NITRIC OXIDE RELEASING PHOSPHODIESTERASE TYPE 5 INHIBITOR

Abstract

The invention relates to nitric oxide releasing phosphodiesterase type 5 (PDE5) inhibitors of formula (I) or (Ib) or (II) or a stereoisomer or a pharmaceutically acceptable salt thereof:

##STR00001## useful for the treatment of ocular condition associated with elevated intraocular pressure such as ocular hypertension, glaucoma or retinopathies. The invention further relates to 3-[(2S)-2,3-bis(nitrooxy)propoxy]propanoic acid and 6-(nitrooxy)hexanoic acid for use in the treatment of ocular condition associated with elevated intraocular pressure such as ocular hypertension, glaucoma or retinopathies.

Claims

1. A compound of formula (I) or (II) or a stereoisomer or a pharmaceutically acceptable salt thereof: ##STR00071## wherein: R.sub.1 is the residue of a nitric oxide releasing molecule having the following formula:
R.sub.1=—C(O)—(O—CH.sub.2).sub.y(CH.sub.2).sub.m—[O—(CH.sub.2).sub.n].sub.p—(CH—ONO.sub.2).sub.q—CH.sub.2—ONO.sub.2 wherein: y is 1 or 0; p is 1 or 0; q is 1 or 0; m is an integer ranging from 1 to 10; preferably m is from 1 to 6; n is an integer ranging from 1 to 6; preferably n is 1 or 2; R.sub.2 is ethyl or n-propyl; R.sub.3 is methyl or ethyl; with the proviso that when in formula (I) R.sub.2 is ethyl, R.sub.3 is methyl or when R.sub.2 is n-propyl, R.sub.3 is ethyl.

2. A compound of formula (I) or (II) or a stereoisomer or a pharmaceutically acceptable salt thereof according to claim 1 wherein y is 0.

3. A compound of formula (I) or (II) or a stereoisomer or a pharmaceutically acceptable salt thereof according to claim 2 wherein R.sub.1 is selected from the group consisting of: ##STR00072##

4. A compound of formula (I) or (II) or a stereoisomer or a pharmaceutically acceptable salt thereof according to claim 1 where y is 1.

5. A compound of formula (I) or (II) or a stereoisomer or a pharmaceutically acceptable salt thereof according to claim 4 wherein R.sub.1 is selected from the group consisting of: ##STR00073##

6. A compound of formula (I) or a stereoisomer or a pharmaceutically acceptable salt thereof according to claim 1 wherein R.sub.2 is n-propyl and R.sub.3 is ethyl.

7. A compound of formula (I) or a stereoisomer or a pharmaceutically acceptable salt thereof according to claim 1 wherein R.sub.2 is ethyl, R.sub.3 is methyl.

8. A compound of formula (Ib) stereoisomers or salts said compounds having formula ##STR00074## wherein: R.sub.1 is the residue of a nitric oxide releasing molecule having the following formula:
R.sub.1=—C(O)—(O—CH.sub.2).sub.y(CH.sub.2).sub.m—[O—(CH.sub.2).sub.n].sub.p—(CH—ONO.sub.2).sub.q—CH.sub.2—ONO.sub.2 wherein: y is 1 or 0; p is 1 or 0; q is 1 or 0; m is an integer ranging from 1 to 10; n is an integer ranging from 1 to 6; R.sub.2 is ethyl or n-propyl; R.sub.3 is methyl or ethyl; with the proviso that when R.sub.2 is ethyl then R.sub.3 is methyl; when R.sub.2 is n-propyl then R.sub.3 is ethyl; and with the proviso that when R.sub.2 is ethyl and R.sub.3 is methyl, y is 0, p and q are 0, then m is not 2.

9. A compound of formula (Ib) according to claim 8 wherein R.sub.1 is selected from the group consisting of the radicals of formula (IIIa)-(IIIh): ##STR00075##

10. A compound of formula (Ib) according to claim 8 wherein R.sub.1 is selected from the group consisting of the radicals of formula (IVa)-(IVh): ##STR00076##

11. A compound of formula (Ib) according to claim 8 wherein R.sub.2 is ethyl and R.sub.3 is methyl or R.sub.2 is n-propyl and R.sub.3 is ethyl.

12. A compound of formula (Ib) according to claim 9 wherein R.sub.2 is ethyl and R.sub.3 is methyl.

13. A compound of formula (Ib) according to claim 9 wherein R.sub.2 is n-propyl and R.sub.3 is ethyl.

14. A compound of formula (Ib) according to claim 8 wherein y is 1.

15. A compound of formula (I) or a stereoisomer or a pharmaceutically acceptable salt thereof according to claim 1 selected from the group consisting of: 2-(4-(3-(5-ethyl-4-oxo-7-propyl-4,5-dihydro-3H-pyrrolo[3,2-d]pyrimidin-2-yl)-4-propoxyphenylsulfonyl)piperazin-1-yl)ethyl 3-(2,3-bis(nitrooxy)propoxy)propanoate (Compound (1)); 2-(4-((3-(5-ethyl-4-oxo-7-propyl-4,5-dihydro-3H-pyrrolo[3,2-d]pyrimidin-2-yl)-4-propoxyphenyl)sulfonyl)piperazin-1-yl)ethyl 2-(2-(nitrooxy)ethoxy)acetate (Compound (2)); 2-(4-(3-(5-ethyl-4-oxo-7-propyl-4,5-dihydro-3H-pyrrolo[3,2-d]pyrimidin-2-yl)-4-propoxyphenylsulfonyl)piperazin-1-yl)ethyl 5,6-bis(nitrooxy)hexanoate (Compound (3)); 2-(4-((3-(5-ethyl-4-oxo-7-propyl-4,5-dihydro-3H-pyrrolo[3,2-d]pyrimidin-2-yl)-4-propoxyphenyl)sulfonyl)piperazin-1-yl)ethyl 6-(nitrooxy)hexanoate (Compound (4)); 2-(4-(3-(5-ethyl-4-oxo-7-propyl-4,5-dihydro-3H-pyrrolo[3,2-d]pyrimidin-2-yl)-4-propoxyphenylsulfonyl)piperazin-1-yl)ethyl 6-(nitrooxy)hexyl carbonate (Compound (5)); 2-(4-(3-(5-ethyl-4-oxo-7-propyl-4,5-dihydro-3H-pyrrolo[3,2-d]pyrimidin-2-yl)-4-propoxyphenylsulfonyl)piperazin-1-yl)ethyl 5,6-bis(nitrooxy)hexyl carbonate (Compound (6)); 2-{4-[4-ethoxy-3-(5-methyl-4-oxo-7-propyl-4,5-dihydro-1H-pyrrolo[3,2-d]pyrimidin-2-yl)benzene-1-sulfonyl]piperazin-1-yl}ethyl 5,6-bis(nitrooxy)hexanoate (Compound (7)) 2-{4-[4-ethoxy-3-(5-methyl-4-oxo-7-propyl-4,5-dihydro-1H-pyrrolo[3,2-d]pyrimidin-2-yl)benzene-1-sulfonyl]piperazin-1-yl}ethyl 6-(nitrooxy)hexanoate (Compound (8));

16. A compound of formula (II) or a stereoisomer or a pharmaceutically acceptable salt thereof according to claim 1 selected from the group consisting of (S)-(1-(4-(3-chloro-4-methoxybenzylamino)-5-(pyrimidin-2-ylmethylcarbamoyl) pyrimidin-2-yl)pyrrolidin-2-yl)methyl 6-(nitrooxy)hexanoate (Compound (9)); [(2S)-1-(4-{[(3-chloro-4-methoxyphenyl)methyl]amino}-5-{[(pyrimidin-2-yl) methyl]carbamoyl}pyrimidin-2-yl)pyrrolidin-2-yl]methyl 5,6-bis(nitrooxy)hexanoate (Compound (10)); (S)-(1-(4-(3-chloro-4-methoxybenzylamino)-5-(pyrimidin-2-ylmethylcarbamoyl) pyrimidin-2-yl)pyrrolidin-2-yl)methyl 2-(2-(nitrooxy)ethoxy)acetate (Compound (11)); [(2S)-1-(4-{[(3-chloro-4-methoxyphenyl)methyl]amino}-5-{[(pyrimidin-2-yl) methyl]carbamoyl}pyrimidin-2-yl)pyrrolidin-2-yl]methyl 3-[2,3-bis(nitrooxy)propoxy]propanoate (Compound (12)); (S)-(1-(4-(3-chloro-4-methoxybenzylamino)-5-(pyrimidin-2-ylmethylcarbamoyl) pyrimidin-2-yl)pyrrolidin-2-yl)methyl 6-(nitrooxy)hexylcarbonate (Compound (13)); ((S)-1-(4-(3-chloro-4-methoxybenzylamino)-5-(pyrimidin-2-ylmethylcarbamoyl) pyrimidin-2-yl)pyrrolidin-2-yl)methyl 5,6-bis(nitrooxy)hexyl carbonate (Compound (14));

