Three-year efficacy and safety of certolizumab pegol for the treatment of plaque psoriasis: results from the randomized phase 3 CIMPACT trial.

BACKGROUND: Certolizumab pegol (CZP) is an Fc-free, PEGylated anti-tumor necrosis factor biologic. OBJECTIVES: To report 3-year outcomes from the CIMPACT (NCT02346240) phase 3, CZP in moderate to severe plaque psoriasis, randomized controlled trial. METHODS: Adults were randomized 3:3:3:1 to CZP 200 mg every other week (Q2W), CZP 400 mg Q2W, etanercept biweekly or placebo. At Week 16, CZP- and etanercept-treated PASI 75 responders were re-randomized to CZP 200 mg Q2W, CZP 400 mg Q4W, CZP 400 mg Q2W or placebo for maintenance treatment; PASI 75 non-responders entered an open-label escape CZP 400 mg Q2W arm. Patients entering the open-label extension (OLE; Weeks 48-144) from blinded treatment received CZP 200 mg Q2W. RESULTS: Double-blinded results have been reported previously. 261 patients received 200 mg Q2W upon OLE entry. PASI 75 response was maintained in patients continuing 200 mg Q2W treatment through Weeks 16-144 (Week 144: 96.2%). In patients dosed down at Week 48 (double-blinded 400 mg to 200 mg Q2W), PASI 75 decreased (Week 48: 98.7%; Week 144: 85.9%). In patients who received placebo through Weeks 16-48, PASI 75 response decreased (Week 48: 60.4%), then increased following Week 48 switch to 200 mg Q2W (Week 144: 95.1%). 48 and 36 patients initially randomized to 200 and 400 mg Q2W, respectively, were Week 16 PASI 75 non-responders and entered the escape arm; at Week 144, 71.8% and 78.2% achieved PASI 75. No new safety signals were identified. CONCLUSIONS: Response to CZP was durable over three years; no new safety signals were identified.

Impact of recurrent COVID-19 disease waves on acute myocardial infarction epidemics: results from a regional network.

Aims: To assess the impact of COVID-19 related public containment measures during recurrent COVID-19 waves on hospital admission rate for acute myocardial infarction (AMI).Methods and results: Clinical characteristics, reperfusion therapy modalities, COVID-19 status and in-hospital mortality of consecutive AMI patients who were admitted in a regional AMI network were recorded during one year starting in March 2020 and were compared with the year before. The COVID-19 study period encompassed two waves: the first in March-May 2020 and the second in October-December 2020. A total of 1349 AMI patients were hospitalised of which 725 during the pre-COVID period and 624 during the COVID period (incidence rate ratio of 1.16, p = 0,006). The impact was predominantly present in the first wave (32% reduction: n = 204 vs 152) and evanished during the second wave (3% increase (152 vs 156). A similar pattern was observed for ACS with cardiac arrest with a 92% reduction (n = 36 vs 3) during the first wave and no change during the second wave (18 vs 18). After correction for temperature and air quality, COVID-19 epidemic remained associated with a decrease of AMI hospitalisation (p = 0.046). Reperfusion strategy for AMI patients, were comparable between both study periods. The in-hospital mortality between the two periods was comparable (2.6% versus 1.9%), but COVID-19 positive ACS patients (n = 7) had a high mortality rate (14%).Conclusion: COVID-19 related public containment measures resulted during the first wave in a 32% reduction of AMI hospitalisation, but this impact was not visible anymore during the second wave.

Lymphocytic bronchiolitis after lung transplantation is associated with daily changes in air pollution.

