The CAP1/Prss8 catalytic triad is not involved in PAR2 activation and protease nexin-1 (PN-1) inhibition.

Serine proteases, serine protease inhibitors, and protease-activated receptors (PARs) are responsible for several human skin disorders characterized by impaired epidermal permeability barrier function, desquamation, and inflammation. In this study, we addressed the consequences of a catalytically dead serine protease on epidermal homeostasis, the activation of PAR2 and the inhibition by the serine protease inhibitor nexin-1. The catalytically inactive serine protease CAP1/Prss8, when ectopically expressed in the mouse, retained the ability to induce skin disorders as well as its catalytically active counterpart (75%, n=81). Moreover, this phenotype was completely normalized in a PAR2-null background, indicating that the effects mediated by the catalytically inactive CAP1/Prss8 depend on PAR2 (95%, n=131). Finally, nexin-1 displayed analogous inhibitory capacity on both wild-type and inactive mutant CAP1/Prss8 in vitro and in vivo (64% n=151 vs. 89% n=109, respectively), indicating that the catalytic site of CAP1/Prss8 is dispensable for nexin-1 inhibition. Our results demonstrate a novel inhibitory interaction between CAP1/Prss8 and nexin-1, opening the search for specific CAP1/Prss8 antagonists that are independent of its catalytic activity.-Crisante, G., Battista, L., Iwaszkiewicz, J., Nesca, V., Mérillat, A.-M., Sergi, C., Zoete, V., Frateschi, S., Hummler, E. The CAP1/Prss8 catalytic triad is not involved in PAR2 activation and protease nexin-1 (PN-1) inhibition.

Inhibition of lymphocyte mediated cytotoxicity by perforin antisense oligonucleotides.

The granule/perforin exocytosis model of CTL mediated cytolysis proposes that CTL, upon recognition of the specific targets, release the cytolytic, pore-forming protein perforin into the intercellular space which then mediates the cytotoxic effect. However, direct evidence for the involvement of perforin is still lacking, and indeed, recent results even seem incompatible with the model. To determine directly the role of perforin in CTL cytotoxicity, perforin antisense oligonucleotides were exogenously added during the stimulation of mouse spleen derived T cells and human peripheral blood lymphocytes (PBL), respectively. Perforin protein expression in lymphocytes was reduced by up to 65%, and cytotoxicity of stimulated T cells by as much as 69% (5.7-fold). These results provide the first experimental evidence for a crucial role of perforin in lymphocyte mediated cytotoxicity.

Hemodynamic and biochemical consequences of renin inhibition by infusion of CGP 38560A in normal volunteers.

Hemodynamic and biochemical effects of the new renin inhibitor CGP 38560A (molecular weight 826) were tested in 15 healthy volunteers after a single-blind, randomized, placebo-controlled protocol. At a 2-week interval, groups of five subjects received a 30-minute infusion of either 5% dextrose or CGP 38560A 50, 125, or 250 micrograms/kg. Blood pressure, heart rate, plasma renin activity, active and total renin, angiotensin-(1-8)octapeptide (angiotensin II), and aldosterone were sequentially measured up to 3 hours from the onset of the infusion. There was no consistent change in blood pressure or heart rate. Plasma renin activity and angiotensin II decreased dose dependently, and peak suppression was observed at the end of the infusion of CGP 38560A and after the 250-micrograms/kg dose. Plasma renin activity fell from 1.0 +/- 0.19 (mean +/- SEM) to less than 0.05 ng/ml/hr in all five subjects (p less than 0.001), and angiotensin II fell from 7.7 +/- 1.2 to 2.6 +/- 0.9 femtomole/ml (p less than 0.01). Active renin rose fourfold from 24 +/- 1.9 to 98 +/- 14 pg/ml (p less than 0.001) at the end of the infusion of the high dose. Plasma angiotensin II returned toward its initial values much faster than plasma renin activity and active renin. In conclusion, CGP 38560A was well tolerated. It induced a dose-dependent decrease in angiotensin II and plasma renin activity and a long-lasting and dose-dependent rise in active renin. The doses used did not reduce plasma angiotensin II maximally despite reduction of plasma renin activity to unmeasurable levels.(ABSTRACT TRUNCATED AT 250 WORDS)

Characterization of the enzyme involved in the processing of big endothelin-1 in human lung epithelial cells.

