HOE 140, a new highly potent and long-acting bradykinin antagonist in conscious rats

Europm~ Jourm7/ up P/rcwnIaco/o~. XI) ( IO91) 17Y- IX’ P 1991 Elsevier Science Put&hers B.V. tH)l~-2999/91/$:)3.j0 ADONIS

0014?Y99Y1005’78

EJP 20857

Short communication

Gang Bao, Fatimunnisa Qadri, Bernhard Stauss, Harald Stauss, Peter Gohlke and Thomas Unger Departrrrettt of Plrarnracology. Utrirersiiy of Heidelberg and GermatI brsrirure for High Blood Pressare Research. Im Neuenheimer Fe/d 366. D-6900

Heidelberg, F. R.G.

Received 29 April 1991. accepted 14 May 1991

The

inhibitory

depressor antagonist

effects

new bradykinin bradykinin

antagonist

were

that this compound

half-life

with catecholamine

140 (D-Arg-Arg-Pro-Hyp-Gly-Thi-Ser-D-Tic-Oic-Arg)

in conscious rats and compared HOE

than B4146. Plasma catecholamines

did not interfere

B,-receptor

HOE

investigated

B4146 (D-Arg-Hyp-Pro-Gly-Thi-Scr-D-Pro-Thi-Arg).

much longer biological bradykinin

of the

responses to exogenous

140 showed a 250-700-fold

higher

were not increased after application

release. HOE

on

with those of the bradykinin

potency in vivo and a of HOE

140, indicating

140 proved to bc a highly potent. specific and long-acting

antagonist. Bradykinin;

Bradykinin

antagonists;

1. Introduction Since the development of the first specific bradykinin &receptors antagonist (Vavrek and Stewart, 198.51, a number of further bradykinin antagonists have been synthesized and bioassayed. However, the use of these bradykinin antagonists has generally been limited by their relatively low potency and their short biological half-life. Moreover, some bradykinin antagonists increase plasma satecholamines, possibly by a partial bradykinin-like agonist effect on the adrenals (Mulinari et al., 1988). A recently presented bradykinin antagonist (CH 2-D-Arg-Arg-Pro-Hyp-Gly-Thi-Ser-D-PheThi-Arg) (L.ammek et al., 1990) has a high potency and does not interact with catecholamine release. However. its duration of action is still very short. Another new bradykinin Bz-receptor antagonist, HOE 140 (D-Arg-Arg-Pro-Hyp-Gly-Thi-Ser-D-TicOic-Arg) has recently been developed (Breipoh! ct al., i990). it has two artificial amino acid residues at position 7 (D-Tic: 1,2,3,4-tetrahydroisoquinoline-3carboxylic acid) and at position 8 (Oic: [3aS,?aS]-octahydroindol-2-carboxylic acid) instead of D-Phe and Thi. This change has increased not only potency and

Correspondence IO: T. Unger. Department Pharmacology. University of Heidelberg. Im Neuenheimer Feld 366, D-h9OO Heidelberg. F.R.G. Tel. ~9.h2.2l.Sh.39.12/0I: fax 49.h2.21.56.39.-l4.

Plasma catecholamines;

(Conscious rat)

specificity but also the duration of inhibition of the effects of bradykinin in vitro and in anesthetized rats (Hock et al., 1991, Wirth et al., 1991). In the present study we investigated the inhibitory potency and the in vivo half-life of HOE 140 in conscious rats in comparison with B4146 (D-Arg-Hyp-ProGly-Thi-Ser-D-Pro-Thi-Arg), a frequently used representative of the bradykinin antagonists developed by Vavrek and Stewart. In addition we investigated whether plasma catecholamines were influenced by treatment with HOE 140.

2. Materials and methods Male Wistar rats (Dr. K. Thomae GmbH. Biberach. Germany) weighing 250 + 25 g were used for the experiments. Z)ne day before experiments all rats were instrumented with catheters p!aced in the abdominal aorta (PP-IO in PP-50, Portex Corp., Hythe, Kent, UK) via the right femoral artery for recording mean arterial pressure (MAP). Two additional catheters (PP-IO in PP-50) were introduced into the descending aorta via the left carotid artery for application of bradykinin and bradykinin antagonists. respecti-<‘s!y, Catheters were brought to the exterior at the nape of the neck. MAP was m~25urec! directly by use of a Statham P23Db pressure transducer, amplified by a Gould Brush pressure computer (both Gould Inc., Oxnard, Calif., USA).

(II = 8) as ;I

BII-lh II0 (11 = S) and 220 pg/min i.a. infusion. Thcsc doses inhibited s,p~m,c.‘s to csogcnous calcufatcd

bradykinin

by ahout

from the dose-rcsponsc

cspcriment

enc.