Acute rejection represents a major problem after organ transplantation, being a recognized risk for chronic rejection and mortality. Recently, it became clear that lymphocytic bronchiolitis (LB, B-grade acute rejection) is more important than previously thought, as it predisposes to chronic rejection. We aimed to verify whether daily fluctuations of air pollution, measured as particulate matter (PM) are related to histologically proven A-grade rejection and/or LB and bronchoalveolar lavage (BAL) fluid cellularity after lung transplantation. We fitted a mixed model to examine the association between daily variations in PM(10) and A-grade rejection/LB on 1276 bronchoscopic biopsies (397 patients, 416 transplantations) taken between 2001 and 2011. A difference of 10 µg/m(3) in PM(10) 3 days before diagnosis of LB was associated with an OR of 1.15 (95% CI 1.04-1.27; p = 0.0044) but not with A-grade rejection (OR = 1.05; 95% CI 0.95-1.15; p = 0.32). Variations in PM(10) at lag day 3 correlated with neutrophils (p = 0.013), lymphocytes (p = 0.0031) and total cell count (p = 0.024) in BAL. Importantly, we only found an effect of PM10 on LB in patients not taking azithromycin. LB predisposed to chronic rejection (p < 0.0001). The risk for LB after lung transplantation increased with temporal changes in particulate air pollution, and this was associated with BAL neutrophilia and lymphocytosis. Azithromycin was protective against this PM effect.

Stronger associations between daily mortality and fine particulate air pollution in summer than in winter: evidence from a heavily polluted region in western Europe.

BACKGROUND: Numerous studies have shown a strong association between daily mortality and small particulate with a diameter of <10 microm (PM10) air pollution, but the effects of season have not always been well characterised. AIM: To study the shape of the association between short-term mortality and PM10 across seasons and quintiles of outdoor temperature. DESIGN, SETTING AND PARTICIPANTS: Daily data on mortality (n = 354 357), outdoor temperature and PM10 in Flanders, Belgium, from January 1997 to December 2003, were analysed across warm versus cold periods of the year (April-September v October-March), with seasons and quintiles of outdoor temperature as possible effect modifiers. RESULTS: There was a significant (p<0.001) interaction between PM10 and period of the year in relation to mortality. To allow for non-linearity, daily mean PM10 concentrations were categorised into quartiles. Season-specific PM10 quartiles showed a strong and steep linear association between mortality and PM10 in summer and a less linear association in spring and autumn, whereas in winter the association was less strong and mortality was only increased in the highest PM10 quartile. The effect sizes expressed as the percentage increase in mortality on days in the highest season-specific PM(10) quartile versus the lowest season-specific PM10 quartile were 7.8% (95% CI 6.1 to 9.6) in summer, 6.3% (4.7 to 7.8) in spring, 2.2% (0.58 to 3.8) in autumn and 1.4% (0.06 to 2.9) in winter. An analysis by quintiles of temperature confirmed these effect sizes. CONCLUSION: The short-term effect of particulate air pollution on mortality strongly depends on outdoor temperature, even in a temperate climate.

Distinction between surmountable and insurmountable selective AT1 receptor antagonists by use of CHO-K1 cells expressing human angiotensin II AT1 receptors.

1. CHO-K1 cells that were stably transfected with the gene for the human AT1 receptor (CHO-AT1 cells) were used for pharmacological studies of non-peptide AT1 receptor antagonists. 2. In the presence of 10 mM LiCl, angiotensin II caused a concentration-dependent and long-lasting increase of inositol phosphates accumulation with an EC50 of 3.4 nM. No angiotensin II responses are seen in wild-type CHO-K1 cells. 3. [3H]-Angiotensin II bound to cell surface AT1 receptors (dissociates under mild acidic conditions) and is subject to rapid internalization. 4. Non-peptide selective AT1 antagonists inhibited the angiotensin II (0.1 microM) induced IP accumulation and the binding of [3H]-angiotensin II (1 nM) with the potency order: candesartan > EXP3174 > irbesartan > losartan. Their potencies are lower in the presence of bovine serum albumin. 5. Preincubation with the insurmountable antagonist candesartan decreased the maximal angiotensin II induced inositol phosphate accumulation up to 94% and, concomitantly, decreased the maximal binding capacity of the cell surface receptors. These inhibitory effects were half-maximal for 0.6 nM candesartan and were attenuated by simultaneous preincubation with 1 microM losartan indicating a syntopic action of both antagonists. 6. Losartan caused a parallel rightward shift of the angiotensin II concentration-response curves and did not affect the maximal binding capacity. EXP3174 (the active metabolite of losartan) and irbesartan showed a mixed-type behavior in both functional and binding studies. 7. Reversal of the inhibitory effect was slower for candesartan as compared with EXP3174 and irbesartan and it was almost instantaneous for losartan, suggesting that the insurmountable nature of selective AT1 receptor antagonists in functional studies was related to their long-lasting inhibition.