17. A compound of formula (I) or (II) according to claim 1 selected from the group consisting of: [(2S)-1-(4-{[(3-chloro-4-methoxyphenyl)methyl]amino}-5-{[(pyrimidin-2-yl) methyl]carbamoyl}pyrimidin-2-yl)pyrrolidin-2-yl]methyl (5S)-5,6-bis(nitrooxy) hexanoate (Compound (10)-(5S)); [(2S)-1-(4-{[(3-chloro-4-methoxyphenyl)methyl]amino}-5-{[(pyrimidin-2-yl) methyl]carbamoyl}pyrimidin-2-yl)pyrrolidin-2-yl]methyl 3-[(2S)-2,3-bis(nitrooxy) propoxy]propanoate (Compound (12)-(2S)); (S)-(1-(4-(3-chloro-4-methoxybenzylamino)-5-(pyrimidin-2-ylmethylcarbamoyl) pyrimidin-2-yl)pyrrolidin-2-yl)methyl 6-(nitrooxy)hexanoate 2-hydroxypropane-1,2,3-tricarboxylate (Compound (9)-citrate salt); ((S)-1-(4-(3-chloro-4-methoxybenzylamino)-5-(pyrimidin-2-ylmethyl carbamoyl)pyrimidin-2-yl)pyrrolidin-2-yl)methyl (5S)-5,6-bis(nitrooxy)hexanoate 2-hydroxypropane-1,2,3-tricarboxylate (Compound (10)-(5S) citrate salt); (S)-(1-(4-(3-chloro-4-methoxybenzylamino)-5-(pyrimidin-2-ylmethylcarbamoyl) pyrimidin-2-yl)pyrrolidin-2-yl)methyl 2-(2-(nitrooxy)ethoxy)acetate 2-hydroxy propane-1,2,3-tricarboxylate (Compound (11) citrate salt); ((S)-1-(4-(3-chloro-4-methoxybenzylamino)-5-(pyrimidin-2-ylmethylcarbamoyl) pyrimidin-2-yl)pyrrolidin-2-yl)methyl 3-((S)-2,3-bis(nitrooxy)propoxy)propanoate 2-hydroxypropane-1,2,3-tricarboxylate (Compound (12)-(2S) citrate salt); (S)-2-(4-(3-(5-ethyl-4-oxo-7-propyl-4,5-dihydro-3H-pyrrolo[3,2-d]pyrimidin-2-yl)-4-propoxyphenylsulfonyl)piperazin-1-yl)ethyl 3-[2,3-bis-(nitrooxy)propoxy)]propanoate citrate (Compound (1)-(2S) citrate salt); 2-(4-(3-(5-ethyl-4-oxo-7-propyl-4,5-dihydro-3H-pyrrolo[3,2-d]pyrimidin-2-yl)-4-propoxyphenylsulfonyl)piperazin-1-yl)ethyl 6-(nitrooxy)hexanoate citrate (Compound (4) citrate salt); 2-hydroxypropane-1,2,3-tricarboxylic acid 2-{4-[3-(5-ethyl-4-oxo-7-propyl-4,5-dihydro-3H-pyrrolo[3,2-d]pyrimidin-2-yl)-4-propoxybenzene-1-sulfonyl]piperazin-1-yl}ethyl[2-(nitrooxy)ethoxy]acetate (Compound (2) citrate salt); (S)-2-(4-(3-(5-ethyl-4-oxo-7-propyl-4,5-dihydro-3H-pyrrolo[3,2-d]pyrimidin-2-yl)-4-propoxyphenylsulfonyl)piperazin-1-yl)ethyl 5,6-bis(nitrooxy)hexanoate citrate (Compound (3)-(5S) citrate salt); (S)-2-(4-(3-(5-ethyl-4-oxo-7-propyl-4,5-dihydro-3H-pyrrolo[3,2-d]pyrimidin-2-yl)-4-propoxyphenylsulfonyl)piperazin-1-yl)ethyl 3-(2,3-bis(nitrooxy)propoxy) propanoate dihydrochloride (Compound (1)-(2S) HCl salt); ((S)-1-(4-(3-chloro-4-methoxybenzylamino)-5-(pyrimidin-2-ylmethylcarbamoyl) pyrimidin-2-yl)pyrrolidin-2-yl)methyl (S)-5,6-bis(nitrooxy)hexyl carbonate (Compound (14)-(5S)); (S)-(1-(4-(3-chloro-4-methoxybenzylamino)-5-(pyrimidin-2-ylmethylcarbamoyl) pyrimidin-2-yl)pyrrolidin-2-yl)methyl 6-(nitrooxy)hexylcarbonate 2-hydroxypropane-1,2,3-tricarboxylate (Compound (13) citrate salt); (S)-(1-(4-(3-chloro-4-methoxybenzylamino)-5-(pyrimidin-2-ylmethylcarbamoyl) pyrimidin-2-yl)pyrrolidin-2-yl)methyl 6-(nitrooxy)hexanoate fumarate (Compound (9) fumarate salt); (S)-(1-(4-(3-chloro-4-methoxybenzylamino)-5-(pyrimidin-2-ylmethylcarbamoyl) pyrimidin-2-yl)pyrrolidin-2-yl)methyl 6-(nitrooxy)hexanoate maleate (Compound (9) maleate salt).

18. A compound of formula (Ib) according to claim 8 selected from the group consisting of: 2-{4-[4-ethoxy-3-(1-methyl-7-oxo-3-propyl-6,7-dihydro-1H-pyrazolo[4,3-d]pyrimidin-5-yl)benzene-1-sulfonyl]piperazin-1-yl}ethyl 5,6-bis(nitrooxy) hexanoate (Compound (7b)); (S)-2-(4-((4-ethoxy-3-(1-methyl-7-oxo-3-propyl-6,7-dihydro-1H-pyrazolo[4,3-d]pyrimidin-5-yl)phenyl)sulfonyl)piperazin-1-yl)ethyl (S)-5,6-bis(nitrooxy)hexanoate (Compound (7b)-(5S)); 2-{4-[4-ethoxy-3-(1-methyl-7-oxo-3-propyl-6,7-dihydro-1H-pyrazolo[4,3-d]pyrimidin-5-yl)benzene-1-sulfonyl]piperazin-1-yl}ethyl 6-(nitrooxy)hexanoate (Compound (8b)); 2-{4-[4-ethoxy-3-(1-methyl-7-oxo-3-propyl-6,7-dihydro-1H-pyrazolo[4,3-d]pyrimidin-5-yl)benzene-1-sulfonyl]piperazin-1-yl}ethyl 3-[2,3-bis(nitrooxy) propoxy]propanoate (Compound 15); (S)-2-(4-((4-ethoxy-3-(1-methyl-7-oxo-3-propyl-6,7-dihydro-1H-pyrazolo[4,3-d]pyrimidin-5-yl)phenyl)sulfonyl)piperazin-1-yl)ethyl-(S)-3-(2,3-bis(nitrooxy) propoxy)propanoate (Compound (15), (S) isomer).

19. A compound of formula (I) or of formula (Ib) or of formula (II) or a stereoisomer or a pharmaceutically acceptable salt thereof according to claim 1 for use as a medicament.

20. A compound of formula (I) or (Ib) or (II) or a stereoisomer or a pharmaceutically acceptable salt thereof according to claim 1 for use in a method of treating a disease or a condition associated with increased ocular pressure.

21. A compound of formula (I) or (Ib) or (II) or a stereoisomer or a pharmaceutically acceptable salt thereof for use according to claim 20 wherein the disease or condition is selected from ocular hypertension, glaucoma, open-angle glaucoma, normal-tension glaucoma, pigmentary glaucoma, pseudoexfoliative glaucoma and drug induced glaucoma.

22. A compound of formula (I) or (Ib) or (II) or a stereoisomer or a pharmaceutically acceptable salt thereof according to claim 1 for use in a method of treating retinopathies including retinopathy of prematurity, retinal vein occlusion or diabetic macular edema.

23. An ophthalmic pharmaceutical composition comprising a compound of formula (I) or of formula (Ib) or of formula (II) or stereoisomer or a pharmaceutical salt thereof according to claim 1 and a pharmaceutical acceptable excipient or vehicle.

24. A composition comprising a compound of formula (I) or of formula (Ib) or of formula (II) according to claim 1 and at least another active agent selected from the following classes of drugs: beta-adrenergic antagonists, prostaglandin analogues, adrenergic agonists, carbonic anhydrase inhibitors, cholinergic agonists, steroids, cholinesterase inhibitors, soluble guanylate cyclase activators, Rho-kinase inhibitors or cinaciguat.

25. The composition according to claim 24 in the form of a kit including a dosage form comprising the compound of formula (I) or of formula (Ib) or of formula (II) and a dosage form comprising the other active agent selected from the following classes of drugs: beta-adrenergic antagonists, prostaglandin analogues, adrenergic agonists, carbonic anhydrase inhibitors, cholinergic agonists, steroids and cholinesterase inhibitors soluble guanylate cyclase activators, Rho-kinase inhibitors or cinaciguat

26. The compound 3-[(2S)-2,3-bis(nitrooxy)propoxy]propanoic acid for use in the treatment of elevated intraocular pressure, ocular hypertension or conditions associated with elevated intraocular pressure.

27. The compound 6-(nitrooxy)hexanoic acid for use in the treatment of elevated intraocular pressure, ocular hypertension or conditions associated with elevated intraocular pressure or retinopathies.

28. A compound 3-[(2S)-2,3-bis(nitrooxy)propoxy]propanoic acid or 6-(nitrooxy)hexanoic acid for use in a method of the treatment of retinopathies including retinopathy of prematurity, retinal vein occlusion or diabetic macular edema.

Description

EXAMPLE 1

(S)-(1-(4-(3-chloro-4-methoxybenzylamino)-5-(pyrimidin-2-ylmethylcarbamoyl) pyrimidin-2-yl)pyrrolidin-2-yl)methyl 6-(nitrooxy)hexanoate (Compound (9))

[0177] ##STR00047##

[0178] To a solution of avanafil (1.00 g, 2.07 mmol) in dry DCM (15 ml), DMAP (252 mg, 2.07 mmol), DCC (512 mg, 2.48 mmol) and 6-(nitrooxy)hexanoic acid (440 mg, 2.48 mmol) were added.

[0179] The mixture was stirred overnight at room temperature and then diluted with DCM and water. The two phases were separated and the aqueous layer was extracted twice with DCM. The combined organic extracts were dried over MgSO.sub.4 and concentrated under reduced pressure. The residue was purified by reversed-phase chromatography (H.sub.2O/CH.sub.3CN with 0.1% formic acid, from 70:30 to 40:60).

[0180] The residue was taken up in DCM and this solution was washed with an aqueous saturated solution of NaHCO.sub.3. Then, the aqueous layer was extracted twice with DCM. The combined organic phases were dried over MgSO.sub.4 and concentrated under reduced pressure, giving 1.23 g of the desired product (Compound (9)) (yield: 93.0%).

[0181] .sup.1H NMR (400 MHz, DMSO-d.sub.6) δ 9.19 (s, 1H), 8.81 (m, 1H), 8.75 (d, J=4.9 Hz, 2H), 8.54 (m, 1H), 7.50-7.14 (m, 3H), 7.06 (s, 1H), 4.54 (m, 4H), 4.37-3.90 (m, 3H), 3.81 (s, 3H), 3.59-3.39 (m, 4H), 2.32 (t, J=7.1 Hz, 2H), 2.09-1.73 (m, 4H), 1.66-1.52 (m, 4H), 1.32 (dt, J=5.2, 7.5 Hz, 2H).

EXAMPLE 2

[(2S)-1-(4-{[(3-chloro-4-methoxyphenyl)methyl]amino}-5-{[(pyrimidin-2-yl)methyl]carbamoyl}pyrimidin-2-yl)pyrrolidin-2-yl]methyl (5S)-5,6-bis(nitrooxy)hexanoate (Compound (10)-(5S)-isomer)

[0182] ##STR00048##

[0183] To a solution of Avanafil (120 mg, 0.248 mmol) in dry DCM (1.5 ml), DMAP (30 mg, 0.248 mmol), DCC (56 mg, 0.273 mmol) and (5S)-5,6-bis(nitrooxy)hexanoic acid (65 mg, 0.273 mmol) were added.

[0184] The mixture was stirred overnight at room temperature, then diluted with DCM and water. The two phases were separated and the aqueous phase extracted twice with DCM. The combined organic phases were dried over MgSO.sub.4 and concentrated under reduced pressure. The residue was purified by reversed-phase chromatography (H.sub.2O/CH.sub.3CN with 0.01% Formic acid, from 70:30 to 40:60).

[0185] After purification DCM and a saturated solution of NaHCO.sub.3 were added to the residue. The two phases were separated and the aqueous phase extracted twice with DCM. The combined organic phases were dried over MgSO.sub.4 and concentrated under reduced pressure, giving 165 mg of the desired product (Compound (10)-(5S)-isomer) (yield: 94.5%).

[0186] .sup.1H NMR (400 MHz, DMSO-d.sub.6) δ 9.16 (t, J=6.0 Hz, 1H), 8.79 (m, 1H), 8.75 (d, J=4.9 Hz, 2H), 8.54 (s, 1H), 7.38 (t, J=4.9 Hz, 1H), 7.34-7.23 (m, 1H), 7.19 (d, J=7.4 Hz, 1H), 7.05 (t, J=9.8 Hz, 1H), 5.41 (m, 1H), 4.91 (m, 1H), 4.71 (m, 1H), 4.54 (m, 4H), 4.16 (m, 3H), 3.98 (s, 3H), 3.50 (m, 2H), 2.38 (t, J=6.7 Hz, 2H), 1.92 (m, 4H), 1.79-1.51 (m, 4H).

