafzelechin, and a novel compound named as cassiabrevone. We report, for the first time, that guibourtinidol-(4α→8)-epiafzelechin and cassiabrevone inhibit HIV-1 entry (IC of 42.47 µM and 30.96 µM, respectively), as well as that piceatannol interacts with cellular membranes. Piceatannol inhibits HIV-1 infection in a dual-chamber assay mimicking the female genital tract, as well as HSV infection, emphasizing its potential as a microbicide. Structure-activity relationships (SAR) showed that pharmacophoric groups of piceatannol are strictly required to inhibit HIV-1 entry. By a ligand-based in silico study, we speculated that piceatannol and norartocarpetin may have a very similar mechanism of action and efficacy because of the highly comparable pharmacophoric and 3D space, while guibourtinidol-(4α→8)-epiafzelechin and cassiabrevone may display a different mechanism. We finally show that cassiabrevone plays a major role of the crude extract of by blocking the binding activity of HIV-1 gp120 and CD4.
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http://dx.doi.org/10.3390/ijms22095052DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8126241PMC
May 2021

Design and Synthesis of Piperazine-Based Compounds Conjugated to Humanized Ferritin as Delivery System of siRNA in Cancer Cells.

Bioconjug Chem 2021 06 12;32(6):1105-1116. Epub 2021 May 12.

Center for Life Nano- & Neuro-Science, Fondazione Istituto Italiano di Tecnologia (IIT), V.le Regina Elena 291, 00161 Rome, Italy.

Gene expression regulation by small interfering RNA (siRNA) holds promise in treating a wide range of diseases through selective gene silencing. However, successful clinical application of nucleic acid-based therapy requires novel delivery options. Herein, to achieve efficient delivery of negatively charged siRNA duplexes, the internal cavity of "humanized" chimeric Archaeal ferritin (HumAfFt) was specifically decorated with novel cationic piperazine-based compounds (PAs). By coupling these rigid-rod-like amines with thiol-reactive reagents, chemoselective conjugation was efficiently afforded on topologically selected cysteine residues properly located inside HumAfFt. The capability of PAs-HumAfFt to host and deliver siRNA molecules through human transferrin receptor (TfR1), overexpressed in many cancer cells, was explored. These systems allowed siRNA delivery into HeLa, HepG2, and MCF-7 cancer cells with improved silencing effect on glyceraldehyde-3-phosphate dehydrogenase (GAPDH) gene expression with respect to traditional transfection methodologies and provided a promising TfR1-targeting system for multifunctional siRNA delivery to therapeutic applications.
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http://dx.doi.org/10.1021/acs.bioconjchem.1c00137DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8253483PMC
June 2021

Design and Synthesis of New Withaferin A Inspired Hedgehog Pathway Inhibitors.

Chemistry 2021 Jun 4;27(32):8350-8357. Epub 2021 May 4.

Department of Chemistry, Università degli Studi di Milano, Via Golgi 19, 20133, Milan, Italy.

Withanolides constitute a well-known family of plant-based alkaloids characterised by widespread biological properties, including the ability of interfering with Hedgehog (Hh) signalling pathway. Following our interest in natural products and in anticancer compounds, we report here the synthesis of a new class of Hh signalling pathway inhibitors, inspired by withaferin A, the first isolated member of withanolides. The decoration of our scaffolds was rationally supported by in silico studies, while functional evaluation revealed promising candidates, confirming once again the importance of natural products as inspiration source for the discovery of novel bioactive compounds. A stereoselective approach, based on Brown chemistry, allowed the obtainment and the functional evaluation of the enantiopure hit compounds.
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http://dx.doi.org/10.1002/chem.202100315DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8251939PMC
June 2021

Sofosbuvir Selects for Drug-Resistant Amino Acid Variants in the Zika Virus RNA-Dependent RNA-Polymerase Complex In Vitro.

Int J Mol Sci 2021 Mar 6;22(5). Epub 2021 Mar 6.

Department of Medical Biotechnologies, University of Siena, 53100 Siena, Italy.

The nucleotide analog sofosbuvir, licensed for the treatment of hepatitis C, recently revealed activity against the Zika virus (ZIKV) in vitro and in animal models. However, the ZIKV genetic barrier to sofosbuvir has not yet been characterized. In this study, in vitro selection experiments were performed in infected human hepatoma cell lines. Increasing drug pressure significantly delayed viral breakthrough ( = 0.029). A double mutant in the NS5 gene (V360L/V607I) emerged in 3 independent experiments at 40-80 µM sofosbuvir resulting in a 3.9 ± 0.9-fold half- maximal inhibitory concentration (IC) shift with respect to the wild type (WT) virus. A triple mutant (C269Y/V360L/V607I), detected in one experiment at 80 µM, conferred a 6.8-fold IC shift with respect to the WT. Molecular dynamics simulations confirmed that the double mutant V360L/V607I impacts the binding mode of sofosbuvir, supporting its role in sofosbuvir resistance. Due to the distance from the catalytic site and to the lack of reliable structural data, the contribution of C269Y was not investigated in silico. By a combination of sequence analysis, phenotypic susceptibility testing, and molecular modeling, we characterized a double ZIKV NS5 mutant with decreased sofosbuvir susceptibility. These data add important information to the profile of sofosbuvir as a possible lead for anti-ZIKV drug development.
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http://dx.doi.org/10.3390/ijms22052670DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7962015PMC
March 2021

A Selective Competitive Inhibitor of Aldehyde Dehydrogenase 1A3 Hinders Cancer Cell Growth, Invasiveness and Stemness In Vitro.

