Angiotensin II Receptor Pharmacology and AT1-Receptor Blockers

SESSION 1 Angiotensin II receptor pharmacology and AT1-receptor blockers

O Chung, T Csiko´s and T Unger Institute of Pharmacology, Christian-Albrechts-University, Kiel, Germany

Angiotensin II (Ang II) has diverse physiological actions tors have been shown to exert counteracting effects on leading, for example, to increases in extracellular vol- cellular growth and differentiation, vascular tone and ume, peripheral vascular resistance and blood pressure, the release of arginine vasopressin (AVP). In each con- and has also been implicated in the regulation of cell dition the AT2 receptor appears to down-modulate growth and differentiation. Molecular cloning and phar- actions mediated by the AT1 receptor, resulting in macological studies have deﬁned two major classes of decreased cellular proliferation, decreased levels of

Ang II receptors, designated as AT1 and AT2. Most serum AVP levels or decreased vasoconstrictor effects of Ang II are mediated by AT1 receptors. Much responses. In addition, in neuronal cell lines, the AT2 less is known about the physiological role of AT2 recep- receptor reportedly exerts antiproliferative effects and tors. Recent evidence suggests involvement of AT2 promotes neurite outgrowth, an effect accompanied by receptors in development, cell differentiation, apoptosis signiﬁcant changes in the gene expression pattern of and regeneration in various tissues. AT1 and AT2 recep- growth- and differentiation-related genes.

Keywords: angiotensin II; receptors; AT1 antagonists; AT1;AT2

Introduction ACE is not the only enzyme leading to Ang II formation, since other enzymes such as chymase5 The octapeptide, angiotensin II (Ang II), is the major (which is believed to substantially contribute to Ang effector molecule of the renin-angiotensin-system II formation in the human heart),6 CAGE, cathepsin (RAS), and exerts a multitude of actions. Besides G,7,8 tPA, elastase,9 tonin,10 and others can also gen- being the classically recognised potent vasoconstric- erate Ang II. Furthermore, ACE is identical to kinin- tor, Ang II is capable of inducing cell proliferation, ase II, an enzyme degrading bradykinin and other and is as such probably contributing, under patho- kinins to inactive metabolites. A blockade of ACE logic conditions, to structural alterations in various is, therefore, associated with a potentiation of organs. Thus, Ang II is considered an important fac- endogenous kinins, a mechanism thought to contrib- tor in cardiovascular pathology, such as cardiac left ute to desirable effects of ACE inhibitors such as the ventricular hypertrophy and fibrosis, vascular media organ protection.11 Unfortunately, this was also hypertrophy, or neointima formation, and structural associated with some unwarranted effects, for alterations of the heart and kidney, like postinfarct example, dry cough (limiting the use of ACE inhibi- remodelling and nephrosclerosis. In recent years it tors in approximately 10% of patients). has become more evident that the RAS also plays a major role in the development and maintenance of arterial hypertension and numerous approaches Development of Ang II-receptor binding have been undertaken to pharmacologically inter- substances fere with the system. In most cases, the primary therapeutic goal was to lower elevated blood press- In an endeavour to block the RAS more specifically ure. However, since the development of angiotensin- at the level of its receptors, peptidergic Ang II-recep- converting enzyme (ACE) inhibitors, experimental tor antagonists were developed. As the first of its class, saralasin (Sar1 Ile8-Ang II) was introduced in and clinical studies have established that, besides 12 the decrease in blood pressure, drugs like ACE 1971. However, saralasin and further antagonists, inhibitors or angiotensin AT -receptor antagonists such as sarilesin or sarmesin, were all based on a 1 peptide structure that precludes oral adminis- can exert