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.     

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.     

A two-state model of antagonist-AT1 receptor interaction: further support by binding studies at low temperature

The molecular mechanism of insurmountable antagonism was investigated to a large extent in Chinese hamster ovary cells transfected with the human angiotensin II receptor type 1 (AT(1)) receptor. It was proposed that AT(1) receptor antagonists interact with their receptor according to a two-state receptor model. Briefly, this theoretical model reveals that antagonist bound AT(1) receptor can adopt a fast and a slow reversible state. The first, fast reversible state is similar for all antagonists, while the slow reversible state displays the characteristics of each antagonist. In the present study, we performed competition experiments with the AT(1) receptor antagonists candesartan, EXP3174, irbesartan, losartan and ligand [3H]-angiotensin II at 0-4 degrees. This gave the opportunity to verify the two-state model for the first time with experimental data.     

Pharmacokinetics and biochemical efficacy after single and multiple oral administration of losartan, an orally active nonpeptide angiotensin II receptor antagonist, in humans.

1. The pharmacokinetics and biochemical efficacy of losartan, an orally active nonpeptide angiotensin II (AII) receptor antagonist, were evaluated in healthy male volunteers after single and multiple oral administration. 2. Plasma and urinary concentrations of losartan and its active metabolite, E-3174, were determined by a specific high performance liquid chromatographic (h.p.l.c.) method. 3. Plasma concentrations of losartan were proportional to dose over the range of 25 to 200 mg and the terminal half-lives (t1/2,z) ranged from 1.5 to 2.5 h. The mean values of Cmax and AUC0-infinity increased in a dose-dependent manner. 4. Plasma concentrations of E-3174 were higher than those of losartan at all dose levels. The values of Cmax and AUC0-infinity for E-3174 were approximately 2 and 5-8 times higher than those for losartan, respectively. Also the value of t1/2,z was 2 times longer than that of losartan. 5. After multiple dosing for 7 days, the pharmacokinetics of losartan and E-3174 each did not change significantly between day 1 and day 7. 6. Plasma renin activity (PRA) and plasma concentrations of AII increased markedly at all dose levels. Plasma aldosterone levels were slightly reduced, but a similar decrease was also observed with placebo. 7. No clinically significant adverse reaction was observed in any of the volunteers during either study. Blood counts, routine laboratory tests, urine analyses, and electrocardiograms were also not modified by losartan. 8. Losartan appears to be a potent orally active angiotensin II antagonist with a relatively long duration of action.     

Candesartan cilexetil: pharmacological properties and protective effects against organ damage of a novel nonpeptide angiotensin II-receptor antagonist

The nonpeptide angiotensin II (AII) subtype-1 (AT1) receptor antagonist candesartan cilexetil is completely converted to its active form, candesartan, during gastrointestinal absorption. In in vitro studies, candesartan has been found to act as an insurmountable antagonist at the AT1 receptor that dose-dependently reduces the maximal contractions induced by AII and, at high concentrations, virtually eliminates the AT1-receptor-mediated effects of AII. Receptor binding studies suggest that insurmountable antagonism may be due to tight binding to the AT1 receptor and slow dissociation from it. The antihypertensive efficacy of candesartan cilexetil has been demonstrated in different animal models of hypertension. Candesartan cilexetil exerts a long-lasting antihypertensive action in spontaneously hypertensive rats in the low dose range of 0.1-10 mg/kg. The long-lasting antihypertensive effect of candesartan cilexetil is confirmed by the trough/peak ratio in hypertensive patients. It has been demonstrated that administration of AII receptor antagonists is followed by a rise in AII levels, and the increased AII levels result in competition with the antagonist for binding to the receptor. Insurmountable antagonists would seem to be more advantageous since they would block more efficiently in the presence of increasing AII levels than surmountable antagonists. A growing number of studies indicate that candesartan cilexetil provides end-organ protection in addition to lowering blood pressure. The utility of AT1 antagonists may extend beyond treatment of hypertension, chronic heart failure and renal diseases, as suggested by the potential usefulness of ACE inhibitors in the treatment or prevention of many other cardiovascular diseases.     