Cancers (Basel) 2021 Jan 19;13(2). Epub 2021 Jan 19.

Department of Pharmaceutical Sciences, University of Piemonte Orientale, A. Avogadro, 28100 Novara, Italy.

Aldehyde dehydrogenase 1A3 (ALDH1A3) belongs to an enzymatic superfamily composed by 19 different isoforms, with a scavenger role, involved in the oxidation of a plethora of aldehydes to the respective carboxylic acids, through a NAD+-dependent reaction. Previous clinical studies highlighted the high expression of ALDH1A3 in cancer stem cells (CSCs) correlated to a higher risk of cancer relapses, chemoresistance and a poor clinical outcome. We report on the structural, biochemical, and cellular characterization of NR6, a new selective ALDH1A3 inhibitor derived from an already published ALDH non-selective inhibitor with cytotoxic activity on glioblastoma and colorectal cancer cells. Crystal structure, through X-Ray analysis, showed that NR6 binds a non-conserved tyrosine residue of ALDH1A3 which drives the selectivity towards this isoform, as supported by computational binding simulations. Moreover, NR6 shows anti-metastatic activity in wound healing and invasion assays and induces the downregulation of cancer stem cell markers. Overall, our work confirms the role of ALDH1A3 as an important target in glioblastoma and colorectal cells and propose NR6 as a promising molecule for future preclinical studies.
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http://dx.doi.org/10.3390/cancers13020356DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7835878PMC
January 2021

Sempervirine inhibits RNA polymerase I transcription independently from p53 in tumor cells.

Cell Death Discov 2020 Oct 28;6(1):111. Epub 2020 Oct 28.

Department of Biomedicine and Prevention, University of Rome Tor Vergata, Rome, Italy.

In the search of small molecules that can target MDM2/p53 pathway in testicular germ cell tumors (TGCTs), we identified sempervirine (2,3,4,13-tetrahydro-1H-benz[g]indolo[2,3-a]quinolizin-6-ium), an alkaloid of Gelsemium sempervirens, that has been previously proposed as an inhibitor of MDM2 that targets p53-wildtype (wt) tumor cells. We found that sempervirine not only affects cell growth of p53-wt cancer cells, but it is also active in p53-mutated and p53-null cells by triggering p53-dependent and independent pathways without affecting non-transformed cells. To understand which mechanism/s could be activated both in p53-wt and -null cells, we found that sempervirine induced nucleolar remodeling and nucleolar stress by reducing protein stability of RPA194, the catalytic subunit of RNA polymerase I, that led to rRNA synthesis inhibition and to MDM2 block. As shown for other cancer cell models, MDM2 inhibition by nucleolar stress downregulated E2F1 protein levels both in p53-wt and p53-null TGCT cells with the concomitant upregulation of unphosphorylated pRb. Finally, we show that sempervirine is able to enter the nucleus and accumulates within the nucleolus where it binds rRNA without causing DNA damage. Our results identify semperivirine as a novel rRNA synthesis inhibitor and indicate this drug as a non-genotoxic anticancer small molecule.
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http://dx.doi.org/10.1038/s41420-020-00345-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7595235PMC
October 2020

Targeting the RdRp of Emerging RNA Viruses: The Structure-Based Drug Design Challenge.

Molecules 2020 Dec 3;25(23). Epub 2020 Dec 3.

Department of Biotechnology, Chemistry and Pharmacy, Department of Excellence 2018-2022, University of Siena, Via Aldo Moro 2, 53100 Siena, Italy.

The RNA-dependent RNA polymerase (RdRp) is an essential enzyme for the viral replication process, catalyzing the viral RNA synthesis using a metal ion-dependent mechanism. In recent years, RdRp has emerged as an optimal target for the development of antiviral drugs, as demonstrated by recent approvals of sofosbuvir and remdesivir against Hepatitis C virus (HCV) and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), respectively. In this work, we overview the main sequence and structural features of the RdRp of emerging RNA viruses such as Coronaviruses, Flaviviruses, and HCV, as well as inhibition strategies implemented so far. While analyzing the structural information available on the RdRp of emerging RNA viruses, we provide examples of success stories such as for HCV and SARS-CoV-2. In contrast, Flaviviruses' story has raised attention about how the lack of structural details on catalytically-competent or ligand-bound RdRp strongly hampers the application of structure-based drug design, either in repurposing and conventional approaches.
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http://dx.doi.org/10.3390/molecules25235695DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7730706PMC
December 2020

Repurposing drugs for the management of COVID-19.

