Nouveaux inducteurs covalents de la voie de signalisation Keap1/Nrf2/ARE

Le stress électrophile et oxydant est un souci grandissant pour la santé avec l'évolution de nos modes de vie. L'exposition aux ultraviolets, à la pollution, aux substances carcinogènes, à la fumée de cigarette et la pratique intensive d'activités sportives sont autant de causes de stress oxydant pour l'organisme. Ces dommages sont associés à plusieurs maladies et conditions pathologiques telles que cancers, diabètes, infections pulmonaires et maladies neurodégénératives. L'élément de réponse antioxydant (ARE) est un des composants principaux des défenses de la cellule contre ce phénomène. Ce promoteur agit sous le contrôle de Nrf2 (Nuclear factor erythroid 2-related factor 2). Une stratégie populaire pour l'activation de ce mécanisme est l'utilisation d'inducteurs covalents. Ces molécules agissent par la formation de liens covalents avec les nombreux résidus cystéine de Keap1 (Kelch-like ECH-associated protein 1), une protéine chaperonne qui contrôle l'activité de Nrf2. Cette thèse présente la synthèse, les propriétés biologiques et l'étude des relations structure-activité d'une librairie d'électrophiles capables d'induire la transcription des gènes cibles de la voie de signalisation Keap1/Nrf2/ARE. Le premier volet fait état de la comparaison d'une variété d'électrophiles simples pour étudier les préférences de la cible. Le deuxième volet montre que la présence d'une seconde fonction capable de piéger un résidu cystéine fournit des analogues très puissants.

L’ischémie des membres inférieurs chez les diabétiques : le rôle du récepteur AT2

Résumé : Les patients diabétiques ont plus de risques d’être amputés d’une jambe en raison d’une plus faible néovascularisation suite à une ischémie. Nous avons montré une association entre une plus faible réponse angiogénique du VEGF chez les souris diabétiques (DM) et une augmentation de l’expression de SHP-1, pouvant être activée par les récepteurs (AT[indice inférieur 1]/AT[indice inférieur 2]). La délétion du récepteur AT[indice inférieur 2] chez des souris favorise l’angiogenèse dans le muscle ischémique, mais son rôle en condition diabétique demeure inconnu. Notre objectif est de vérifier si la délétion du récepteur AT[indice inférieur 2] chez des souris DM favorise l’angiogenèse suivant l’induction d’une ischémie. Des souris DM de type 1 déficientes (KO) ou non pour le récepteur AT[indice inférieur 2] ont été utilisées. L’ischémie a été induite par la ligature de l'artère fémorale. La perfusion sanguine a été mesurée pendant 2 ou 4 semaines avant la récolte des tissus. Les effets de l’ischémie sur l’expression des récepteurs AT[indice inférieur 1] et AT[indice inférieur 2], des phosphatases SHP-1, SHP-2 et PTP1B, ainsi que l’état de la voie de signalisation du VEGF ont été mesurés. Un essai phosphatase a aussi été effectué suite à l’immunoprécipitation de SHP-1 chez des BAECs stimulés au CGP42112A. Quatre semaines après la chirurgie, le flot sanguin dans le muscle ischémique des souris DM AT[indice inférieur 2]KO s’est rétabli plus rapidement (80%) comparativement à une récupération de 47% chez les souris DM contrôles. L’expression des facteurs pro-angiogéniques (HIF-1α et VEGF) était similaire dans tous les groupes après 2 semaines d’ischémie, mais diminuée chez les DM et retournait à un niveau basal chez les DM-AT[indice inférieur 2]KO après 4 semaines, suggérant un reperfusion plus rapide chez ces souris. La phosphorylation de Akt était aussi plus faible chez les souris DM contrôles mais était rétablie chez les souris AT[indice inférieur 2]KO après 4 semaines d’ischémie. L'expression de SHP-1 était doublée dans le muscle ischémique des souris DM, en comparaison aux souris non DM, un effet absent chez les souris DM AT[indice inférieur 2]KO. L’expression de SHP-2 et PTP1B ne variait pas chez les souris DM sauvages et AT[indice inférieur 2]KO. De plus, l’expression des récepteurs AT[indice inférieur 1] et AT[indice inférieur 2] est augmentée chez les souris DM sauvages en comparaison aux souris NDM. La stimulation du récepteur AT[indice inférieur 2] chez les BAECs a permis d’augmenter l’activité phosphatase de SHP-1. Nos résultats suggèrent que l’expression élevée d’AT[indice inférieur 2] chez les souris DM mène à la surexpression et/ou l’activation de SHP-1, inhibant le signal angiogénique issu du VEGF et empêchant la reperfusion sanguine suite à l’ischémie.