further beneficial actions independently of 13,14 blood pressure. These include a reduction in vascu- tration. Besides their lack of bioavailability, they lar1 and post-myocardial-infarction remodelling2 or also featured the disadvantage of having a short dur- ation of action and a high intrinsic activity by them- preservation of kidney function in (diabetic) nephro- 13 pathy.1,3,4 selves. Furthermore, since they were unable to dis- criminate between the different Ang II-receptor subtypes, they unselectively block all available Ang Correspondence: Prof. Dr. med. Thomas Unger, Institute of II receptors. Pharmacology, University of Kiel, Hospitalstr. 4, D-24105 Kiel, In 1982, imidazole-5-acetic acid derivatives were Germany issued,15 and it was soon recognised that these com- Ang II receptor pharmacology O Chung et al S12 pounds could antagonise Ang II-induced vaso- des transcripts of approximately 2.4 kb in size.51 Its constriction in isolated vessels. This was the basis coding region is located entirely in the third exon. for the further development of highly specific and Different groups have reported that the AT1 recep- selective angiotensin AT1-receptor binding sub- tor interacts with various G-proteins and is coupled 16–19 20–23 stances such as losartan, valsartan, irbesar- to one of the two heteromeric G-proteins: Gq␣ or Gi␣. tan,24–27 candesartan,28 eprosartan,29 telmisartan,30 Ang II binding to specific sites on the extracellular 31 and others (overview ), and AT2-receptor and membrane-spanning portions of the AT1- ligands/antagonists, for example, PD123177, receptor results in subsequent activation of phos- 32,33 PD123319, and CGP42112. pholipases C, D, and A2 via Gq or inhibition of aden- ylate cyclase via Gi, respectively. The phospholipase C activation results in the generation of 1, 4, 5- Ang II receptor subtypes trisphosphate and diacylglycerol, leading to an activation of protein kinase C and an increase in In the past, both the short- and long-term effects of intracellular calcium levels via L-type calcium chan- Ang II were thought to be mediated by a single nels.52–54 The rise in intracellular calcium is associa- receptor. The development of highly specific and ted with typical AT1-receptor–associated responses selective AT1-receptor antagonists was the basis for such as vasoconstriction and secretion of aldos- the identification of the Ang II-receptor subtypes. terone. The cascade induced by phospholipase D Generally recognised are the AT1 and AT2 receptors, and A2 leads to the formation of fatty acids. Phos- which display a heterogeneous distribution in per- pholipase D hydrolyses phosphatidylcholine into ipheral tissues and the brain.34–36 In rodents and phosphatidic acid and choline. Phosphatidic acid Xenopus, the AT1-receptor subtype exists as two iso- and diacylglycerol can be inter-converted allowing 37,38 forms, namely the AT1a and the AT1b receptor. the degradation of the latter to fatty acids and gly- Sequence information from humans also suggests cerol by diacylglycerol lipase. Phospholipase A2 39–41 the existence of a second AT1 receptor, but so hydrolyses phosphatidylcholine and phosphatidyle- far only a single AT1 gene has been detected on chro- thanolamine into the corresponding lysophospho- mosome 3. While AT1a and AT1b subtypes are more lipids (C, E) and fatty acids. A fatty acid frequently or less equally expressed in spleen, liver and kid- released by phospholipase A2 and diacylglycerol 37,38 neys, the AT1a receptor seems to be predominant lipase is arachidonic acid, which is a precursor of in vascular smooth muscle, heart, lung, ovary, and leukotrienes and prostaglandins. hypothalamus.31,37,38 In vascular smooth muscle, Phospholipase C exists in several isoforms that mainly AT1a receptors are found, suggesting that this can be subdivided in three classes, namely PLC-b, subtype plays a role in vasoconstriction. Because the PLC-g, and PLC-d.55 The four b-isoforms are thought AT1b-receptor subtype seems to prevail in