Expert Opin Ther Pat 2021 Apr 31;31(4):295-307. Epub 2020 Dec 31.

Department of Biotechnology, Chemistry and Pharmacy, University of Siena, Siena, Italy.

: The coronavirus disease 2019 (COVID-19) caused by SARS-CoV-2 represents a serious health issue worldwide, with more than 61 million cases and more than 1.4 million deaths since the beginning of the epidemic near the end of 2019. The scientific community strongly responded to this emergency situation with massive research efforts, mostly focused on diagnosis and clinical investigation of therapeutic solutions. In this scenario, drug repurposing played a crucial role in accelerating advanced clinical testing and shortening the time to access the regulatory review.: This review covers the main and most successful drug repurposing approaches from a design, clinical, and regulatory standpoint. Available patents on repurposed drugs are also discussed.: Drug repurposing proved highly successful in response to the current pandemic, with remdesivir becoming the first specific antiviral drug approved for the treatment of COVID-19. In parallel, a number of drugs such as corticosteroids and low molecular weight heparin (LMWH) are used to treat hospitalized COVID-19 patients, while clinical testing of additional therapeutic options is ongoing. It is reasonably expected that these research efforts will deliver optimized and specific therapeutic tools that will increase the preparedness of health systems to possible future epidemics.
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http://dx.doi.org/10.1080/13543776.2021.1861248DOI Listing
April 2021

Glabrescione B delivery by self-assembling micelles efficiently inhibits tumor growth in preclinical models of Hedgehog-dependent medulloblastoma.

Cancer Lett 2021 02 26;499:220-231. Epub 2020 Nov 26.

Department of Molecular Medicine, University La Sapienza, Roma, Italy; Laboratory Affiliated to Istituto Pasteur Italia-Fondazione Cenci Bolognetti, Department of Molecular Medicine, University La Sapienza, Roma, Italy. Electronic address:

Aberrant activation of the Hedgehog (Hh) pathway leads to the development of several tumors, including medulloblastoma (MB), the most common pediatric brain malignancy. Hh inhibitors acting on GLI1, the final effector of Hh signaling, offer a valuable opportunity to overcome the pitfalls of the existing therapies to treat Hh-driven cancers. In this study, the toxicity, delivery, biodistribution, and anticancer efficacy of Glabrescione B (GlaB), a selective GLI1 inhibitor, were investigated in preclinical models of Hh-dependent MB. To overcome its poor water solubility, GlaB was formulated with a self-assembling amphiphilic polymer forming micelles, called mPEG-cholane. mPEG-cholane/GlaB showed high drug loading and stability, low cytotoxicity, and long permanence in the bloodstream. We found that mPEG-cholane efficiently enhanced the solubility of GlaB, thus avoiding the use of organic solvents. mPEG-cholane/GlaB possesses favorable pharmacokinetics and negligible toxicity. Remarkably, GlaB encapsulated in mPEG-cholane micelles was delivered through the blood-brain barrier and drastically inhibited tumor growth in both allograft and orthotopic models of Hh-dependent MB. Our findings reveal that mPEG-cholane/GlaB is a good candidate for the treatment of Hh-driven tumors and provide relevant implications for the translation of GlaB into clinical practice.
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http://dx.doi.org/10.1016/j.canlet.2020.11.028DOI Listing
February 2021

Inhibitory Effect of Lithospermic Acid on the HIV-1 Nucleocapsid Protein.

Molecules 2020 Nov 20;25(22). Epub 2020 Nov 20.

Laboratory of Pharmacognosy, School of Pharmacy, Aristotle University of Thessaloniki, 54124 Thessaloniki, Greece.

The HIV-1 nucleocapsid protein (NC) is a desirable target in antiretroviral therapy due to its high conservation among HIV-1 strains, and to its multiple and crucial roles in the HIV-1 replication cycle. Natural products represent a valuable source of NC inhibitors, with the catechol group being a privileged scaffold in NC inhibition. By coupling molecular modeling with NMR spectroscopy and fluorescence-based assays, we disclosed lithospermic acid, a catechol derivative extracted from , as a potent and chemically stable non-covalent inhibitor of the NC. Being different from other catechol derivative reported so far, lithospermic acid does not undergo spontaneous oxidation in physiological conditions, thus becoming a profitable starting point for the development of efficient NC inhibitors.
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http://dx.doi.org/10.3390/molecules25225434DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7699738PMC
November 2020

Experimental and Computational Druggability Exploration of the 14-3-3ζ/SOS1pS PPI Interface.

J Chem Inf Model 2020 12 2;60(12):6555-6565. Epub 2020 Nov 2.

Department of Biotechnology, Chemistry and Pharmacy, Department of Excellence 2018-2022, University of Siena, Via Aldo Moro 2, 53100 Siena, Italy.

