Drugs of the Future 1993, 18(6): 529-549
`Copyright PROUS SCIENCE
`
`13 3 - A d renergic agonists
`
`Review Article
`
`Ralph Howe
`Research Department, ZENECA Pharmaceuticals, Alderfey
`Park, Macclesfield, Cheshire, SK10 4TG, United Kingdom
`
`CONTENTS
`Introduction (cid:9)
`The adipocyte atypical beta-adrenoceptor (cid:9)
`Distribution of atypical beta-adrenoceptors (cid:9)
`beta3-Adrenoceptor agonists as potential drugs (cid:9)
`Thermogenic drugs for the treatment of obesity
`and diabetes (cid:9)
` 533
`Drugs for the treatment of intestinal hypermotility disorders 542
`Conclusions (cid:9)
` 544
`Acknowledgements (cid:9)
` 544
`References (cid:9)
` 544
`
` 529
` 530
` 531
` 533
`
`Introduction
`Classification of receptor systems which are activated by
`sympathomimetic amines has engaged the attention of
`many investigators.for many years, and continues to do so.
`In 1948, Ahlquist suggested the convenient designations al-
`pha (a) and beta (13) to distinguish major differences in the
`responses elicited in various organ systems by adrenergic
`agents (1). The concept of two kinds of adrenoceptor was
`based on the relative potencies of six sympathomimetic
`'amines to relax the ureters and uterus, contract the nictitat-
`ing membrane, inhibit the gut and stimulate the myocar-
`dium. Thirty years later Ahlquist looked back at adrenocep-
`tors (2) and in 1984 Carrier and Shlafer looked back at the
`man and his work (3).
`"a-Adrenoceptors were generally associated with con-
`traction of smooth muscle. Some blood vessels are con-
`stricted, the radial muscle of iris contracts to produce my-
`driasis and the ureters and splenic capsule are contracted.
`The only organ system that did not fit the scheme that 'all al-
`pha-mediated responses are excitatory' was the gut, which
`was relaxed by what we now call a-agonists.
`The j3-adrenoceptor was generally associated with inhibi-
`tory responses: blood vessels are dilated, bronchial smooth
`muscle is relaxed, and, as was the case with a-agonists, In-
`testinal activity is inhibited. Just as the gut served to demon-
`strate that all a-mediated responses are not excitatory, the
`heart served to demonstrate that not all responses that are
`classified as '13' were of an inhibitory nature" (3).
`In 1967,*Lands et al. obtained results which could not be
`explained easily on the assumption that the 13 type desig-
`nates a single receptor population (4). Their studies showed
`that two distinct receptor subtypes could at that time be in-
`
`eluded within the 13 type, They found a marked similarity be-
`tween the rank order of potencies of fifteen catecholamine
`sympathomimetic amines for lipolysis in white adipose tis-
`sue and cardiac stimulation and also a similarity for bron-
`chodilator and vasodepressor activity. There was a lack of
`correlation for other combinations of the four effects. The re-
`ceptor mediating responses in the heart and lipolysis was
`deeignated 131 and that mediating vasodepressor activity
`and bronchodilation was labelled 132. Further studies based
`on the potencies of sympathomimetic amines classified the
`receptor responsible for inhibition in the rabbit small intes-
`tine as Pi and that In the rat uterus and diaphragm muscle
`as 132 (5).
`*While pharmacologists were engaged in the classification
`of adrenoceptors, medicinal chemists had been producing
`compounds which would act as agonists or antagonists of
`the various adrenoceptor types. Some of these agents were
`non-selective for the adrenoceptor subtypes but others had
`varying degrees of selectivity fora particular subtype. Ex-
`amples are isoprenaline, a selective agonist for 13-adreno-
`ceptors rather than a-adrenoceptors but non-selective for
`either Pr or132- adrenoceptors (6), and propranolol, a selec-
`tive antagonist for 13- rather than a-adrenoceptors but
`non-selective for either Pr or 132-adrenoceptors (7). Exam-
`ples of agents with selectivity for 131- rather than 132-adreno-
`ceptors are the agonist prenalterol (8, 9) and the antagonist
`atenolol (10); agents with selectivity for 132: rather than
`131-adrenoceptors are the agonist salbutamol (11) and the
`'antagonist ICI 118551 (12, 13). Such agents were key to
`confirming or questioning the classification and to extending
`it. In particular, the classification of the white adipose tissue
`13-adrenoceptor as the 131 subtype has been questioned by
`many groups and this is discussed in the next section.