Angiotensin II type 1 receptor antagonists. Why do some of them produce insurmountable inhibition?

Chinese hamster ovary (CHO) cells expressing human recombinant angiotensin II type 1 (AT(1)) receptors offer a useful experimental system in which antagonist binding and inhibition of AT-induced inositol mono-, bis-, and trisphosphate accumulation can be measured under identical experimental conditions. The major conclusions of the current work are: All investigated AT(1) antagonists are competitive with respect to AT. They bind to a common or overlapping binding site on the receptor in a mutually exclusive way. Reduction of the maximal angiotensin II response, i.e. insurmountable inhibition, is observed only when the cells are preincubated with candesartan, EXP3174, or irbesartan and is strictly related to the dissociation rate of the antagonist-receptor complex. On the other hand, inhibition by losartan is fully surmountable by AT, and its dissociation is very rapid. With respect to the binding kinetics, the antagonist-receptor complex can adopt a fast and a slow reversible state. The equilibrium between both states, which is dependent upon the nature of the antagonists, determines the extent of insurmountable inhibition. Consequently, the dissociation rate of the different antagonists correlates with the amount of insurmountable inhibition. In addition to the relatively slow dissociation of candesartan, reassociation to the receptor, which is measurable in CHO-AT(1) cells, likely contributes to its long-lasting blood pressure lowering effect in vivo.

Reversible and syntopic interaction between angiotensin receptor antagonists on Chinese hamster ovary cells expressing human angiotensin II type 1 receptors.

Evidence for a competitive type of interaction between angiotensin II type 1 (AT(1)) antagonists on Chinese hamster ovary cells expressing the human AT(1) receptor (CHO-AT(1)) was obtained by analyzing the binding of [(3)H]-2-ethoxy-1-[(2'-(1H-tetrazol-5-yl)biphenyl-4-yl)methyl]-1H- ben zimidazoline-7-carboxylic acid ([(3)H]candesartan) and by measuring the AT-induced production of inositol phosphates. The AT(1) antagonists candesartan, 2-n-butyl-4-chloro-1-[(2'-(1H-tetrazol-5-yl)biphenyl-4-yl)methyl]+ ++imid azole-5-carboxylic acid (EXP3174), or 2-n-butyl-4-chloro-5-hydroxymethyl-1-[(2'-(1H-tetrazol-5-yl)bip hen yl- 4-yl)methyl]imidazole (losartan) produced a concentration-dependent increase in the apparent K(d) values of [(3)H]candesartan in saturation binding experiments, while the B(max) values were unchanged. Furthermore, the dissociation rate of the radioligand initiated by 1 microM unlabelled candesartan was not changed in the presence of 10 microM losartan, 10 microM EXP3174, or 10 microM irbesartan (2-n-butyl-4-spirocyclopentane-1-[(2'-(1H-tetrazol-5-yl)b iph enyl-4-yl) methyl]2-imidazolin-5-one)). Preincubation of the CHO-AT(1) cells with candesartan, EXP3174, and irbesartan caused a reduction in the maximal AT-induced inositol mono-, bis-, and trisphosphate production. This insurmountable effect was reversed in the presence of 1 microM losartan. In line with this finding, the insurmountable antagonist concentration-inhibition curves at 10 microM AT were shifted to the right in the presence of losartan. For candesartan this effect was concentration-dependent, yielding a pK(B) value for losartan of 7.7, which is similar to the pK(B) from previously obtained AT concentration-response curves. Finally, the dissociation rate of candesartan, EXP3174, irbesartan, and losartan was determined by measuring the recovery of AT responses after antagonist pretreatment and washing of the cells with medium containing 1 microM losartan to prevent re-association of the insurmountable antagonists. In addition, similar kinetic data were obtained from the slowing of the [(3)H]candesartan association rate to antagonist preincubated cells.

A two-state receptor model for the interaction between angiotensin II type 1 receptors and non-peptide antagonists.