Biosynthesis of active endothelin-1 (ET-1) implies an enzymatic processing of the inactive precursor Big ET-1 (1-39) into the mature, 21 amino acid peptide. The aim of this study was to characterize in airway and alveolar epithelial cells the enzymes responsible for this activation. BEAS-2B and A549 cells, which both produce ET-1, were studied in vitro as models for bronchiolar and alveolar cells, respectively. Both cell lines were able to convert exogenously added Big ET-1 (0.1 microM) into ET-1, suggesting a cell surface or an extracellular processing. The conversion was inhibited by phosphoramidon in both cell lines with an IC50 approximately 1 microM, but not by thiorphan, a specific inhibitor of neutral endopeptidase 24.11 (NEP). The endogenous production of serum-stimulated BEAS-2B and A549 cells was not inhibited by thiorphan, and phosphoramidon showed inhibition only at high concentration (>100 microM). Western blotting following electrophoresis in reducing conditions demonstrated a protein of MR 110 corresponding to the ECE-1 monomer in both BEAS-2B and A549 cells, as well as in whole lung extracts. By RT-PCR we revealed the mRNA encoding for the ECE-1b and/or -1c subtype, but not ECE-1a, in both cell lines. We conclude that BEAS-2B and A549 cells are able to process either endogenous or exogenous Big ET-1 by ECE-1 and that isoforms 1b and 1c could be involved in this processing with no significant role of NEP.

Study of a sialyltransferase ST3GAL6, in adipogenesis and obesity

RésuméL'obésité et les maladies métaboliques qui lui sont associées tels que le diabète ou les maladies cardiovasculaires ont un impact épidémiologique croissant. Ainsi, les mécanismes moléculaires se produisant dans le tissu adipeux en expansion font l'objet de nombreuses investigations. Dans ce contexte, nous nous sommes particulièrement intéressés à l'adipogénèse, le procédé permettant la formation d'adipocytes matures et fonctionnels. Le gène St3gal6 code pour une enzyme appelée β-galactosidase a2,3-sialyltransferase 6 et participant à la voie de glycosylation. Cette protéine appartient à la famille des a2,3- sialyltransferases dont la fonction principale est de transférer un acide sialique à l'extrémité de chaînes glycosidiques présentes sur les glycoprotéines et les glycolipides. Dans une précédente étude de transcriptomique réalisée chez la souris, St3gal6 a été décrit comme un gène dont l'expression est augmentée dans le tissu adipeux blanc d'animaux en surpoids et dont l'expression est normalisée après une perte de poids. Afin d'étudier le rôle potentiel de St3gal6 dans le développement du tissu adipeux, nous nous sommes intéressés à la régulation de son expression en cas d'obésité ainsi qu'à ses effets sur l'adipogénèse. Nous avons d'abord montré que St3gal6 s'exprime aussi bien dans le tissu adipeux blanc que dans le tissu adipeux brun. Puis nous avons confirmé dans deux différents modèles animaux que l'expression de St3gal6 dans le tissu adipeux était augmentée en cas d'obésité. Nous avons aussi observé in vitro une induction de St3gal6 dans des adipocytes traités par des cytokines pro-inflammatoires sécrétées dans le tissu adipeux d'individus obèses. Enfin, parmi les six membres que compte la famille des a2,3-sialyltransferases, St3gal6 est celui dont l'expression est la plus significativement induite en situation d'obésité. En outre, au cours de la différenciation des adipocytes blancs et bruns, l'expression de St3gal6 est augmentée et son inhibition réduit le potentiel de maturation des adipocytes qui accumulent moins de lipides. A l'inverse, la surexpression de St3gal6 dans des préadipocytes blancs augmente leur taux de différenciation in vitro; la formation de gouttelettes lipidiques et l'expression de genes spécifiques de l'adipocyte mature sont accrues. Enfin, le traitement d'adipocytes blancs in vitro avec un inhibiteur pharmacologique des a2,3-sialyltransferases ou une sialidase clivant les résidus sialylés montre qu'un défaut de a2,3-sialylation affectant les adipocytes diminue leur potentiel adipogénique. Par conséquent, ces résultats suggèrent que St3gal6 est impliqué dans la voie de différenciation des adipocytes et que cette a2,3-sialylation joue un rôle dans le remodelage du tissu adipeux induit par l'obésité.AbstractIn order to better understand molecular events occurring in obesity and leading to its associated complications, we were interested in the biology of adipose tissue and particularly in the study of adipogenesis, the process by which new mature adipocytes develop and accumulate lipids.The β-galactosidase a2,3-sialyltransferase 6 (St3gal6) gene encodes for an enzyme involved in post-translational protein glycosylation. Thereby, St3gal6 enzyme belongs to the a2,3sialyltransferase family whose function is to add sialic acids at outer position on glycosidic chain of glycoproteins or glycolipids. Previously, in mouse, St3gal6 has been described as a gene whose expression in white adipose tissue is increased in overweighted animals and normalized after weight loss. Therefore, we have assumed that St3gal6 may play a role in adipose tissue development and in tissue remodelling triggered by obesity. First we show that St3gal6 is expressed in white but also in brown adipose tissue. St3gal6 upregulation upon weight gain was confirmed in two mouse models of obesity namely diet- induced and genetically-induced obesity. We also report that St3gal6 is induced by pro¬inflammatory cytokines known to be oversecreted in adipose tissue during obesity. Furthermore, St3gal6 is the a2,3-sialyltransferase whose expression is more markedly induced in adipose tissue. In addition, we demonstrate that St3gl6 expression is progressively increased in late stages of white and brown adipogenesis while St3gal6 knockdown inhibits adipocyte differentiation in vitro. Conversely, St3gal6 overexpression in a white preadipocyte cell line increases lipid accumulation during differentiation process and enhances gene expression of mature white adipocyte markers. Finally, using an a2-3 sialyltransferase inhibitor and a sialidase treatment on white adipocyte cell line, we observe that a decreased a2,3-sialylation impairs adipocyte differentiation in vitro. Altogether, these result suggest that St3gal6 plays a role in adipogenesis and in tissue remodelling associated with obesity likely through its enzymatic activity of a2,3-sialylation. Thus, a2,3-sialylation appears as a novel pathway of interest whose precise molecular mechanisms remain to be elucidated in the context of adipose tissue development and adipocyte functions.

