APH Inhibitors that Reverse Aminoglycoside Resistance in Enterococcus casseliflavus.

Aminoglycoside-phosphotransferases (APHs) are a class of bacterial enzymes that mediate acquired resistance to aminoglycoside antibiotics. Here we report the identification of small molecules counteracting aminoglycoside resistance in Enterococcus casseliflavus. Molecular dynamics simulations were performed to identify an allosteric pocket in three APH enzymes belonging to 3' and 2'' subfamilies in which we then screened, in silico, 12,000 small molecules.

Third-Generation CD73 Inhibitors Based on a 4,6-Disubstituted-2-Thiopyridine Scaffold.

Various series of 4,6-disubstituted-2-thiopyridine derivatives were synthesized and evaluated as potential ecto-5'-nucleotidase (CD73) inhibitors. Altogether, about ninety compounds were prepared using a general synthetic pathway involving one or two steps (eventually one-pot) procedures. Variation of the nature of the substituents in positions 4 and 6 (methyl, trifluoromethyl or phenyl) of the thiopurine ring, as well as on the thiol function, was examined and led to marked differences both in term of reactivity and ability to interfere with the putative target protein.

A new class of capsid-targeting inhibitors that specifically block HIV-1 nuclear import.

HIV-1 capsids cross nuclear pore complexes (NPCs) by engaging with the nuclear import machinery. To identify compounds that inhibit HIV-1 nuclear import, we screened drugs in silico on a three-dimensional model of a CA hexamer bound by Transportin-1 (TRN-1). Among hits, compound H27 inhibited HIV-1 with a low micromolar IC.

The Oncoprotein Fra-2 Drives the Activation of Human Endogenous Retrovirus Env Expression in Adult T-Cell Leukemia/Lymphoma (ATLL) Patients.

Human endogenous retroviruses (HERVs) are retroviral sequences integrated into 8% of the human genome resulting from ancient exogenous retroviral infections. Unlike endogenous retroviruses of other mammalian species, HERVs are mostly replication and retro-transposition defective, and their transcription is strictly regulated by epigenetic mechanisms in normal cells. A significant addition to the growing body of research reveals that HERVs' aberrant activation is often associated with offsetting diseases like autoimmunity, neurodegenerative diseases, cancers, and chemoresistance.

Structural basis of the mycobacterial stress-response RNA polymerase auto-inhibition via oligomerization.

Self-assembly of macromolecules into higher-order symmetric structures is fundamental for the regulation of biological processes. Higher-order symmetric structure self-assembly by the gene expression machinery, such as bacterial DNA-dependent RNA polymerase (RNAP), has never been reported before. Here, we show that the stress-response σ factor from the human pathogen, Mycobacterium tuberculosis, induces the RNAP holoenzyme oligomerization into a supramolecular complex composed of eight RNAP units.

An evolutionarily conserved N-terminal leucine is essential for MX1 GTPase antiviral activity against different families of RNA viruses.

Myxovirus resistance protein 1 (MX1) and MX2 are homologous, dynamin-like large GTPases, induced upon interferon exposure. Human MX1 (HsMX1) is known to inhibit many viruses, including influenza A virus, by likely acting at various steps of their life cycles. Despite decades of studies, the mechanism(s) of action with which MX1 proteins manage to inhibit target viruses is not fully understood.

Identification of a non-canonical G3BP-binding sequence in a Mayaro virus nsP3 hypervariable domain.

Ras-GTPase-activating SH3 domain-binding-proteins 1 (G3BP1) and 2 (G3BP2) are multifunctional RNA-binding proteins involved in stress granule nucleation, previously identified as essential cofactors of Old World alphaviruses. They are recruited to viral replication complexes formed by the Chikungunya virus (CHIKV), Semliki Forest virus (SFV), and Sindbis virus (SINV) an interaction with a duplicated FGxF motif conserved in the hypervariable domain (HVD) of virus-encoded nsP3. According to mutagenesis studies, this FGxF duplication is strictly required for G3BP binding and optimal viral growth.

Singular Interface Dynamics of the SARS-CoV-2 Delta Variant Explained with Contact Perturbation Analysis.