the to be activated by subunits of G-proteins, whereas anterior pituitary, adrenal gland, uterus, and several the g isoforms have to be activated by tyrosine phos- 31,38,42,43 periventricular brain areas, this receptor phorylation. AT1 receptor activation leads to the may be involved in hormonal secretion and central tyrosine phosphorylation of PLC-g1 in both rat osmotic control. smooth-muscle and glomerular mesangial cells.56,57 Sequence information obtained after polymerase In smooth-muscle cells it has been shown that, for chain reaction cloning suggests the existence of a this purpose, the AT1 receptor has to recruit the sol- 44 c-src third AT1 (AT1c) subtype in the rat, but like the uble tyrosine kinase pp60 , since the receptor 58 putative AT1-receptor subtype in humans, no itself lacks intrinsic kinase activity. PLC-g1 phos- chromosomal localisation has been identified for the phorylation leads to the PIP2 hydrolysis into 1,4,5- rat AT1c-receptor subtype. The AT1 receptor belongs IP3 and diacylglycerol. The induction of 1,4,5-IP3 to the superfamily of 7-transmembrane domain, G- leads to intracellular calcium release, which in turn protein coupled receptors. AT1-receptor cDNA has induces smooth-muscle cell contraction and other been obtained from a number of tissues, such as signaling events. heart, liver, and placenta. The majority of the open Ang II-induced cell hypertrophy and hyperplasia reading frames encode for 359-amino acid receptors. are both mediated through the AT1 receptor. Protein The single human AT1-receptor gene has a mini- kinase C, as well as elevated intracellular calcium mum size of 60 kb and contains five exons, with the ion (Ca2+) levels, have been shown to promote the entire coding region within the fifth exon.45–47 In rat expression of the growth-related inducible tran- and mouse, the AT1a and AT1b subtypes are highly scription factors (ITFs) such as c-fos,c-myc and c- homologous, and exhibit a 91% similarity on a jun.59 The proteins encoded by the growth-related nucleic acid, and a 96% similarity on an amino ITFs act as transcription factors for various target acid, level. genes, which may be involved in the stimulation of The human AT1- and rat AT1a-receptor nucleic mitogenesis. Ang II also induces, by stimulation of acid and amino acid sequences share an 86% and a its AT1 receptors, the transcription of platelet- ␣ ␣ ␤ 94% homology, respectively. The differences are not derived growth factor- (PDGF- ) chain and TGF- 1 located in a particular region of the receptor, but are and, via this mechanism, is directly coupled to the evenly spread throughout the coding region. The rat expression of growth factors.60,61 AT1a gene has a size of at least 84 kb and is com- Recent reports link Ang II, through the AT1 recep- posed of four exons48–50 with the entire coding tor, to the Jak/STAT signalling pathway.62 This is a sequence located in its last exon. The AT1b gene is significant observation since the Jak/STAT pathway at least 15 kb long, consists of three exons, and enco- has been previously identified for cytokine receptors Ang II receptor pharmacology O Chung et al

77 S13 which leads to transcriptional activation of early et al, who demonstrated that the G protein Gi (but 63 growth-response genes, and is now possibly also not Go) is involved in AT2-receptor by mediated an important signalling pathway through which Ang modulation of K+ channels in rat primary cultures II mediates its proliferative effects.64 Interaction of of neuronal origin. Ang II with the Jak/STAT pathway has been Conversely, Mukoyama et al72 reported that the rat 65 observed in rat aortic smooth muscle cells. Ang II AT2 receptor shares a seven transmembrane domain is able to rapidly phosphorylate tyrosine of the kin- topology that may belong to a unique class of seven ases Jak2 and Tyk2. Stimulation of Jak activity also transmembrane receptors for which G-protein coup- leads to the phosphorylation of both STAT1 and ling has not been demonstrated. In