`Arch has stressed the serious limitations of classifying re-
`ceptors solely on the basis of rank order of potencies of ago-
`nists (14). The approach can be criticized if only a limited
`range of agonists is used and it can be misleading if some
`of the agonists have a lower efficacy than others and abso-
`lute responses are compared. Rather than use agonists,
`whose efficacies may differ, pharmacologists now prefer to
`classify receptors using antagonists, which have zero effica-
`cy. It Is necessary to use agonists selective for each receptor
`
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`530 (cid:9)
`
`P-Adrenergic agonists
`
`subtype in combination with antagonists of various selectivi-
`ties. A novel receptor type can onfy be detected using both
`an agonist that acts, at legst in part, via this receptor and an
`antagonist that has a different affinity for the novel receptor
`compared with known receptors. He pointed out that most
`studies fall far short of these stringent criteria.
`
`. The adipocyte atypical 13-adrenocepter
`
`The classification of the lipolytic 13-adrenoceptor as a
`subtype was questioned by Harms et al. in 1974 (15, 16),
`who showed that antagonism of lipolysis could correlate
`with either fl radrenoceptor antagonism or132-adrenoceptor
`antagonism depending upon the structural type of the an-
`tagonist used. They proposed a hybrid receptor having
`characteristics of both the 131- and 132-adrenoceptor sub-
`types. Later studies In Which isoprenaline was used as the
`13-adrenoceptor agonist and two series of analogs,of tole-
`molol as antagonists supported the idea of a dualistic 13 and
`- 132 Character of the rat adipocyte fl-adrenoceptor which
`)could not be explained by the presence of two different pop-
`- .ulations of pi- ahdl3i-adrenoceptors. Further, the rat 'adipo-
`cyte13-adrenoceptor population was shown to be homoge-
`neous (17). These views on the hybrid nature of the
`13-adrenoceptor of the white adipose cell•of the rat were sup-
`ported by Tan and Curtis-Prior who, in 1983, proposed that
`It be termed a 'beta-hybrid' or '133' adrenoceptor (18).
`The hypothesis of a new- and discrete P-adrenoceptor
`subtype (133) remained tenuous in the-absence of agonists
`or antagonists selective for it. In 1984, however, Wilson et
`al. of Beecham Pharmaceuticals reported on the activities
`of three novel arylethanolamine 13-adrenoceptor agonists,
`BRL 28410, BRL 35113 and BRL 35135*, which showed se-
`lectivity for rat white adipose tissue lipolysis, supporting the
`view that the rat lipolytic 13-adrenoceptor is atypical, that is,
`it is distinct from either the 131- or 132-adrenoceptor subtype
`(19). Appropriate studies were carried out using available
`13-adrenoceptor antagonists; there was no antagonist with
`selectivity for the atypical 13-adrenoceptor. Wilson et al.
`•oointed out that the hybrid 111/132 receptor proposed by
`.harms eta!, and De Vente et al. adequately explained the
`results obtained by those authors but that the model did not
`readily explain their own result's. It is difficult to envisage a
`lipolytic receptor with 13i- and, 132-adrenoceptor characteris-
`tics when, for example, BRL 35113 is a poor agonist at both
`pi- and132-adrenoceptors yet is a potent agonist at the lipo-
`lytic 13-adrenoceptor.
`Concurrent studies by Arch et al. of the Beecham Group
`(20), who used BRL 28410, BRL 35113 and BRL 37344 (the
`carboxylic acid related to the methyl ester BRL 35135) in a
`
`• BRL compounds, including BRL 35135, occasionally have the
`suffix 'A' after the number. It is understood that this simply indi-
`cates that the substance is a salt and so the 'A' is omitted in this
`review. The pharmacological properties of these compounds and
`the clinical results on two of them are considered in a later section.
`Chemical structures are also shown in a later section.