EXAMPLE 3

(S)-(1-(4-(3-chloro-4-methoxybenzylamino)-5-(pyrimidin-2-ylmethylcarbamoyl) pyrimidin-2-yl)pyrrolidin-2-yl)methyl 2-(2-(nitrooxy)ethoxy) acetate (Compound (11))

[0187] ##STR00049##

[0188] To a solution of Avanafil (160 mg, 0.331 mmol) in dry DCM (1.5 ml), DMAP (40 mg, 0.327 mmol), DCC (82 mg, 0.397 mmol) and 2-(2-(nitrooxy)ethoxy)acetic acid (82 mg, 0.497 mmol) were added.

[0189] The mixture was stirred over weekend at room temperature and then diluted with DCM and water. The two phases were separated and the aqueous phase extracted twice with DCM. The combined organic phases were dried over MgSO.sub.4 and concentrated under reduced pressure. The residue was purified by reversed-phase chromatography (H.sub.2O/CH.sub.3CN with 0.01% formic acid, from 90:10 to 65:35).

[0190] After purification DCM and a saturated solution of NaHCO.sub.3 were added to the residue. The two phases were separated and the aqueous phase extracted twice with DCM. The combined organic phases were dried over MgSO.sub.4 and concentrated under reduced pressure, giving 109 mg of the desired product (Compound (11)) (yield: 52%).

[0191] .sup.1H NMR (400 MHz, DMSO-d.sub.6) δ 9.16 (t, J=6.0 Hz, 1H), 8.79 (s, 1H), 8.75 (d, J=4.9 Hz, 2H), 8.54 (s, 1H), 7.38 (t, J=4.9 Hz, 1H), 7.24 (m, 1H), 7.06 (s, 1H), 4.70 (m, 2H), 4.54 (m, 4H), 4.26 (dd, J=10.5, 3.4 Hz, 1H), 4.18 (s, 2H), 3.80 (s, 3H), 3.78 (m, 2H), 3.50 (m, 2H), 2.02-1.79 (m, 4H).

EXAMPLE 4

[(2S)-1-(4-{[(3-chloro-4-methoxyphenyl)methyl]amino}-5-{[(pyrimidin-2-yl)methyl]carbamoyl}pyrimidin-2-yl)pyrrolidin-2-yl]methyl 3-[(2S)-2,3-bis(nitrooxy)propoxy]propanoate (Compound (12)-(2S)-isomer)

[0192] ##STR00050##

[0193] To a solution of Avanafil (160 mg, 0.331 mmol) and 4-nitrophenyl 3-[(2S)-2,3-bis(nitrooxy)propoxy]propanoate (82 mg, 0.497 mmol) in dry DCM (1.5 ml), DMAP (40 mg, 0.327 mmol) was added.

[0194] The mixture was stirred over weekend at room temperature, then diluted with DCM and water. The two phases were separated and the aqueous phase extracted twice with DCM. The combined organic phases were dried over MgSO.sub.4 and concentrated under reduced pressure. The residue was purified by reversed-phase chromatography (H.sub.2O/CH.sub.3CN with 0.01% formic acid, from 85:15 to 55:45).

[0195] After purification DCM and a saturated solution of NaHCO.sub.3 were added to the residue. The two phases were separated and the aqueous phase extracted twice with DCM. The combined organic phases were dried over MgSO.sub.4 and concentrated under reduced pressure, giving 227 mg of the desired product (Compound (12)-(2S)-isomer) (yield: 95.0%).

[0196] .sup.1H NMR (400 MHz, DMSO-d.sub.6) δ 9.16 (t, J=6.0 Hz, 1H), 8.78 (s, 1H), 8.75 (d, J=4.9 Hz, 1H), 8.53 (s, 1H), 7.38 (t, J=4.9 Hz, 1H), 7.34-7.16 (m, 1H), 7.05 (t, J=9.4 Hz, 1H), 5.55 (q, 1H), 4.89 (d, J=12.9 Hz, 1H), 4.75 (m, 1H), 4.54 (m, 4H), 4.41-3.93 (m, 3H), 3.80 (s, 3H), 3.78-3.63 (m, 4H), 3.51 (m, 2H), 2.57 (t, J=6.1 Hz, 2H), 1.92 (m, 4H).

Synthesis of 4-nitrophenyl 3-[(2S)-2,3-bis(nitrooxy)propoxy]propanoate

Step 1: Synthesis of (S)-4-(allyloxymethyl)-2,2-dimethyl-1,3-dioxolane

[0197] NaH (60%) (3.4 g; 85.54 mmol) was suspended in dry THF (140 ml) together with S(+)-1,2-isopropylidenglycerol (5.65 g; 42.75 mmol) and 15-Crown-5 (0.9 g; 4.28 mmol).

[0198] The mixture was cooled to 0° C. and allyl bromide (7.2 ml; 85.54 mmol) was added dropwise. The suspension was stirred for 6h at rt, than NH.sub.4CI saturated solution (100 ml) was added dropwise at 0° C. and then the mixture extracted with Et.sub.2O (3×100 ml). The combined organic layers were washed once with brine and concentrated under reduced pressure carefully. The residue was purified by flash chromatography (Biotage SP4 instrument, SNAP 100 column, isocratic elution Et.sub.2O/Cyclohexane 1:9), affording 5.8 g, (yield 78.5%) of the title compound ((S)-4-(allyloxymethyl)-2,2-dimethyl-1,3-dioxolane).

[0199] .sup.1HNMR (300 MHz, Chloroform-d) δ 6.00-5.78 (m, 1H), 5.35-5.13 (m, 2H), 4.35-4.21 (m, 1H), 4.11-3.98 (m, 3H), 3.73 (m, 1H), 3.57-3.40 (m, 2H), 1.51-1.32 (m, 6H).

Step 2: Synthesis of (S)-3-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)propan-1-ol

[0200] To a stirred solution of (S)-4-(allyloxymethyl)-2,2-dimethyl-1,3-dioxolane (2.0 g, 10.64 mmol) in dry THF (180 ml) cooled at 0° C., 9BBN 0.5M (241.8 ml, 121 mmol) was added dropwise. The reaction was stirred 30 minutes at 0° C. and overnight at room temperature. The reaction mixture was cooled at 0° C. and NaOH 2N (84 ml) was added together with H.sub.2O.sub.2 30% (58 ml). The mixture was diluted with Et.sub.2O (100 ml) and NaOH 1N (100 ml). The 2 phases were separated and aqueous layer was extracted with Et.sub.2O (3×120 ml). The combined organic layers were washed once with brine, dried over Na.sub.2SO.sub.4 and concentrated under reduced pressure. The residue was purified by flash chromatography (Biotage SP4 instrument, SNAP 100 column, EtOAC in cyclohexane, from 30% to 80% in 10 c.v. affording 6.4 g (yield: 100%) of the title compound ((S)-3-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)propan-1-ol).

[0201] .sup.1HNMR (300 MHz, Chloroform-d) δ 4.35-4.20 (m, 1H), 4.11-4.01 (m, 1H), 3.85-3.63 (m, 5H), 3.58-3.46 (m, 2H), 1.97-1.80 (m, 2H), 1.45 (s, 3H), 1.38 (s, 3H).

Step 3: Synthesis of (S)-3-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)propanoic Acid

[0202] To a solution of (S)-3-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)propan-1-ol (2.03 g; 10.51 mmol) in acetone (50 ml) cooled at 0° C., NaHCO.sub.3 sat solution (56 ml), NaBr (0.45 g, 4.20 mmol) and TEMPO (0.34 g, 2.10 mmol) were added. Trichloroisocyanuric acid (4.91 g, 21.02 mmol) was then added portionwise. The mixture was allowed to reach room temperature and stirred for 3h. The mixture was then cooled at 0° C. and isopropanol (20 ml) was added slowly. The mixture was stirred at 0° C. 30 minutes. The formation of a white solid was observed. The precipitate was filtered off and the solvent concentrated. NaOH 2N was added to the residue (pH=12) and the aqueous solution was washed twice with EtOAc. To the aqueous phase HCl 1N was added until pH 2-3 and it was extracted with EtOAc (5×50 ml). The combined organic phases were dried over Na.sub.2S0.sub.4 and then evaporated affording 0.82 g (Yield: 38%) of the title compound ((S)-3-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)propanoic acid).

[0203] .sup.1HNMR (300 MHz, Chloroform-d) δ 4.34-4.16 (m, 1H), 4.11-3.97 (m, 1H), 3.85-3.68 (m, 3H), 3.63-3.42 (m, 2H), 2.74-2.55 (m, 2H), 1.41 (s, 3H), 1.35 (s, 3H).

Step 4: Synthesis of (S)-4-nitrophenyl 3-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy) propanoate

[0204] To a solution of (S)-3-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy)propanoic acid (0.82 g, 4.02 mmol), DCC (0.83 mg, 4.02 mmol) and DMAP (0.1 g, 0.80 mmol) in DCM (15 ml), 4-nitrophenol (0.56 g; 4.02 mmol) was added portionwise. The mixture was stirred overnight at room temperature, then the precipitate was filtered off and the solvent evaporated. The residue was purified by flash chromatography (Biotage SP4 instrument, EtOAc in cyclohexane from 5% to 50% in 12 CV) affording 0.97 g of the title compound ((S)-4-nitrophenyl 3-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy) propanoate) (Yield: 74%).

[0205] .sup.1H NMR (300 MHz, Chloroform-d) δ 8.33-8.21 (m, 2H), 7.37-7.20 (m, 2H), 4.37-4.20 (m, 1H), 4.09-4.00 (m, 1H), 3.96-3.84 (m, 2H), 3.80-3.66 (m, 1H), 3.63-3.47 (m, 2H), 2.96-2.78 (m, 2H), 1.42 (s, 3H), 1.36 (s, 3H).

Step 5: Synthesis of (R)-4-nitrophenyl 3-(2,3-dihydroxypropoxy) propanoate

[0206] To a stirred solution of (S)-4-nitrophenyl 3-((2,2-dimethyl-1,3-dioxolan-4-yl)methoxy) propanoate (0.97 g, 2.97 mmol) in THF (10 ml), HCl 3N (2 ml) was added and the solution stirred for 4 h at room temperature. Then EtOAc (5 ml) and H.sub.2O (5 ml) were added and the two phases were separated. The aqueous phase was extracted with EtOAc (2×5 ml). The combined organic layers were washed with brine, dried over Na.sub.2SO.sub.4 and concentrated under reduced pressure affording 0.89 g of the title compound ((R)-4-nitrophenyl 3-(2,3-dihydroxypropoxy) propanoate), which was used in the next step without further purification.

Step 6: Synthesis of 4-nitrophenyl 3-[(2S)-2,3-bis(nitrooxy)propoxy]propanoate

[0207] To a solution of Ac.sub.2O (0.76 ml, 8.07 mmol) in DCM (5 ml) at −40° C., fuming HNO3 (0.38 ml, 9.32 mmol) was added dropwise. A solution of (R)-4-nitrophenyl 3-(2,3-dihydroxypropoxy) propanoate (0.89 g, 3.11 mmol) in DCM (7 ml) was then added dropwise. The mixture was allowed to reach 0° C. and stirred for 4 hours. The mixture was then poured into ice and NaHCO.sub.3 was added portionwise. The two phases were separated and the aqueous phase washed twice with DCM. The combined organic phases were dried over Na.sub.2S0.sub.4 and concentrated affording 0.56 g of the title compound (4-nitrophenyl 3-[(2S)-2,3-bis(nitrooxy)propoxy]propanoate) (yield of two steps: 38%).