Identificazione di un nuovo meccanismo microRNA-dipendente di resistenza all'inibizione della via di segnale PI3K/AKT nel carcinoma della prostata

Benché le alterazioni della via PI3K/AKT siano molto sudiate a causa del loro ruolo nella tumorigenesi, e rappresentino pertanto un importante bersaglio terapeutico, i risultati di numerosi studi clinici con inibitori di PI3K o AKT sono finora deludenti, in parte a causa dell’insorgenza di resistenza provocata dall'interruzione dei circuiti di feedback negativo. In questo studio, abbiamo scoperto che l’inattivazione farmacologica di AKT in cellule di carcinoma prostatico PC3 porta alla down-regolazione di un microRNA con funzione di oncosoppressore, il miR-145-5p, e ad un drammatico aumento di espressione di uno dei suoi geni target, cioè N/KRas. E’ interessante sottolineare che questo microRNA è considerato un marker di progressione metastatica nel carcinoma prostatico, il cui livello di espressione aiuta a discriminare tra pazienti con iperplasia prostatica benigna e cancro alla prostata. Inoltre, la bassa espressione di miR-145 aumenta il rischio di progressione della malattia da localizzata a metastatica. La conferma che l’aumento di Ras, osservato sia in termini di mRNA che di proteina, è dipendente dalla caduta del miR-145-5p, è stata poi ottenuta tramite un modello di PC3 ingegnerizzate per ottenere il silenziamento inducibile del miR-145-5p. Tramite un array di fosfoproteine siamo poi stati in grado di verificare che l’aumento di Ras provoca la riattivazione della cascata di PI3K/AKT e di ERK. Dal punto di vista meccanicistico, quindi, lo studio ha portato all’identificazione di un nuovo meccanismo di resistenza adattativa, in cui l’inattivazione di AKT provoca una caduta del miR-145-5p che, a sua volta, aumenta l’espressione di Ras e riattiva il signaling di PI3K, rendendo inefficace il trattamento farmacologico. Questi risultati sono particolarmente rilevanti alla luce di recenti studi (NCT04493853; NCT03072238; NCT02525068) e di trial clinici in corso (NCT04737109; NCT03673787), basati sulla somministrazione combinata di inibitori della sintesi degli androgeni con gli inibitori di AKT capitasertib o ipatasertib.

Ikkε Is Key To Induction Of Insulin Resistance In The Hypothalamus, And Its Inhibition Reverses Obesity.

IKK epsilon (IKKε) is induced by the activation of nuclear factor-κB (NF-κB). Whole-body IKKε knockout mice on a high-fat diet (HFD) were protected from insulin resistance and showed altered energy balance. We demonstrate that IKKε is expressed in neurons and is upregulated in the hypothalamus of obese mice, contributing to insulin and leptin resistance. Blocking IKKε in the hypothalamus of obese mice with CAYMAN10576 or small interfering RNA decreased NF-κB activation in this tissue, relieving the inflammatory environment. Inhibition of IKKε activity, but not TBK1, reduced IRS-1(Ser307) phosphorylation and insulin and leptin resistance by an improvement of the IR/IRS-1/Akt and JAK2/STAT3 pathways in the hypothalamus. These improvements were independent of body weight and food intake. Increased insulin and leptin action/signaling in the hypothalamus may contribute to a decrease in adiposity and hypophagia and an enhancement of energy expenditure accompanied by lower NPY and increased POMC mRNA levels. Improvement of hypothalamic insulin action decreases fasting glycemia, glycemia after pyruvate injection, and PEPCK protein expression in the liver of HFD-fed and db/db mice, suggesting a reduction in hepatic glucose production. We suggest that IKKε may be a key inflammatory mediator in the hypothalamus of obese mice, and its hypothalamic inhibition improves energy and glucose metabolism.