Pharmacokinetics and blood pressure response of losartan in end-stage renal disease

BACKGROUND: Losartan is a selective angiotensin AT1 receptor antagonist currently employed in the management of essential hypertension. This compound is in common use in populations with renal failure and end-stage renal disease (ESRD). OBJECTIVE: To investigate the pharmacokinetics and pharmacodynamics of losartan in patients with ESRD in order to establish administration guidelines. METHODS: Patients were administered losartan 100 mg/day for 7 days, and after the seventh and final dose pharmacokinetic parameters were determined for both losartan and its active metabolite E-3174. During the study, the haemodialytic clearances of losartan and E-3174 were measured during a standard 4-hour dialysis session. Neurohumoral and biochemical changes were assessed during losartan administration. RESULTS: The pharmacokinetics of losartan and E-3174 in haemodialysis patients did not alter to a clinically significant level. Losartan administration was accompanied by a decline in plasma aldosterone level as well as by an increase in plasma renin activity. Losartan administration resulted in a decline in plasma uric acid level, despite the fact that the study participants had no residual renal function. Losartan and E-3174 were not dialysable. CONCLUSIONS: The pharmacokinetics of losartan and E-3174 are minimally altered in ESRD; thus, dosage adjustment is not required in the presence of advanced dialysis-dependent renal failure. In addition, postdialysis supplementation is not required for losartan because of the negligible dialysability of losartan and E-3174.     

Losartan: a selective angiotensin II type 1 (AT1) receptor antagonist for the treatment of heart failure

Losartan (COZAAR) is the prototype of a new class of potent and selective angiotensin II (AII) type 1 (AT(1)) receptor antagonists with the largest published preclinical and clinical data base. Since all of the AII antagonists are selective for the AT(1) receptor, these drugs should exhibit similar cardiovascular effects. However, since the pharmacokinetic/pharmacodynamic profiles of these agents and their degree of affinity for the AT(1) receptor differ, it is likely that differences in clinical profiles between these drugs exist and will require investigation. Losartan (parent compound), has moderate affinity for the AT(1) receptor (competitive inhibition). Losartan is well-absorbed orally as an active drug and is rapidly converted via oxidation in the human liver to a more potent metabolite (designated E3174) with an affinity 20- to 30-times greater for the AT(1) receptor (non-competitive inhibition). E3174 has a half-life of 6 - 9 h; elimination is via renal and hepatic routes. Antihypertensive and, in heart failure patients, haemodynamic activity is observed over a 24 h period with once daily dosing. Over 6 million patients have been treated for hypertension with continued excellent tolerability. Clinical experience in heart failure is growing, and recent data suggest an improved survival with losartan versus captopril, a drug from the angiotensin-converting-enzyme inhibitor class with proven benefit in this population. The current comprehensive losartan clinical end-point programme (4 large scale morbidity/mortality trials) should provide evidence regarding the efficacy of direct blockade of the AT(1) receptor with losartan compared to standard therapy: 1) The Losartan Heart Failure Survival Study - ELITE II, 2) The Losartan Post-Myocardial Infarction Survival Study - OPTIMAAL, 3) The Losartan Hypertension Survival Study - LIFE and 4) The Losartan Renal Protection Study - RENAAL.     

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.     

Functional interaction between losartan and central tachykinin NK3 receptors in the conscious rat.