their studies, STAT2 proteins, and subsequent translocation to the stimulation of the cloned AT2 receptor, transiently nucleus of the STAT1 protein. In cultured neonatal expressed in COS-7 cells, failed to increase IP3 or fibroblasts, Bhat et al66 showed that Ang II induces intracellular calcium, and no apparent effects on STAT protein phosphorylation and translocation of cAMP and cGMP levels and phosphotyrosine phos- STAT proteins, leading to initiation of gene tran- phatase activity could be observed. In NG 108–15 scription. In neonatal rat cardiac myocytes cells, cells, which constitutively express AT2 receptors, 54 STAT1 and STAT3 are rapidly phosphorylated in Buisson et al found that AT2 receptor stimulation response to Ang II, and coimmunoprecipitation results in the inhibition of T-type calcium channels 52–54 experiments have shown that the AT1 receptor is through an as yet undefined pathway. In another directly linked to components of the Jak/STAT path- cell line from rat pheochromocytoma, PC12W, 67 way. Also, antibodies against STAT proteins in which only expresses AT2 receptors, Bottari and col- vascular smooth muscle cells inhibit the Ang II leagues60,78 reported that Ang II stimulates a mem- mediated proliferative response.68 A physical associ- brane-associated phosphotyrosine phosphatase and ation between Jak2 kinase and a part of the intra- inhibits ANP-sensitive particulate guanylate cyclase cellular carboxy tail of the AT1 receptor has been via a G-protein-independent pathway. 69 shown. Very recently it has been shown that Ang The relationship between AT2-receptor by II mediates the Jak/STAT pathway under pathologi- mediated signalling and tyrosine phosphoryl- 70 71,78 cal conditions. ation, as well as the fact that the AT2-receptor In contrast to the AT1 receptor, much less is subtype is highly and transiently expressed in foetal known about the structural and functional proper- tissues, followed by a dramatic drop in most organs 79 ties of the AT2 receptor, which has been cloned just after birth, suggests that this receptor plays a recently from the mouse, rat, and human.71,72 In all role in physiologic processes involving cellular three species, the AT2-receptor cDNA encodes for a growth, differentiation, and adhesion. Studies from 363-amino acid receptor, and sequence comparison our laboratory have demonstrated that angiotensin reveals an expectedly high homology on both the peptides can exert an antimitogenic action on rat nucleic and amino acid level (human and rat AT2 and bovine endothelial cells of different origins via receptor cDNA share a homology of 89% and 91% the AT2 receptor, suggesting that Ang II exerts differ- on nucleic acid and amino acid level, respectively). ential growth-modulating actions depending on the Both mouse and human AT2 receptor genes are presence or absence of Ang II-receptor subtypes on 73,74 80–82 located on the x chromosome. The human AT2- a given cell. receptor gene consists of three exons with its entire In further studies in PC12W cells, we also demon- coding region located in the third exon.75,76 strated that Ang II inhibits serum- and epidermal The AT2-receptor cDNA has been isolated from a growth factor-induced proliferation and potentiates number of tissues and from whole rat and mouse the nerve growth factor- and epidermal growth fac- fetuses. Comparing the AT1- and AT2-nucleic acid tor-mediated growth inhibition through its AT2 sequences reveals a lack of homology (only a 34% receptor.83 Our results are supported by recent find- 84 identity). In the adult organism, the AT2 receptor is ings by Nakajima et al who attempted to character- present at high concentrations in the adrenal med- ise the role of the AT2 receptor in the model of neo- ulla, uterus, and ovary, and is also found in vascular intima formation in the balloon-injured rat carotid 14,20 endothelium and in distinct brain areas. The fact artery. In this in vivo gene transfer study, the AT2 that it is expressed at high levels in