IL-4 receptor α blockade prevents sensitization and alters acute and long-lasting effects of allergen-specific immunotherapy of murine allergic asthma.

BACKGROUND: Allergen-specific immunotherapy (AIT) is the only causal treatment for allergy. However, success rates vary depending on the type of allergy and disease background of the patient. Hence, strategies targeting an increased therapeutic efficacy are urgently needed. Here, the effects of blockade of IL-4 and IL-13 signaling on different phases of AIT were addressed. METHODS: The impact of the recombinantly produced IL-4 and IL-13 antagonist IL-4 mutein (IL-4M) on allergic sensitization and AIT outcome in experimental allergic asthma were analyzed in a murine model. The effects of IL-4M administration were assessed prior/during sensitization, immediately after AIT under allergen challenge, and two weeks post-treatment. RESULTS: Intervention with IL-4M prior/during sensitization led to strong induction of IgG1, IgG2a, IgG2b, and IgG3, decrease of specific and total IgE, as well as of IL-5 in serum. Similar effects on the serum immunoglobulin levels were observed immediately after IL4M-supplemented AIT during allergen challenge. Additionally, IL4M markedly suppressed type-2 cytokine secretion of splenocytes beyond the effect of AIT alone. These effects were equaled to those of AIT alone two weeks post-treatment. Intriguingly, here, IL-4M induced a sustained decrease of Th2-biased Tregs (ST2+ FOXP3+ GATA3intermediate ). CONCLUSIONS: IL-4 and IL-13 blockade during experimental AIT demonstrates beneficial effects on immunological key parameters such as immunoglobulin and cytokine secretion immediately after AIT. Although two weeks later these effects were dropped to those of AIT alone, the number of potentially disease-triggering Th2-biased Tregs was further significantly decreased by IL-4M treatment. Hence, IL-4/IL13-targeting therapies prime the immune memory in therapy success-favoring manner.

Pollen-derived adenosine is a necessary cofactor for ragweed allergy.

BACKGROUND: Ragweed (Ambrosia artemisiifolia) is a strong elicitor of allergic airway inflammation with worldwide increasing prevalence. Various components of ragweed pollen are thought to play a role in the development of allergic responses. The aim of this study was to identify critical factors for allergenicity of ragweed pollen in a physiological model of allergic airway inflammation. METHODS: Aqueous ragweed pollen extract, the low molecular weight fraction or the major allergen Amb a 1 was instilled intranasally on 1-11 consecutive days, and allergic airway inflammation was evaluated by bronchoalveolar lavage, lung histology, serology, gene expression in lung tissue, and measurement of lung function. Pollen-derived adenosine was removed from the extract enzymatically to analyze its role in ragweed-induced allergy. Migration of human neutrophils and eosinophils toward supernatants of ragweed-stimulated bronchial epithelial cells was analyzed. RESULTS: Instillation of ragweed pollen extract, but not of the major allergen or the low molecular weight fraction, induced specific IgG1 , pulmonary infiltration with inflammatory cells, a Th2-associated cytokine signature in pulmonary tissue, and impaired lung function. Adenosine aggravated ragweed-induced allergic lung inflammation. In vitro, human neutrophils and eosinophils migrated toward supernatants of bronchial epithelial cells stimulated with ragweed extract only if adenosine was present. CONCLUSIONS: Pollen-derived adenosine is a critical factor in ragweed-pollen-induced allergic airway inflammation. Future studies aim at therapeutic strategies to control these allergen-independent pathways.

Development and function of intestinal innate lymphoid cells.

Innate lymphoid cells (ILCs) are generated from common lymphoid precursors, like lymphocytes, but do not express an antigen receptor. ILCs include Natural Killer (NK) cells, first described 38 years ago, as well as the more recently discovered lymphoid tissue inducer (LTi) cells, NK(22) cells and ILC2s. ILCs reflect many functions of CD4(+) T helper cells by expressing IFNγ, IL-17, IL-22 or IL-13. However, in contrast to T cells, they are not selected on the basis of antigen specificity, and expand and act shortly after stimulation. Therefore, ILCs play fundamental roles early in responses to infection and injury, in the maintenance of homeostasis, and possibly in the regulation of adaptive immunity. Here, we review the recent data on the development and role of RORγt(+) ILCs and ILC2s in intestinal homeostasis and defense.