`
`study of lipolysis in rat interscapular brown adipocytes,
`showed that in the rat the brown adipocyte receptor is nei-
`ther ,a or nor a 132-adrenoceptor and that brown and white
`adipocytes "have similar, though not necessarily identical,"
`13-adrenoceptors. The most potent compound, BRL 37344,
`stimulated lipolysis with 400- and 20-fold selectivity com-
`pared with atria1131 ancitrachea1132 responses; respectively.
`Stock and Sudera studied rat brown adipocyte respiration,
`rather than lipolysia, using isoprenalina, BRL 37344 and ICI
`201651 as agonists • and p'ropranolol, atenolol and ICI
`118551 as antagonists, and concluded that the interaction
`of the novel agonists with brown adipocyte13-adrenoceptors
`differed from that of isoprenaline, giving support to the work
`.
`of the Beecham group (21). (cid:9)
`Receptor binding studies using conventional13-adrenerg-
`ic agents had failécl to reveal the atypical13-adrenoceptor in
`brown adipocytes. Most studies suggested that the brown
`adipocyte receptor is of the 131-subtyPei but two groirps had
`reported13,-: 132-adrenoceptór populations of 59:41 (22) and
`80:20 (23) for rat brown adipose tissue. Arch (14) haS re-
`viewed the binding studies and has pointed to the problems
`that affect them; First, the receptors stridied by binding
`methods may not be the ones that mediate the functional re-
`sponse. Second, just as functional studies using antago-
`nists can only detect receptors through which the ligand.
`acts, binding studies cannot detect receptors that do not
`bind the labelled ligand. Detection of the atypical receptor by
`binding studies requires the use of a labelled ligand that
`binds selectively to the atypical receptor. Recently, Hollenga
`and Zaagsma have investigated the effects of the selective
`antagonists CGP 20712A (131) and ICI 118551 (32) on BRL
`37344- and isoprenaline-induced lipolysis in rat white adipo-
`cytes (24). Their results show that the selective 133-agonist
`BRL 37344 acts solely through atypical 13-adrenoceptors,
`whereas isoprenaline, a non-selective 13-agonist, acts pre-
`dominantly through atypical 13-adrenoceptors; Pradreno-
`ceptors, detected in binding studies, play at most a small
`and subordinate functional role in isoprenaline-induced li-
`polysis. Similarly, adenyl cyClase activation in rat adipocytes
`•by BRL 37344 is solely mediated by atypical (133 subtype) re-
`ceptors and isoprenaline is predominantly mediated by
`atypical (33 subtype) receptors (25).
`An important contribution was made by Emorine et al. in
`1989 when they reported the isolation of a gene which coded
`for the human 133-adrenoceptor (26). A human genomic
`DNA library was screened with the entire coding regions of
`the genes for the turkey 131- and the human 132-adrenocep-
`tors. Sequences complementary to both receptor probes
`were found in fragments from 14 clones, one of which was
`entirely sequenced and shown to contain a gene coding for
`a polypeptide with an amino acid sequence 51% and 46%
`identical to those of 131- and 132-adrenoceptors, respectively.
`The receptor protein, which the authors called '133-adreno-
`ceptor', had clear atypical 13-adrenoceptor properties when
`expressed in Chinese hamster ovary cells (CHO), which
`normally contain no 13-adrenoceptor activity (26, 27). The
`CHO-133 cells were analyzed for ligand binding and cAMP
`production and compared with the • similarly prepared
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`Drugs Fut 1993, 18(6)
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`531
`
`CH0-131 and CHID-132 cells (28). They used P-adrenergic li-
`gands known to interact selectively with p,, 132 or atypical
`receptors and confirmed 'that the human 133-adrenoceptor,
`when expressed in CHO cells, is indeed related to the atypi-
`cal P-adrenoceptors Charebterized in rodent'adipoie tissue.