The exploration of the druggability of certain protein-protein interactions (PPIs) still remains a challenging task in drug discovery. Here, we present a case study using the 14-3-3-PPI, showing how small molecules can be located that are able to modulate this key oncogenic pathway. A workflow embracing biophysical techniques and MD simulations was developed to evaluate the potential of a 14-3-3ζ PPI system to bind new tool compounds. The significance of the use of computational approaches to compensate for the limitations of experimental techniques is demonstrated.
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http://dx.doi.org/10.1021/acs.jcim.0c00722DOI Listing
December 2020

Inhibition of Melanoma Cell Migration and Invasion Targeting the Hypoxic Tumor Associated CAXII.

Cancers (Basel) 2020 Oct 17;12(10). Epub 2020 Oct 17.

Department of Medical Biotechnologies, Cellular and Molecular Physiology Unit, University of Siena, 53100 Siena, Italy.

Background: Intratumoral hypoxia contributes to cancer progression and poor prognosis. Carbonic anhydrases IX (CAIX) and XII (CAXII) play pivotal roles in tumor cell adaptation and survival, as aberrant Hedgehog (Hh) pathway does. In malignant melanoma both features have been investigated for years, but they have not been correlated before and/or identified as a potential pharmacological target. Here, for the first time, we demonstrated that malignant melanoma cell motility was impaired by targeting CAXII via either CAs inhibitors or through the inhibition of the Hh pathway.

Methods: We tested cell motility in three melanoma cell lines (WM-35, SK-MEL28, and A375), with different invasiveness capabilities. To this end we performed a scratch assay in the presence of the smoothened (SMO) antagonist cyclopamine (cyclo) or CAs inhibitors under normoxia or hypoxia. Then, we analyzed the invasiveness potential in the cell lines which were more affected by cyclo and CAs inhibitors (SK-MEL28 and A375). Western blot was employed to assess the expression of the hypoxia inducible factor 1α, CAXII, and FAK phosphorylation. Immunofluorescence staining was performed to verify the blockade of CAXII expression.

Results: Hh inhibition reduced melanoma cell migration and CAXII expression under both normoxic and hypoxic conditions. Interestingly, basal CAXII expression was higher in the two more aggressive melanoma cell lines. Finally, a direct CAXII blockade impaired melanoma cell migration and invasion under hypoxia. This was associated with a decrease of FAK phosphorylation and metalloprotease activities.

Conclusions: CAXII may be used as a target for melanoma treatment not only through its direct inhibition, but also through Hh blockade.
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http://dx.doi.org/10.3390/cancers12103018DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7602957PMC
October 2020

What Makes Thienoguanosine an Outstanding Fluorescent DNA Probe?

J Am Chem Soc 2020 10 23;142(40):16999-17014. Epub 2020 Sep 23.

Laboratoire de Bioimagerie et Pathologies, UMR 7021 CNRS Université de Strasbourg, Faculté de Pharmacie, 74 route du Rhin, 67401 Illkirch, France.

Thienoguanosine (G) is an isomorphic guanosine (G) surrogate that almost perfectly mimics G in nucleic acids. To exploit its full potential and lay the foundation for future applications, 20 DNA duplexes, where the bases facing and neighboring G were systematically varied, were thoroughly studied using fluorescence spectroscopy, molecular dynamics simulations, and mixed quantum mechanical/molecular mechanics calculations, yielding a comprehensive understanding of its photophysics in DNA. In matched duplexes, G's hypochromism was larger for flanking G/C residues but its fluorescence quantum yield (QY) and lifetime values were almost independent of the flanking bases. This was attributed to high duplex stability, which maintains a steady orientation and distance between nucleobases, so that a similar charge transfer (CT) mechanism governs the photophysics of G independently of its flanking nucleobases. G can therefore replace any G residue in matched duplexes, while always maintaining similar photophysical features. In contrast, the local destabilization induced by a mismatch or an abasic site restores a strong dependence of G's QY and lifetime values on its environmental context, depending on the CT route efficiency and solvent exposure of G. Due to this exquisite sensitivity, G appears ideal for monitoring local structural changes and single nucleotide polymorphism. Moreover, G's dominant fluorescence lifetime in DNA is unusually long (9-29 ns), facilitating its selective measurement in complex media using a lifetime-based or a time-gated detection scheme. Taken together, our data highlight G as an outstanding emissive substitute for G with good QY, long fluorescence lifetimes, and exquisite sensitivity to local structural changes.
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http://dx.doi.org/10.1021/jacs.0c06165DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7544670PMC
October 2020

14-3-3 binding creates a memory of kinase action by stabilizing the modified state of phospholamban.

Sci Signal 2020 09 1;13(647). Epub 2020 Sep 1.

Department of Molecular Biology, Universitätsmedizin Göttingen, Humboldtallee 23, 37073 Göttingen, Germany.