Dcprcssor

5-min

the dcprcssor

rc-

YW.

as

curves obtained

in

responses to i.a. injected

bradykinin wcrc rccordcd prior to and 5. IO and I5 min after B-II-W and HOE IJO application. Rats treated with

HOE

1-W rcccived

tions at ISmin

additional up to

intcwals

hradykinin

injec-

YO min after bradykinin

application.

anti~gOlliSt

In cspcriment

three.

plaxma catccholamincs an i.a. bolus

the effect

of HOE

\\‘as investigated.

injection

of either

(n = 5) or 3.2 pg HOE by 0.1 ml O.Yci NKI.

140 on

Rats received

0.3 ml O.Y%

NaCl

I40 (n = 6) in 0.1 ml followed

Five minutes after the injections,

I ml blood \vas collected from the aorta via a catheter in the carotid artery. The blood samples were centrifuged for 20 min at 2.500 rpm and 4°C. The plasma \vas scparatcd The antagonistic potcncics of the brudykinin nists arc csprc
antago-

doacs (ED,,,

and

ED,,) of the bradykinin antagonists that inhibited 110 snd WC; of the dcprcssoi rcsr >nscs to i.;t. injections of 250 ng hradukinin. On the &rg of the sspcrimcnt.

the rats \vcrc

at-

tached to the monitoring cyuipmsnt. Espcrimcnts \vcrc started iIfttX 3 stahifization period of 60 min. In cspcrimcnt one. bradykinin \\as injected t\\icc at 5-min intcrvals before

bradykinin

antagonist

treatment.

and the

average of the t~va deprcsbor responses tcr bradykinin was used as a control ~afuc.

chota~lincs

and stored

at

were dctcrmined

Plasma

cate-

by high-performance

-80’

C.

tiq-

uid chromatography (HPLC) with electrochemical detcction according to the method described by Eriksson and Persson (1982). Means + SEM are reported. mcasurcments were subjected ance (ANOVA)

followed

Data for catecholamine to an analysis of vari-

by a multiple.

pairwise com-

parison (Bonferroni). Differcnccs were considered significant if P < 0.05. Percent inhibitions were calculated from

the

respective

control

hradykinin antagonists tistical analysis.

values

before

use

and were not subjected

of

to sta-

In group la. B-II% \vas injcctcd i.a. as a holus 5 min after the kccond bradykinin injection at doses of 0.8. t.6. 3.2. 6.4. IO. 40. IOO. 200. -I(H) or X00 pg (n = 8 per dose) follo\vd by 3 third hrsdykinin injection after IO-15 s. Only one dose of the bradgkinin antagonist ws given per animal. Thirty minutes later. when the

3. Results Control reduced

i.a. bolus injections

MAP

of 250 ng bradykinin

by 5.86 + 0.58 mmHg * min (AU0

The

blood prcssurc had returned to control Icvcls. the procedure was repcatcd but with HOE l-10 at doses of 6.25 12.5. 25. 50. IW. X0. 100. 600 ng: 1.6, 3.2. 6.4 or

antagonistic potcncics of the bradykinin antagonists HOE 149 and B4143 against the depressor actions of hradykinin

in conscious

I2.S ~(8 (n = S per dose) instead of B1116.

comparison

of ED,,,

In group lb. the bradykinin antagonists were adminis*.>-_* ,,,,d iiS 5-min i.a. infusions instead of i.a. bolus

the dose-response strates that HOE

injections. An infusion of BJ146 at doses of 0.61. 2. IO.

potent

-10. 100 or 200 pg/min (n = 8 per dose) was started shortly after the second control injection of bradykinin

HOE ’ V and 84146 were 2.1 and 1000 pg after an i.a. bolus injection and 0.84 and 220 pg/min after a 5-min

and was stopped immediately after the third hradykinin . . . InJectIon. Only one dose of the bradyhinin antagonists

ws

given per animal. Thirty minutes fatcr. the proccdurc was repeated but with HOE I40 (1.35. 3.5, 5. to. 20. 40. X0, 160. 320, 640 ng/min; I.28 or 2.56 gg/min; n = 8 per dose) instead of B4tlh. In experiment two. the kinetics of the two bradykinin antagonists were compared. using cquipotent doses of 2-I pg HOE 140 (n = 8) and 1000 pg 84146 (n = X) &II QS an i.a. holus Injection md 0.84 pg/min HOE

than

B4146.

in fig.

1. A

and ED,$,, values calculated

rats are shown

from

curves of experiment one demon:40 was about 250-700-fold more The

respective

ED,,,,

infusion. The ED,,, values were calculated

vdtues for

as I4 ng and

3.1 ng/min for HOE 140. and 3.4 kg and 3.0 pg/min for B4 146. respect ivcly. Marked differences in the duration of action were obscrvcd between the two bradykinin antagonists (fig. 2). HOE 140 inbibited the bradykinin effect by 76.4 and 70.5% 15 min after an i.a. injection and 15 min after a 5-min i.a. infusion. respectively, and was still inhibit+w !!f!rr ..E ? 39.55; inhibition) jn ._-- 6!! ...* ml...a.. (21 ,.! ..I,0 CIIU” contrast,

the inhibitory

action

of B4!46

had virtually

4. Discussion

30 TIME

60

90

6mln)