`The potency orderof 13-adrenergic agonists for the 03-adre-
`noceptor was clearly different from that for the Pradreno-
`ceptor and for the 112-edrenoberitói and reseMbled that for
`lipolysis stimulation in rodent brown adipose tissue. Also,
`classical 13-antagonists displayed very low affinity for the
`N-adrenoceptor expressed in CHO cells. However, differ-
`ences between the cloned human P3-adrenoceptor and ro-
`dent atypical adrenoceptors led Zaagsma and Nahorski to
`question whether they.were, in fact; homologous proteins
`(29). Also, experiments with the cloned yet N-adrenoceptor
`(30, 31) indicate that its pharmacological properties are vir-
`tually identical to those of the atypical adrenoceptor in rat
`brown fat but differ in eeyerai respects from those reported
`for the oforiéd hutilen*03.2atIOèn'cibiiptór (26-.24.,;; (cid:9)
`•-•
`Récentlj/„Liggett (32)hPS expressed the rat and the hu-
`. (cid:9).. -
`man P3-adrenoceptor in CHO cells and has determined key
`pharmacological properties in parallel studies. Typical cate-
`cholamine agonists were fund to have a similar low affinity
`for both rat and human N-adrenoceptors. In contrast to
`catecholamine agonists, differences In agonist efficacy and/
`or potency were noted for each of the non-catecholamine
`atypical agonists tested. .For example, BRL 37344* was a
`full agonist (intrinsic activity 1.0 relative.to isoprenaline) Ion
`the rat 133-adrenóceplor, but its intrinsiO activity was i7nly:
`0.60 for the human 133:adrenoceptor; elk, it as About 15
`times less potent ter the 'fnitnin than for thé fat 1334dreno-
`ceptor. The data indicate that the molecular actions of atypi-
`cal (3-adrenoceptor agonists differ markedly between spe-
`cies and so the action of atypical agonists at rodent
`133-adrenoceptors may not be predictive of therapeutic po-
`tential in humans.
`A feature of brown adipose tissue is that the response of
`the tissue to (33-agonists ihcreases on chrohie agoniat expo-.
`sure (33,34) in contrast to tissues containing predominantly
`f31- or f32-adrenobeptors which, become desensitized (35;
`36). In line with this is the recent finding that whereas both
`13i- and 112-adrenoceptOr subtypes Undergo agonist-depen-
`dent decreases in receptor expression during long-term ag-
`onist exposure, cells expressing the 133-adrenoceptor sub-
`type fail to undergo a decrease in receptor number after
`prolonged agonist exposure and, in fact, display increases
`in number over time (37). However, Revelli etal. have dem-
`onstrated that administration of the thermogianic P-adren-
`ergic agonist Ro 16-8714 (see. later) to lean and' obese
`Zucker rats induces a marked down-regulation of the
`N-adrenoceptor of the interscapular brown adipose tissue
`over a time course of 72 hours. It was proposed that the
`down-regulation of the 03-adrenoceptor observed in this
`
`.*It would appear that BRL 34377 was reported in error when BRL
`37344 was meant.
`
`study 72 hours after beginning treatment with Ro 16-8714
`is compensated in the whole animal by the trophic effect of
`the drug on the brown adipode tissue (38).
`
`Distribution of atypical f3-adrenoceptors
`
`It is now generally accepted that the (3-adrenoceptor me-
`diating lipolysis in rat white adipose tissue and oxygen con-
`sumption in brown adipose tissue is of neither the p, nor the
`52 subtype, that it is atypical, and it is being referred to.as the
`133-adrenoceptor. It is characterized by a marked respon-
`siveness to atypical 137agbnists like BRL 37344 compared
`with classical pi- and P2-agonists, and by having a low affin-
`ity for propranolol and standard Pi- and 32-antagonists:.
`In this section,' atypical P-adrenoceptors in other tissues'
`are reviewed and inevitably the question is raised as to
`whether they should be designated P3-adrenoceptors. It had
`been suggested earlier (39)' that the p, and (32 classification
`of Lands may represent two extremes ofa.varlablespec-
`trurn of different "isorepeptors", in the same way as specific./ )
`enzymes can exist in isoenzymic forms. The other atypical '
`13-adrenoceptors may be similar but not :identical to.those
`designated 03. Some authors have expressed caution at us-
`ing the specific nomenclature' J33-adrenoceptor because of
`the complexities, involved in making the assignment (40).