Cis association of Ly49A with MHC class I restricts natural killer cell inhibition.

Natural killer (NK) cell function is negatively regulated by inhibitory receptors interacting with major histocompatibility complex class I molecules expressed on target cells. Here we show that the inhibitory Ly49A NK cell receptor not only binds to its H-2D(d) ligand expressed on potential target cells (in trans) but also is constitutively associated with H-2D(d) in cis (on the same cell). Cis association and trans interaction occur through the same binding site. Consequently, cis association restricts the number of Ly49A receptors available for binding of H-2D(d) on target cells and reduces NK cell inhibition through Ly49A. By lowering the threshold at which NK cell activation exceeds NK cell inhibition, cis interaction allows optimal discrimination of normal and abnormal host cells.

The inhibition of MAPK potentiates the anti-angiogenic efficacy of mTOR inhibitors.

The mammalian target of rapamycin (mTOR) which is part of two functionally distinct complexes, mTORC1 and mTORC2, plays an important role in vascular endothelial cells. Indeed, the inhibition of mTOR with an allosteric inhibitor such as rapamycin reduces the growth of endothelial cell in vitro and inhibits angiogenesis in vivo. Recent studies have shown that blocking mTOR results in the activation of other prosurvival signals such as Akt or MAPK which counteract the growth inhibitory properties of mTOR inhibitors. However, little is known about the interactions between mTOR and MAPK in endothelial cells and their relevance to angiogenesis. Here we found that blocking mTOR with ATP-competitive inhibitors of mTOR or with rapamycin induced the activation of the mitogen-activated protein kinase (MAPK) in endothelial cells. Downregulation of mTORC1 but not mTORC2 had similar effects showing that the inhibition of mTORC1 is responsible for the activation of MAPK. Treatment of endothelial cells with mTOR inhibitors in combination with MAPK inhibitors reduced endothelial cell survival, proliferation, migration and tube formation more significantly than either inhibition alone. Similarly, in a tumor xenograft model, the anti-angiogenic efficacy of mTOR inhibitors was enhanced by the pharmacological blockade of MAPK. Taken together these results show that blocking mTORC1 in endothelial cells activates MAPK and that a combined inhibition of MAPK and mTOR has additive anti-angiogenic effects. They also provide a rationale to target both mTOR and MAPK simultaneously in anti-angiogenic treatment.

Neutral endopeptidase versus angiotensin converting enzyme inhibition in essential hypertension.