[0208] .sup.1H NMR (300 MHz, Chloroform-d) δ 8.35-8.22 (m, 2H), 7.35-7.22 (m, 2H), 5.48-5.34 (m, 1H), 4.88-4.73 (m, 1H), 4.73-4.56 (m, 1H), 3.94-3.85 (m, 2H), 3.85-3.77 (m, 2H), 2.88 (t, 2H).

Synthesis of 3-[(2S)-2,3-bis(nitrooxy)propoxy]propanoic Acid (Compound (IIIg)-OH)

[0209] A solution of 4-nitrophenyl 3-[(2S)-2,3-bis(nitrooxy)propoxy]propanoate (Step 6) (500 mg, 1.33 mmol) in 5 ml of a 1/1 (vol/vol) mixture of EtOH/THF was cooled to 0° C. Then 1.33 ml (2.66 mmol) of 2N NaOH solution was dropwise added to the stirred solution in 45 minutes.

[0210] The mixture was left under stirring for 1h. Then the organic solvents were removed under vacuum and EtOAc (25 ml) and water (25 ml) were added. The organic phase was removed and the pH of the aqueous phase was set to 1 using HCl 2N. The aqueous phase was extracted with DCM (2×15 ml) dried over sodium sulfate and evaporated under vacuum.

[0211] The obtained raw product was purified on a 4 g 25μ C18 cartridge using a gradient of (ACN)/(H.sub.2O+0.1% HCOOH) from 5/95 to 30/70 over 30 minutes.

[0212] The fractions containing clean product were pooled and lyophilized to give 210 mg of the title compound (3-[(2S)-2,3-bis(nitrooxy)propoxy]propanoic acid as a clear oil) (yield 61%).

[0213] .sup.1H NMR (400 MHz, DMSO-d.sub.6) δ ppm 2.43-2.48 (m, 2H) 3.61-3.78 (m, 4H) 4.76 (dd, J=12.89, 6.54 Hz, 1H) 4.92 (dd, J=12.89, 2.93 Hz, 1H) 5.54-5.59 (m, 1H) 12.23 (br s, 1H)

EXAMPLE 5

(S)-(1-(4-(3-chloro-4-methoxybenzylamino)-5-(pyrimidin-2-ylmethylcarbamoyl) pyrimidin-2-yl)pyrrolidin-2-yl)methyl 6-(nitrooxy)hexanoate 2-hydroxypropane-1,2,3-tricarboxylate (Compound (9) Citrate Salt)

[0214] ##STR00051##

[0215] To a solution of compound (9), prepared as described in Example 1, (100 mg, 0.155 mmol) in Methanol (1.5 ml), citric acid monohydrate (33 mg, 0.155 mmol) was added. The mixture was stirred for 10 minutes at room temperature, then concentrated and the solid washed with MTBE. The solid was filtered and dried under reduced pressure, giving 125 mg of the desired product (Compound (9) citrate salt) (crude yield: 96.6%).

[0216] .sup.1H NMR (600 MHz, DMSO-d6) δ 12.39-12.36 (m, 3H), 9.19 (s, 1H), 8.80 (m, 1H), 8.75 (d, J=4.9 Hz, 2H), 8.53 (m, 1H)), 7.42-7.15 (m, 3H), 7.06 (m, 1H), 4.65-4.38 (m, 6H), 4.37-4.11 (m, 3H), 3.80 (s, 3H) 3.59-3.39 (m, 2H), 2.75 (d, J=15.4 Hz, 2H), 2.65 (d, J=15.4 Hz, 2H), 2.32 (m, 2H), 2.05-1.80 (m, 4H), 1.47-1.08 (m, 4H), 1.32 (q, 2H)

EXAMPLE 6

((S)-1-(4-(3-chloro-4-methoxybenzylamino)-5-(pyrimidin-2-ylmethylcarbamoyl) pyrimidin-2-yl)pyrrolidin-2-yl)methyl (5S)-5,6-bis(nitrooxy) hexanoate 2-hydroxypropane-1,2,3-tricarboxylate (Compound (10)-(5S)-isomer, Citrate Salt)

[0217] ##STR00052##

[0218] To a solution of Compound (10)-(5S)-isomer, prepared as described in Example 2, (165 mg, 0.234 mmol) in Methanol (1.5 ml), citric acid monohydrate (49 mg, 0.233 mmol) was added. The mixture was stirred for 10 minutes at room temperature, then concentrated and the residue was washed with diethyl ether. The solid was filtered and dried under reduced pressure, giving 205 mg of the desired product (Compound (10)-(5S)-isomer-citrate salt) (crude yield: 97.8%).

[0219] .sup.1H NMR (600 MHz, DMSO-d6) δ 12.37 (s, 3H), 9.18 (s, 1H), 8.85-8.70 (m, 3H), 8.53 (m, 1H), 7.42-7.36 (m, 2H), 7.34-7.15 (m, 1H), 7.10-7.01 (m, 1H), 5.41 (m, 1H), 4.91 (d, J=12.4 Hz, 1H), 4.70 (m, 1H), 4.63-4.42 (m, 4H), 4.38-3.92 (m, 3H), 3.81 (s, 3H), 3.51 (m, 2H), 2.75 (d, J=15.4 Hz, 2H), 2.66 (d, J=15.4 Hz, 2H), 2.38 (m, 2H), 1.92 (m, 4H), 1.79-1.58 (m, 4H).

EXAMPLE 7

(S)-(1-(4-(3-chloro-4-methoxybenzylamino)-5-(pyrimidin-2-ylmethylcarbamoyl) pyrimidin-2-yl)pyrrolidin-2-yl)methyl 2-(2-(nitrooxy)ethoxy) acetate 2-hydroxy propane-1,2,3-tricarboxylate (Compound (11) Citrate Salt)

[0220] ##STR00053##

[0221] To a solution of Compound 11, prepared as described in Example 3, (109 mg, 0.173 mmol) in Methanol (1.0 ml), citric acid monohydrate (36 mg, 0.171 mmol) was added. The mixture was stirred for 10 minutes at room temperature, then concentrated and the residue was washed with diethyl ether. The solid was filtered and dried under reduced pressure, giving 139 mg of the desired product (Compound (11) citrate salt) (crude yield: 97.6%).

[0222] .sup.1H NMR (600 MHz, DMSO-d6) δ 12.36 (s, 3H), 9.18 (s, 1H), 8.85-8.72 (m, 3H), 8.53 (m, 1H), 7.41-7.35 (m, 2H), 7.34-7.15 (m, 1H), 7.06 (m, 1H), 4.70-4.64 (m, 2H), 4.62-4.46 (m, 4H), 4.40-3.99 (m, 5H), 3.80 (s, 3H), 3.78 (m, 2H), 3.58-3.43 (m, 2H), 2.75 (d, J=15.4 Hz, 2H), 2.66 (d, J=15.4 Hz, 2H), 1.93 (m, 4H).

EXAMPLE 8

((S)-1-(4-(3-chloro-4-methoxybenzylamino)-5-(pyrimidin-2-ylmethylcarbamoyl) pyrimidin-2-yl)pyrrolidin-2-yl)methyl 3-((S)-2,3-bis(nitrooxy)propoxy)propanoate 2-hydroxypropane-1,2,3-tricarboxylate (Compound (12)-(2S)-isomer, Citrate Salt)

[0223] ##STR00054##

[0224] To a solution of compound 12, (2S)-isomer, prepared as described in Example 4 (227 mg, 0.315 mmol) in Methanol (2 ml), citric acid monohydrate (66 mg, 0.315 mmol) was added. The mixture was stirred for 10 minutes at room temperature, then concentrated and the residue was washed with diethyl ether. The solid was filtered and dried under reduced pressure, giving 280 mg of the desired product (Compound (12)-(2S)-isomer, citrate salt) (crude yield: 97.4%).

[0225] .sup.1H NMR (600 MHz, DMSO-d6) δ 12.35 (s, 3H), 9.18 (s, 1H), 8.77 (m, 2H), 8.55 (m, 1H), 7.46-7.33 (m, 1H), 7.34-7.17 (m, 1H), 7.06 (dd, J=16.6, 8.3 Hz, 1H), 5.55 (s, 1H), 4.81 (m, 4H), 4.63-4.42 (m, 4H), 4.38-3.96 (m, 3H), 3.81 (s, 3H), 3.71 (m, 2H), 3.57-3.42 (m, 2H), 2.75 (d, J=15.4 Hz, 2H), 2.65 (d, J=15.4 Hz, 2H), 2.57 (t, J=5.9 Hz, 2H), 1.92 (m, 4H).

EXAMPLE 9

(S)-2-(4-(3-(5-ethyl-4-oxo-7-propyl-4,5-dihydro-3H-pyrrolo[3,2-d]pyrimidin-2-yl)-4-propoxyphenylsulfonyl)piperazin-1-yl)ethyl 3-[2,3-bis(nitrooxy)propoxy]propanoate citrate (Compound (1)-(2S)-isomer, Citrate Salt)

[0226] ##STR00055##

[0227] To a solution of Mirodenafil (500 mg, 0.940 mmol) and 4-nitrophenyl 3-[(2S)-2,3-bis(nitrooxy)propoxy]propanoate (see Example 4) (423 mg, 1.128 mmol) in dry DCM (5 ml), DMAP (115 mg, 0.940 mmol) was added.

[0228] The mixture was stirred at room temperature for 48h, then it was diluted with DCM and water. The two phases were separated and the aqueous phase extracted twice with DCM. The combined organic phases were dried over MgSO.sub.4 and concentrated under reduced pressure. The residue was purified by reversed-phase chromatography (H.sub.2O/CH.sub.3CN with 0.01% formic acid, from 70:30 to 20:80).

[0229] After purification DCM and a saturated solution of NaHCO.sub.3 were added to the residue. The two phases were separated and the aqueous phase extracted twice with DCM. The combined organic phases were dried over MgSO.sub.4 and concentrated under reduced pressure, giving 540 mg of (S)-2-(4-(3-(5-ethyl-4-oxo-7-propyl-4,5-dihydro-3H-pyrrolo[3,2-d]pyrimidin-2-yl)-4-propoxyphenylsulfonyl)piperazin-1-yl)ethyl 3-(3-nitro-2-(nitrooxy)propoxy)propanoate (compound (1), yield: 75.0%).

[0230] Compound (1) (295 mg, 0.384 mmol) was re-dissolved in Methanol (1.5 ml), and citric acid monohydrate (81 mg, 0.384 mmol) was added. The mixture was stirred for 10 minutes at room temperature, then concentrated and the solid washed with diethyl ether. The solid was filtered and dried under reduced pressure, giving 361 mg of the title compound (Compound (1)-(2S)-isomer, citrate salt) (crude yield: 98%).

[0231] .sup.1H NMR (400 MHz, DMSO-d6) δ 2.30 (bs, 1H), 7.89 (d, J=2.4 Hz, 1H), 7.80 (dd, J=8.8, 2.5 Hz, 1H), 7.38 (d, J=8.9 Hz, 1H), 7.31 (s, 1H), 5.53 m, J=10.3, 6.7, 3.9 Hz, 1H), 4.88 (dd, J=12.8, 3.0 Hz, 1H), 4.72 (dd, J=12.9, 6.5 Hz, 1H), 4.37 (q, J=7.1 Hz, 2H), 4.10 (dt, J=11.4, 6.0 Hz, 4H), 3.77-3.58 (m, 4H), 2.90 (s, 4H), 2.75 (d, J=15.4 Hz, 2H), 2.65 (d, J=15.4 Hz, 2H), 2.61-2.52 (m, 10H), 1.80-1.58 (m, 4H), 1.36 (t, J=7.1 Hz, 3H), 0.97 (t, 3H), 0.94 (t, 3H).