`On the other hand Arch (14) has suggested that this group
`of atypical .PredrenoceptorS,:having the characteristics de-
`scribed above,, will obtain their due recognition only if, they
`are described as 133-adrenoceptors. In this review the origi-
`nal authors' nomenclature of "atypical" or 133" is used. How-
`ever, the present author is clearly following Arch in grouping
`together atypical P-adrenoceptors, that is, those which do
`not fit into either the 131 or the 132 classification. Some further
`subdivision may prove necessary in the future. It is recom-
`mended that in each case the data on which the assignment
`was made be examined. The situation is complicated by the
`co-occurrence of 13.1-, 132- and atypical adrenoceptors in cer-
`tain tissues, by species differences, and, as mentioned ear-
`lier by the agonists used perhapS having different efficacies
`in different tissueS. MacKay (41) has indicated the problems( )
`that can arise in the determination of PA2 Values of antago-
`nists, and in making comparisons of values obtained by dif-
`ferent 'workers, while Jenkinson (42) his pointed out that
`there is still some confusion over terms and terminology.
`The'presence of an undefined adranobeptOr which func-
`tions to inhibit histamine-induced longitudinal muscle ten-
`sion development in the guinea pig ileum had been evident
`from the failure of propranolol to block the action of p-adre-
`noceptbr agonists. The question arose as to whether the
`adrenoceptor in the guinea pig ileum was the same as the
`P3-adrenoceptor on adipocytes. Bond and Clarke (40)
`showed that the order and relative potency of agonists at the
`ileum receptor was BRL 37344 (20) >(-)- isoprenaline (8) >
`noradrenaline (1) > adrenaline (0.5) > fenoterol (0.35) >
`(+)-isoprenaline (0.27), similar to the more restricted series
`examined by Arch et al. (20) for lipolysis in brown fat, BRL
`37344 (1) > isoprenaline (0.2) > fenoterol (0.008) and bý
`
`
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`(cid:9)
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`532 (cid:9)
`
`3-Adrenergic agonists
`
`Wilson etal. (19) for lipolysisin whitept, BRL 35135 (which
`• is hydrolyzed to the active free acid BRL 37344) (1) = isopre-
`naline (1) > fenoterol (0.03). Further, the guinea pig ileum
`adrenoceptor was totally resistant to propranolol at up to 10
`p.M. Bond and Clarke refer to the guinea pig Ileum p-adreno-
`ceptor as atypical (see their expressed caution above, refer-
`ence 40) rather than fla. One important piece *of evidence
`which argues against the notion that the adipocyte and the
`Ileal receptor are the same comes from data obtained with
`propranolol. The atypical 3-adrenoceptor on fat cells is sen-
`sitive to blockade by propranolol but abnormally low pA2val-
`ues (6.2-6.6) have been reported. In contrast, the adreno-
`ceptor on guinea pig ileum is resistant to propranolol at
`concentrations that are at least 8-16 times its equilibrium
`dissociation constant for the adipocyte receptor. However,
`Bond and Clarke point out that it would be 'Premature to dis-
`criminate tile two recéptore solely wen the basis of the pro-
`pranolol result.. Blue et al. have further characterized the
`atypical13.-adrenoceptor in guinea pig ileum by showing that
`while it is resistant to antagonism by propranolol, it iš corn-
`.,_oetitively antagonized by (-)-alprenolol and (-)-dih.ydroal-
`• .`enolol with pA2 values of 6.47 and 6.43, respectively.
`nese values are much lower than those, 8.2 and 8.81, re-
`Ppectively, for antagonism at the Pradrenoceptor. (-)-Alpre-
`nolol also exerted agonist activity at the atypical 3-adreno-
`ceptor in guinea pig ileum (43). Further, in the presence of
`sufficient prcipranolol (51.1.M)to saturate 01-actrenoceptors,
`cyanopindolol gave a pA2 value of 7.63. Thus, cyanopindo-
`Id is the most potenfantagonist so far identified for the atypi-
`_ (cid:9)
`.
`
`cal P-acIFenciceptór and, in the pie. aerie-et:if PripPrario161, may
`serve as a useful probe for studies at the atypical P-adreno-
`ceptor (44).