Emerging SARS-CoV-2 variants raise concerns about our ability to withstand the Covid-19 pandemic, and therefore, understanding mechanistic differences of those variants is crucial. In this study, we investigate disparities between the SARS-CoV-2 wild type and five variants that emerged in late 2020, focusing on the structure and dynamics of the spike protein interface with the human angiotensin-converting enzyme 2 (ACE2) receptor, by using crystallographic structures and extended analysis of microsecond molecular dynamics simulations. Dihedral angle principal component analysis (PCA) showed the strong similarities in the spike receptor binding domain (RBD) dynamics of the Alpha, Beta, Gamma, and Delta variants, in contrast with those of WT and Epsilon.

Cytotoxic and Antitumoral Activity of N-(9H-purin-6-yl) Benzamide Derivatives and Related Water-soluble Prodrugs.

Background: The development of small molecules as cancer treatments is still of both interest and importance.

Objective: Having synthesized and identified the initial cytotoxic activity of a series of chemically related N-(9H-purin-6-yl) benzamide derivatives, we continued their evaluation on cancer cell models. We also synthesized water-soluble prodrugs of the main compound and performed in vivo experiments.

4-Substituted-1,2,3-triazolo nucleotide analogues as CD73 inhibitors, their synthesis, in vitro screening, kinetic and in silico studies.

Three series of nucleotide analogues were synthesized and evaluated as potential CD73 inhibitors. Nucleobase replacement consisted in connecting the appropriate aromatic or purine residues through a triazole moiety that is generated from 1,3-dipolar cycloaddition. The first series is related to 4-substituted-1,2,3-triazolo-β-hydroxyphosphonate ribonucleosides.

Mitigation of heterocyclic aromatic amines in cooked meat. Part I: Informed selection of antioxidants based on molecular modeling.

This work aimed to develop a method permitting an informed choice of antioxidants to reduce carcinogenic heterocyclic aromatic amine (HAA) formation during proteinaceous food cooking. Therefore, a three-step approach was developed. First, the most promising antioxidants were selected using molecular modeling approaches.

Extracellular NAD enhances PARP-dependent DNA repair capacity independently of CD73 activity.

Changes in nicotinamide adenine dinucleotide (NAD) levels that compromise mitochondrial function trigger release of DNA damaging reactive oxygen species. NAD levels also affect DNA repair capacity as NAD is a substrate for PARP-enzymes (mono/poly-ADP-ribosylation) and sirtuins (deacetylation). The ecto-5'-nucleotidase CD73, an ectoenzyme highly expressed in cancer, is suggested to regulate intracellular NAD levels by processing NAD and its bio-precursor, nicotinamide mononucleotide (NMN), from tumor microenvironments, thereby enhancing tumor DNA repair capacity and chemotherapy resistance.

Dihydropyrimidinase protects from DNA replication stress caused by cytotoxic metabolites.

Imbalance in the level of the pyrimidine degradation products dihydrouracil and dihydrothymine is associated with cellular transformation and cancer progression. Dihydropyrimidines are degraded by dihydropyrimidinase (DHP), a zinc metalloenzyme that is upregulated in solid tumors but not in the corresponding normal tissues. How dihydropyrimidine metabolites affect cellular phenotypes remains elusive.

Transportin-1 binds to the HIV-1 capsid via a nuclear localization signal and triggers uncoating.

The initial steps of HIV replication in host cells prime the virus for passage through the nuclear pore and drive the establishment of a productive and irreparable infection. The timely release of the viral genome from the capsid-referred to as uncoating-is emerging as a critical parameter for nuclear import, but the triggers and mechanisms that orchestrate these steps are unknown. Here, we identify β-karyopherin Transportin-1 (TRN-1) as a cellular co-factor of HIV-1 infection, which binds to incoming capsids, triggers their uncoating and promotes viral nuclear import.

Synthesis of Substituted 5'-Aminoadenosine Derivatives and Evaluation of Their Inhibitory Potential toward CD73.

Derivatives of 5'-aminoadenosine containing methyl carboxylate, methyl phosphonate, gem-bisphosphonate, bis(methylphosphonate), and α-carboxylmethylphosphonate or phosphonoacetate moieties were synthesized from key intermediate 5'-aminonucleoside. These nucleotide analogues were envisaged as 5'-mono- or diphosphate nucleoside mimics. All compounds were evaluated for CD73 inhibition in a cell-based assay (MDA-MB-231) and toward the purified recombinant protein.

