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The binding of mineralocorticoid hormones to the mineralocorticoid receptor is the first step in a cascade of events leading to the stimulation of Na ϩ reabsorption by renal cortical collecting duct (CCD) principal cells. The agonist properties of mineralocorticoid hormones are linked to contacts between their 21-hydroxyl group and Asn770, a residue of the ligand-binding domain of the human mineralocorticoid receptor (hMR). Here, we investigate whether the presence of a hydroxyl group at position 11, 17, or 20 could also alter the activity of progesterone (P), a mineralocorticoid antagonist without the 21-hydroxyl group. Both 17␣-hydroxyprogesterone (17OHP) and 20␣-hydroxyprogesterone (20OHP) antagonized the aldosterone-induced trans-activation activity (IC 50 : 17OHP, 10 Ϫ7 M; 20OHP, 10 Ϫ8 M) of the hMR transiently expressed in COS-7 cells lacking steroid receptors. In cultured mouse mpkCCD cl4 principal cells, 17OHP and 20OHP also prevented the aldosterone-stimulated amiloride-sensitive component of the short-circuit current (Ams I sc ), reflecting Na ϩ absorption mediated by the epithelial Na ϩ channel (ENaC). In contrast, 11␤-hydroxyprogesterone (11OHP) activated the transiently expressed hMR in COS-7 cells in a dose-dependent manner (ED 50 : 10 Ϫ8 M) and, like aldosterone, stimulated Ams I sc in mpkCCD cl4 cells. Docking 11OHP within the hMR-ligand-binding domain homology model revealed that the agonist activity of 11OHP is caused by contacts between its 11␤-hydroxyl group and Asn770. Furthermore, 11OHP was unable to activate the mutant hMR/N770A, in which Ala is substituted for Asn at position 770. These findings demonstrate that in the absence of the 21-hydroxyl group, the 11␤-hydroxyl group can produce the contact with the hMRAsn770 required for the hMR activation leading to stimulated Na ϩ absorption.In the kidney, the collecting duct is the main site of Na ϩ reabsorption and is subjected to a fine hormonal control by aldosterone and vasopressin (Rossier and Palmer, 1992). In the principal cells of the cortical collecting duct (CCD), sodium enters via the amiloride-sensitive epithelial sodium channel (ENaC) and exits via the basolaterally located Na,KATPase pump. The regulation of sodium reabsorption by aldosterone requires it to be bound to the mineralocorticoid receptor (MR), a member of the nuclear receptor superfamily (Arriza et al., 1987). These receptors share a common modular structure with three major functional domains. The N-terminal region contains a constitutive trans-activation function. The central DNA-binding domain consists of two zinc fingers that are involved in DNA binding and receptor dimerization. The ligand-binding domain (LBD) lies in the C-terminal region and is involved in several functions, including nuclear localization, interaction with the 90-kDa heat-shock protein, homo-and/or hetero-dimerization, and a ligand-dependent activation function (Evans, 1988;Tsai and O'Malley, 1994;Mangelsdorf et al., 1995;Ribeiro et al., 1995).The crystal structure of the MR-LBD has not ...

Nucleic acid aptamers are often referred to as chemical antibodies. Because they possess several advantages, like their smaller size, temperature stability, ease of chemical modification, lack of immunogenicity and toxicity, and lower cost of production, aptamers are promising tools for clinical applications. Aptamers against cell surface protein biomarkers are of particular interest for cancer diagnosis and targeted therapy. In this study, we identified and characterized RNA aptamers targeting cells expressing integrin α5β1. This αβ heterodimeric cell surface receptor is implicated in tumor angiogenesis and solid tumor aggressiveness. In glioblastoma, integrin α5β1 expression is associated with an aggressive phenotype and a decrease in patient survival. We used a complex and original hybrid SELEX (selective evolution of ligands by exponential enrichment) strategy combining protein-SELEX cycles on the recombinant α5β1 protein, surrounded by cell-SELEX cycles using two different cell lines. We identified aptamer H02, able to differentiate, in cyto- and histofluorescence assays, glioblastoma cell lines, and tissues from patient-derived tumor xenografts according to their α5 expression levels. Aptamer H02 is therefore an interesting tool for glioblastoma tumor characterization.

Sixteen cationic prodrugs of the antitumora lkylphospholipid (APL) erufosine were rationally synthesized to provideo riginal gene delivery reagents with improvedc ytotoxicityp rofile. The DNA complexationp roperties of these cationic lipids were determined and associated transfection rates were measured. Furthermore, the self-assembly properties of the pro-erufosine compounds were investigated and their criticala ggregation concentration was determined. Their hydrolytic stability underp Hc onditions mimickingt he extracellulare nvironment and the late endosome milieu was measured. Hemolytic activity and cytotoxicityo ft he compoundsw ere investigated. The results obtained in various cell lines demonstrate that the prodrugs of erufosined isplay antineoplastic activity similar to that of the parent antitumor drug but are not associated with hemolytic toxicity,w hich is ad ose-limiting side effect of APLs and am ajor obstacle to their use in anticancer therapeutic regimen. Furthermore, by using lipoplexes prepared from ap rodrug of erufosine and a plasmid DNAe ncoding ap ro-apoptotic protein (TRAIL), evidencew as provided for selective cytotoxicityt owards tumor cells while nontumorc ells were resistant. This study demonstratest hat the combination approachi nvolving well tolerated erufosine cationic prodrugs and cancerg ene therapy holds significant promiseint umor therapy.Supporting information and the ORCID identification number(s) for the author(s) of this articlecan be found under: https://doi.