OBJECTIVE: To evaluate the antihypertensive efficacy of sinorphan, an orally active inhibitor of neutral endopeptidase EC 3.4.24.11. DESIGN: The ability of sinorphan (100 mg twice a day) to lower blood pressure was compared with that of the angiotensin converting enzyme (ACE) inhibitor captopril (25 mg twice a day) using a randomized-sequence, double-blind crossover design in 16 patients with essential hypertension. Each treatment was administered for 4 weeks and treatments were separated by a 3-week placebo period. At the end of the last phase of treatment sinorphan was combined with captopril for a further 4-week period. The changes in systolic (SBP) and diastolic blood pressure (DBP) were monitored using repeated ambulatory blood pressure monitoring. RESULTS: When given as monotherapy for 4 weeks, neither sinorphan nor captopril significantly reduced the 24-h or the 14-h daytime mean SBP or DBP. However, a significant decrease in DBP was observed during the first 6 h after the morning administration of captopril. With sinorphan only a significant decrease in night-time SBP was found. With the combined therapy of sinorphan and captopril, significant decreases both in SBP and in DBP were observed, which were sustained over 24 h. After 4 weeks of sinorphan alone or in combination with captopril, no change in plasma atrial natriuretic peptide level was found. However, urinary cyclic GMP excretion increased transiently after administration of the neutral endopeptidase inhibitor. CONCLUSIONS: Neutral endopeptidase inhibition with sinorphan has a limited effect on blood pressure in hypertensive patients when given alone. However, simultaneous neutral endopeptidase and ACE inhibition induces a synergistic effect, and might therefore represent an interesting new therapeutic approach to the treatment of essential hypertension.

ENaC inhibition stimulates Cl- secretion in the mouse cortical collecting duct through an NKCC1-dependent mechanism.

In cortical collecting ducts (CCDs) perfused in vitro, inhibiting the epithelial Na(+) channel (ENaC) reduces Cl(-) absorption. Since ENaC does not transport Cl(-), the purpose of this study was to determine how ENaC modulates Cl(-) absorption. Thus, Cl(-) absorption was measured in CCDs perfused in vitro that were taken from mice given aldosterone for 7 days. In wild-type mice, we observed no effect of luminal hydrochlorothiazide on either Cl(-) absorption or transepithelial voltage (V(T)). However, application of an ENaC inhibitor [benzamil (3 μM)] to the luminal fluid or application of a Na(+)-K(+)-ATPase inhibitor to the bath reduced Cl(-) absorption by ∼66-75% and nearly obliterated lumen-negative V(T). In contrast, ENaC inhibition had no effect in CCDs from collecting duct-specific ENaC-null mice (Hoxb7:CRE, Scnn1a(loxlox)). Whereas benzamil-sensitive Cl(-) absorption did not depend on CFTR, application of a Na(+)-K(+)-2Cl(-) cotransport inhibitor (bumetanide) to the bath or ablation of the gene encoding Na(+)-K(+)-2Cl(-) cotransporter 1 (NKCC1) blunted benzamil-sensitive Cl(-) absorption, although the benzamil-sensitive component of V(T) was unaffected. In conclusion, first, in CCDs from aldosterone-treated mice, most Cl(-) absorption is benzamil sensitive, whereas thiazide-sensitive Cl(-) absorption is undetectable. Second, benzamil-sensitive Cl(-) absorption occurs by inhibition of ENaC, possibly due to elimination of lumen-negative V(T). Finally, benzamil-sensitive Cl(-) flux occurs, at least in part, through transcellular transport through a pathway that depends on NKCC1.

Acetylcholine-induced vasodilation and reactive hyperemia are not affected by acute cyclo-oxygenase inhibition in human skin