Identification Of Target Genes Using Gene Expression Profile Of Granulocytes From Patients With Chronic Myeloid Leukemia Treated With Tyrosine Kinase Inhibitors.

Differential gene expression analysis by suppression subtractive hybridization with correlation to the metabolic pathways involved in chronic myeloid leukemia (CML) may provide a new insight into the pathogenesis of CML. Among the overexpressed genes found in CML at diagnosis are SEPT5, RUNX1, MIER1, KPNA6 and FLT3, while PAN3, TOB1 and ITCH were decreased when compared to healthy volunteers. Some genes were identified and involved in CML for the first time, including TOB1, which showed a low expression in patients with CML during tyrosine kinase inhibitor treatment with no complete cytogenetic response. In agreement, reduced expression of TOB1 was also observed in resistant patients with CML compared to responsive patients. This might be related to the deregulation of apoptosis and the signaling pathway leading to resistance. Most of the identified genes were related to the regulation of nuclear factor κB (NF-κB), AKT, interferon and interleukin-4 (IL-4) in healthy cells. The results of this study combined with literature data show specific gene pathways that might be explored as markers to assess the evolution and prognosis of CML as well as identify new therapeutic targets.

Acute Exercise Decreases Trb3 Protein Levels In The Hypothalamus Of Obese Rats.

To evaluate the effects of acute exercise on the TRB3 protein levels and interaction between TRB3/Akt proteins in the hypothalamus of obese rats. In addition, we evaluated the relationship between TRB3 and endoplasmic reticulum stress (ER stress) and verified whether an acute exercise session is able to influence these processes. In the first part of the study, the rats were divided into three groups: control (lean) - fed with a standard rodent chow, DIO - fed with a high fat diet and DIO submitted to a swimming acute exercise protocol (DIO-EXE). In the second part of the study, we used other three groups: control (lean) receiving an intracerebroventricular (i.c.v.) infusion of vehicle, lean receiving an i.c.v. infusion of thapsigargin, and lean receiving an i.c.v infusion of thapsigargin and performing an acute exercise session. Four hours after the exercise session, the food intake was measured and the hypothalamus was dissected and separated for subsequent protein analysis by immunoblotting and Real Time PCR. The acute exercise session reduced the TRB3 protein levels, disrupted the interaction between TRB3/Akt proteins, increased the phosphorylation of Foxo1 and restored the anorexigenic effects of insulin in the hypothalamus of DIO rats. Interestingly, the suppressive effects of acute exercise on TRB3 protein levels may be related, at least in part, to the decrease of ER stress (evaluated though pancreatic ER kinase phosphorylation - pPERK and C/EBP homologous protein - CHOP protein levels) in the hypothalamus. In conclusion, the reduction of hypothalamic TRB3 protein levels mediated by exercise may be associated with the reduction of ER stress. These data provided a new mechanism by which an acute exercise session improves insulin sensitivity in hypothalamus and restores food intake control in obesity.

Effect Of Atorvastatin On Wound Healing In Rats.