1. The cardiovascular and behavioural effects elicted by the intracerebroventricular (i.c.v.) injection of substance P (SP), neurokinin A (NKA), [MePhe7]neurokinin B ([MePhe7]NKB) or angiotensin II (AII) in the conscious rat were assessed before and 5 min after i.c.v. pretreatment with antagonists selective for angiotensin AT1 (losartan and its active metabolite EXP 3174), angiotensin AT2 (PD 123,319) or tachykinin NK3 (R 486) receptors. 2. I.c.v. administration of 25 pmol AII evoked an increase in mean arterial blood pressure (MAP) and water intake behaviour, accompanied by a transient bradycardia, whereas 25 pmol [MePhe7]NKB caused a transient increase in MAP and heart rate (HR) concurrently with marked wet dog shake behaviour. At the same dose, SP and NKA were more potent than [MePhe7]NKB in increasing MAP and HR, but did not produce water intake or wet dog shake behaviours. 3. Losartan (650 pmol, i.c.v.) reduced significantly the cardiovascular and behavioural responses to AII or [MePhe7]NKB, but not to SP or NKA. While 65 pmol losartan was inactive, 260 pmol inhibited selectively the central effects of AII. Whereas EXP 3174 (6.5 nmol) blocked both AII and [MePhe7]NKB-mediated responses, the dose of 650 pmol blocked only the responses to AII. 4. The central responses to AII and [MePhe7]NKB were not affected by PD 123,319 (650 pmol). On the other hand, the [MePhe7]NKB-induced central effects were significnatly reduced by R 486 (650 pmol). The NK3-selective antagonist had no effect against AII. 5. This study provides functional evidence, to support earlier binding data, that losartan (and to some extent its active metabolite EXP 3174) interact with the tachykinin NK3 receptor in rat brain.(ABSTRACT TRUNCATED AT 250 WORDS)     

Drug-disease interactions: losartan effect is not downregulated by rheumatoid arthritis

Inflammatory conditions, such as rheumatoid arthritis, reduce response to calcium channel and beta-adrenergic antagonists but not the angiotensin II type 1 receptor (AT(1)R) antagonist valsartan. Inflammation also reduces clearance of some drugs or active metabolite, thereby reducing response. Active (n = 14) and controlled rheumatoid arthritis (n = 12) and healthy subjects (n = 12) received losartan (100 mg). Blood pressures were measured, and samples were taken for pharmacokinetic and inflammatory mediator concentration determination. Active disease significantly increased arthritic index, nitric oxide, and Creactive protein. Although no between-group difference in plasma losartan concentration-time curves was observed, concentrations of the active metabolite, EXP 3174, were significantly reduced by arthritis. This, however, was not accompanied by reduced clinical response. One subject produced no detectable concentrations of EXP 3174 likely due to insufficient CYP2C9 activity. Despite reduced concentrations of the active metabolite, AT1R antagonists potency does not appear to be reduced by inflammation.     

Population pharmacokinetic of losartan and its active metabolite E-3174 in five different ethnic populations of China

The aim of this study was to develop a combined population pharmacokinetic (PPK) model for losartan and its active metabolite E-3174 in five Chinese ethnicities for individualized drug therapy in clinical practice. HPLC method was used to determine the blood levels of losartan and E-3174 simultaneously. One-, two- and three-compartment models were fitted to plasma concentration time data of 50 Chinese healthy subjects (including Han, Mongolian, Korean, Hui and Uigur) using nonlinear mixed-effect modeling (NONMEM). From the basic model of losartan, the effects of demography and biochemical covariates were investigated, which were added one by one by the forward inclusion and backward elimination. The final models of losartan and E-3174 were connected by first order or transit compartment model. Pharmacokinetic parameters of losartan and its active metabolite E-3174 were assessed simultaneously in one integrated model with the plausible covariates on the key pharmacokineticparameters of E-3174. Nonparametric bootstrap was used for the model stability validation. The data of losartan were best described using a two-compartment model with linear elimination. The time to reach C_max of losartan and E-3174 were obtained to be 0.9 and 3.8 h, respectively. Two transit compartments were chosen with adequate fit of the delayed T_max of E-3174. The population estimates for transformation of losartan to E-3174 was about 73.9%. Ethnicity factor showed significant influence on the non-metabolizing E-3174 clearance CL₁₀, the peripheral compartment clearance CL₂ and the central compartment volume V₁of losartan and also has a significant effect on the transit rate (Kt). A total of 925 out of 1000 iterations succeeded in minimization.The PPK models were steady and reliable. Ethnicity factor showed significant influence on both losartan clearance and the transition from losartan to E-3174, no covariate influencing the PK parameters of E-3174 was identified.     