Fig. 1. Dose-Jcpendent inhibition of the depressor responses to exogenous bradykinin by an i.a. bolus injection or 5-min La. infusion of the bradykinin antagonist HOE 140 (open and solid circles) and Blllh (open and solid triangles). Bradykinin (Ytt ngl was injected i.a. hefore and IO-15 s after administration of the respective bradykinin antagonist.

disappeared 15 min after injection and infusion (4.5 and 5.4% inhibition). Plasma catecholamines were not significantly altered by an i.a. injection of 3.2 pg HOE 140. Plasma noradrenaline concentrations in HOE 140- and vehicle-treated rats were not significantly different (164 f 66 pg/ml vs. 129 f 61 pg/ml). Plasma adrenaline tended to be reduced in HOE 140-treated rats (253 + 73 pg/ml vs. 584 f 181 pg/ml in control); however, this difference did not reach statistical signit’icance.

0.0 1

I

BK-ANTAGONIST

100 (06)

Fig. 2. Timr course of the effect of HOE IJO and BJI46 on depressor responses to Rradykinin C5tl ng. i.a. ho!us). Bradykinin antagonists were given as i.a. holus injections a! ~+oscsof 2.1 pg (HOE 110. open circles) and ItNHl /~g tB4146, open triangles) or as S-min infusions at doses of 0.X4 wg/min (HOE 140. solid circles) and 220 pg/min (BJllh. solid triangles). Bradykinin was injected before. and at 5.min or 15min intervals after application of the bradykfn;t~ xrtagonists. 7;~ Lcro represents the inhibition of bradykinin responses IO-IS s after administration of the respective hradykinin antagonist.

Our rwtlts in conscious rats demonstrate that HOE 140 is a highly potent and long-acting bradykinin B.-rrccptor antagonist. Comparison of ED,, and ED,,,-values, calculated after an i.a. bobs injection or after a 5-min infusion of the two bradykinin antagonists, revealed that HOE 140 is 250-700 times more potent than B4146 in antagonizing the depressor responses to exogenously applied bradykinin. Furthermore, comparison of the ED,,,, values calculated after the 5-min infusion showed that HOE 140 is more than IO-fold more potent than the novel bradykinin antagonist (CH ,-D-Arg-Arg-Pro-Hyp-Gly-Thi-Ser-D-Phe-ThiArgl introduced recently by Lammeck et al. (1990). The difference in potency between these two bradykinin antagonist was even more obvious (> 200 fold) when lower doses (ED,,, values) were compared. A further important feature of HOE 140 is its long-lasting inhibitory action (half-life of more than one hour after an i.a. bolus injection), which has already been observed by Wirth et al. (1991) in anesthetized rats. In contrast, the half-life of B4146 was estimated to be less than 5 min. The longer half-life in vivo and the higher potency of HOE 140 cor-ipared to B4146 may be the result of a higher affinity for Bz-receptors as well as a higher stability against enzymatic degradation (Hock et al.. 1991). Previous studies have demonstrated that some bradykinin antagonists stimulate catecholamine release from the adrenal gland. For instance, Mulinari et al. (1988) showed a 2-fold increase of the plasma noradrenaline concentrations and a 6-fold increase of plasma adrenaline concentrations after a 5-min infusion of the bradykinin antagonist B3852 (Arg-Pro-Hyp-Gly-PheScr-D-Phe-Phe-Arg. 250 pg/min). Our results demonstrate that HOE 140 at a dose of 3.2 pg, which inhibited the depressor responses to 250 ng bradykinin more than 90%, did not significantly alter plasma catecholamine concentrations. Thus, this compound does not appear to influence the release of catecholamines from the adrenal gland. In summary, our data in conscious rats show that the novel bradykinin B,-receptor antagonist HOE 140 possesses a high potency and long duration of action and is devoid of a bradykinin-like agonistic effect on catecholamine release. Thus. HOE 140 may become a valuable tool to investigate further the kallikrein-kinin system under physiological and pathophysiok?gical conditions.

Brripohl. G.. S. Henke. 1. Knolle. H. Anagnostopoulos and B.A. Schiilkens. J(iVt;, fioe i40: A new and highly po:ent bradykinin

Mulinari. R.. A. Benrtos. I. Gavras and H. Gavras. 1988. Vascular and +mpathoadrrnal responses to hradykinin and a hradykinin an2!0guc. Hyper!rnsion ! I. 754. Stauss. B.. K. Itoi. H. Swuss and Th. Unger. 1990. A novel inexpensi\e computer +*tem IO record and analyse hemodynamic data in conscious rats. European J. Pharmacol. 183, X63. Vavrek. R.J. and J.M. Stewart. 1985. Competitive antagonists of hradykinin. Peptides 6. 161. \C’irth. K.. F.J. Hock. U. Albus. W. Linz. H. Anagnostopoulos, St. Henke. G. Breipohl. W. Kiinig. J. Knolle. and B.A. Schiilkens, 1991. Hoc I41 a new potent and long acting bradykinin-antagonist: in \ivo studies. Br. J. Pharmacol. 102. 774.