Skin-wound healing is a complex and dynamic biological process involving inflammation, proliferation, and remodeling. Recent studies have shown that statins are new therapeutical options because of their actions, such as anti-inflammatory and antioxidant activity, on vasodilation, endothelial dysfunction and neoangiogenesis, which are independent of their lipid-lowering action. Our aim was to investigate the effect of atorvastatin on tissue repair after acute injury in healthy animals. Rats were divided into four groups: placebo-treated (P), topical atorvastatin-treated (AT), oral atorvastatin-treated (AO), topical and oral atorvastatin-treated (ATO). Under anesthesia, rats were wounded with an 8-mm punch in the dorsal region. Lesions were photographed on Days 0, 1, 3, 7, 10, 12, and 14 post-injury and samples taken on Days 1, 3, 7, and 14 for protein-expression analysis of insulin receptor substrate (IRS)-1, phosphatidylinositol 3-kinase (PI3K), protein kinase B (Akt), glycogen synthase kinase (GSK)-3, endothelial nitric oxide synthase (eNOS), vascular endothelial growth factor (VEGF), extracellular signal-regulated kinase (ERK), interleukin (IL)-10, IL-1β, IL-6, and tumor necrosis factor (TNF)-α. Upon macroscopic examination, we observed significant reductions of lesion areas in groups AT, AO, and ATO compared to the P group. Additionally, AT and AO groups showed increased expression of IRS-1, PI3K, Akt, GSK-3, and IL-10 on Days 1 and 3 when compared with the P group. All atorvastatin-treated groups showed higher expression of IRS-1, PI3K, Akt, GSK-3, IL-10, eNOS, VEGF, and ERK on Day 7. On Days 1, 3, and 7, all atorvastatin-treated groups showed lower expression of IL-6 and TNF-α when compared with the P group. We conclude that atorvastatin accelerated tissue repair of acute lesions in rats and modulated expressions of proteins and cytokines associated with cell-growth pathways.

Leucine Supplementation Does Not Affect Protein Turnover And Impairs The Beneficial Effects Of Endurance Training On Glucose Homeostasis In Healthy Mice.

Endurance exercise training as well as leucine supplementation modulates glucose homeostasis and protein turnover in mammals. Here, we analyze whether leucine supplementation alters the effects of endurance exercise on these parameters in healthy mice. Mice were distributed into sedentary (C) and exercise (T) groups. The exercise group performed a 12-week swimming protocol. Half of the C and T mice, designated as the CL and TL groups, were supplemented with leucine (1.5 % dissolved in the drinking water) throughout the experiment. As well known, endurance exercise training reduced body weight and the retroperitoneal fat pad, increased soleus mass, increased VO2max, decreased muscle proteolysis, and ameliorated peripheral insulin sensitivity. Leucine supplementation had no effect on any of these parameters and worsened glucose tolerance in both CL and TL mice. In the soleus muscle of the T group, AS-160(Thr-642) (AKT substrate of 160 kDa) and AMPK(Thr-172) (AMP-Activated Protein Kinase) phosphorylation was increased by exercise in both basal and insulin-stimulated conditions, but it was reduced in TL mice with insulin stimulation compared with the T group. Akt phosphorylation was not affected by exercise but was lower in the CL group compared with the other groups. Leucine supplementation increased mTOR phosphorylation at basal conditions, whereas exercise reduced it in the presence of insulin, despite no alterations in protein synthesis. In trained groups, the total FoxO3a protein content and the mRNA for the specific isoforms E2 and E3 ligases were reduced. In conclusion, leucine supplementation did not potentiate the effects of endurance training on protein turnover, and it also reduced its positive effects on glucose homeostasis.

Mechanisms Of Insulin Resistance In The Amygdala: Influences On Food Intake.