The cardiac membrane protein phospholamban (PLN) is targeted by protein kinase A (PKA) at Ser and by Ca/calmodulin-dependent protein kinase II (CaMKII) at Thr β-Adrenergic stimulation and PKA-dependent phosphorylation of Ser acutely stimulate the sarcoplasmic reticulum calcium pump (SERCA) by relieving its inhibition by PLN. CaMKII-dependent phosphorylation may lead to longer-lasting SERCA stimulation and may sustain maladaptive Ca handling. Here, we demonstrated that phosphorylation at either Ser or Thr converted PLN into a target for the phosphoadaptor protein 14-3-3 with different affinities. 14-3-3 proteins were localized within nanometers of PLN and endogenous 14-3-3 coimmunoprecipitated with pentameric PLN from cardiac membranes. Molecular dynamics simulations predicted different molecular contacts for peptides phosphorylated at Ser or Thr with the binding groove of 14-3-3, resulting in varied binding affinities. 14-3-3 binding protected either PLN phosphosite from dephosphorylation. β-Adrenergic stimulation of isolated adult cardiomyocytes resulted in the membrane recruitment of endogenous 14-3-3. The exogenous addition of 14-3-3 to β-adrenergic-stimulated cardiomyocytes led to prolonged SERCA activation, presumably because 14-3-3 protected PLN pentamers from dephosphorylation. Phosphorylation of Ser was disrupted by the cardiomyopathy-associated ∆Arg mutation, implying that phosphorylation of Thr by CaMKII may become crucial for 14-3-3 recruitment to ∆Arg PLN. Consistent with PLN acting as a dynamic hub in the control of Ca handling, our results identify 14-3-3 binding to PLN as a contractility-augmenting mechanism.
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http://dx.doi.org/10.1126/scisignal.aaz1436DOI Listing
September 2020

-Beyerane Diterpenes as a Key Platform for the Development of ArnT-Mediated Colistin Resistance Inhibitors.

J Org Chem 2020 08 10;85(16):10891-10901. Epub 2020 Aug 10.

Department of Chemistry and Technology of Drugs, "Department of Excellence 2018-2022", Sapienza University of Rome, P.le Aldo Moro 5, 00185 Rome, Italy.

Colistin is a last-resort antibiotic for the treatment of multidrug resistant Gram-negative bacterial infections. Recently, a natural -beyerene diterpene was identified as a promising inhibitor of the enzyme responsible for colistin resistance mediated by lipid A aminoarabinosylation in Gram-negative bacteria, namely, ArnT (undecaprenyl phosphate-alpha-4-amino-4-deoxy-l-arabinose arabinosyl transferase). Here, semisynthetic analogues of hit were designed, synthetized, and tested against colistin-resistant strains including clinical isolates to exploit the versatility of the diterpene scaffold. Microbiological assays coupled with molecular modeling indicated that for a more efficient colistin adjuvant activity, likely resulting from inhibition of the ArnT activity by the selected compounds and therefore from their interaction with the catalytic site of ArnT, an -beyerane scaffold is required along with an oxalate-like group at C-18/C-19 or a sugar residue at C-19 to resemble L-Ara4N. The -beyerane skeleton is identified for the first time as a privileged scaffold for further cost-effective development of valuable colistin resistance inhibitors.
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http://dx.doi.org/10.1021/acs.joc.0c01459DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8009527PMC
August 2020

Potency and Selectivity Optimization of Tryptophanol-Derived Oxazoloisoindolinones: Novel p53 Activators in Human Colorectal Cancer.

ChemMedChem 2021 01 2;16(1):250-258. Epub 2020 Sep 2.

Department of Therapeutic and Pharmaceutical Chemistry, University of Lisbon, Av. Prof. Gama Pinto, 1649-003, Lisboa, Portugal.

To search for novel p53 activators, four series of novel (S)- and (R)-tryptophanol-derived oxazoloisoindolinones were synthesized in a straightforward manner and their antiproliferative activity was evaluated in the human colorectal cancer HCT116 cell line. Structural optimization of the hit compound SLMP53-1 led to the identification of a (R)-tryptophanol-derived isoindolinone that was found to be six-fold more active, with increased selectivity for HCT116 cells with p53 and with low toxicity in normal cells. Binding studies with MDM2 showed that the antiproliferative activity of tryptophanol-derived isoindolinones does not involve inhibition of the main negative regulator of the p53 protein. Molecular docking simulations showed that although these molecules establish hydrophobic interactions with MDM2, they do not possess the required features to bind MDM2.
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http://dx.doi.org/10.1002/cmdc.202000522DOI Listing
January 2021

A chalcone derivative binds a putative allosteric site of YopH: Inhibition of a virulence factor of Yersinia.

Bioorg Med Chem Lett 2020 08 12;30(16):127350. Epub 2020 Jun 12.

Centro de Biologia Molecular Estrutural, Departamento de Bioquímica, Universidade Federal de Santa Catarina, Campus Trindade, Florianópolis, SC 88040-900, Brazil; Universidade Federal do Piauí, CPCE, Bom Jesus, PI 64900-000, Brazil. Electronic address:

Identification of allosteric inhibitors of PTPs has attracted great interest as a new strategy to overcome the challenge of discover potent and selective molecules for therapeutic intervention. YopH is a virulence factor of the genus Yersinia, validated as an antimicrobial target. The finding of a second substrate binding site in YopH has revealed a putative allosteric site that could be further exploited. Novel chalcone compounds that inhibit PTPs activity were designed and synthesized. Compound 3j was the most potent inhibitor, interestingly, with different mechanisms of inhibition for the panel of enzymes evaluated. Further, our results showed that compound 3j is an irreversible non-competitive inhibitor of YopH that binds to a site different than the catalytic site, but close to the well-known second binding site of YopH.
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http://dx.doi.org/10.1016/j.bmcl.2020.127350DOI Listing
August 2020

Natural Products as an Important Source in Drug Discovery.