`In rat gastric fundus, the resistance of P-adrenoceptor re-
`laxant responses to propranolol and the antagonism by cya-
`nopindolol of BRL 37344- and isoprenaline-induced re-
`sponses led McLaughlin and MacDonald to conclude that
`atypical p-adrenoceptors were present (45). The relatively
`low potency of BRL 37344 at this site compared with other
`atypical p-adrenoceptor sites may be due to differences in •
`coupling efficiericy or may indicate receptor heterogeneity.
`In guinea pig gastric fundus the relatively high potency of
`7L 35135, together with the relatively weak antagonism of
`fi-"adrenoceptor agonists by propranolol, support the pres-
`ence of atypical 3-adrenoceptors similar to those in rat adi-
`pocytes (46, 47).
`Evidence for an atypical, or 33-adrenoceptor, in ferret tra-
`cheal epitheliuni has been 'reported by Webber and Stock '
`(48). BRL 37344 was more potent by 4-5 orders of magni-
`tude than the f31-(prenalterol) and P2-(salbutamol) adreno-
`ceptor selective agonists in stimulating methacholine-in-
`' duced albumin transport. The weak antagonism óf the
`response to BRL 37344 by ICI 118551 was consistent with
`this response being mediated by an atypical 13-adrenocep-
`tor. The use of BRL 37344 has shown the presence of atypi-
`cal P-adrenoceptors on rat jejunum (49). Several (31- or
`132-agonists and antagonists were tested and it was found
`that non-selective • and selective antagonists for' Pr or
`Pradrenoceptors showed a relatively low affinity, cempared
`
`to their affinity for Pi- or Pradrenoceptors. BRL 37344 was
`more potent, although a partial agonist (60% maximal re-
`sponse), compared to isoprenaline; whereas it was clearly
`less potent than isoprenaline on pi- or 32-adrenoceptors.
`BRL 37344 and BRL 35135 have been used to provide ev-
`idence that an atypical P-adrenoceptor is present in guinea
`pig bronchi (50). BRL 35135 reduced non-adrenergic
`non-cholinergic contractions induced by electrical field stim-
`ulation and this inhibitory effect of BRL 35135 was not signif-
`icantly altered by non-selective or PI-selective antagonists.
`While BRL 35135 inhibited the contractile response to elec-
`trical field stimulation, it did not inhibit the contractile re-
`sponse to exogenous substance P, which led to the sugges-
`tion that it elicits its inhibitor/ effects pre-junctionally on
`neuronal terminals rather than-post-junctionally on the ta-
`chykinin receptors of airway smooth muscles.
`Just as the Beecham ..BRL. compounds have contributed
`to the identification of, atypical P-adrenoceptors, so has a
`group of phenylethanolaminotetralines reported by the Sa-
`nofi Group, e.g. SR 58306A (51), SR' 58539B (52) and SR
`58611A (53). In vitro, SR 58306A, 'unlike isoprenaline and
`the 32-selective agonists saibutarnol and ritodrine, potently
`inhibited rat colon spontaneous contraction at concentra-
`tions substantially lower than those only partially relaxing
`the guinea pig trachea; also, it had no chronotropic action on
`guinea pig atria. 3-Selective agonists were used as compa-
`rators because they had shown proniise for treating condi-
`tions of abnormally enhanced gastrointestinal motility; they
`could not be used therapeutically because of their concur-,
`rent cardiovascular effects. Non-selective -a renergic an-
`tagonists competitively antagonized the action of SR
`58306A on the colon, which was not prevented by selective
`pi- or 32-antagonists. Only alprenolol competitively antago-
`nized the action of isoprenalindon the.colon; antagonism by
`either pindolol or propranolol was non-competitive. The re-
`'suits suggested that compounds such as SR 58306A inhibit
`colonic motility by selectively stimulating atypical p-adreno-
`ceptors, while isoprenaline interacts also with (31- and
`132-adrenoceptors which coexist with the atypical P-adreno-
`ceptor in rat colon. In vivo, the minimal effective i.v. dose of
`SF1'58539B for raising heart rate was five times its ED50 for
`Inhibition of colon motility; at this multiple of ED 50 it caused
`a rise in blood pressure. SR 58611A was the most selective
`potent compound of those studied in vitro on rat colon, rat
`uterus and guinea pig atrium.