Résumé Il est actuellement reconnu que l'endothélium vasculaire joue un rôle primordial dans la genèse des maladies cardiovasculaires, notamment l'artériosclérose. Dès lors, il est important de pouvoir investiguer la fonction endothéliale en clinique. Pour ce faire, il est particulièrement simple d'examiner la microcirculation cutanée, car celle-ci est très simplement accessible, de manière non-invasive, par fluxmétrie laser-Doppler. Pratiquement, on mesure l'augmentation du flux sanguin dermique en réponse à des stimuli connus pour agir via l'endothélium vasculaire. Les stimuli endothélium-dépendants les plus courants sont l'interruption temporaire du flux sanguin qui est suivie d'une hyperémie réactive, et l'administration transcutanée d'acétylcholine (Ach) par iontophorèse. La iontophorèse consiste à obtenir le transfert d' une substance ionisée, telle l'Ach, par l'application d'un courant électrique de polarité appropriée. L'objectif du présent travail était de déterminer le rôle des prostaglandines dans ces réponse vasodilatatrices dépendante de l'endothélium, rôle actuellement peu clair. 23 jeunes hommes volontaires non fumeurs et en bonne santé habituelle ont été examinés lors de deux visites séparées par 1 à 3 semaines. Lors de chaque visite, l'hyperémie réactive et la réponse vasodilatatrice à l'Ach ont été déterminées dans la peau de l'avant bras après administration soit d'un placebo, soit d'un inhibiteur de la cyclooxygénase (COX, enzyme qui contrôle la synthèse des prostaglandines). Chez certains sujets, l'inhibiteur était de l'acétylsalicylate de lysine (900 mg par voie intraveineuse). Chez d'autres sujets, il s'agissait d'indométhacine. (75 mg par voie orale). Comme la stimulation nociceptive liée au courant iontophorétique peut influencer la réponse à l'Ach, celle-ci a été déterminée en présence et en l'absence d'anesthésie de surface (crème de lidocaine). La réponse à l'Ach a été obtenue pour 4 doses différentes de cet agent (exprimées sous la forme de la densité de charge iontophorétique appliquée : 0.28, 1.4, 7, et 14 millicoulombs par cm2 de peau exposée). Le flux sanguin dermique était mesuré par imagerie laser-Doppler, une variante de la fluxmétrie laser-Doppler classique permettant l'exploration d'une surface de peau de taille arbitraire. Quelle que soit la condition testée, nous n'avons jamais observé la moindre influence de l'inhibition de la COX sur l'hyperémie réactive, ni sur la réponse à l'Ach. Cette dernière était augmentée significativement par l'anesthésie cutanée, que les sujets aient reçu ou non de l'acétylsalicylate de lysine ou de l'indométhacine . Par exemple, la réponses moyenne (±SD) à la plus haute dose d'Ach (testée sur 6 sujets, et exprimée en unités de perfusion, comme il est d'usage en fluxmétrie laser-Doppler ) était la suivante : en l'absence d'anesthésie : acétylsalicylate de lysine 339 ± 105, placebo 344 ± 68 ; avec l'anesthésie : acétylsalicylate de lysine 453 ± 76 , placebo 452 ± 65 (p * 0.001 pour les effets de l'anesthésie). En conclusion, nos résultats infirment une contribution des prostaglandines à l'hyperémie réactive ou à la vasodilatation induite par l'acétylcholine dans la microcirculation cutanée. Dans ce lit vasculaire, l'anesthésie locale accroît la vasodilatation induite par l'acétylcholine par un mécanisme indépendant des prostaglandines.

Angiotensin-converting enzyme inhibition improves vascular function in rheumatoid arthritis.

BACKGROUND: The excess in cardiovascular risk in patients with rheumatoid arthritis provides a strong rationale for early therapeutical interventions. In view of the similarities between atherosclerosis and rheumatoid arthritis and the proven benefit of angiotensin-converting enzyme inhibitors in atherosclerotic vascular disease, it was the aim of the present study to delineate the impact of ramipril on endothelial function as well as on markers of inflammation and oxidative stress in patients with rheumatoid arthritis. METHODS AND RESULTS: Eleven patients with rheumatoid arthritis were included in this randomized, double-blind, crossover study to receive ramipril in an uptitration design (2.5 to 10 mg) for 8 weeks followed by placebo, or vice versa, on top of standard antiinflammatory therapy. Endothelial function assessed by flow-mediated dilation of the brachial artery, markers of inflammation and oxidative stress, and disease activity were investigated at baseline and after each treatment period. Endothelial function assessed by flow-mediated dilation increased from 2.85+/-1.49% to 4.00+/-1.81% (P=0.017) after 8 weeks of therapy with ramipril but did not change with placebo (from 2.85+/-1.49% to 2.84+/-2.47%; P=0.88). Although systolic blood pressure and heart rate remained unaltered, diastolic blood pressure decreased slightly from 78+/-7 to 74+/-6 mm Hg (P=0.03). Tumor necrosis factor-alpha showed a significant inverse correlation with flow-mediated dilation (r=-0.408, P=0.02), and CD40 significantly decreased after ramipril therapy (P=0.049). CONCLUSIONS: Angiotensin-converting enzyme inhibition with 10 mg/d ramipril for 8 weeks on top of current antiinflammatory treatment markedly improved endothelial function in patients with rheumatoid arthritis. This finding suggests that angiotensin-converting enzyme inhibition may provide a novel strategy to prevent cardiovascular events in these patients.