embryonic receptor was transfected to the injured vessel, and tissues, but to a much lesser extent in normal adult the formation of neointima was studied in the pres- tissues, has raised speculations as to its possible role ence or absence of the AT2 receptor. Morphometric in development and cell differentiation. analysis performed 14 days after balloon injury The molecular structure and signal transduction revealed that myointima size was reduced by 70% pathways of the AT2 receptor are far from being in the presence of the AT2 receptor. This effect could completely understood, and it is still controversial be reversed by the AT2 antagonist, PD 123319, sug- whether or not the AT2 receptor is coupled to G- gesting that the expressed AT2 receptor mediated the proteins, and how it signals. Kambayashi et al71 inhibiting effect on neointima formation. It appears reported that the rat AT2 receptor inhibits a phos- from these studies that, under physiologic con- photyrosine phosphatase in COS-7 cells, kidney ditions, one of the functions of the AT2 receptor in cells from African green monkeys, stably expressing the vasculature consists of the inhibition of angio- the rat AT2 receptor. This effect is dependent on a gensis. pertussis-toxin-sensitive, G-protein-coupled mech- Under pathophysiologic conditions, the expression anism. Further evidence in support of AT2 receptors of the AT2 receptor may be increased to control coupling to G proteins has been provided by Kang excessive growth mediated via the AT1 receptor or Ang II receptor pharmacology O Chung et al S14 via other growth factors. This may be relevant to such as losartan and eprosartan, are strong competi- 85 neointima proliferation, post-myocardial infarc- tive antagonists for Ang II at the AT1 receptor, and, tion adaptation, and left ventricular hypertro- therefore, can be more readily displaced by high 86,87 phy. Recent reports indicate a role for the AT2 concentrations of Ang II at their binding sites. receptor in blood pressure regulation as well. Ang II causes an enhanced depressor response in rats when Losartan AT1 receptors are blocked. This effect is blocked by PD 123177.88 Also, coinfusion of rats with Ang II Losartan is a imidazole-derivative with a biphenyl- and PD 123199 results in an enhanced blood press- tetrazol side chain (Figure 1a), which is considered ure response, as compared to rats infused with Ang to be essential for the Ang II-receptor binding II alone.89 properties.19 Losartan, in a first-pass metabolisation Although the AT2 receptor seems to be involved in the liver, is in part oxidized at the imidazole ring in growth and differentiation, lack of a functional to the active metabolite, EXP3174 (Figure 1b),92 receptor still allows for development of an organ- which substantially contributes to most of the phar- ism, as illustrated by the establishment of AT2 macodynamic and pharmacokinetic properties knockout mice strains. Data obtained from these observed. mice links the AT2 receptor to behaviour and to alt- Losartan has a 10-times lower affinity to the AT1 90,91 ered blood pressure regulation. receptor than does Ang II. Its selectivity for the AT1 receptor is 3000-fold higher than the that for the AT2 19 receptor, with a binding affinity (IC50) to the AT1 Pharmacologic properties of AT1 19 antagonists receptor of 19 nmol/L. Despite the fact that losartan binding to the AT1 receptor is sufficient to result The AT1 receptor antagonists, the ‘sartans’ (Figure in an effective receptor blockade, it is lower when 1a–g), are novel in regard to at least two aspects: compared to some other AT1 antagonists, whose IC50 first, they afford a highly specific and selective values range from 1.5 to 4 nmol/L.21,26,93,94 However, blockade of the angiotensin AT1 receptor subtype the binding affinity of EXP3174 ( IC50 of 3.7 and are void of any intrinsic activity;87 second, they nmol/L),19 is substantially higher, and is within the are, in contrast to saralasin and other earlier range of