Discovery of novel, orally active dual NK1/NK2 antagonists.

Exploration of the SAR around selective NK2 antagonists, SR48968 and ZD7944, led to the discovery that naphth-1-amide analogues provide potent dual NK1 and NK2 antagonists. ZD6021 inhibited binding of [3H]-NKA or [3H]-SP to human NK1 and NK2 receptors, with high-affinity (K(i)=0.12 and 0.62nM, respectively). In functional assays ZD6021 had, at 10(-7)M, in human pulmonary artery pK(B)=8.9 and in human bronchus pK(B)=7.3, for NK1 and NK2, respectively. Oral administration of ZD6021 to guinea pigs dose-dependently attenuated ASMSP induced extravasation of plasma proteins, ED(50)=0.5mg/kg, and NK2 mediated bronchoconstriction, ED(50)=13mg/kg.

Behavioral approach to nondyskinetic dopamine antagonists: identification of seroquel.

A great need exists for antipsychotic drugs which will not induce extrapyramidal symptoms (EPS) and tardive dyskinesias (TDs). These side effects are deemed to be a consequence of nonselective blockade of nigrostriatal and mesolimbic dopamine D2 receptors. Nondyskinetic clozapine (1) is a low-potency D2 dopamine receptor antagonist which appears to act selectively in the mesolimbic area. In this work dopamine antagonism was assessed in two mouse behavioral assays: antagonism of apomorphine-induced climbing and antagonism of apomorphine-induced disruption of swimming. The potential for the liability of dyskinesias was determined in haloperidol-sensitized Cebus monkeys. Initial examination of a few close cogeners of 1 enhanced confidence in the Cebus model as a predictor of dyskinetic potential. Considering dibenzazepines, 2 was not dyskinetic whereas 2a was dyskinetic. Among dibenzodiazepines, 1 did not induce dyskinesias whereas its N-2-(2-hydroxyethoxy)ethyl analogue 3 was dyskinetic. The emergence of such distinctions presented an opportunity. Thus, aromatic and N-substituted analogues of 6-(piperazin-1-yl)-11H-dibenz[b,e]azepines and 11-(piperazin-1-yl)dibenzo[b,f][1,4]thiazepines and -oxazepines were prepared and evaluated. 11-(4-[2-(2-Hydroxyethoxy)ethyl]piperazin-1-yl)dibenzo[b,f][1,4]thiazepine (23) was found to be an apomorphine antagonist comparable to clozapine. It was essentially nondyskinetic in the Cebus model. With 23 as a platform, a number of N-substituted analogues were found to be good apomorphine antagonists but all were dyskinetic.

Pharmacological characterization of ZD6021: a novel, orally active antagonist of the tachykinin receptors.

The tachykinins, substance P, neurokinin A, and neurokinin B, have been implicated in many diseases. The present study evaluated the pharmacological properties of a novel tachykinin antagonist ZD6021 [3-cyano-N-((2S)-2-(3,4-dichlorophenyl)-4-[4-[2-(methyl-(S)-sulfinyl)-phenyl]piperidino]butyl)-N-methyl-]-napthamide]. The affinity (K(i)) of ZD6021 for the cloned human neurokinin (NK)1, NK2, and NK3 receptors was 0.12 +/- 0.01, 0.64 +/- 0.08, and 74 +/- 13 nM, respectively. Mucin secretion by Chinese hamster ovary cells transfected with the human NK1 receptor was dose dependently inhibited by ZD6021: pIC(50) = 7.6 +/- 0.1. For NK1 and NK2 receptors, the agonist concentration-response curves using isolated tissues were displaced rightward in the presence of ZD6021: rabbit pulmonary artery, pA2 = 8.7 and 8.5; human pulmonary artery and bronchus, pKB = 8.9 +/- 0.4 and 7.5 +/- 0.2, at 10(-7) M, respectively. Senktide-induced contractions of isolated guinea pig ileum were also blocked by low concentrations of ZD6021. Oral administration of ZD6021 to guinea pigs dose dependently attenuated tracheal extravasation of plasma proteins induced by the NK1 receptor agonist Ac-[Arg6,Sar9,Met(O2)11]-SP(6-11), ED50 = 0.8 micromol/kg, and bronchoconstriction, elicited by the NK2 receptor agonist [beta-Ala8]-NKA(4-10), ED50 = 20 micromol/kg. Potency was unaffected by feeding. After oral administration of ZD6021, the time to peak activity was 150 min for the NK1 receptor and 60 min for the NK2 receptor with pharmacodynamic half-lives of 280 and 458 min, respectively. These data indicate that ZD6021 is a potent, orally active antagonist of all three tachykinin receptors. This compound may be useful for future studies of tachykinin-related pathology such as asthma.