Lead optimization and biological evaluation of fragment-based cN-II inhibitors.

The development of cytosolic 5'-nucleotidase II (cN-II) inhibitors is essential to validate cN-II as a potential target for the reversion of resistance to cytotoxic nucleoside analogues. We previously reported a fragment-based approach combined with molecular modelling, herein, the selected hit-fragments were used again in another computational approach based on the Ilib-diverse (a software enabling to build virtual molecule libraries through fragment based de novo design) program to generate a focused library of potential inhibitors. A molecular scaffold related to a previously identified compound was selected and led to a novel series of compounds.

CD73 inhibition by purine cytotoxic nucleoside analogue-based diphosphonates.

The ecto-5'-nucleotidase CD73 has emerged as an important drug target in oncoimmunology as well as in other diseases. We describe new ADP analogues as CD73 inhibitors based on the replacement of the adenosine moiety, in the reference inhibitor APCP, by purine nucleoside analogues. Compounds were assessed for CD73 inhibition both on purified recombinant protein and on CD73-expressing cancer cells.

Identification of allosteric inhibitors of the ecto-5'-nucleotidase (CD73) targeting the dimer interface.

The ecto-5'-nucleotidase CD73 plays an important role in the production of immune-suppressive adenosine in tumor micro-environment, and has become a validated drug target in oncology. Indeed, the anticancer immune response involves extracellular ATP to block cell proliferation through T-cell activation. However, in the tumor micro-environment, two extracellular membrane-bound enzymes (CD39 and CD73) are overexpressed and hydrolyze efficiently ATP into AMP then further into immune-suppressive adenosine.

The Glutaminase-Dependent System Confers Extreme Acid Resistance to New Species and Atypical Strains of .

Neutralophilic bacteria have developed specific mechanisms to cope with the acid stress encountered in environments such as soil, fermented foods, and host compartments. In , the glutamate decarboxylase (Gad)-dependent system is extremely efficient: it requires the concerted action of glutamate decarboxylase (GadA/GadB) and of the glutamate (Glu)/γ-aminobutyrate antiporter, GadC. Notably, this system is operative also in new strains/species of , among which , but not in the "classical" species, with the exception of marine mammals strains.

Neurotoxicity of a Biopesticide Analog on Zebrafish Larvae at Nanomolar Concentrations.

Despite the ever-increasing role of pesticides in modern agriculture, their deleterious effects are still underexplored. Here we examine the effect of A6, a pesticide derived from the naturally-occurring α-terthienyl, and structurally related to the endocrine disrupting pesticides anilinopyrimidines, on living zebrafish larvae. We show that both A6 and an anilinopyrimidine, cyprodinyl, decrease larval survival and affect central neurons at micromolar concentrations.

Kinetic characterization and molecular docking of novel allosteric inhibitors of aminoglycoside phosphotransferases.

Background: Bacterial antibiotic resistance often leads to treatment failure which may have serious consequences, especially in critically sick patients. Resistance to aminoglycosides is mainly due to the expression of antibiotic-modifying enzymes. One important mechanism of aminoglycoside modification is the ATP/GTP-dependent O-phosphorylation catalyzed by aminoglycoside phosphotransferases, APHs.

Beta-hydroxyphosphonate ribonucleoside analogues derived from 4-substituted-1,2,3-triazoles as IMP/GMP mimics: synthesis and biological evaluation.

A series of seventeen β-hydroxyphosphonate ribonucleoside analogues containing 4-substituted-1,2,3-triazoles was synthesized and fully characterized. Such compounds were designed as potential inhibitors of the cytosolic 5'-nucleotidase II (cN-II), an enzyme involved in the regulation of purine nucleotide pools. NMR and molecular modelling studies showed that a few derivatives adopted similar structural features to IMP or GMP.

An evolutionary conserved zinc finger protein is involved in Toxoplasma gondii mRNA nuclear export.

Apicomplexan parasites are responsible for some of the most deadly parasitic diseases affecting humans and livestock. There is an urgent need for new medicines that will target apicomplexan-specific pathways. We characterized a Toxoplasma gondii C2H2 zinc finger protein, named TgZNF2, which is conserved among eukaryotes.