The interaction between non-peptide antagonists and the human angiotensin II type 1 (AT1) receptor in CHO-K1 cells was investigated by incubating the cells with antagonist, followed by a brief exposure to angiotensin II and measurement of the resulting inositol phosphate accumulation. The experimental data, expressed either as angiotensin II concentration-response curves or as antagonist concentration-inhibition curves, were in good agreement with computer-generated data according to a single-state model for the surmountable antagonist losartan and according to a two-step, two-state receptor model for the insurmountable antagonists candesartan, EXP3174, and irbesartan. Experimental and computer-generated data concerning the simultaneous exposure of the receptors to EXP3174 and losartan indicated that losartan produced a concentration-dependent restoration of the maximal response (angiotensin II concentration-response curves) as well as a rightward shift of the insurmountable portion of the EXP3174 inhibition curves, thus counteracting the higher-affinity EXP3174 binding. In conclusion, these findings provide further support for the concept that insurmountable and surmountable AT1 antagonists are mutually competitive and that insurmountable antagonist-receptor complexes may adopt different states.

Binding of the antagonist [3H]candesartan to angiotensin II AT1 receptor-transfected [correction of tranfected] Chinese hamster ovary cells.

Binding of the non-peptide angiotensin II AT1 antagonist [3H](2-ethoxy-1-[(2'-(1H-tetrazol-5-yl)biphenyl-4-yl)methyl]- H-benzimidazoline-7-carboxylic acid ([3H]candesartan) to human angiotensin II AT1 receptor-transfected Chinese hamster ovary (CHO-AT1) cells was inhibited to the same extent by angiotensin II and non-peptide angiotensin II AT1 antagonists. No binding was observed in control CHO-K1 cells. Dissociation was slow (k(-1) = 0.0010+/-0.0001 min(-1)) after removal of the free [3H]candesartan but increased 5-fold upon addition of supramaximal concentrations of angiotensin II AT1 antagonists. Angiotensin II responses recovered equally slow from candesartan-pretreatment. When washed and further incubated, these angiotensin II responses also recovered more rapidly in the presence of 2-n-butyl-4-chloro-5-hydroxymethyl-1-[(2'-(1H-tetrazol-5-yl)biphen yl-4-yl)methyl]imidazole (losartan), indicating that unlabelled ligands prevented reassociation. [3 H]candesartan saturation binding experiments required a long time to reach equilibrium. Therefore, the equilibrium dissociation constant (Kd = 51+/-8 pM) was calculated from the association and dissociation rate constants. Our findings indicate that the insurmountable nature of candesartan in functional studies is related to its slow dissociation from the receptor.

Insurmountable angiotensin AT1 receptor antagonists: the role of tight antagonist binding.

Angiotensin II increased the inositol phosphates production (EC50 = 3.4+/-0.7 nM) in Chinese hamster ovary (CHO) cells expressing the cloned human angiotensin AT1 receptor (CHO-AT1 cells). Coincubation with angiotensin AT1 receptor antagonists produced parallel rightward shifts of the concentration-response curve without affecting the maximal response. The potency order is 2-ethoxy-1-[(2'-(1H-tetrazol-5-yl)biphenyl-4-yl)methyl]-1H-benz imidazoline-7-carboxylic acid (candesartan) > 2-n-butyl-4-chloro-1-[(2'-(1H-tetrazol-5-yl)biphenyl-4-yl)methyl]i midazole-5-carboxylic acid (EXP3174) > 2-n-butyl-4-spirocyclopentane-1-[(2'-(1H-tetrazol-5-yl)biphe nyl-4-yl)methyl]2-imidazolin-5-one (irbesartan)> of 2-n-butyl-4-chloro-5-hydroxymethyl-1-(2'-(1H-tetrazol-5-yl)bipheny l-4-yl)methyl]imidazole (losartan). Additionally, preincubation with these antagonists depressed the maximal response, i.e., 95%, 70%, 30% of the control response for candesartan, EXP3174 and irbesartan and not detectable for losartan. Increasing the antagonist concentration or prolonging the preincubation time did not affect this depression. Furthermore, these values remained constant for candesartan and EXP3174, when the angiotensin II incubation time varied between 1 and 5 min. Our data indicate that antagonist-receptor complexes are divided into a fast reversible/surmountable population and a tight binding/insurmountable population at the very onset of the incubation with angiotensin II.