EXAMPLE 10

2-(4-(3-(5-ethyl-4-oxo-7-propyl-4,5-dihydro-3H-pyrrolo[3,2-d]pyrimidin-2-yl)-4-propoxyphenylsulfonyl)piperazin-1-yl)ethyl 6-(nitrooxy)hexanoate citrate (Compound (4) Citrate Salt)

[0232] ##STR00056##

[0233] To a solution of Mirodenafil (500 mg, 0.940 mmol) in dry DCM (5 ml), DMAP (115 mg, 0.940 mmol), DCC (233 mg, 1.128 mmol) and 6-(nitrooxy)hexanoic acid (200 mg, 1.128 mmol) were added.

[0234] The mixture was stirred overnight at room temperature and then diluted with DCM and water. The two phases were separated and the aqueous layer was extracted twice with DCM. The combined organic extracts were dried over MgSO.sub.4 and concentrated under reduced pressure. The residue was purified by reversed-phase chromatography (H.sub.2O/CH.sub.3CN with 0.1% Formic acid, from 70:30 to 20:80).

[0235] After purification DCM and a saturated solution of NaHCO.sub.3 were added to the residue. The two phases were separated and the aqueous phase extracted twice with DCM. The combined organic phases were dried over MgSO.sub.4 and concentrated under reduced pressure, giving 450 mg of 2-(4-(3-(5-ethyl-4-oxo-7-propyl-4,5-dihydro-3H-pyrrolo[3,2-d]pyrimidin-2-yl)-4-propoxyphenylsulfonyl)piperazin-1-yl)ethyl 6-(nitrooxy)hexanoate (compound (4), yield: 69.0%).

[0236] To a solution of Compound (4) (200 mg, 0.290 mmol) in Methanol (1.5 ml), citric acid monohydrate (61 mg, 0.290 mmol) was added. The mixture was stirred for 10 minutes at room temperature, then concentrated and the solid washed with diethyl ether. The solid was filtered and dried under reduced pressure, giving 253 mg of the Title compound (Compound (4) citrate salt) (yield 98.8%).

[0237] .sup.1H NMR (400 MHz, DMSO-d6) δ 11.67 (s, 1H), 7.90 (d, J=2.4 Hz, 1H), 7.80 (dd, J=8.8, 2.5 Hz, 1H), 7.38 (d, J=8.9 Hz, 1H), 7.31 (s, 1H), 4.47-4.32 (m, 4H), 4.09 (dt, J=11.3, 6.0 Hz, 4H), 2.90 (s, 4H), 2.75 (d, J=15.4 Hz, 2H), 2.65 (d, J=15.4 Hz, 2H), 2.55 (ddd, J=11.8, 8.4, 4.7 Hz, 8H), 2.26 (t, J=7.3 Hz, 2H), 1.81-1.69 (m, 2H), 1.69-1.55 (m, 4H), 1.50 (m, 2H), 1.39-1.33 (m, 3H), 1.29 (q, 2H), 0.94 (m, 6H).

EXAMPLE 11

2-hydroxypropane-1,2,3-tricarboxylic acid 2-{4-[3-(5-ethyl-4-oxo-7-propyl-4,5-dihydro-3H-pyrrolo[3,2-d]pyrimidin-2-yl)-4-propoxybenzene-1-sulfonyl]piperazin-1-yl}ethyl [2-(nitrooxy)ethoxy]acetate (Compound (2) Citrate Salt)

[0238] ##STR00057##

[0239] To a solution of Mirodenafil (500 mg, 0.940 mmol) in dry DCM (5 ml), DMAP (115 mg, 0.940 mmol), DCC (233 mg, 1.128 mmol) and [2-(nitrooxy)ethoxy]acetic acid (186 mg, 1.128 mmol) were added.

[0240] The mixture was stirred at room temperature for 48h then it was diluted with DCM and water. The two phases were separated and the aqueous phase extracted twice with DCM. The combined organic phases were dried over MgSO.sub.4 and concentrated under reduced pressure. The residue was purified by reversed-phase chromatography (H.sub.2O/CH.sub.3CN with 0.1% formic acid, from 70:30 to 40:60).

[0241] After purification DCM and a saturated solution of NaHCO.sub.3 were added to the residue. The two phases were separated and the aqueous phase extracted twice with DCM. The combined organic phases were dried over MgSO.sub.4 and concentrated under reduced pressure, giving 338 mg of 2-{4-[3-(5-ethyl-4-oxo-7-propyl-4,5-dihydro-3H-pyrrolo[3,2-d]pyrimidin-2-yl)-4-propoxybenzene-1-sulfonyl]piperazin-1-yl}ethyl [2-(nitrooxy) ethoxy]acetate (compound (2), yield: 53.0%).

[0242] To a solution of Compound (2) (172 mg, 0.253 mmol) in Methanol (1.5 ml), citric acid monohydrate (53 mg, 0.276 mmol) was added. The mixture was stirred 10 minutes at room temperature, then concentrated and the solid washed with diethyl ether. The solid was filtered and dried under reduced pressure, giving 220 mg of the title compound (Compound (2) citrate salt) (crude yield: 99.8%).

[0243] .sup.1H NMR (400 MHz, DMSO-d6) δ 11.68 (s, 1H), 7.90 (t, J=5.8 Hz, 1H), 7.80 (m, 1H), 7.38 (d, J=8.9 Hz, 1H), 7.31 (s, 1H), 4.67-4.59 (m, 2H), 4.45-4.27 (m, 2H), 4.20-4.04 (m, 6H), 3.78-3.71 (q, 2H), 2.90 (s, 4H), 2.75 (t, J=15.4 Hz, 2H), 2.65 (t, J=15.4 Hz, 2H), 2.61-2.52 (m, 8H), 1.81-1.55 (m, 4H), 1.36 (t, J=7.1 Hz, 3H), 0.94 (t, 6H).

EXAMPLE 12

(S)-2-(4-(3-(5-ethyl-4-oxo-7-propyl-4,5-dihydro-3H-pyrrolo[3,2-d]pyrimidin-2-yl)-4-propoxyphenylsulfonyl)piperazin-1-yl)ethyl 5,6-bis(nitrooxy)hexanoate Citrate (Compound (3)-(5S) Isomer, Citrate Salt)

[0244] ##STR00058##

[0245] To a solution of Mirodenafil (500 mg, 0.940 mmol) in dry DCM (5 ml), DMAP (115 mg, 0.940 mmol), DCC (233 mg, 1.128 mmol) and (5S)-5,6-bis(nitrooxy)hexanoic acid (270 mg, 1.128 mmol) were added.

[0246] The mixture was stirred overnight at room temperature, then diluted with DCM and water. The two phases were separated and the aqueous phase extracted twice with DCM. The combined organic phases were dried over MgSO.sub.4 and concentrated under reduced pressure. The residue was purified by reversed-phase chromatography (H.sub.2O/CH.sub.3CN with 0.1% formic acid, from 70:30 to 40:60).

[0247] After purification DCM and a saturated solution of NaHCO.sub.3 were added to the residue. The two phases were separated and the aqueous phase extracted twice with DCM. The combined organic phases were dried over MgSO.sub.4 and concentrated under reduced pressure, giving 550 mg of (S)-2-(4-(3-(5-ethyl-4-oxo-7-propyl-4,5-dihydro-3H-pyrrolo[3,2-d]pyrimidin-2-yl)-4-propoxyphenylsulfonyl)piperazin-1-yl)ethyl 5,6-bis(nitrooxy)hexanoate (Compound (3), yield: 77.8%).

[0248] To a solution of Compound (3) (207 mg, 0.275 mmol) in Methanol (1.5 ml), citric acid monohydrate (58 mg, 0.302 mmol) was added. The mixture was stirred for 10 minutes at room temperature, then concentrated and the solid washed with diethyl ether. The solid was filtered and dried under reduced pressure, giving 254 mg of the title compound (Compound (3)-(5S) isomer, citrate salt) (yield: 97.8%).

[0249] .sup.1H NMR (400 MHz, DMSO-d6) δ 11.67 (s, 1H), 7.89 (d, J=2.4 Hz, 1H), 7.80 (dd, J=8.8, 2.5 Hz, 1H), 7.38 (d, J=8.9 Hz, 1H), 7.31 (s, 1H), 5.38 (m, J=13.1, 5.9, 2.6 Hz, 1H), 4.89 (dd, J=12.9, 2.6 Hz, 2H), 4.70 (m, 2H), 4.37 (q, J=7.1 Hz, 2H), 4.09 (dt, J=11.3, 6.0 Hz, 4H), 2.90 (s, 4H), 2.75 (t, 2H), 2.65 (d, J=15.4 Hz, 2H), 2.61-2.52 (m, 8H), 2.32 (t, J=7.1 Hz, 2H), 1.82-1.55 (m, 8H), 1.36 (t, J=7.1 Hz, 3H), 0.94 (t, 6H).

EXAMPLE 13

2-(4-(3-(5-ethyl-4-oxo-7-propyl-4,5-dihydro-3H-pyrrolo[3,2-d]pyrimidin-2-yl)-4-propoxyphenylsulfonyl)piperazin-1-yl)ethyl 6-(nitrooxy)hexyl Carbonate (Compound (5))

[0250] ##STR00059##

Step 1: 2-(4-((3-(5-ethyl-4-oxo-7-propyl-4,5-dihydro-3H-pyrrolo[3,2-d]pyrimidin-2-yl)-4-propoxyphenyl)sulfonyl)piperazin-1-yl)ethyl (4-nitrophenyl) Carbonate

[0251] To a solution of Mirodenafil (290 mg, 0.545 mmol) in dry DCM (5 ml), 4-Nitrophenyl chloroformate (121 mg, 0.600 mmol) and DMAP (74 mg, 0.600 mmol) were added. The mixture was stirred overnight at room temperature, then it was diluted with DCM and water. The two phases were separated and the aqueous phase extracted twice with DCM. The combined organic phases were dried over MgSO.sub.4 and concentrated under reduced pressure. The residue was purified by flash chromatography (Cyclohexane:EtOAc:MeOH 5:5:0.1%). Then the fraction was collected and dried under reduced pressure, affording 310 mg of the title compound (2-(4-((3-(5-ethyl-4-oxo-7-propyl-4,5-dihydro-3H-pyrrolo[3,2-d]pyrimidin-2-yl)-4-propoxyphenyl)sulfonyl) piperazin-1-yl)ethyl (4-nitrophenyl) carbonate) (yield: 81.7%) as a colorless oil.

Step 2: 2-(4-(3-(5-ethyl-4-oxo-7-propyl-4,5-dihydro-3H-pyrrolo[3,2-d]pyrimidin-2-yl)-4-propoxyphenylsulfonyl)piperazin-1-yl)ethyl 6-(nitrooxy)hexyl Carbonate

[0252] To a solution of 2-(4-((3-(5-ethyl-4-oxo-7-propyl-4,5-dihydro-3H-pyrrolo[3,2-d]pyrimidin-2-yl)-4-propoxyphenyl)sulfonyl)piperazin-1-yl)ethyl (4-nitrophenyl) carbonate (150 mg, 0.21 mmol)) in dry DCM (1 ml), DMAP (30 mg, 0.24 mmol) and 6-hydroxyhexyl nitrate (39 mg, 0.24 mmol) were added. The mixture was stirred overnight at room temperature, then it was diluted with DCM and water. The two phases were separated and the aqueous phase extracted twice with DCM. The combined organic phases were dried over MgSO.sub.4 and concentrated under reduced pressure. The residue was purified by reversed-phase chromatography (H.sub.2O/CH.sub.3CN with 0.1% formic acid, from 90:10 to 20:80). Dichloromethane and a saturated solution of NaHCO.sub.3 were added to the residue after purification. The two phases were separated and the aqueous phase extracted twice with DCM. Then the fraction was collected and dried under reduced pressure, affording 150 mg of the title compound (Compound (5)) (yield: 97%) as a sticky oil.