other AT1 antagonists. The affinity of developed peptide antagonists, based on a nonpep- EXP3174 for the AT1 receptor is 30 000-fold higher 19 tide structure that renders them suitable for oral than that for the AT2 receptor. While losartan itself 87 administration. AT1 antagonists are generally well acts as a pure competitive antagonist, its active tolerated, and have a low profile of side effects, metabolite, EXP3174, also suppresses the effect- which in most studies is not significantly different maximum of Ang II, and therefore displays noncom- from placebo. However, even though all of the sart- petitive properties.95,96 Thus, losartan features com- ans seem to bear overwhelming similarities, it is still petitive as well as non-competitive antagonism. The necessary to scrutinise their chemical background oral bioavailability of losartan is approximately 18,97 and their pharmacologic properties, because subtle 33%. Peak plasma concentrations (Tmax) of losar- differences may have an important clinical impact. tan after oral administration are observed after 1 h, For example, as these molecules were derived from and after 3 to 4 h for EXP3174.18,97 The terminal half- the same initially described imidazole derivatives, life of losartan is 2 h, that of EXP is 6 to 9 h.18,97 Ten most of them feature two heterocyclic structures at hours after oral losartan administration, no signifi- a biphenyl structure. cant plasma concentrations can be measured, while However, some of the more recently developed EXP3174 still reveals measurable concentrations AT1 antagonists display significant differences from after 24 h. Therefore, it is likely that EXP3174, more this concept in their chemical structure: valsartan, than losartan itself, contributes to the 24-h effective- for example, is a tetrazole-biphenyl-valine deriva- ness of losartan. Both losartan and EXP3174 display tive with only one heterocyclic structure at position high binding to plasma proteins (mainly albumin) of 4Ј, and eprosartan lacks the biphenyl structure altog- 98.7% and 99.8%, respectively.98 The distribution ether. While some of the sartans, such as irbesartan, volume for losartan was calculated to be 34 L, that valsartan, eprosartan, and telmisartan can be admin- of EXP3174 is 12 L.97 Approximately 14% of an istered in their active forms, while others, for applied losartan dose is actually converted into example candesartan, are administered as an inac- EXP3174 in the liver.18,97 In addition, inactive oxid- tive prodrug, which then is converted in vivo into ized and glucuronic acid coupled metabolites are the active form. Losartan, which features a some- equally produced. what reduced AT1-receptor blocking property by The metabolism includes the pathway of the P 450 itself, is converted in vivo to its more potent metab- isoenzymes 2C9 and 3A4. Total clearance of losartan olite EXP3174,92 which then appears to be respon- is 600 ml/min, that of EXP3174 is 50 ml/min. The sible for most of the clinical effects. renal clearance of losartan is 75 ml/min, that of EXP The mode of action at the receptor site can also is 25 ml/min. Radioactively labelled losartan is be different among the sartans. Some, as for example excreted to 35–43% in urine and to 55–60% in candesartan, valsartan, irbesartan, and EXP3174, faeces.99 Unmetabolised losartan in urine accounts 99 achieve AT1-receptor blockade with an antagonism for approximately 5% of the administered dose. In that has been designated ‘insurmountable’,21,92,93 healthy volunteers and hypertensive patients, an meaning that they cannot be readily displaced, even uricosuric effect of losartan was observed.100 This by very high concentrations of Ang II, while others, effect might be due to a benzbromarone-structure in Ang II receptor pharmacology O Chung et al S15

Figure 1 Chemical structures of the AT1-receptor antagonists: (a) losartan, (b) losartan metabolite, (c) valsartan, (d) irbesartan, (e) candesartan cilexetil, (f) candesartan and (g) eprosartan.