Obesity is increasing worldwide and is triggered, at least in part, by enhanced caloric intake. Food intake is regulated by a complex mechanism involving the hypothalamus and hindbrain circuitries. However, evidences have showing that reward systems are also important in regulating feeding behavior. In this context, amygdala is considered a key extra-hypothalamic area regulating feeding behavior in human beings and rodents. This review focuses on the regulation of food intake by amygdala and the mechanisms of insulin resistance in this brain area. Similar to the hypothalamus the anorexigenic effect of insulin is mediated via PI3K (phosphoinositide 3-kinase)/Akt (protein kinase B) pathway in the amygdala. Insulin decreases NPY (neuropeptide Y) and increases oxytocin mRNA levels in the amygdala. High fat diet and saturated fatty acids induce inflammation, ER (endoplasmic reticulum) stress and the activation of serine kinases such as PKCθ (protein kinase C theta), JNK (c-Jun N-terminal kinase) and IKKβ (inhibitor of nuclear factor kappa-B kinase beta) in the amygdala, which have an important role in insulin resistance in this brain region. Overexpressed PKCθ in the CeA (central nucleus of amygdala) of rats increases weight gain, food intake, insulin resistance and hepatic triglycerides content. The inhibition of ER stress ameliorates insulin action/signaling, increases oxytocin and decreases NPY gene expression in the amygdala of high fat feeding rodents. Those data suggest that PKCθ and ER stress are main mechanisms of insulin resistance in the amygdala of obese rats and play an important role regulating feeding behavior.

Intravesical Bacillus Calmette-guérin Efficiently Reduces P70s6k1 But Not 4e-bp1 Phosphorylation In Nonmuscle Invasive Bladder Cancer.

We characterized the functional consequences of intravesical bacillus Calmette-Guérin on the molecular mechanism of the AKT/mTOR signaling pathway in nonmuscle invasive bladder cancer. To our knowledge this has not been reported previously. At age 7 weeks female Fischer 344 rats received 1.5 mg/kg MNU intravesically every other week for 6 weeks. They were randomized at 10 per group to MNU (0.2 ml vehicle), bacillus Calmette-Guérin (10(6) cfu Connaught strain), rapamycin (15 μg/ml) and bacillus Calmette-Guérin plus simultaneous rapamycin, each intravesically for 6 weeks. At week 15 the bladders were collected for histopathology, immunohistochemistry and immunoblot to determine p-AKT, Rictor, Raptor, p-4E-BP1, p-p70S6K1, p-AMPK-α, p-mTOR and p-p53. Papillary carcinoma (pTa) and high grade intraepithelial neoplasia (pTis) predominated in the MNU group while normal urothelium, papillary and flat hyperplasia were more common in treated groups. Nonmuscle invasive bladder cancer treated with bacillus Calmette-Guérin showed suppression of p70S6K1 but not 4E-BP1 phosphorylation. This suggests that 4E-BP1 is regulated differently than p70S6K1, escaping the bacillus Calmette-Guérin action that occurs in a mTOR independent manner. The association of bacillus Calmette-Guérin with rapamycin but not rapamycin monotherapy affected p70S6K1 and 4E-BP1 phosphorylation with no features of in situ carcinoma (pTis). The activation status of p70S6K1 and 4E-BP1 might be used to stratify patients who could benefit from targeting such molecular elements with multitarget/multidrug intravesical therapy. In the future 4E-BP1 might be a worthwhile new target for bacillus Calmette-Guérin refractory nonmuscle invasive bladder cancer.

Jnk And Ikkβ Phosphorylation Is Reduced By Glucocorticoids In Adipose Tissue From Insulin-resistant Rats.

Peripheral insulin resistance (IR) is one of the main side effects caused by glucocorticoid (GC)-based therapies, and the molecular mechanisms of GC-induced IR are not yet fully elucidated. Thus, we aimed to investigate the effects of dexamethasone treatment on the main components of insulin and inflammatory signaling in the adipose tissue of rats. Male Wistar rats received daily injections of dexamethasone (1mg/kg body weight (b.w.), intraperitoneally (i.p.)) for 5 days (DEX), whereas control rats received saline (CTL). The metabolic status was investigated, and the epididymal fat fragments were collected for lipolysis and western blot analyses. The DEX rats became hyperglycemic, hyperinsulinemic, insulin resistant and glucose intolerant, compared with the CTL rats (P<0.05). The basal glycerol release in the fat fragments was 1.5-fold higher in the DEX rats (P<0.05). The phosphorylation of protein kinase B (PKB) at ser(473) decreased by 44%, whereas, the phosphorylation of insulin receptor substrate (IRS)-1 at ser(307) increased by 93% in the adipose tissue of the DEX rats after an oral bolus of glucose (P<0.05). The basal phosphorylation of c-jun-N-terminal kinase (JNK) and inhibitor of nuclear factor kappa-B (IKKβ) proteins was reduced by 46% and 58%, respectively, in the adipose tissue of the DEX rats (P<0.05). This was paralleled with a significant reduction (47%) in the glucocorticoid receptor (GR) protein content in the adipose tissue of the DEX rats (P<0.05). The insulin-resistant status of rats induced by dexamethasone administration have PKB and IRS-1 activity attenuated in epididymal fat without increases in the phosphorylation of the proinflammatory signals JNK and IKKβ.