[0253] .sup.1H NMR (600 MHz, DMSO-d6) δ 11.65 (s, 1H), 7.89 (d, J=2.4 Hz, 1H), 7.80 (dd, J=8.8, 2.4 Hz, 1H), 7.38 (d, J=8.9 Hz, 1H), 7.31 (s, 1H), 4.47 (t, J=6.6 Hz, 2H), 4.37 (q, J=7.1 Hz, 2H), 4.13 (t, J=5.9 Hz, 2H), 4.00 (t, J=6.6 Hz, 2H), 2.88 (t, 2H), 2.60-2.51 (m, 8H), 1.79-1.70 (m, 2H), 1.68-1.57 (m, 4H), 1.56-1.48 (m, 2H), 1.36 (t, J=7.2 Hz, 3H), 1.33-1.22 (m, 4H), 0.97 (t, J=7.4 Hz, 3H), 0.92 (t, J=7.3 Hz, 3H).

EXAMPLE 14

(S)-5,6-bis (nitrooxy)hexyl 2-(4-(3-(5-ethyl-4-oxo-7-propyl-4,5-dihydro-3H-pyrrolo [3,2-d]pyrimidin-2-yl)-4-propoxyphenylsulfonyl)piperazin-1-yl)ethyl Carbonate (Compound (6))

[0254] ##STR00060##

[0255] To a solution of 2-(4-((3-(5-ethyl-4-oxo-7-propyl-4,5-dihydro-3H-pyrrolo[3,2-d]pyrimidin-2-yl)-4-propoxyphenyl)sulfonyl)piperazin-1-yl)ethyl (4-nitrophenyl) carbonate, prepared as described in Example 13, Step 1 (200 mg, 0.29 mmol)) in dry DCM (2 ml), DMAP (39 mg, 0.32 mmol) and 6-hydroxyhexane-1,2-diyl dinitrate (72 mg, 0.32 mmol) were added.

[0256] The mixture was stirred overnight at room temperature then it was diluted with DCM and water. The two phases were separated and the aqueous phase extracted twice with DCM. The combined organic phases were dried over MgSO.sub.4 and concentrated under reduced pressure. The residue was purified by reversed-phase chromatography (H.sub.2O/CH.sub.3CN with 0.1% formic acid, from 85:15 to 20:80). Dichloromethane and a saturated solution of NaHCO.sub.3 were added to the residue after purification. The two phases were separated and the aqueous phase extracted twice with DCM. Then the fraction was collected and dried under reduced pressure, affording 150 mg of the desired product (Compound (6)) (yield: 73.5%) as a sticky oil.

[0257] .sup.1H NMR (400 MHz, DMSO-d6) δ 11.66 (s, 1H), 7.88 (d, J=2.4 Hz, 1H), 7.80 (dd, J=8.8, 2.5 Hz, 1H), 7.38 (d, J=8.9 Hz, 1H), 7.31 (s, 1H), 5.38 (dt, J=6.2, 4.3 Hz, 1H), 4.90 (dd, J=12.9, 2.6 Hz, 1H), 4.67 (dd, J=12.9, 6.1 Hz, 1H), 4.37 (q, J=7.2 Hz, 2H), 4.20-3.96 (m, 6H), 2.75 (m, 6H), 2.62-2.51 (m, 8H), 1.82-1.52 (m, 8H), 1.46-1.29 (m, 3H), 0.94 (t, J=7.4 Hz, 6H).

EXAMPLE 15

(S)-(1-(4-(3-chloro-4-methoxybenzylamino)-5-(pyrimidin-2-ylmethylcarbamoyl) pyrimidin-2-yl)pyrrolidin-2-yl)methyl 6-(nitrooxy)hexyl Carbonate (Compound (13))

[0258] ##STR00061##

Step 1: (S)-(1-(4-((3-chloro-4-methoxybenzyl)amino)-5-((pyrimidin-2-ylmethyl) carbamoyl)pyrimidin-2-yl)pyrrolidin-2-yl)methyl (4-nitrophenyl) Carbonate

[0259] To a solution of Avanafil (500 mg, 1.033 mmol) in dry DCM (10 ml), 4-Nitrophenyl chloroformate (416 mg, 2.066 mmol) and Pyridine (184 μL, 2.346 mmol) were added. The mixture was stirred for 2h at room temperature then diluted with DCM and water. The two phases were separated and the aqueous layer extracted twice with DCM, giving 1.02 g of the desired product ((S)-(1-(4-((3-chloro-4-methoxybenzyl)amino)-5-((pyrimidin-2-ylmethyl) carbamoyl)pyrimidin-2-yl)pyrrolidin-2-yl)methyl (4-nitrophenyl) carbonate).

Step 2: (S)-(1-(4-(3-chloro-4-methoxybenzylamino)-5-(pyrimidin-2-ylmethyl carbamoyl)pyrimidin-2-yl)pyrrolidin-2-yl)methyl 6-(nitrooxy)hexyl Carbonate

[0260] To a solution of (S)-(1-(4-((3-chloro-4-methoxybenzyl)amino)-5-((pyrimidin-2-ylmethyl)carbamoyl)pyrimidin-2-yl)pyrrolidin-2-yl)methyl (4-nitrophenyl) carbonate (1.02 g, 1.57 mmol) in dry DCM (20 ml), DMAP (211 mg, 1.73 mmol) and 6-hydroxyhexyl nitrate (282 mg, 1.73 mmol) were added. The mixture was stirred overnight at room temperature, then it was diluted with DCM and water. The two phases were separated and the aqueous phase extracted twice with DCM. The combined organic phases were dried over MgSO.sub.4 and concentrated under reduced pressure. The residue was purified by flash chromatography (DCM/iPrOH from 100:0 to 90:10). Then the fraction was collected and dried under reduced pressure, affording 405 mg of title compound (Compound (13)) (yield: 38%) as a sticky oil.

[0261] .sup.1H NMR (600 MHz, DMSO-d6) δ 9.17 (t, J=6.0 Hz, 1H), 8.79 (m, 1H), 8.75 (d, J=4.9 Hz, 2H), 8.55 (m, 1H), 7.45-7.15 (m, 3H), 7.04 (d, J=8.3 Hz, 1H), 4.61-4.43 (m, 6H), 4.29 (m, 3H), 4.05 (t, J=6.4 Hz, 2H), 3.81 (s, 3H), 3.56-3.42 (m, 2H), 2.05-1.80 (m, 4H), 1.60 (m, 4H), 1.32 (t, 4H).

EXAMPLE 16

(S)-2-(4-(3-(5-ethyl-4-oxo-7-propyl-4,5-dihydro-3H-pyrrolo[3,2-d]pyrimidin-2-yl)-4-propoxyphenylsulfonyl)piperazin-1-yl)ethyl 3-(2,3-bis(nitrooxy)propoxy)propanoate Dihydrochloride (Compound (1)-(2S) Dihydrochloride Salt)

[0262] ##STR00062##

[0263] To a solution of Compound (1), obtained in Example 9 (72 mg, 0.094 mmol) in Methanol (1.0 ml), HCl 3M methanolic solution (63 uL, 0.188 mmol) was added. The mixture was stirred for 10 minutes at room temperature, then concentrated and the solid washed with Et.sub.2O. The solid was filtered and dried under reduced pressure, giving 68 mg of the title compound (Compound (1)-(2S) dihydrochloride salt) (yield: 86%).

[0264] .sup.1H NMR (600 MHz, DMSO-d6) δ 11.73 (s, 1H), 11.05 (s, 1H), 8.07-7.76 (m, 2H), 7.42 (d, J=8.7 Hz, 1H), 7.32 (s, 1H), 5.59-5.52 (m, 1H), 4.90 (dd, J=12.8, 2.8 Hz, 1H), 4.75 (dd, J=12.8, 6.5 Hz, 1H), 4.38 (q, J=7.1 Hz, 3H), 4.14 (t, J=6.3 Hz, 2H), 3.77-3.66 (m, 6H), 2.60-2.49 (m, 14H), 1.80-1.72 (m, 2H), 1.69-1.60 (m, 2H), 1.36 (t, J=7.2 Hz, 3H), 0.97 (t, J=7.4 Hz, 3H), 0.93 (t, J=7.3 Hz, 3H).

EXAMPLE 17

((S)-1-(4-(3-chloro-4-methoxybenzylamino)-5-(pyrimidin-2-ylmethylcarbamoyl) pyrimidin-2-yl)pyrrolidin-2-yl)methyl (S)-5,6-bis(nitrooxy)hexyl Carbonate (Compound (14)-(5S) Isomer)

[0265] ##STR00063##

[0266] To a solution of (S)-(1-(4-((3-chloro-4-methoxybenzyl)amino)-5-((pyrimidin-2-ylmethyl)carbamoyl)pyrimidin-2-yl)pyrrolidin-2-yl)methyl (4-nitrophenyl) carbonate (prepared as described in Example 15, Step 1 (758 mg, 1.167 mmol) in dry DCM (10 ml), DMAP (157 mg, 1.284 mmol) and (2S)-6-hydroxyhexane-1,2-diyl dinitrate (288 mg, 1.284 mmol) were added.

[0267] The mixture was stirred overnight at room temperature and then diluted with DCM and water. The two phases were separated and the aqueous layer extracted twice with DCM. The combined organic phases were dried over MgSO.sub.4 and concentrated under reduced pressure. The residue was purified by flash chromatography (DCM/iPrOH from 100:0 to 90:10) and then by reversed-phase chromatography (H.sub.2O/CH.sub.3CN, from 100:0 to 0:100), giving 334 mg of the desired product (Compound (14)-(5S) isomer) (yield: 39%) as a sticky solid.

[0268] .sup.1H NMR (600 MHz, DMSO-d6) δ 9.17 (d, J=5.5 Hz, 1H), 8.79 (m, 1H), 8.75 (d, J=4.9 Hz, 2H), 8.55 (s, 1H), 7.44-7.16 (m, 3H), 7.04 (d, J=7.7 Hz, 1H), 5.41 (m, 1H), 4.92 (d, J=12.8 Hz, 1H), 4.69 (dd, J=12.7, 6.0 Hz, 1H), 4.64-4.42 (m, 4H), 4.39-4.14 (m, 3H), 4.06 (m, 2H), 3.81 (m, 3H), 3.49 (m, 2H), 1.92 (m, 4H), 1.66 (m, 4H), 1.42 (m, 2H).