the losartan molecule, which is also contained in Valsartan substances used for uricosuric therapy. So far, this uricosuric property has only been described for los- Valsartan is a tetrazol-biphenyl-valine derivative artan, but not for EXP3174 or other AT1 antagonists. and features only one heterocyclic structure in its Ang II receptor pharmacology O Chung et al S16 molecule (Figure 1c).20 It displays non-competitive via the P450 isoenzyme 2C9, and that oxidation via antagonism at the AT1 receptor. The Ki for the AT1 the isoenzyme 3A4 plays only a minor role. receptor in smooth-muscle cells is 2.4 nmol/L, the So far, eight metabolites of irbesartan could be one for the AT2 receptor in human myometrium identified, including 6% glucuronide metabolites membranes is 57.7 ␮mol/L, therefore, the selectivity and to 9% COOH-derivatives, which were found in for the AT1 over the AT2 receptor is about 30 000- the plasma. No active metabolite of irbesartan has 21 fold. In human adrenal glands, the IC50 was been described. Binding to plasma proteins is 90– determined to be 2.6 nmol/L, a value comparable to 92%, comparatively low. A dose linearity for the 22 102 the other mentioned AT1 antagonists. After oral AUC has been shown in therapeutic doses. Total administration, valsartan is absorbed quickly, with clearance is 157–176 ml/min, mean renal clearance aTmax at 2 h. Bioavailability is 25%, and is reduced after oral administration is only 3–3.5 ml/min. A by 46% with concomitant food intake.101 Valsartan distribution volume of 53 L has been reported. has a protein binding of 94% to 97%, mostly to albumin.101 The distribution volume was calculated to Candesartan-cilexetil/candesartan be 17 L. Fully 30% of an intravenous dose is excreted unchanged in urine, and about 70% is Candesartan-cilexetil is a prodrug suitable for oral excreted in the faeces. After oral administration, use (Figure 1e). It is rapidly converted to the active 13% of the administered dose appears unchanged in drug, candesartan (Figure 1f), by ester hydrolysis urine, the rest (83%) is excreted via the faeces.101 during absorption from the gastrointestinal tract. So far, in contrast to all other AT1 antagonists, no Candesartan displays high selectivity for AT1 recep- metabolites could be found for valsartan, which tors, with tight binding to and slow dissociation might be due to its unique chemical structure. Val- from the receptor.103 In displacement studies using sartan displays no clear dose linearity for plasma rabbit aortic membranes, the affinity of candesartan concentration, as the area under the plasma concen- to the AT1 receptor was approximately 80 and 10 tration-time curve (AUC) is less than proportional times higher than that of losartan and EXP3174, to the administered dose.101 Total clearance is 36.7 respectively.104 Candesartan displays an ‘insur- ml/min. Renal clearance accounts only for about mountable’ antagonism at the AT1 receptor, in 30% of the total plasma clearance. In patients with experiments using inhibition of Ang II-induced con- hepatic insufficiency, the dosage of valsartan has to traction of rabbit aorta.105 After oral administration, be reduced, as in these patients an increased Cmax candesartan-cilexetil is converted to candesartan, and a two-fold increased AUC have been described. with an average absolute bioavailability of candesartan of approximately 40%. The AUC of candesartan is not significantly affected by food intake.28,106 T Irbesartan max is reached 3–5 h after oral administration. Serum Like losartan, irbesartan is also an imidazole-deriva- concentrations (AUC) displayed dose linearity with tive with a biphenyl-tetrazol-side chain (Figure increasing doses in the therapeutic dose range.107 No 1d).26 In human smooth-muscle cells, irbesartan has sex-related differences in the pharmacokinetics have an about 10 times higher affinity for the AT1 recep- so far been observed. tor, as compared to losartan. The IC50 was measured Candesartan is highly bound to plasma protein in various isolated tissues, for example, aorta, ileum, (Ͼ99%).28,106 Candesartan is mainly eliminated and in cultured cells, and values ranged from 0.8 unchanged via urine and bile, and is only to a minor 26,102 to 4.1 nmol/L. The IC50 for the AT2 receptor is extent inactivated by hepatic metabolism. So far, reported to be 10 ␮mol/L, the selectivity factor for only one inactive metabolite (CV15959) has been the AT1 receptor over the