N-aryl-3,3,3-trifluoro-2-hydroxy-2-methylpropanamides: KATP potassium channel openers. Modifications on the western region.

A subset of antiandrogen compounds, the N-aryl-3,3,3-trifluoro-2-hydroxy-2-methylpropanamides 1, were found to activate ATP sensitive potassium channels (KATP) and represent a new class of potassium channel openers (PCOs). A structure-activity relationship was carried out on the western region of this series with the goal of obtaining an activator of the ATP sensitive potassium channel suitable for use in the treatment of urge urinary incontinence. In particular three large 4-(N-aryl) substituents, the (N-phenyl-N-methylamino)sulfonyl, benzoyl, and 4-pyridylsulfonyl moieties, yielded non-antiandrogen, KATP potassium channel openers (39, 41, and 64, respectively) that are bladder selective in an in vivo rat model that simultaneously measures bladder contractions, heart rate, and blood pressure. Substitutions of the aryl rings of 41 and 64 gave several derivatives that also display selectivity in the in vivo rat model; however, none appear to offer a substantial advantage over 41 and 64. The PCO activity of 41 and 64 resides in the (S)-(-) enantiomers. ZD6169, 41(S), has been selected into development for the treatment of urge urinary incontinence.

Studies of the K(ATP) channel opening activity of the new dihydropyridine compound 9-(3-cyanophenyl)-3,4,6,7,9,10-hexahydro-1,8-(2H,5H)-acridined ione in bladder detrusor in vitro.

The potassium (K+) channel opening activity of ZM244085 (9-(3-cyanophenyl)-3,4,6,7,9,10-hexahydro-1,8-(2H,5H)-acridined ione, CAS 149398-59-4), a novel dihydropyridine (DHP), was ascertained. In a set of functional assays, its mechanoinhibitory effect on myogenic activity of guinea pig bladder detrusor muscles, either mildly or highly depolarized with 15 or 80 mmol/l KCl, was measured. ZM244085 had negligible effect on the tone of the detrusor contracted with 80 mmol/l KCl but reduced the myogenic activity induced with 15 mmol/l KCl (IC50=4.2 +/- 0.4 mumol/l). Glibenclamide, an ATP-sensitive K+ (KATP) channel blocker, competitively antagonized this action of ZM244085 with a pA2 value of 7.6. This functional profile of ZM244085 is similar to that of the prototypic K+ channel opener cromakalim but stands in contrast to that of typical DHP Ca2+ channel blockers such as nifedipine and nimodipine. The membrane potential of the guinea pig detrusor, recorded with intracellular microelectrodes, was hyperpolarized 6.8 +/- 3.1 mV by ZM244085 (10 mumol/l). This hyperpolarization was completely blocked by glibenclamide but not affected by apamin (10 mumol/l), a toxin blocking specifically small conductance and Ca2+ dependent K+ (SKCa) channels. ZM244085 (10 mumol/l) increased the whole cell KATP current in isolated guinea pig detrusor cells by 8.8 +/- 2.5 pA, but failed to activate large conductance and Ca2+ dependent K+ (BKCa) channels in excised inside-out membrane patches from those cells. The results from these studies showed that ZM244085 is a K+ channel opener which activates predominantly KATP channels in vitro to relax bladder detrusors.

Calcium dependent K-channels in guinea pig and human urinary bladder.

This study provides evidence for the presence of large conductance Ca(2+)-dependent K-channels in guinea pig and human urinary bladder smooth muscle. A23187, a Ca(2+)-ionophore, increased charybdotoxin and iberiatoxin sensitive 42K efflux in human urinary bladder smooth muscle cells, suggesting that large conductance Ca(2+)-dependent K-channels are present in these cells. NS004, a large conductance Ca(2+)-dependent K-channel opener, relaxed guinea pig bladder strips precontracted with 15 mM KCl which is inhibited by iberiatoxin. In addition, NS004 also evoked an iberiatoxin sensitive increase in 86Rb/42K efflux in guinea pig and human urinary bladder smooth muscle cells, demonstrating that NS004 activates large conductance Ca(2+)-dependent K-channels to achieve its relaxation effect in the bladder.