Antitumor Activity Of Irradiated Riboflavin On Human Renal Carcinoma Cell Line 786-o.

Riboflavin (vitamin B2) is a precursor for coenzymes involved in energy production, biosynthesis, detoxification, and electron scavenging. Previously, we demonstrated that irradiated riboflavin (IR) has potential antitumoral effects against human leukemia cells (HL60), human prostate cancer cells (PC3), and mouse melanoma cells (B16F10) through a common mechanism that leads to apoptosis. Hence, we here investigated the effect of IR on 786-O cells, a known model cell line for clear cell renal cell carcinoma (CCRCC), which is characterized by high-risk metastasis and chemotherapy resistance. IR also induced cell death in 786-O cells by apoptosis, which was not prevented by antioxidant agents. IR treatment was characterized by downregulation of Fas ligand (TNF superfamily, member 6)/Fas (TNF receptor superfamily member 6) (FasL/Fas) and tumor necrosis factor receptor superfamily, member 1a (TNFR1)/TNFRSF1A-associated via death domain (TRADD)/TNF receptor-associated factor 2 (TRAF) signaling pathways (the extrinsic apoptosis pathway), while the intrinsic apoptotic pathway was upregulated, as observed by an elevated Bcl-2 associated x protein/B-cell CLL/lymphoma 2 (Bax/Bcl-2) ratio, reduced cellular inhibitor of apoptosis 1 (c-IAP1) expression, and increased expression of apoptosis-inducing factor (AIF). The observed cell death was caspase-dependent as proven by caspase 3 activation and poly(ADP-ribose) polymerase-1 (PARP) cleavage. IR-induced cell death was also associated with downregulation of v-src sarcoma (Schmidt-Ruppin A-2) viral oncogene homologue (avian)/protein serine/threonine kinase B/extracellular signal-regulated protein kinase 1/2 (Src/AKT/ERK1/2) pathway and activation of p38 MAP kinase (p38) and Jun-amino-terminal kinase (JNK). Interestingly, IR treatment leads to inhibition of matrix metalloproteinase-2 (MMP-2) activity and reduced expression of renal cancer aggressiveness markers caveolin-1, low molecular weight phosphotyrosine protein phosphatase (LMWPTP), and kinase insert domain receptor (a type III receptor tyrosine kinase) (VEGFR-2). Together, these results show the potential of IR for treating cancer.

Proteasome Inhibition Represses Unfolded Protein Response and Nox4, Sensitizing Vascular Cells to Endoplasmic Reticulum Stress-Induced Death