Evidence that nitric oxide regulates AT1-receptor agonist and antagonist efficacy in rat injured carotid artery

Vascular injury stimulates AT1-receptor expression and nitric oxide (NO) production in smooth muscle cells (SMCs). We examined the ability of AT1 agonists and antagonists to regulate vascular tone ex vivo in injured arteries and the possible modulation by SMC-derived NO. Rings of rat carotid arteries were isolated at day 7 after endothelial denudation and stimulated with angiotensin (Ang) II in the absence or presence of the AT1 antagonists losartan, L-158,809, or EXP-3174. Freshly denuded contralateral arteries were used as controls. AngII-induced contractions were similar in control and injured arteries. Losartan caused an insurmountable inhibition of AngII-induced contractions in injured but not control arteries. Enhanced inhibition of AngII in injured arteries also was observed in the presence of L-158,809 and EXP-3174. In the presence of the NO synthesis inhibitor nitromonomethyl-L-arginine (L-NMMA), maximal contractions to AngII were greater in injured than in control vessels, and AT1-receptor blockade with losartan was surmountable in all vessels. Mechanical removal of superficial neointimal SMCs attenuated NO production and normalized the efficacy of losartan in injured arteries. These results suggest a role for NO in reducing the biologic effects of AT1-receptor agonists and potentiating the efficacy of AT1 antagonists in vessels undergoing remodeling after injury.     

Nitric oxide-dependent antiplatelet action of AT1-receptor antagonists in a pulmonary thromboembolism in mice

The aim of this study was to determine whether AT1-receptor antagonists could inhibit platelet activation-dependent pulmonary thromboembolism in mice and to investigate the involvement of nitric oxide in this action. Losartan, its active metabolite EXP3174, and valsartan given intraperitoneally 1 hour before the thrombotic challenge (in doses of 3, 10, or 30 mg/kg) protected mice from death or hind-limb paralysis in response to intravenous injection of a mixture of collagen and epinephrine; losartan was effective in all doses used, whereas EXP3174 and valsartan reduced mortality only in the two higher doses. The protective action of EXP3174 and valsartan was abolished when nitric oxide synthase was inhibited with l-NAME, whereas that of losartan was only partially reduced. Moreover, only losartan protected mice from death caused by intravenous injection of the thromboxane A2 mimetic U46619 and this action was preserved in l-NAME-pretreated animals. Our results demonstrate the ability of AT1-receptor antagonists to inhibit platelet activation in vivo in a nitric oxide-dependent mechanism. Stronger antiplatelet activity of losartan, most likely due to its blockade of thromboxane A2/prostaglandin H2 receptor, could be of potential clinical relevance, particularly in conditions in which synthesis of endogenous nitric oxide is impaired.     

Comparative effects of angiotensin II AT-1-type receptor antagonists in vitro on human platelet activation