ZENECA ZD6169: a novel KATP channel opener with in vivo selectivity for urinary bladder.

(S)-N-(4-benzoylphenyl)-3,3,3-trifluoro-2-hydroxy-2-methyl-propionamide (ZD6169) is a novel ATP-sensitive potassium channel opener. Bladder activity and selectivity after oral dosing were studied in conscious, normotensive rats and dogs by monitoring cystometric and cardiovascular (CV) parameters. The reference ATP-sensitive K+ channel opener cromakalim was also evaluated in this study. ZD6169 significantly reduced micturition frequency in rats (ED50 = 0.16 mg/kg), but its effect on CV parameters was minimal (ED20 = 30 mg/kg), yielding a selectivity dose ratio of 187. The duration of action was between 7 and 24 hr at doses of 0.3 and 3 mg/kg, but it was more than 24 hr at 10 mg/kg. The ED50 value for bladder activity in dogs was less than 1.0 mg/kg, and the ED20 value for CV activity was slightly greater than 15 mg/kg but less than 20 mg/kg; the selectivity ratio was greater than 15. A significant improvement in bladder compliance was noted in dogs with ZD6169, and the bladder activity in rats was blocked by i.v. glibenclamide (3 mg/kg). Cromakalim had a bladder profile similar to that of ZD6169 but appeared to be more selective for CV parameters. In conclusion, ZENECA ZD6169 is a unique ATP-sensitive K+ channel opener with in vivo selectivity of relaxing bladder smooth muscle. This agent has the potential for treating patients with urge incontinence.

K-channel opening activity of ZD6169 and its analogs: effect on 86Rb efflux and 3H-P1075 binding in bladder smooth muscle.

Zeneca ZD6169, (S)-N-(4-benzoylphenyl)-3,3,3-trifluoro- 2-hydroxy-2-methylpropionamide, is a novel compound which relaxes urinary bladder smooth muscle in vitro. The effect of ZD6169 and two of its analogs on 86Rb efflux and 3H-P1075 binding in guinea pig bladder strips was investigated to characterize the K-channel opening properties of this compound. ZD6169 concentration dependently increased the rate of 86Rb efflux from guinea pig bladder strips. 86Rb efflux evoked by ZD6169 and its analogs was blocked by glibenclamide (30 muM) but not by charybdotoxin, apamin or alpha-dendrotoxin, suggesting that this compound activates KATP channels in guinea pig bladder. In addition, interaction of ZD6169 with KATP channels was also confirmed in human bladder smooth muscle cells. Specific binding of 3H-P1075, a potent opener of KATP channels, to guinea pig urinary bladder strips was observed. 3H-P1075 binding was inhibited by known KATP openers. ZD6169 inhibited binding of 3H-P1075 to urinary bladder strips like other structurally different KATP openers, e.g. cromakalim and pinacidil. Potencies for inhibition of 3H-P1075 binding by ZD6169 and other potassium channel openers correlate well with potencies for increase in 86Rb efflux and bladder muscle relaxation studies. It is concluded that Zeneca ZD6169 is a potassium channel opener which activates ATP-sensitive K-channels in guinea pig urinary bladder strips as well as in human bladder cells. Furthermore, binding studies suggest that the effects of ZD6169 and its analogs are mediated by binding to the site labeled by 3H-P1075 in guinea pig bladder strips.

Zeneca ZD6169 and its analogs from a novel series of anilide tertiary carbinols: in vitro KATP channel opening activity in bladder detrusor.