EXAMPLE 18

(S)-(1-(4-(3-chloro-4-methoxybenzylamino)-5-(pyrimidin-2-ylmethylcarbamoyl) pyrimidin-2-yl)pyrrolidin-2-yl)methyl 6-(nitrooxy) hexylcarbonate 2-hydroxypropane-1,2,3-tricarboxylate (Compound (13) Citrate Salt)

[0269] ##STR00064##

[0270] To a solution of (S)-(1-(4-(3-chloro-4-methoxybenzylamino)-5-(pyrimidin-2-ylmethylcarbamoyl)pyrimidin-2-yl)pyrrolidin-2-yl)methyl 6-(nitrooxy)hexyl carbonate (Compound (13), prepared as described in Example 15 (218 mg, 0.324 mmol) in Methanol (1.0 ml), citric acid monohydrate (68 mg, 0.324 mmol) was added. The mixture was stirred for 10 minutes at room temperature, then concentrated and the solid washed with MTBE. The solid was filtered and dried under reduced pressure, giving 265 mg of the desired product (Compound (13) citrate salt) (crude yield: 94.5%).

[0271] .sup.1H NMR (400 MHz, DMSO-d6) δ 9.19 (t, J=5.8 Hz, 1H), 8.80 (m, 1H), 8.75 (d, J=4.9 Hz, 2H), 8.54 (m, 1H), 7.38 (t, J=4.9 Hz, 1H), 7.35-7.15 (m, 2H), 7.06 (s, 1H), 4.64-4.43 (m, 6H), 4.29 (m, 3H), 4.05 (t, J=6.5 Hz, 2H), 3.81 (s, 3H), 3.49 (t, J=7.1 Hz, 2H), 2.76 (d, J=15.4 Hz, 2H), 2.65 (d, J=15.4 Hz, 2H), 2.06-1.81 (m, 4H), 1.61 (m, 4H), 1.33 (m, 4H).

EXAMPLE 19

2-{4-[4-ethoxy-3-(1-methyl-7-oxo-3-propyl-6,7-dihydro-1H-pyrazolo[4,3-d]pyrimidin-5-yl)benzene-1-sulfonyl]piperazin-1-yl}ethyl 6-(nitrooxy)hexanoate (Compound (8b))

[0272] ##STR00065##

[0273] To a solution of Lodenafil (400 mg, 0.793 mmol) in dry DCM (5 ml), DMAP (97 mg, 0.793 mmol), DCC (196 mg, 0.951 mmol) and 6-(nitrooxy)hexanoic acid (168 mg, 0.951 mmol) were added.

[0274] The mixture was stirred overnight at room temperature then it was diluted with DCM and water. The two phases were separated and the aqueous phase extracted twice with DCM. The combined organic phases were dried over MgSO.sub.4 and concentrated under reduced pressure. The residue was purified by reversed-phase chromatography (H.sub.2O/CH.sub.3CN with 0.01% formic acid, from 90:10 to 20:80). Dichloromethane and a saturated solution of NaHCO.sub.3 were added to the residue after purification. The two phases were separated and the aqueous phase extracted twice with DCM. The combined organic phases were dried over MgSO.sub.4 and concentrated under reduced pressure, affording 150 mg of the desired product (Compound (8b)) (yield: 69.0%).

[0275] .sup.1H NMR (400 MHz, DMSO-d6) δ 12.17 (s, 1H), 7.81 (m, 2H), 7.36 (d, J=8.7 Hz, 1H), 4.43 (t, J=6.6 Hz, 2H), 4.19 (q, J=6.9 Hz, 2H), 4.14 (s, 3H), 4.03 (t, J=5.7 Hz, 2H), 2.86 (bs, 4H), 2.75 (t, J=7.5 Hz, 2H), 2.58-2.37 (m, 5H), 2.23 (t, J=7.3 Hz, 2H), 1.72 (dd, J=14.9, 7.4 Hz, 2H), 1.63-1.53 (m, 2H), 1.53-1.40 (m, 2H), 1.35-1.20 (m, 6H), 0.92 (t, J=7.4 Hz, 3H).

EXAMPLE 20

(S)-2-(4-((4-ethoxy-3-(1-methyl-7-oxo-3-propyl-6,7-dihydro-1H-pyrazolo[4,3-d]pyrimidin-5-yl)phenyl)sulfonyl)piperazin-1-yl)ethyl (S)-5,6-bis(nitrooxy)hexanoate (Compound (7b)-(5S)) Isomer)

[0276] ##STR00066##

[0277] To a solution of Lodenafil (400 mg, 0.793 mmol) in dry DCM (5 ml), DMAP (97 mg, 0.793 mmol), DCC (196 mg, 0.951 mmol) and (5S)-5,6-bis(nitrooxy)hexanoic acid (226 mg, 0.951 mmol) were added. The mixture was stirred overnight at room temperature, then it was diluted with DCM and water. The two phases were separated and the aqueous phase extracted twice with DCM. The combined organic phases were dried over MgSO.sub.4 and concentrated under reduced pressure. The residue was purified by reversed-phase chromatography (H.sub.2O/CH.sub.3CN with 0.01% Formic acid, from 85:15 to 10:90). Dichloromethane and a saturated solution of NaHCO.sub.3 were added to the residue after purification. The two phases were separated and the aqueous phase extracted twice with DCM. The combined organic phases were dried over MgSO.sub.4 and concentrated under reduced pressure, affording 150 mg of the desired product (Compound (7b)-(5S)) isomer) (yield: 78.0%).

[0278] .sup.1H NMR (400 MHz, DMSO-d6) δ 12.17 (s, 1H), δ 7.85-7.76 (m, 2H), 7.36 (d, J=8.7 Hz, 1H), 5.36 (m, J=6.0, 3.4 Hz, 1H), 4.87 (dd, J=12.9, 2.6 Hz, 1H), 4.65 (dd, J=12.9, 5.9 Hz, 1H), 4.26-4.12 (m, 5H), 4.02 (t, J=5.7 Hz, 2H), 2.87 (bs, 4H), 2.75 (t, J=7.6 Hz, 2H), 2.59-2.41 (m, 3H), 2.30 (t, J=7.1 Hz, 2H), 2.08 (s, 2H), 1.78-1.33 (m, 7H), 1.32 (t, J=6.9 Hz, 3H), 0.92 (t, J=7.4H).

EXAMPLE 21—(COMPARATIVE EXAMPLE)—PDE5 INHIBITOR NITRATE ESTER

2-(4-(3-(5-ethyl-4-oxo-7-propyl-4,5-dihydro-3H-pyrrolo[3,2-d]pyrimidin-2-yl)-4-propoxyphenylsulfonyl)piperazin-1-yl)ethyl Nitrate

[0279] ##STR00067##

[0280] The nitrate ester of mirodenafil was prepared from mirodenafil following procedure described in WO 2017/085056 (Example 29): the hydroxyl group of mirodenafil was first transformed into the corresponding bromide with known method (PPh.sub.3/NBS) and then nitrated with Silver nitrate in acetonitrile.

[0281] .sup.1H NMR (600 MHz, CHLOROFORM-d) δ ppm 10.71 (br s, 1H) 7.80 (dd, J=8.65, 2.49 Hz, 1H) 7.15 (d, J=8.80 Hz, 1H) 5.30 (s, 1H) 4.41-4.55 (m, 5H) 4.25 (t, J=6.45 Hz, 3H) 3.11 (br s, 5H) 2.72 (t, J=7.48 Hz, 4H) 2.53-2.69 (m, 5H) 2.00-2.09 (m, 3H) 1.74 (sxt, J=7.39 Hz, 4H) 1.48 (t, J=7.19 Hz, 5H) 1.20 (t, J=7.33 Hz, 5H) 1.00 (t, J=7.33 Hz, 5H).

EXAMPLE 22

[0282] Intraocular Pressure (IOP) Lowering Effect in Laser-Induced Ocular Hypertensive Non-Human Primates

[0283] In this study the efficacy in lowering intraocular pressure of a compound of formula (Ia), compound (1)-(2S)-citrate salt (disclosed in Example 9), the NO-donor 3-[(2S)-2,3-bis(nitrooxy)propoxy]propanoic acid ((IIIg)-OH) and the reference PDE5 inhibitor (mirodenafil) were evaluated in laser-induced ocular hypertensive non-human primates.

Experimental Procedure

[0284] Compound (1)-(2S)-citrate salt (0.48 μmoles/eye equal to 1.59% w/v, 30 μL), Mirodenafil (0.48 μmoles/eye equal to 1% w/v, 30 μL), NO-donor ((IIIg)-OH=3-[(2S)-2,3-bis(nitrooxy)propoxylpropanoic acid; 1.17 μmoles/eye equal to 1% w/v, 30 μL) or vehicle (phosphate buffer pH 6, cremophor EL 5% w/v, DMSO 0.3% w/v, benzalkonium chloride 0.02% w/v) were administered to laser induced ocular hypertensive non-human primates in a masked fashion. The contralateral eye was not treated.

[0285] Intraocular pressure was measured at selected time points using a pneumatonometer (Model 30™ Reichert, Depew, N.Y., USA) prior to (basal) and at selected time point following ocular dosing. One topical drop of proparacaine HCl 0.13% was applied to the eye prior to each intraocular pressure measurement.

[0286] Results

[0287] Data are reported as mean±SEM of n=13. Efficacy (E) as IOP change was calculated versus basal and vehicle values using the following equation: (IOP.sub.drug−IOP.sub.basal drug)−(IOP.sub.veh−IOP.sub.basal veh). Values reported refer to maximum IOP change (E.sub.max) and the IOP lowering effect detected at 300 min post-dose (E.sub.300min).

[0288] The results demonstrated that the nitric oxide releasing phosphodiesterase type 5 inhibitor of the invention (compound (1)-(2S)-citrate salt) showed higher maximum intraocular-pressure lowering effect (E.sub.max) and a longer duration with respect to the single administrations of the compound (IIIg)-OH (NO-donor) and mirodenafil (reference PDE5 inhibitor). Of note, the NO-donor (IIIg-OH) was administered at doses 2.4 times higher than the compound of the invention ((1)-(2S) citrate salt).

TABLE-US-00001 TABLE 1 IOP lowering activity following topical dosing in laser-induced ocular hypertensive non-human primates Amount administered IOP change (mmHg) Compound (μmoles/eye) E.sub.max/time post-osing E.sub.300 min (1)-(2S)- 0.48 −7.0 ± 0.9 (60 min) −5.3 ± 1.9 citrate salt Mirodenafil 0.48 −5.0 ± 2.1 (180 min) −3.4 ± 2.3 (IIIg)-OH 1.17 −6.0 ± 2.7 (180 min) −2.8 ± 2.8

EXAMPLE 23

[0289] Intraocular Pressure (IOP) Lowering Effect in Laser-Induced Ocular Hypertensive Non-Human Primates

[0290] In independent experiments the efficacies in lowering intraocular pressure of a compound of formula (II), compound (9)-citrate salt (disclosed in Example 5), and the NO-donor 6-(nitrooxy)hexanoic acid) ((IIIa)-OH) were evaluated in laser-induced ocular hypertensive non-human primates.

Experimental Procedure

[0291] Compound (9)-citrate salt (0.79 μmoles/eye equal to 2.2% w/v, 30 μL) or the NO-donor ((IIIa)-OH=6-(nitrooxy)hexanoic acid; (0.79 μmoles/eye equal to 0.47% w/v, 30 μL) or vehicle (phosphate buffer pH 6, cremophor EL 5% w/v, DMSO 0.3% w/v, benzalkonium chloride 0.02% w/v) were administered to laser induced ocular hypertensive non-human primates. The contralateral eye was not treated.

[0292] Intraocular pressure was measured using a pneumatonometer (Model 30™ Reichert, Depew, N.Y., USA) prior to (basal) and at selected time point following ocular dosing. One topical drop of proparacaine HCl 0.13% was applied to the eye prior to each intraocular pressure measurement.