Background: Endoplasmic reticulum (ER) stress has pathophysiological relevance in vascular diseases and merges with proteasome function. Proteasome inhibition induces cell stress and may have therapeutic implications. However, whether proteasome inhibition potentiates ER stress-induced apoptosis and the possible mechanisms involved in this process are unclear. Methodology/Principal Findings: Here we show that proteasome inhibition with MG132, per se at non-lethal levels, sensitized vascular smooth muscle cells to caspase-3 activation and cell death during ER stress induced by tunicamycin (Tn). This effect was accompanied by suppression of both proadaptive (KDEL chaperones) and proapoptotic (CHOP/GADD153) unfolded protein response markers, although, intriguingly, the splicing of XBP1 was markedly enhanced and sustained. In parallel, proteasome inhibition completely prevented ER stress-induced increase in NADPH oxidase activity, as well as increases in Nox4 isoform and protein disulfide isomerase mRNA expression. Increased Akt phosphorylation due to proteasome inhibition partially offset the proapoptotic effect of Tn or MG132. Although proteasome inhibition enhanced oxidative stress, reactive oxygen species scavenging had no net effect on sensitization to Tn or MG132-induced cell death. Conclusion/Relevance: These data indicate unfolded protein response-independent pathways whereby proteasome inhibition sensitizes vascular smooth muscle to ER stress-mediated cell death. This may be relevant to understand the therapeutic potential of such compounds in vascular disease associated with increased neointimal hyperplasia.

Genes up- and down-regulated by dermcidin in breast cancer: a microarray analysis

Dermcidin (DCD) is a human gene mapped to chromosome 12q13 region, which is co-amplified with multiple oncogenes with a well-established role in the growth, survival and progression of breast cancers. Here, we present a summary of a DNA microarray-based study that identified the genes that are up- and down-regulated in a human MDA-361 pLKO control clone and three clones expressing short hairpin RNA against three different regions of DCD mRNA. A list of 235 genes was differentially expressed among independent clones (> 3-fold change and P < 0.005). The gene expression of 208 was reduced and of 27 was increased in the three DCD-RNAi clones compared to pLKO control clone. The expression of 77 genes (37%) encoding for enzymes involved in amino acid metabolism, glucose metabolism and oxidoreductase activity and several genes required for cell survival and DNA repair were decreased. The expression of EGFR/ErbB-1 gene, an important predictor of outcome in breast cancer, was reduced together with the genes for betacellulin and amphiregulin, two known ligands of EGFR/ErbB receptors. Many of the 27 genes up-regulated by DCD-RNAi expression have not yet been fully characterized; among those with known function, we identified the calcium-calmodulin-dependent protein kinase-II delta and calcineurin A alpha. We compared 132 up-regulated and 12 down-regulated genes in our dataset with those genes up- and down-regulated by inhibitors targeting various signaling pathway components. The analysis showed that the genes in the DCD pathway are aligned with those functionally influenced by the drugs sirolimus, LY-294002 and wortmannin. Therefore, DCD may exert its function by activating the PI3K/AKT/mTOR signaling pathway. Together, these bioinformatic approaches suggest the involvement of DCD in the regulation of genes for breast cancer cell metabolism, proliferation and survival.

Potent Cardioprotective Effect of the 4-Anilinoquinazoline Derivative PD153035: Involvement of Mitochondrial K(ATP) Channel Activation

Background: The aim of the present study was to evaluate the protective effects of the 4-anilinoquinazoline derivative PD153035 on cardiac ischemia/reperfusion and mitochondrial function. Methodology/Principal Findings: Perfused rat hearts and cardiac HL-1 cells were used to determine cardioprotective effects of PD153035. Isolated rat heart mitochondria were studied to uncover mechanisms of cardioprotection. Nanomolar doses of PD153035 strongly protect against heart and cardiomyocyte damage induced by ischemia/reperfusion and cyanide/aglycemia. PD153035 did not alter oxidative phosphorylation, nor directly prevent Ca(2+) induced mitochondrial membrane permeability transition. The protective effect of PD153035 on HL-1 cells was also independent of AKT phosphorylation state. Interestingly, PD153035 activated K(+) transport in isolated mitochondria, in a manner prevented by ATP and 5-hydroxydecanoate, inhibitors of mitochondrial ATP-sensitive K(+) channels (mitoK(ATP)). 5-Hydroxydecanoate also inhibited the cardioprotective effect of PD153035 in cardiac HL-1 cells, demonstrating that this protection is dependent on mitoK(ATP) activation. Conclusions/Significance: We conclude that PD153035 is a potent cardioprotective compound and acts in a mechanism involving mitoK(ATP) activation.