Curr Pharm Des 2020 ;26(24):2805-2806

Department of Biotechnology, Chemistry and Pharmacy, University of Siena, Via Aldo Moro 2, 53100 Siena, Italy.

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http://dx.doi.org/10.2174/138161282624200625104355DOI Listing
January 2021

Alvaxanthone, a Thymidylate Synthase Inhibitor with Nematocidal and Tumoricidal Activities.

Molecules 2020 Jun 23;25(12). Epub 2020 Jun 23.

Nencki Institute of Experimental Biology, 3 Pasteur Street, 02-093 Warsaw, Poland.

With the aim to identify novel inhibitors of parasitic nematode thymidylate synthase (TS), we screened in silico an in-house library of natural compounds, taking advantage of a model of nematode TS three-dimensional (3D) structure and choosing candidate compounds potentially capable of enzyme binding/inhibition. Selected compounds were tested as (i) inhibitors of the reaction catalyzed by TSs of different species, (ii) agents toxic to a nematode parasite model ( grown in vitro), (iii) inhibitors of normal human cell growth, and (iv) antitumor agents affecting human tumor cells grown in vitro. The results pointed to alvaxanthone as a relatively strong TS inhibitor that causes population growth reduction with nematocidal potency similar to the anthelmintic drug mebendazole. Alvaxanthone also demonstrated an antiproliferative effect in tumor cells, associated with a selective toxicity against mitochondria observed in cancer cells compared to normal cells.
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http://dx.doi.org/10.3390/molecules25122894DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7356228PMC
June 2020

A novel colistin adjuvant identified by virtual screening for ArnT inhibitors.

J Antimicrob Chemother 2020 09;75(9):2564-2572

Department of Sciences, 'Department of Excellence 2018 - 2022', Roma Tre University, Rome, Italy.

Background: Colistin is a last-resort treatment option for many MDR Gram-negative bacteria. The covalent addition of l-aminoarabinose to the lipid A moiety of LPS is the main colistin resistance mechanism in the human pathogen Pseudomonas aeruginosa.

Objectives: Identification (by in silico screening of a chemical library) of potential inhibitors of ArnT, which catalyses the last committed step of lipid A aminoarabinosylation, and their validation in vitro as colistin adjuvants.

Methods: The available ArnT crystal structure was used for a docking-based virtual screening of an in-house library of natural products. The resulting putative ArnT inhibitors were tested in growth inhibition assays using a reference colistin-resistant P. aeruginosa strain. The most promising compound was further characterized for its range of activity, specificity and cytotoxicity. Additionally, the effect of the compound on lipid A aminoarabinosylation was verified by MS analyses of lipid A.

Results: A putative ArnT inhibitor (BBN149) was discovered by molecular docking and demonstrated to specifically potentiate colistin activity in colistin-resistant P. aeruginosa isolates, without relevant effect on colistin-susceptible strains. BBN149 also showed adjuvant activity against colistin-resistant Klebsiella pneumoniae and low toxicity to bronchial epithelial cells. Lipid A aminoarabinosylation was reduced in BBN149-treated cells, although only partially.

Conclusions: This study demonstrates that in silico screening targeting ArnT can successfully identify inhibitors of colistin resistance and provides a promising lead compound for the development of colistin adjuvants for the treatment of MDR bacterial infections.
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http://dx.doi.org/10.1093/jac/dkaa200DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7443731PMC
September 2020

5,6-Dihydroxypyrimidine Scaffold to Target HIV-1 Nucleocapsid Protein.

ACS Med Chem Lett 2020 May 19;11(5):766-772. Epub 2020 Mar 19.

Department of Biotechnology, Chemistry and Pharmacy, University of Siena, via Aldo Moro 2, 53100 Siena, Italy.

The HIV-1 nucleocapsid (NC) protein is a small basic DNA and RNA binding protein that is absolutely necessary for viral replication and thus represents a target of great interest to develop new anti-HIV agents. Moreover, the highly conserved sequence offers the opportunity to escape the drug resistance (DR) that emerged following the highly active antiretroviral therapy (HAART) treatment. On the basis of our previous research, nordihydroguaiaretic acid acts as a NC inhibitor showing moderate antiviral activity and suboptimal drug-like properties due to the presence of the catechol moieties. A bioisosteric catechol replacement approach led us to identify the 5-dihydroxypyrimidine-6-carboxamide substructure as a privileged scaffold of a new class of HIV-1 NC inhibitors. Hit validation efforts led to the identification of optimized analogs, as represented by compound , showing improved NC inhibition and antiviral activity as well as good ADME and PK properties.
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http://dx.doi.org/10.1021/acsmedchemlett.9b00608DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7236274PMC
May 2020

Structural Elucidation and Antimicrobial Characterization of Novel Diterpenoids from var. .

