On the multiplicity of platelet prostaglandin receptors. II. The use of N-0164 for distinguishing the loci of action for PGI2, PGD2, PGE2 and hydantoin analogs.

The omega-chain variant analogs of prostacyclin (PGI2) and PGD2 in which n-amyl side-chain has been replaced by a cyclohexyl group have been prepared and their cardiovascular activities have been compared to those of BW-245C(Fig. 1) a potent anti-aggregatory vasodilator bearing a cyclohexyl-terminated side-chain on a hydantoin skeleton. The cyclohexyl group has little effect on PGI2, but converts PGD2 to a long lasting hypotensive agent and increases the platelet anti-aggregatory potency of PGD2 by a factor of 8. The prostaglandin antagonist N-0164 selectively blocks the anti-aggregatory actions of PGD2, cyclohexyl-PGD2, and BW-245C; with essentially no effect on PGI2, cyclohexyl-PGI2 and PGE2 at comparably effective doses. The latter observation is contrary to an earlier report by MacIntyre, but supports the view that the anti-aggregatory effect of high doses of PGE2 (EC50=50 microM) is mediated by the PGI2 receptor. The hydantoin acts at the platelet PGD2 receptor.

A single platelet aggregometric assay for pro-and antiaggregatory prostanoids.

A method is described for characterizing disaggregatory and proaggregatory prostanoids in a single assay. This "post-addition assay" consists of adding the test agent to PRP containing PGI2 (4-6 nM) 90 sec after an ADP challenge. The relative potencies of known inhibitors of ADP-induced aggregation were identical to the relative disaggregatory potencies determined by post-addition. The proaggregatory relative potencies for 16, 16-Me2-PGE2,PGE2,PGH2 and 11 alpha,9 alpha-epoxymethano-15 alpha-hydroxy-prostadienoic acid were 5.8 :2 : 1.8 : 1, respectively, revealing the significant thrombosis promoting activity of PGE2 and its analogs.

Molecular basis for prostaglandin potency. III. Tests of the significance of the "hairpin conformation" in biorecognition phenomena.

Prostaglandin analogs of the PGF2 alpha, 15-epi-PGF2 alpha, and PGE2 type bearing the following methyl substitution patterns -- 15-Me, 16, 16-(Me)2, 17, 17-(Me)2, and 18, 18, 20-(Me)3 -- and analogs constrained to "hairpin" alignment [via 1, (omega-1)-olide formation] and to "non-hairpin" arrangements [via 1, 9- and 1, 15-olide formation] are compared in the following biological assays: contraction of uterine and gastro-intestinal smooth muscle strips, luteolytic antifertility potency in the hamster, binding affinity to two different PGF2 alpha-receptor preparations from bovine corpora lutea, binding to the PGE-specific receptors from rat kidney and liver, inhibition of ADP- induced aggregation of human platelet-rich-plasma, and the effect on rat blood blood pressure. The methylated prostaglandins were also concerted to the corresponding prostacyclins and examined as to action on the platelet and on rat blood pressure. All evidence points to topographically distinct receptors for F2 alpha-, E- and I2- type prostaglandins. Cross-reactivity is reduced in most of the analogs examined. Independent of the target organ or tissue, the receptors show common features based on the functional class of PG recognized. "Hairpin" alignment improves binding (and potency) only for the PGF2 alpha specific assays. PGE-specific binding and potency is disrupted to an increasing extent as the chain branching point is moved further from the 15-hydroxyl center. In contrast 16, 16-dimethylation is particularly disruptive for the PGI2/E1 platelet receptor interaction.

Methyl ethers of prostaglandins F2 alpha and I2.

Regiospecific monomethyl prostaglandin f2 alpha ethers (at 0-9, 0-11, and 0-15) have been prepared by total synthesis. The 9, 15-bis-ether was also prepared. The 11- and 15-monoethers have been converted to the corresponding prostacyclins. Nuclear Magnetic Resonance (NMR) spectral comparisons indicate conformational changes associated with ether formation; nonetheless, the PGF2 alpha monoethers all retain significant biological activity: 3-420% of natural PGF2 alpha. The 9- and 15- menthyl ethers show increased selectivity for luteolytic activity as measured in the hamster antifertility (HAF) assay. In contrast the prostacyclin ethers are essentially devoid of agonist activity on both the platelet and vasculature. Prostacyclin diastereomers [5a] also lack activity and it appears that any modification at or of the C-11 or C-15 functions reduces receptor binding by at least a factor of 100.