`The phenylethanolaminotetralines were tested for their
`ability to induce lipolysis in rat white adipocyles (54, 55).
`There was a good correlation (r = 0.97) between the lipolytic
`EC50s of (-)- and (+)-isoprenaline and several phenyletha-
`nolaminotetralines and their inhibition of rat colon motility,
`suggesting that receptors of the same type, may account for
`their actions on both preparations. McLaughlin and MacDo-
`nald (56) investigated the responses to noradrenaline, iso-
`prenaline and BRL 37344 in the rat distal colon in vitro and
`provided a further link between the effects of BRL 37344 and
`the Sanofi compounds. Relaxant responses to BRL 37344
`were only weakly antagonized by propranolol. Responses
`to isoprenaline, resistant to propranolol antagonism, were
`antagonized in the presence of propranolol by cyanopindo-
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`
`33-A
`
`Tt
`33-a
`(20)
`infla
`they
`Pate
`may
`glau
`• perc
`
`Sawai Ex. 1004
`Page 4 of 21
`
`

`

`Drugs Fut 1993, 18(6)
`
`533
`
`lol (pA2 7.12), which had earlier been suggested as a probe
`for atypical 0-adrenoceptors. (44). Manara et al. (57) have
`compared inter alla SR 58539B with BRL 37344 and shown
`that the former has the greater rat colon to rat uterus or guin-
`ea pig atrium selectivitY. The same group has carried out
`binding assays with [31-1]-dihYdroalprenolol and has failed to
`identify sites cOrrespcindind to the atypical P-adrenoceptors
`clearly evidenced by theirfunctional studies in rat Colon (58).
`The identification of a radioligand with sufficiently high affin-
`ity for atypical receptors is of paramount importance.
`An atypical P-adrerrocé0tor in rat esophagus has been
`characterized using agonist and antagonist probes (59, 60).
`The order and relativity of agonist potency was BRL 37344
`(36) > (-)-isoprenaline (7) > SR 58611A (5) > (-)-adrenaline •
`(2) > (-)- noradrenaline (1)>•(+)-iioprénaline (0.2) > ferioter-'
`ol(0.02). The receptcir site in rat eiephagüi exhibited an'un-
`usually low 'affinity ter Propren-oloi cOMPared with that nor-
`mally obtained at cladsical P-adrenoCeptbrd. The failure of
`cyanopindolol to antagonize (-)-isoprénalina suggested that
`.
`.. (cid:9)
`the site;may differ frcirn PtherItyPicel -ad
`Both SR 58611A and BRL 37344 stimulated acid séCre:
`tion in rat sternach in vitrò (61). The response to SR 58611A
`was reduced by pr'Oprarrolol but riot by ß1- Or N-Selective
`antagon fits. The lack 6f effect of alprenolol was considered
`surprising:Thid eigéiii had togen" piit'ted earlier to be Partic-
`ularly effective (but not 'selective for) atypical P-adrenocep-
`tors (43). In a siMilar, type of •Study, the samegroup showed
`that stimulation of biCarbonate Secretion in cecUrn In vitro
`is mediated by atYpical f3'adri3n6Ceritors (62). In this case
`the response t6 SR 68611A WaS reduced bY alprenolol but
`•
`.
`• not by Proprenolol, indicatingffiet the atypicel13"-dretiócep- .
`tors mediating the two secretory processes, although simi-
`lar, may not be identical.
`SR 58611A has been reported to have an antidepres-
`sant-like effect in rodents (63). The effect was not antago-
`nized by selective (cid:9)
`a-Afib- Onisii, but
`was blocked by high doses of proPranolol and alprenolol,
`suggesting the presence of atypical P-adrenoceptors in ro-
`dent brain.
`Atypical P-adrenoceptors may also 'occur in rat liver (26),
`.in rat soleus muscle (64), In the heart (65), in the circular
`smooth muscle layers of the human colon (66) and in the
`dog hind limb (67). The antinocicéptive action of several
`fš-adrenoceptor agonists in the' mouse abdominal constric-
`tion test has been attributed to stimulation of atypical
`13-adrenoceptors (68).