Interaction between the partially insurmountable antagonist valsartan and human recombinant angiotensin II type 1 receptors.

The interaction between the AT1 receptor-selective antagonist valsartan, and its human receptor, was investigated by direct radioligand binding as well as by its inhibition of angiotensin II induced inositol phosphate accumulation in CHO cells expressing human recombinant AT1 receptors. Specific binding of [3H]-valsartan rapidly reached equilibrium at 37 degrees C. It was saturable and occurred to a homogeneous class of sites with a K(D) of 0.88+/-0.076. It was inhibited by other AT1 receptor antagonists with the same potency order as previously described for the binding of [3H]-angiotensin II and [3H]-candesartan to human AT1 receptors (i.e. candesartan > or = EXP3174 > valsartan = irbesartan = angiotensin II > losartan). When valsartan and angiotensin II were applied simultaneously to the CHO-AT1 cells. the antagonist caused a rightward shift of the angiotensin II concentration response curve. Hence, valsartan interacts with the AT1 receptor in a manner that is competitive with angiotensin II. Pre-incubation of the cells with 0.5, 5 and 50 nM valsartan caused an additional, concentration-dependent, up to 55% decline of the maximal response. The partial nature of this insurmountable inhibition by valsartan was confirmed by biphasic antagonist concentration-inhibition curves. These data reflect the co-existence of a fast reversible/surmountable as well as a tight binding/insurmountable valsartan receptor complex. In agreement, pre-incubation of the CHO-AT1 cells with 5 and 50 nM valsartan produced a partial inhibition of the angiotensin II induced increase of the free intracellular calcium concentration. [3H]-Valsartan dissociated from its receptors with a half-life of 17 min. In functional recovery experiments with valsartan-pre-treated cells, the angiotensin II-mediated response was half-maximally restored within approximately 30 min. These kinetic data suggest that the insurmountable inhibition by valsartan is related to its relatively slow dissociation from the human AT1 receptors.

Binding characteristics of [(3)H]-irbesartan to human recombinant angiotensin type 1 receptors.

The aim of the present work was to investigate the binding properties of the selective AT(1)-receptor antagonist irbesartan to human AT(1)-receptors by direct radioligand binding. For this purpose the specific binding of [(3)H]-irbesartan to intact Chinese Hamster Ovary (CHO) cells expressing human recombinant AT(1)-receptors was determined. Specific binding of [(3)H]-irbesartan rapidly reached equilibrium and was saturable with a KD of 1.94 +/- 0.12 to a homogeneous class of binding sites. Its binding was inhibited by other AT(1) antagonists (AIIAs) with the same potency order as previous results from [(3)H]-angiotensin II and [(3)H]-candesartan binding to human AT(1)-receptors. Whereas the dissociation rate of [(3)H]-irbesartan was essentially independent of the radioligand concentration, it was much slower at 12 degrees C when compared with 37 degrees C. Moreover, the dissociation rate was similar, as determined in washout experiments in the absence or presence of unlabelled AT(1) antagonists. At 37 degrees C the dissociation rate constant corresponded to a half-life of approximately seven minutes, which is sufficient to explain the partially insurmountable inhibition by irbesartan in previous studies. In contrast, other phenomena such as the plasma half life and tissue-related factors are necessary to explain its sustained in vivo antihypertensive effect.

Lys(199) mutation of the human angiotensin type 1 receptor differentially affects the binding of surmountable and insurmountable non-peptide antagonists.

Many slow dissociating (insurmountable) non-peptide angiotensin type 1 receptor (AT1) antagonists contain,besides the acidic biphenyltetrazole substructure of losartan, a second acidic group to stabilise antagonist-receptor complexes. To investigate the involved basic amino-acids of the human AT1-receptor, wild-type and mutant receptors were transiently transfected in CHO-K1 cells and characterised by [3H]candesartan binding. Lys199-->Gln substitution decreased the affinity 45-fold for candesartan (95% insurmountable),18-fold for EXP3174 (70% insurmountable), 10-fold for irbesartan (40% insurmountable) and 5-fold for losartan (surmountable). His256 -->Ala substitution had only minor effects. This suggests that Lys199 is important for the tight binding of non-peptide antagonists.