Ly49E-dependent inhibition of natural killer cells by urokinase plasminogen activator.

The Ly49 natural killer (NK)-cell receptor family comprises both activating and inhibitory members, which recognize major histocompatibility complex (MHC) class I or MHC class I-related molecules and are involved in target recognition. As previously shown, the Ly49E receptor fails to bind to a variety of soluble or cell-bound MHC class I molecules, indicating that its ligand is not an MHC class I molecule. Using BWZ.36 reporter cells, we demonstrate triggering of Ly49E by the completely distinct, non-MHC-related protein urokinase plasminogen activator (uPA). uPA is known to be secreted by a variety of cells, including epithelial and hematopoietic cells, and levels are up-regulated during tissue remodeling, infections, and tumorigenesis. Here we show that addition of uPA to Ly49E-positive adult and fetal NK cells inhibits interferon-gamma secretion and reduces their cytotoxic potential, respectively. These uPA-mediated effects are Ly49E-dependent, as they are reversed by addition of anti-Ly49E monoclonal antibody and by down-regulation of Ly49E expression using RNA interference. Our results suggest that uPA, besides its established role in fibrinolysis, tissue remodeling, and tumor metastasis, could be involved in NK cell-mediated immune surveillance and tumor escape.

Inhibition of Mitogen-Activated Protein Kinase Erk1/2 Promotes Protein Degradation of ATP Binding Cassette Transporters A1 and G1 in CHO and in HuH7 cells.

Signal transduction modulates expression and activity of cholesterol transporters. We recently demonstrated that the Ras/mitogen-activated protein kinase (MAPK) signaling cascade regulates protein stability of Scavenger Receptor BI (SR-BI) through Proliferator Activator Receptor (PPARα) -dependent degradation pathways. In addition, MAPK (Mek/Erk 1/2) inhibition has been shown to influence liver X receptor (LXR) -inducible ATP Binding Cassette (ABC) transporter ABCA1 expression in macrophages. Here we investigated if Ras/MAPK signaling could alter expression and activity of ABCA1 and ABCG1 in steroidogenic and hepatic cell lines. We demonstrate that in Chinese Hamster Ovary (CHO) cells and human hepatic HuH7 cells, extracellular signal-regulated kinase 1/2 (Erk1/2) inhibition reduces PPARα-inducible ABCA1 protein levels, while ectopic expression of constitutively active H-Ras, K-Ras and MAPK/Erk kinase 1 (Mek1) increases ABCA1 protein expression, respectively. Furthermore, Mek1/2 inhibitors reduce ABCG1 protein levels in ABCG1 overexpressing CHO cells (CHO-ABCG1) and human embryonic kidney 293 (HEK293) cells treated with LXR agonist. This correlates with Mek1/2 inhibition reducing ABCG1 cell surface expression and decreasing cholesterol efflux onto High Density Lipoproteins (HDL). Real Time reverse transcriptase polymerase chain reaction (RT-PCR) and protein turnover studies reveal that Mek1/2 inhibitors do not target transcriptional regulation of ABCA1 and ABCG1, but promote ABCA1 and ABCG1 protein degradation in HuH7 and CHO cells, respectively. In line with published data from mouse macrophages, blocking Mek1/2 activity upregulates ABCA1 and ABCG1 protein levels in human THP1 macrophages, indicating opposite roles for the Ras/MAPK pathway in the regulation of ABC transporter activity in macrophages compared to steroidogenic and hepatic cell types. In summary, this study suggests that Ras/MAPK signaling modulates PPARα- and LXR-dependent protein degradation pathways in a cell-specific manner to regulate the expression levels of ABCA1 and ABCG1 transporters.

Rational design of indoleamine 2,3-dioxygenase inhibitors.

Indoleamine 2,3-dioxygenase (IDO) is an important therapeutic target for the treatment of diseases such as cancer that involve pathological immune escape. We have used the evolutionary docking algorithm EADock to design new inhibitors of this enzyme. First, we investigated the modes of binding of all known IDO inhibitors. On the basis of the observed docked conformations, we developed a pharmacophore model, which was then used to devise new compounds to be tested for IDO inhibition. We also used a fragment-based approach to design and to optimize small organic molecule inhibitors. Both approaches yielded several new low-molecular weight inhibitor scaffolds, the most active being of nanomolar potency in an enzymatic assay. Cellular assays confirmed the potential biological relevance of four different scaffolds.