`
`•
`
`j33-Adrinoceptor agonistd as potential drugs
`
`Therapeutic areas where it has been suggested that
`133-adrenoceptor agonists may provide drugs are obesity
`(20), diabetes (69), intestinal hypermotility disorders (54),
`inflammatory airways disease (48) and depression .(63);
`they may. also provide locally acting analgesic drugs (68).
`Patent applications suggest that 33-adrenergic agonists
`may be used in the treatment of ocular hypertension and
`glaircoma (70), hypertension (71), hypertriglyceridemia, hy-
`percholesterolemia and atherosclerosis (72), and may also
`
`be used as topical antiinflammatory agents and platelet ag-
`gregation Inhibitors (73). Work in the first three of these
`areas has yielded novel compounds with selectivity for the
`133-adreneceptor which have not only been used as tools to
`help characterize thei33-adrenoceptor, as discussed above,
`but have been considered worthy of clinical investigation.
`None is as yet generally available for hunian use. Thé plan
`is now to discuss these compoundd principally under the
`heading of the therapeutic area in which they originated.
`
`Thermogenid drugs for the treatment of obesity and
`diabetes
`
`A thermogenic function Was first ascribed to brown adjs.,
`pose tiésue (BAT) in 1963 (74). BAT appeers to be present
`in most, .if .nol all 'mamrrials, but rarely accOunts for r+tore
`than 2% of body weight (7). Initially its heat Producing func-
`tion was believed to be of importance only in certain species
`as part of the process . of non-shivering thermogenesis
`(NST) during arousal from hibernation, during acclimation
`to cold or duririgthe neonatal period (76-78): It was not until
`1978 that BAT was established as the major but not neces-
`sarily the. exclusive : site of . NST in rodents- .(79-81):. In
`cold-adapted rats it was shown that during maximal nora-
`drenaline stimulation the blood flow to BAT is'equiyaleht to
`one-third of the cardiac output and, from measurements of
`oxygen extraction by BAT, it was shown that this tissue Could
`account for over 60% 6f NST. Cold-adepted rate have an in-
`creased turnover and urinary exCretion 'of' nóradrenaline,
`show an enhanced thermogenic reSponse to catechola-
`Mines, and hypertrophy and hyperplasia of • BAT is ob-
`served. The high metabolic rate of these animals can be In-
`hibited by I3-adrenergic blockade (82, 83). NST has been
`observed in larger adult mammals, including man (84).
`Non-shivering heat en:Ai:lotion depends on the oxidation
`of prirriarily fettY acidd bý BAT: The Capacity of the tissue for
`thermogenesls is dependent upon a 'proton conductance'
`pathway. which is associated with an inner mitochondria'
`membrane 'uncoupling' protein unique to brown fat. Ther-
`mogenesis in BAT is initiated by the sympathetic.rel ease of
`noradrenaline which acts predominantly via j3-adrenocep-
`tors, now designated 133-adrenoceptors, to cause an activa-
`tion of adenyl cyclase which, by increasing the concentra-
`tion of cAMP, stimulates a hormone sensitive lipase thereby
`releasing free fatty acids. These act both as the substrate for
`13-oxidation and to override the control exerted on respire- .
`• tion by purine nucleotides binding to the GDP.- binding pro-
`tein thermogenin (uncoupling protein), thereby uncoupling
`ATP synthesis from respiration.
`Uncoupling protein has a high affinity for purine nucleo-
`tides and the specific binding of GDP is used to Monitor the
`activity of the proton conductance pathway. In rats, where
`the activity of the BAT mitochondrial proton conductance
`pathway is low, a proportion of the uncoupling protein is
`'masked' and unable to bind GDP. However, on activation of
`the proton conductance pathway, GDP-binding sites are
`'unmasked'. Therefore, GDP-binding In isolated mitochon-
`dria reflects the thermogenic status of BAT (85).
`A phenomenon which shares many of the features of
`cold-induced NST is diet-induced thermogéneSis (DIT),
`
`Sawai Ex. 1004
`Page 5 of 21
`
`

`

`. (cid:9)
`
`534 (cid:9)
`
`13-Adrencirgi