Air pollution-associated procoagulant changes: the role of circulating microvesicles.

BACKGROUND: Epidemiological studies suggest an association between exposure to particulate matter (PM) in air pollution and the risk of venous thromboembolism (VTE). OBJECTIVES: To investigate the underlying pathophysiological pathways linking PM exposure and VTE. PATIENTS AND METHODS: We assessed potential associations between PM exposure and coagulation and inflammation parameters, including circulating microvesicles, in a group of 233 patients with diabetes. RESULTS: The numbers of circulating blood platelet-derived and annexin V-binding microvesicles were inversely associated with the current levels of PM(2.5) or PM(10), measured on the day of sampling. Recent past exposure to PM(10), up to 1 week prior to blood sampling, estimated at the patients' residential addresses, was associated with elevated high-sensitivity C-reactive protein (CRP), leukocytes and fibrinogen, as well as with tissue factor (TF)-dependent procoagulant changes in thrombin generation assays. When longer windows of past exposure were considered, up to 1 year preceding blood sampling, procoagulant changes were evident from the strongly increased numbers of red blood cell-derived circulating microvesicles and annexin V-binding microvesicles, but they no longer associated with TF. Past PM exposure was never associated with activated partial thromboplastin time (aPTT), prothrombin time (PT), or factor (F) VII, FVIII, FXII or D-dimers. Residential distance to a major road was only marginally correlated with procoagulant changes in FVIII and thrombin generation. CONCLUSIONS: Increases in the number of microvesicles and in their procoagulant properties, rather than increases in coagulation factors per se, seem to contribute to the risk of VTE, developing during prolonged exposure to air pollutants.

Inhibition of angiotensin II-induced inositol phosphate production by triacid nonpeptide antagonists in CHO cells expressing human AT1 receptors.

PURPOSE: The aim of the present work is to describe the inhibitory properties of LY301875 and LY303336, two polysubstituted 4-aminoimidazole AT1 receptor antagonists, on CHO cells expressing human recombinant AT1 receptors. METHODS: The binding of [3H]-angiotensin II to intact cells as well as to angiotensin II induced inositol phosphate accumulation is measured. RESULTS: Both antagonists inhibit specific [3H]-angiotensin II binding to AT1 receptors in these cells, with IC50 values of 5.9 and 5.2 nM, respectively. Preincubation of the cells with LY301875 results in a decline of up to 80% of the maximal angiotensin II-stimulated inositol phosphate (IP) production. A near complete decline of the maximal response is observed for LY303336. This insurmountable inhibition is attenuated for both antagonists when losartan is included during the preincubation of the cells. CONCLUSIONS: Functional recovery experiments, in which antagonist-preincubated cells are washed and exposed to fresh media, suggest that the insurmountable inhibition by LY301875 and LY303336 is related to their relatively slow dissociation from the AT1 receptors. As already described for losartan and the derived insurmountable AT1 antagonists candesartan. EXP3174, and irbesartan, coincubation experiments reveal that LY301875 and LY303336 interact with the AT1 receptor in a manner that is competitive with angiotensin II.

The effects of candesartan on human AT1 receptor-expressing Chinese hamster ovary cells.

Chinese hamster ovary cells expressing the cloned human angiotensin II receptor of the AT1 subtype (CHO-AT1 cells) were used as an 'in vitro" model system to investigate the action mechanism of the nonpeptide AT1 receptor blocker candesartan. In the presence of 10 mM LiCl, angiotensin II causes a long-lasting increase in the production of inositol phosphates in these cells. This effect is dose-dependent with half-maximal stimulation (EC50) at 3 nM angiotensin II. Pre-incubation of the cells for 30 min at 37 degrees C with candesartan decreases the maximal response to angiotensin II by up to 90%, with a half-maximal decrease at 0.5 nM candesartan. At this concentration, candesartan only produces a slight rightward shift of the angiotensin II dose-response curve. Recovery experiments on CHO-AT1 cells reveal that the inhibitory effect of candesartan is only slowly reversed after removal of the blocker. The insurmountable effect of candesartan can therefore be ascribed to its long-lasting inhibition of the AT1 receptor.