A recent study has shown that losartan, an AT-1-receptor antagonist, interacts with thromboxane A2 (TxA2)/prostaglandin H2 (PGH2) receptors in human platelets. The aim of this study was to analyze the ability of different angiotensin II (Ang II) AT-1-receptor antagonists to inhibit TxA2-dependent human platelet activation. Platelets were obtained from healthy volunteers. Platelets were stimulated with the TxA2 analogue, U46619 (10(-6) M). U46619-stimulated platelet activation was significantly reduced by both losartan and irbesartan in a dose-dependent manner. Only maximal doses of valsartan (5 x 10(-6) M) and the main metabolite of losartan, EXP3174 (5 x 10(-6) M), reduced U46619-induced platelet activation. Whereas the active form of candesartan cilexetil (candesartan, CV-11974) failed to modify platelet activation involved by TxA2, telmisartan showed a higher effect than valsartan and EXP3174 but lower than either losartan and irbesartan. Losartan or irbesartan reduced the binding of [3H]-U46619 to platelets, an effect that was observed with lower ability with the other AT-1 antagonists. Although platelets expressed AT-1-type receptors, exogenous Ang II did not modify platelet activation. This effect was not modified by blocking the AT-2 receptor with PD123319. These results suggest that some AT-1-receptor antagonists reduce TxA2-dependent activation independent of Ang II involvement.     

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.     

Telmisartan has the strongest binding affinity to angiotensin II type 1 receptor: comparison with other angiotensin II type 1 receptor blockers

There is a growing body of evidence that the renin-angiotensin system (RAS) plays a pivotal role in the pathogenesis of cardiovascular diseases. Indeed, large clinical trials have demonstrated a substantial benefit of the blockade of this system for cardiovascular-organ protection. Although several types of angiotensin II type 1 (AT1) receptor blockers (ARBs) are commercially available for the treatment of patients with hypertension, comparisons of the binding affinity to AT1 receptor among them remain to be elucidated. In this study, we examined the dissociation rate of several ARBs from AT1 receptor in vitro. Angiotensin II time-dependently dissociated telmisartan, olmesartan, candesartan, valsartan, losartan and an active metabolite of losartan, EXP3174, from membrane components containing human AT1 receptor The dissociation rate constant of each ARB was 0.003248, 0.004171, 0.005203, 0.009946, 0.01027 and 0.008561 min(-1), with corresponding half-lives of 213, 166, 133, 70, 67 and 81 min, respectively. These results demonstrate that telmisartan has the strongest binding affinity to AT1 receptor among various ARBs examined herein. The rank order of affinity was telmisartan > olmesartan > candesartan > EXP3174 > or = valsartan > or = losartan. The present findings suggest that telmisartan (Micardis) may have long-lasting blood pressure-lowering effects and superior cardioprotective properties in patients with hypertension due to its strongest AT1 receptor antagonistic ability.     

Antagonist interaction with endogenous AT(1) receptors in human cell lines

Using Chinese Hamster Ovary cells expressing human AT(1) receptors cells (CHO-hAT(1)), it was previously shown that insurmountable inhibition of the angiotensin II response by non-peptide antagonists is related to the duration of their receptor occupancy. In the present study it was shown that these antagonists displayed similar binding characteristics to endogenously expressed AT(1) receptors in human adrenal cortex cells (NCI-h295) and renal vascular smooth muscle cells (HVSMC). Competition binding studies with [(3)H]candesartan for NCI-h295 cells, with [(125)I]Sar(1)-Ile(8) angiotensin II for HVSMC and with both radioligands for CHO-hAT(1) cells displayed the same potency order for unlabelled antagonists: candesartan>EXP3174>irbesartan>losartan. The AT(2) receptor antagonist PD123319 displayed low potency in all instances. The apparent half-lives of the antagonist-AT(1) receptor complexes in NCI-h295 cells and HVSMC were comparable to those obtained under identical conditions with CHO-hAT(1) cells. Angiotensin II increased the inositol phosphate accumulation dose dependently with half-maximal response at 17.4+/-1.6nM for NCI-h295 cells and 4.5+/-0.8nM for HVSMC. Pre-incubation of the cells with losartan only produced concentration-dependent rightward shifts of the angiotensin II concentration-response curve. The maximal response was decreased by 85-92% with candesartan, 70-88% with EXP3174 and 60% with irbesartan. The similar binding and inhibitory properties of these antagonists among the investigated cell types validates the use of CHO-hAT(1) cells for investigating pharmacological properties of human AT(1) receptors.     