ACS Med Chem Lett 2020 May 30;11(5):760-765. Epub 2020 Jan 30.

Department of Biochemical Sciences, Laboratory Affiliated to Pasteur Italia-Fondazione Cenci Bolognetti, Sapienza University of Rome, P.le Aldo Moro 5 00185, Rome, Italy.

Novel diterpenoids were isolated from the extracts of var. and found to display a selective activity against Gram-positive bacterial strains with negligible cytotoxicity toward human keratinocytes. This study highlighted the role played by the acidic group at C18 of the tetracyclic ent-beyerene scaffold for antibacterial effects and how the length and flexibility of the alkyl chain between the two carbonyl groups are crucial factors to increase the antimicrobial activity of the molecules, supporting the development of natural products from and their derivatives for treatment of microbial infections.
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http://dx.doi.org/10.1021/acsmedchemlett.9b00605DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7236223PMC
May 2020

A Class of Potent Inhibitors of the HIV-1 Nucleocapsid Protein Based on Aminopyrrolic Scaffolds.

ACS Med Chem Lett 2020 May 27;11(5):698-705. Epub 2020 Jan 27.

Laboratoire de Bioimagerie et Pathologies, UMR 7021 CNRS, Faculté de Pharmacie, Université de Strasbourg, 74 route du Rhin, 67401 Illkirch, France.

The HIV-1 nucleocapsid protein 7 (NC) is a potential target for effective antiretroviral therapy due to its central role in virus replication, mainly linked to nucleic acid (NA) chaperone activity, and low susceptibility to drug resistance. By screening a compounds library, we identified the aminopyrrolic compound CN14_17, a known carbohydrate binding agent, that inhibits the NC chaperone activity in the low micromolar range. Different from most of available NC inhibitors, CN14_17 fully prevents the NC-induced annealing of complementary NA sequences. Using fluorescence assays and isothermal titration calorimetry, we found that CN14_17 competes with NC for the binding to NAs, preferentially targeting single-stranded sequences. Molecular dynamics simulations confirmed that binding to cTAR occurs preferably within the guanosine-rich single stranded sequence. Finally, CN14_17 exhibited antiretroviral activity in the low micromolar range, although with a moderate therapeutic index. Overall, CN14_17 might be the progenitor of a new promising class of NC inhibitors.
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http://dx.doi.org/10.1021/acsmedchemlett.9b00558DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7236033PMC
May 2020

In Memory of Maurizio Botta: His Vision of Medicinal Chemistry.

ACS Med Chem Lett 2020 May 14;11(5):611. Epub 2020 May 14.

Department of Biotechnology, Chemistry and Pharmacy, University of Siena.

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http://dx.doi.org/10.1021/acsmedchemlett.0c00210DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7236028PMC
May 2020

Dual SMO/BRAF Inhibition by Flavonolignans from .

Antioxidants (Basel) 2020 May 5;9(5). Epub 2020 May 5.

Department of QSAR and Molecular Modelling, Institute of Biophysics and Biomedical Engineering, Bulgarian Academy of Sciences, 1113 Sofia, Bulgaria.

Silymarin is the standardized extract from the fruits of (L.) Gaertn., a well-known hepatoprotectant and antioxidant. Recently, bioactive compounds of silymarin, i.e., silybins and their 2,3-dehydro derivatives, have been shown to exert anticancer activities, yet with unclear mechanisms. This study combines in silico and in vitro methods to reveal the potential interactions of optically pure silybins and dehydrosilybins with novel protein targets. The shape and chemical similarity with approved drugs were evaluated in silico, and the potential for interaction with the Hedgehog pathway receptor Smoothened (SMO) and BRAF kinase was confirmed by molecular docking. In vitro studies on SMO and BRAF V600E kinase activity and in BRAF V600E A-375 human melanoma cell lines were further performed to examine their effects on these proteins and cancer cell lines and to corroborate computational predictions. Our in silico results direct to new potential targets of silymarin constituents as dual inhibitors of BRAF and SMO, two major targets in anticancer therapy. The experimental studies confirm that BRAF kinase and SMO may be involved in mechanisms of anticancer activities, demonstrating dose-dependent profiles, with dehydrosilybins showing stronger effects than silybins. The results of this work outline the dual SMO/BRAF effect of flavonolignans from Silybum marianum with potential clinical significance. Our approach can be applied to other natural products to reveal their potential targets and mechanism of action.
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http://dx.doi.org/10.3390/antiox9050384DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7278695PMC
May 2020

A deadly spillover: SARS-CoV-2 outbreak.

Expert Opin Ther Pat 2020 Jul 29;30(7):481-485. Epub 2020 Apr 29.

Dipartimento Neurofarba, Sezione Di Scienze Farmaceutiche E Nutraceutiche, Università Degli Studi Di Firenze , Sesto Fiorentino (Florence), Italy.