The potassium (K+) channel opening activity of Zeneca ZD6169 and one of its pyridylsulfonyl analogs from the anilide tertiary carbinol series was ascertained. Their mechanoinhibitory effects on the myogenic activity of the guinea pig bladder detrusor muscle were measured in a set of functional assays. Elevating the K+ concentration in the tissue bath from 15 to 80 mmol/l increased the IC50 value of ZD6169 from 1.61 +/- 0.22 223 +/- 37 mumol/l. This result suggests that ZD6169 may act as a K+ channel opener. Similar to the prototypic ATP-sensitive K+ (KATP) channel opener cromakalim, the K+ channel openers from the anilide tertiary carbinol series displayed stereoselective mechanoinhibitory activity only in the test protocol in which the detrusor was stimulated with 15 mmol/l KCl. Being the active enantiomer, ZD6169 has an activity more than 30-fold higher than the less active enantiomer. ZD6169 at 10 mumol/l hyperpolarized the guinea pig detrusor membrane potential by 6.1 +/- 1.2 mV and increased the whole cell KATP current in isolated guinea pig smooth muscle cells by 34.9 +/- 7.9 pA. This is comparable to the increase of 26.8 +/- 5.0 pA obtained with 10 mumol/l of lemakalim, the active enantiomer of cromakalim. The K+ channel opening activity of ZD6169 and the pyridylsulfonyl analog was competitively antagonized by the KATP channel blocker glibenclamide in the guinea pig detrusor with a pA2 value of 7.2. This activity, however, was unaffected by blockers of small and large conductance Ca-dependent K+ channels, such as apamin and charybdotoxin, respectively. The present study showed that Zeneca ZD6169 and its analog from the anilide tertiary carbinol series are K+ channel openers that activate KATP channels in vitro to relax bladder detrusors.

K-channel opening activity of dihydropyridine ZM244085: effect on 86Rb efflux and 3H-P1075 binding in urinary bladder smooth muscle.

Zeneca ZM244085, 9-(3 cyanophenyl)hexahydro-1,8 acridinedione, is a novel dihydropyridine (DHP) which relaxes KCl precontracted urinary bladder smooth muscle in vitro. The effect of ZM244085 on low and high KCl induced contractions, 86Rb efflux and [3H]-P1075 binding in guinea pig bladder strips was investigated to characterize the K-channel opening properties of this compound. Since ZM244085 is a dihydropyridine its effect on DHP binding sites on Ca2+ channels was also investigated. ZM244085 was found to be more potent in relaxing detrusor strips precontracted with 15 mM KCl than strips precontracted with 80 mM KCl (Li et al., 1995). This functional profile of ZM244085 is similar to that exhibited by typical K-channel openers (PCO). In addition, inhibition of ZM244085 induced relaxation of detrusor strips by glibenclamide suggests that ZM244085 opens ATP sensitive K-channel (KATP) in urinary bladder (Li et al., 1995). Since the glibenclamide sensitive smooth muscle relaxation activity of ZM244085 could still be an indirect effect of this compound on KATP channels we carried out 86Rb efflux studies and [3H]-P1075 binding studies to further confirm these findings. The 86Rb efflux assay is a direct method for monitoring the movement of K+ ions across the cell membranes. Displacement of [3H]-P1075 binding to bladder membranes supports a direct action of the compound on the KATP channel. The present study demonstrates that ZM244085 in a concentration dependent manner increases the rate of 86Rb efflux from guinea pig bladder strips. This effect was inhibited by glibenclamide (30 microM), a known KATP channel blocker. In addition, interaction of ZM244085 with KATP channels was also confirmed in human bladder smooth muscle cells using a 42K efflux assay. Furthermore, we were able to demonstrate that ZM244085, structurally distinct PCO, inhibited the binding of 3H-P1075 to urinary bladder strips in a manner similar to other KATP openers such as cromakalim and pinacidil. Inhibition of 3H-P1075 binding by ZM244085 and other PCO's correlates well with increases in 86Rb efflux and bladder muscle relaxation studies. Finally, ZM244085 did not exhibit any significant affect on VSCC as evidenced by very weak inhibition of [3H]-PN200,110 binding to bladder membranes by ZM244085. It is concluded that Zeneca ZM244085 is a PCO which activates KATP channels in urinary bladder.

Antimalarials. 10. Substituted 3-halo- and 3-methoxy-2-aryl-4-quinoline(di-n-butylaminomethyl)methanols.

Four 2-aryl-4-quinoline(di-n-butylaminomethyl)methanols with Br, Cl, F, or OMe in position 3 were synthesized by modifications of standard reactions. The antimalarial activity decreased with increased size of the 3-substituent. The 3-F-4',6,8-Cl3 compound was the most active (at 2.5 mg/kg) and was completely curative at 80 mg/kg against P. berghei in mice.