A new class of diacidic nonpeptide angiotensin II receptor antagonists: candesartan cilexetil.

Blockade of the action of angiotensin II (AII) has long been a target for development of novel antihypertensive agents. We recently discovered a novel class of potent nonpeptide AII receptor antagonists, benzimidazole-7-carboxylic acids including candesartan. Candesartan is a highly potent and insurmountable angiotensin II type-1 receptor (AT1)-selective antagonist. Structure-activity relationship (SAR) studies revealed that the adjacent arrangement of a lipophilic substituent, a tetrazolylbiphenylmethyl moiety and a carboxyl group was the important structural requirement for potent AII antagonistic activity. The benzimidazole ring was found to be one of the most suitable templates arranging these three essential components in correct direction. Especially, the presence of a carboxyl group at the 7-position was found to be essential for insurmountable antagonism. Although candesartan is a very potent AII antagonist, it was found to be absorbed rather inefficiently upon oral administration. To improve bioavailability (BA) of candesartan, chemical modification was examined to yield candesartan cilexetil, a prodrug of candesartan. Candesartan cilexetil is a potent and long-acting blocker that provides effective 24 hr blood pressure control. Our alternative research efforts to improve oral BA was performed by replacement of the tetrazole ring in candesartan by other new acidic bioisosteric heterocyclic rings to find the nonprodrug AII antagonist TAK-536, bearing 5-oxo-1,2,4-oxadiazole ring, which was as potent and orally active as candesartan cilexetil.     

Functional comparison of the antagonistic properties of some Angiotensin II type 1 receptor blockers on the contraction elicited by Angiotensin II and thromboxane A2 on human saphenous veins

We previously described on human vascular preparations that, in addition to its antagonistic properties on Angiotensin II type 1 (AT1) receptor, losartan could also inhibit the contraction elicited by the stable thromboxane A2 mimetic U46619. The present study was designed (1) to investigate, in human vascular preparations (the saphenous veins) whether these antagonistic properties on thromboxane A2/prostaglandin H2 (TP) receptor were shared by some other AT1 receptor antagonists (irbesartan and valsartan) and the active metabolite of losartan EXP3174, and (2) to compare their antagonistic properties on TP receptors to their antagonistic properties on AT1 receptors. In the presence of indomethacin (10 microM) and Nomega-nitro-L-arginine (100 microM), irbesartan, valsartan, and EXP3174 induced a rightward shift of U46619- and angiotensin II-induced contraction. The inhibitory effect of irbesartan, valsartan, and EXP3174 on U46619-induced contraction was significant from 100 microM while their inhibitory effect on the contraction elicited by angiotensin II was significant from 1 nM. With regard to the plasma therapeutic concentrations of irbesartan, valsartan, and EXP3174, these data suggest that TP receptor blockade does not account for the antihypertensive effects of these AT1 receptor blockers.     

Angiotensin II receptor antagonists: candesartan cilexetil

Blockade of the action of angiotensin II (AII) has long been a target for the development of novel antihypertensive agents. We recently discovered a novel class of potent nonpeptide AII receptor antagonists, benzimidazole-7-carboxylic acids including candesartan. Candesartan is a highly potent and insurmountable AII type-1 receptor (AT1)-selective antagonist. Structure-activity relationship (SAR) studies revealed that the adjacent arrangement of a lipophilic substituent, a tetrazolylbiphenylmethyl moiety and a carboxyl group was the important structural requirement for potent AII antagonistic activity. Especially, the presence of a carboxyl group at the 7-position was found to be essential for insurmountable antagonism. To improve bioavailability of candesartan, chemical modification was examined to yield candesartan cilexetil, a prodrug of candesartan. Candesartan cilexetil is a potent and long-acting blocker that, when given once a day to patients, provides effective 24 hr blood pressure control.