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http://dx.doi.org/10.1080/13543776.2020.1760838DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7196921PMC
July 2020

Imidazo[1,2-]pyridine Derivatives as Aldehyde Dehydrogenase Inhibitors: Novel Chemotypes to Target Glioblastoma Stem Cells.

J Med Chem 2020 05 20;63(9):4603-4616. Epub 2020 Apr 20.

Department of Pharmacy, University of Pisa, Via Bonanno 6, 56126 Pisa, Italy.

Glioblastoma multiforme (GBM) is the deadliest form of brain tumor. It is known for its ability to escape the therapeutic options available to date thanks to the presence of a subset of cells endowed with stem-like properties and ability to resist to cytotoxic treatments. As the cytosolic enzyme aldehyde dehydrogenase 1A3 turns out to be overexpressed in these kinds of cells, playing a key role for their vitality, treatments targeting this enzyme may represent a successful strategy to fight GBM. In this work, we describe a novel class of imidazo[1,2-]pyridine derivatives as aldehyde dehydrogenase 1A3 inhibitors, reporting the evidence of their significance as novel drug candidates for the treatment of GBM. Besides showing an interesting functional profile, in terms of activity against the target enzyme and selectivity toward highly homologous isoenzymes, representative examples of the series also showed a nanomolar to picomolar efficacy against patient-derived GBM stem-like cells, thus proving the concept that targeting aldehyde dehydrogenase might represent a novel and promising way to combat GBM by striking its ability to divide immortally.
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http://dx.doi.org/10.1021/acs.jmedchem.9b01910DOI Listing
May 2020

Identification of Phosphate-Containing Compounds as New Inhibitors of 14-3-3/c-Abl Protein-Protein Interaction.

ACS Chem Biol 2020 04 20;15(4):1026-1035. Epub 2020 Mar 20.

Department of Biotechnology, Chemistry and Pharmacy-Department of Excellence 2018-2022, Università degli Studi di Siena, via Aldo Moro 2, 53100 Siena, Italy.

The 14-3-3/c-Abl protein-protein interaction (PPI) is related to carcinogenesis and in particular to pathogenesis of chronic myeloid leukemia (CML). Previous studies have demonstrated that molecules able to disrupt this interaction improve the nuclear translocation of c-Abl, inducing apoptosis in leukemia cells. Through an X-ray crystallography screening program, we have identified two phosphate-containing compounds, inosine monophosphate (IMP) and pyridoxal phosphate (PLP), as binders of human 14-3-3σ, by targeting the protein amphipathic groove. Interestingly, they also act as weak inhibitors of the 14-3-3/c-Abl PPI, demonstrated by NMR, SPR, and FP data. A 37-compound library of PLP and IMP analogues was investigated using a FP assay, leading to the identification of three further molecules acting as weak inhibitors of the 14-3-3/c-Abl complex formation. The antiproliferative activity of IMP, PLP, and the three derivatives was tested against K-562 cells, showing that the parent compounds had the most pronounced effect on tumor cells. PLP and IMP were also effective in promoting the c-Abl nuclear translocation in c-Abl overexpressing cells. Further, these compounds demonstrated low cytotoxicity on human Hs27 fibroblasts. In conclusion, our data suggest that 14-3-3σ targeting compounds represent promising hits for further development of drugs against c-Abl-dependent cancers.
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http://dx.doi.org/10.1021/acschembio.0c00039DOI Listing
April 2020

Hedgehog signaling pathway inhibitors: an updated patent review (2015-present).

Expert Opin Ther Pat 2020 Apr 19;30(4):235-250. Epub 2020 Feb 19.

Department of Biotechnology, Chemistry and Pharmacy, Department of Excellence 2018-2022, University of Siena, Siena, Italy.

: Hedgehog (Hh) signaling plays a pivotal role in tissue development and stemness, and its deregulation is found in many different tumors. Several efforts have been devoted to discovery of Hh inhibitors, including three drugs approved by the Food and Drug Administration (FDA), targeting the upstream receptor smoothened (SMO). However, SMO mutations or SMO-independent Hh pathway activation raise the need for novel Hh inhibitors.: This review describes Hh inhibitors with anticancer potential patented in the period 2015-present.: Despite the initial enthusiasm in SMO antagonists, drug-resistant mutations, and SMO-independent Hh activation limited their clinical application. A growing number of therapeutic strategies are currently focusing on downstream Hh effectors (i.e. glioma-associate oncogenes (GLI) proteins) or other signaling pathways related to Hh, in addition to drug repositioning. Given the heterogenic nature of cancers, a terrific clinical impact is expected by multi-targeting approaches able to modulate simultaneously SMO and GLI, and/or additional targets that act as regulators of Hh signaling. It is expected that these alternative strategies might be investigated in clinical trials in the next years against a wide variety of tumor types, and that they provide improved outcomes compared to current SMO antagonists or other single-agent anticancer drugs.
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http://dx.doi.org/10.1080/13543776.2020.1730327DOI Listing
April 2020
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