AT2 receptor is, therefore, described. The terminal half-life of candesartan is in the range of 10 000-fold.102 In the rabbit aorta, 5–9 h, and no significant accumulation after mul- irbesartan (1–10 nmol/L) leads to a dose-dependent tiple doses was observed (plasma concentrations blockade of Ang II-induced contractions.102 As these +3% to +17%).107 Total plasma clearance of candes- effects cannot be reversed, even with high concen- artan is about 0.37 ml/min/kg, with a renal clearance trations of Ang II, an insurmountable antagonism of about 0.19 ml/min/kg. After an oral dose of cande- is present. sartan cilexetil, about 20–30% is excreted in urine, Resorption after oral administration is fast. and 60–70%, in faeces. The apparent volume of dis- Bioavailability is 60–80%, and thus higher as that tribution of candesartan is 0.1 L/kg. of most other AT1 antagonists. The terminal half-life after oral application is 11–15 h.102 After intra- Eprosartan venous administration, half-life was calculated as 13 h. Tmax was measured to be in the range of 1–3.5 h, Eprosartan is an imidazole-5-acrylic acid derivative, 102 depending on the dose administered per os. and contains, in contrast to the other AT1 antagon- Approximately 60% to 80% of an orally adminis- ists, no biphenyle structure (Figure 1g). It has a high tered irbesartan dose is excreted via faeces, while affinity to the AT1 receptor with an IC50 of 1.5 about 25% of an orally administered and 23% of an nmol/L (rat mesenteric artery) and 9.2 nmol/L in intravenously administered dose appear in urine. membrane preparations from the adrenal gland.29 However, as irbesartan is strongly metabolised via Eprosartan is already present in its active form, and hepatic glucuronidation and oxidation, only about features competitive antagonism for Ang II at the 29 1% is excreted as the unchanged molecule. In vitro AT1 receptor. studies suggest that irbesartan is mainly oxidized Oral resorption is fast, and maximal plasma con- Ang II receptor pharmacology O Chung et al S17 centrations of eprosartan occur 1–2 h after adminis- 2 Schieffer B et al. Comparative effects of chronic angiot- 30 tration. However, compared to the other AT1 ensin-converting enzyme inhibition and Ang II type 1 antagonists (bioavailability, 25–80%), eprosartan receptor blockade on cardiac remodeling after myocar- features a low bioavailability of only 13%, limited dial infarction in the rat. Circulation 1994; 89: 2273– by a low absorption after oral administration rather 2282. 3 Remuzzi A et al. Comparison of the effects of angioten- than by a high first-pass metabolism. After intra- sin-converting enzyme inhibition and angiotensin II venous administration, 61% is excreted via the receptor blockade on the evolution of spontaneous faeces, and 37%, via urine, indicating biliar and glomerular injury in male MWF/Ztm rats. Exp Nephrol renal excretion. Twenty percent is excreted in the 1996; 4: 19–25. urine as an acylglucuronide metabolite, and 80% is 4 Lafayette RA, Mayer G, Park SK, Meyer TW. Angioten- excreted unchanged. Due to the low absorption after sin II receptor blockade limits glomerular injury in rats oral administration, 90% is excreted via the faeces, with reduced renal mass. J Clin Invest 1992; 90: 766– and only 7% via urine. 771. No active metabolite of eprosartan has so far been 5 Urata H et al. Ang II-forming pathways in normal and described. In young healthy volunteers, the terminal failing human hearts. Circ Res 1990; 66: 883–890. 6 Urata H et al. identification of a highly specific chym- half-life after intravenous administration was about ase as the major angiotensin-II-forming enzyme in the 2 h, and after oral administration, 4.5–9 h. Binding human heart. J Biol Chem 1990; 265: 22348–22357. to plasma proteins is high, and described to be 7 Dzau VJ et al. Human neutrophils release serine pro- between 97.9–98.6% up to plasma concentrations of teases capable of activating prorenin. Circ Res 1987; 10 000 ng/ml, independent of concentration. A dose 60: 595–601. linearity for plasma concentrations has been shown 8 Wintroub BU, Klickstein LB, Watt KW. A human neu- for lower doses, however, a nonlinear relationship trophil dependent pathway for generation of Ang II. is found in higher doses. The distribution volume J Clin Invest 1981; 68: 484–490. is 13 L. Total clearance is 130 ml/min, mean renal 9 Dzau VJ, Sasamura H, Hein L. 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