3-HYDROXY ANALOGUES OF THE p-SELECTIVE OPIOID ANTAGONIST CYPRODIME Schmidhammer H*, Jennewein HK*, Bell K +, Patel D +, Traynor JR + * Institute of Pharmaceutical Chemistry, University of Innsbruck, Innrain 52a, A-6020 Innsbruck, Austria; + Department of Chemistry, Loughborough University of Technology, Loughborough, Leicestershire, LE11 3 TU, U. K.

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Several 3-hydroxy analogues of the p-selective, competitive opioid antagonist cyprodime have been prepared starting from naltrexone or naloxone. The novel compounds have been evaluated in opioid receptor binding assays (using homogenates of guinea-pig brain) and in isolated tissue bioessay (mouse vas deferens preparation, MVD). No agonist activity was detected with any compound in the MVD up to concentrations of 3000 nM. The !~ affinities are similar in both binding assays and MVD supporting full antagonist nature of the compounds at !~ receptors. Both at K and G receptors a difference between binding and MVD is seen. In most cases higher affinity is seen in the bioassay. This could indicate differences in degree of agonism of the compounds or receptor differences in the two tissues. The findings suggest that the 3-hydroxy derivatives of cyprodime can distinguish between K and also G opioid receptors in the MVD and guinea-pig brain, which may indicate differences between the receptor populations in these tissues.

The 3-hydroxy analogues of cyprodime (compounds 3 - 6) have been prepared starting from naltrexone or naloxone in several steps.

R4" OR 3 '0

1 R 1 = CPM, R 2 = R 3 = Me, R 4 = H (CYPRODIME)

2 R 1 = CPM, R 2 = R 3 = Me, R 4 = OH (3-HYDROXYCYPRODIME) 3 R 1 = CPM, R 2 = Et, R 3 = Me, R 4 = OH

4 R 1 = CPM, R 2 = Me, R 3 = nBu, R 4 = OH 5 R 1 = allyl, R 2 = R 3 = Me, R 4 = OH 6 R 1 = CPM, R 2 = H, R 3 = Me, R 4 = OH

CPM = cyclopropylmethyl

The novel compounds were studied using opioid receptor binding assays (Table 1) and isolated tissue bioassay (mouse vas deferens preparation, MVD; Table 2). In each case results were compared with cyprodime (1) (1, 2) and 3-hydroxycyprodime (2) (3).

Table 1: Receptor Binding Data Determined in Homogenates of Guinea-pig Brain

Ki (riM) selectivity ratio

compound p K 6 ,¢/p 6/p

3 2.8 11.0 41.6 3.9 149 4 81.2 60.7 274 0.7 3.4 5 10.1 90.3 32.1 8.9 3.2 6 54.4 23.0 336 0.4 6.2 2 (3-hydroxycycprodime) 6.15 4.18 13.8 0.7 2.2 Cyprodime 23. i 62.3 112 2.7 4.8 Naloxone* 1.8 17.2 27 9.6 15

p Sites were labelled with [3HIDAMGO, 6 sites with [3HIDPDPE, and K sites with [3HIU69,593. * Values were taken from ref. 4.

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Table 2 : Antagonist Potency Determined in the Mouse Vas Deferens Preparation (MVD)

Ke (nM) selectivity ratio

compound ~t ~ 6 K/~ 6/~

3 1.84 223 176 121 96 4 93.8 174 10124 1.9 108 $ 4.60 4.67 2272 1.0 494 6 24.9 243 8922 9.8 358 2 (3-hydroxycyprodime) 4.61 471 380 102 82 Cyprodime* 55.4 1551 6108 28 l 10 Naloxone# 2.7 14.6 20.4 5.4 7.5

Agonists used: DAMGO (~t), C1977 (K), and DPDPE (6). * Cyprodime Ke values determined against normorphine (~t), ethylketocyclazocine (~:) and DADLE (6). # Naloxone Ke values for K receptors determined against U50,488.

No agonist activity was detected with any compound in the MVD at concentrations up to 3000 nM. The introduction of a 14-ethoxy group in 3-hydroxycyprodime (to produce compound 3) enhances the affinity at all three types of opioid receptors measured in both binding assays and isolated tissue bioassays. At !~ receptors similar affinities were obtained by binding and MVD for all compounds tested. However, at ~ receptors a difference between binding and isolated tissues is seen. Affinity determined in binding is between 3 and 100 fold higher than affinity determined in the MVD, except for compound 5 where higher affinity is seen in the bioassay. This could indicate differences in partial agonism of the compounds or receptor differences in the two tissues. At b receptors the compounds have also higher affinity in binding than in the MVD. This may suggest either that the compounds are partial agonists at 6 receptors or differences between receptors in the MVD and guinea-pig brain. The findings suggest that 3-hydroxycyprodime and its analogues 3 - 6 can distinguish between K and also

opioid receptors in the MVD and guinea-pig brain, which may indicate differences between the receptor populations in these tissues.

REFERENCES 1. H. Schmidhammer, W. P. Burkard, L. Eggstein-Aeppli, C. F. C. Smith (1989) J. Med.

Chem. 32, 418-421. 2. H. Schmidhammer, C. F. C. Smith, D. Erlach, M. Koch, R. Krassnig, W. Schwetz, C.

Wechner (1990) J. Med. Chem. 33, 1200-1206. 3. D. Patel, H. K. Jennewein, H. Schmidhammer, C. F. C. Smith, J. R. Traynor (1992) Br. J.

Pharmacol. 105, 142P. 4. I. Magnan, S. I. Paterson, A. Tavani, H. W. Kosterlitz (1982) N. S. Arch. Pharmakol. 319,

197.