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Nowadays, the need for therapeutic biomaterials displaying anti-inflammatory properties to fight against inflammation-related diseases is continuously increasing. Compact polyelectrolyte complexes (CoPECs) represent a new class of materials obtained by ultracentrifugation of a polyanion/polycation complex suspension in the presence of salt. Here, a noncytotoxic β-cyclodextrin-functionalized chitosan/alginate CoPEC was formulated, characterized, and described as a promising drug carrier displaying an intrinsic anti-inflammatory property. This new material was successfully formed, and due to the presence of cyclodextrins, it was able to trap and release hydrophobic drugs such as piroxicam used as a model drug. The intrinsic anti-inflammatory activity of this CoPEC was analyzed in vitro using murine macrophages in the presence of lipopolysaccharide (LPS) endotoxin. In this model, it was shown that CoPEC inhibited LPS-induced TNF-α and NO release and moderated the differentiation of LPS-activated macrophages. Over time, this kind of bioactive biomaterial could constitute a new family of delivery systems and expand the list of therapeutic tools available to target inflammatory chronic diseases such as arthritis or Crohn's disease.

Spirolactones harboring various C7 substituents are aldosterone antagonists, and some of them are used in the treatment of essential hypertension. They bind to the human mineralocorticoid receptor and render it transcriptionally inactive. Structural analysis using a three-dimensional homology model of the ligand-binding domain of the receptor has revealed that the Met852 residue of the ligand-binding cavity faces the C7 substituent of spirolactones. We therefore tested the binding capacities of C7-substituted spirolactones in an in vitro system expressing either the mutant receptor, in which Met852 was replaced by alanine, or the wild-type receptor. The M852A mutation had almost no effect on the binding of C7-substituted spirolactones to mineralocorticoid receptor but dramatically reduced the capacity of the receptor to bind steroids with no C7 substituent (aldosterone, cortisol, deoxycorticosterone, and canrenone). cis-trans Cotransfection assays revealed that two spirolactones characterized by having a propyl group [7␣-propyl-17␣-hydroxy-3-oxo-preg-4-ene-21-carboxylic acid ␥-lactone (RU26752)] or a thioacetyl group (spironolactone) at the C7 position acquired agonist properties when bound to the mutant receptor. In contrast, mexrenone and eplerenone, both of which harbor an acetyl group at the C7 position, retained antagonist properties when bound to the mutant receptor. Overall, these findings indicate that Met852 acts as an organizer residue that plays two major roles: 1) it allows steroids with no substituent at the C7 position to be accommodated within the ligand-binding cavity; and 2) it is involved in the steric hindrance that prevents C7-substituted spirolactones from folding the receptor in its active state.

DR4 (Death Receptor 4) and DR5 (Death Receptor 5) are two potential targets for cancer therapy due to their ability to trigger apoptosis of cancer cells, but not normal ones, when activated by their cognate ligand TRAIL (TNF related apoptosis-inducing ligand). Therapies based on soluble recombinant TRAIL or agonist antibodies directed against one of the receptors are currently under clinical trials. However, TRAIL-R positive tumor cells are frequently resistant to TRAIL induced apoptosis. The precise mechanisms of this resistance are still not entirely understood. We have previously reported on synthetic peptides that bind to DR5 (TRAILmim/DR5) and induce tumor cell apoptosis in vitro and in vivo. Here, we showed that while hexameric soluble TRAIL is able to efficiently kill the DR5 positive lymphoma Jurkat or the carcinoma HCT116, these cells are resistant to apoptosis induced by the divalent form of TRAILmim/DR5 and are poorly sensitive to apoptosis induced by an anti-DR5 agonist monoclonal antibody. This resistance can be restored by the cross-linking of anti-DR5 agonist antibody but not by the cross-linking of the divalent form of TRAILmim/DR5. Interestingly, the divalent form of TRAILmim/DR5 that induced apoptosis of DR5 positive BJAB cells, acts as an inhibitor of TRAIL-induced apoptosis on Jurkat and HCT116 cells. The rapid internalization of DR5 observed when treated with divalent form of TRAILmim/DR5 could explain the antagonist activity of the ligand on Jurkat and HCT116 cells but also highlights the independence of the mechanisms responsible for internalization and activation when triggering the DR5 apoptotic cascade.