[0293] Results

[0294] Efficacy (E) as TOP change was calculated versus basal and vehicle values using the following equation: (IOP.sub.drug−IOP.sub.basal drug)−(IOP.sub.veh−IOP.sub.basal veh) where TOP values are taken at 30, 60, 180, 300 and 480 minutes post-dosing.

[0295] The results reported in Table 2 demonstrated that the nitric oxide releasing phosphodiesterase type 5 inhibitor of the invention (compound (9)-citrate salt) showed higher maximum intraocular-pressure lowering efficacy (compare E.sub.max of compound (IIIa)-OH of −5.2±1.0 mmHg at 60 min to the E.sub.max of compound (9)-citrate salt of −6.0±2.1 at 480 min) and a longer duration (compare E.sub.480 of compound (IIIa)-OH of −2.7±1.5 mmHg to that of compound (9)-citrate salt of −6.0±2.1 mmHg at the same time-point).

TABLE-US-00002 TABLE 2 IOP lowering activity following topical dosing in laser-induced ocular hypertensive non-human primates Amount administered IOP change (mmHg) Compound (μmoles/eye) E.sub.max/time post dosing E.sub.480 (9)- 0.79 −6.0 ± 2.1 −6.0 ± 2.1 citrate salt (480 min) (IIIa)-OH 0.79 −5.2 ± 1.0 −2.7 ± 1.5 (60 min)

[0296] Data reported are mean±SEM of three independent experiments each performed on n=8 eye for compound 6-(nitrooxy)hexanoic acid ((IIIa)-OH) and mean±SEM of one experiment performed on n=8 eye for compound (9)-citrate salt.

EXAMPLE 24

[0297] Intraocular Pressure (IOP) Lowering Effects in Ocular Normotensive New Zealand White Rabbits

[0298] In this study the efficacy in lowering intraocular pressure of the NO-donor that is compound 3-[(2S)-2,3-bis(nitrooxy)propoxylpropanoic acid ((IIIg)-OH) (disclosed in Example 4) and the reference PDE5 inhibitor (mirodenafil) were assessed in ocular normotensive New Zealand white rabbits.

[0299] Experimental Procedure

[0300] Male New Zealand white (NZW) rabbits were topically administered (50 μL) with vehicle (phosphate buffer pH 6.0, cremophor EL 5% w/v, DMSO 0.3% w/v, benzalkonium chloride 0.02% w/v), 0.5 μmoles/eye equal to 0.63% w/v mirodenafil (50 μL dose), 0.5 μmoles/eye equal to 0.26% w/v compound (IIIg-OH) (50 μL dose) alone or concomitantly with 0.5 μmoles/eye of mirodenafil (50 μL dose).

[0301] Intraocular pressure was determined using a pneumatonometer (Model 30™ Reichert, Depew, N.Y., USA) prior to (basal) and at selected time point following ocular dosing. One topical drop of NOVESINA 0.4% (purchased from Thea Pharma) was applied to the eye prior to each intraocular pressure measurement.

[0302] Results Data are expressed as mean±SEM of n=6-9. Efficacy (E) as intraocular pressure (TOP) change was calculated versus basal and vehicle values using the following equation:

(IOP.sub.drug−IOP.sub.basal drug)−(IOP.sub.veh−IOP.sub.basal veh)

[0303] The IOP values are taken at maximal effect and at 300 min post-dosing.

[0304] The results reported in Table 3 demonstrated that the combined instillation of mirodenafil (reference PDE5 inhibitor) with compound (3-[(2S)-2,3-bis(nitrooxy)propoxy]propanoic acid ((IIIg)-OH) did not show any improvement of the intraocular pressure lowering effect compared with single instillation of each compound; indeed the efficacy (E.sub.max) of Mirodenafil is comparable to the efficacy of the combined instillation of mirodenafil and compound (IIIg)-OH.

TABLE-US-00003 TABLE 3 IOP lowering activity following topical dosing in ocular normotensive New Zealand white rabbits Amount IOP change (mmHg) administered E.sub.max/ Compound (μmoles/eye) time post−dosing E.sub.300 min Mirodenafil 0.5 −3.68 ± 0.89 −1.46 ± 0.79 120 min (IIIg)-OH 0.5 −2.88 ± 0.36 −0.08 ± 0.23  30 min Mirodenafil + 0.5 −3.20 ± 0.46 −0.19 ± 0.27 (IIIg)-OH each  30 min (coadministration) compound

EXAMPLE 25

[0305] Intraocular Pressure (IOP) Lowering Effects in Ocular Normotensive New Zealand White Rabbits

[0306] In this study the efficacies in lowering intraocular pressure of the compound of the invention (1)-(2S)-citrate salt (disclosed in Example 9), that is a NO-releasing derivative of Mirodenafil, and of Mirodenafil nitrate ester in which the nitrooxy group (ONO.sub.2) is directly linked to the hydroxyl group of the Mirodenafil molecule (PDE5-nitrate ester disclosed in the comparative example 21) were assessed in ocular normotensive New Zealand white rabbits.

Experimental Procedure

[0307] Male New Zealand white (NZW) rabbits were topically administered (50 μL) with vehicle (phosphate buffer pH 6.0, cremophor EL 5% w/v, DMSO 0.3% w/v, benzalkonium chloride 0.02% w/v), 0.5 μmoles/eye equal to 1% w/v (1)-(2S)-citrate salt (504 dose), 0.5 μmoles/eye equal to 0.6% w/v the PDE5 nitrate ester (504 dose).

[0308] Intraocular pressure was determined using a pneumatonometer (Model 30™ Reichert, Depew, N.Y., USA) prior to (basal) and at selected time point following ocular dosing. One topical drop of NOVESINA 0.4% (purchased from Thea Pharma) was applied to the eye prior to each intraocular pressure measurement.

[0309] Results

[0310] Data are expressed as mean±SEM of n=6-9. Efficacy (E) as intraocular pressure (TOP) change was calculated versus basal and vehicle values as follow:

(IOP.sub.drug−IOP.sub.basal drug)−(IOP.sub.veh−IOP.sub.basal veh)

The IOP values are taken at min at 30, 60 and 180 minutes post-dosing.

[0311] The results reported in Table 4 demonstrated that the nitric oxide releasing phosphodiesterase type 5 inhibitor of the invention (compound (1)-(2S)-citrate salt) showed higher intraocular pressure-lowering efficacy (compare the E.sub.max of compound (1)-(2S)-citrate salt to the corresponding E.sub.max of PDE5 nitrate ester) and a longer duration (compare IOP change at 180 min of compound (1)-(2S)-citrate salt to that of PDE5 nitrate ester) with respect to the correspondent PDE5 nitrate ester.

TABLE-US-00004 TABLE 4 IOP lowering activity following topical dosing in ocular normotensive New Zealand white rabbits IOP change (mmHg) Compound E.sub.max/time post dosing E.sub.180 min (1)-(2S)-citrate salt −3.8 ± 0.5 (60min) −1.8 ± 0.8 PDE5 nitrate ester −2.6 ± 0.5 (60min)  0.3 ± 0.8

EXAMPLE 26

[0312] Intraocular Pressure (IOP) Lowering Effect in Laser-Induced Ocular Hypertensive Non-Human Primates

[0313] In this experiment the efficacy in lowering intraocular pressure of a compound of formula (II), compound (9)-fumarate (disclosed in Example 29), was evaluated in laser-induced ocular hypertensive non-human primates.

Experimental Procedure

[0314] Compound (9)-fumarate (2.0% w/v, 30 μL) or vehicle (phosphate buffer pH 6, cremophor EL 5% w/v, DMSO 0.3% w/v, benzalkonium chloride 0.02% w/v) were administered to laser induced ocular hypertensive non-human primates. The contralateral eye was not treated.

[0315] Intraocular pressure was measured using a pneumatonometer (Model 30™ Reichert, Depew, N.Y., USA) prior to (basal) and at selected time point following ocular dosing. One topical drop of proparacaine HCl 0.13% was applied to the eye prior to each intraocular pressure measurement.

[0316] Results

[0317] Efficacy (E) as TOP change was calculated versus basal and vehicle values using the following equation: (IOP.sub.drug−IOP.sub.basal drug)−(IOP.sub.veh−IOP.sub.basal veh) where IOP values are taken at 30, 60, 300, 480 and 1440 minutes post-dosing.

[0318] The results reported in Table 5 demonstrated that compound (9)-fumarate salt showed maximum intraocular pressure-lowering efficacy, E.sub.max of −4.1±2.7 mmHg, at 300 min. Moreover, the IOP-lowering effect of compound (9)-fumarate salt persisted for 24 hours post-dosing time at which the IOP change was still −2.8±2.0 mmHg.

TABLE-US-00005 TABLE 5 IOP lowering activity following topical dosing in laser- induced ocular hypertensive non-human primates IOP change (mmHg) Compound 30 min 60 min 300 min 480 min 1440 min (9)- −4.0 ± 1.3 −3.5 ± 1.6 −4.1 ± 2.7 −3.0 ± 2. 8 −2.8 ± 2.0 fumarate salt

[0319] Data reported are mean±SEM of one experiment performed on n=8 eye.

EXAMPLE 27

[0320] Intraocular Pressure (IOP) Lowering Effect in Laser-Induced Ocular Hypertensive Non-Human Primates

[0321] In this study the efficacies in lowering intraocular pressure of compound (1)-(2S)-citrate salt (disclosed in Example 9), the NO-donor 3-[(2S)-2,3-bis(nitrooxy)propoxy]propanoic acid (compound (IIIg)-OH, disclosed in Example 4), the reference PDE5 inhibitor (Mirodenafil) and of the combined administration of Mirodenafil and the NO-donor 3-[(2S)-2,3-bis(nitrooxy)propoxy]propanoic acid were evaluated in laser-induced ocular hypertensive non-human primates.

Experimental Procedure

[0322] Compound (1)-(2S)-citrate salt (0.31 μmoles/eye equal to 1.0% w/v, 30 μL), Mirodenafil (0.31 μmoles/eye equal to 0.63% w/v, 30 μL), (IIIg)-OH (0.31 μmoles/eye equal to 0.26% % w/v, 30 μL), Mirodenafil concomitantly dosed with (IIIg)-OH or vehicle (phosphate buffer pH 6, cremophor EL 5% w/v, DMSO 0.3% w/v, benzalkonium chloride 0.02% w/v) were administered to laser induced ocular hypertensive non-human primates in a masked fashion. The contralateral eye was not treated.

[0323] Intraocular pressure was measured at selected time points using a pneumatonometer (Model 30™ Reichert, Depew, N.Y., USA) prior to (basal) and at selected time point following ocular dosing. One topical drop of proparacaine HCl 0.13% was applied to the eye prior to each intraocular pressure measurement.

[0324] Results

[0325] Data are reported as mean±SEM of n=13. Efficacy (E) as IOP change was calculated versus basal and vehicle values using the following equation: (IOP.sub.drug−IOP.sub.basal drug)−(IOP.sub.veh−IOP.sub.basal veh). Values reported refer to maximum TOP change (E.sub.max) and the TOP lowering effect detected at 300 min post-dose (E.sub.300min).

[0326] The results reported in Table 6 demonstrated that the nitric oxide releasing phosphodiesterase type 5 inhibitor of the invention (compound (1)-(2S)-citrate salt) showed higher intraocular-pressure lowering effect compared to equimolar doses of the NO-donor ((IIIg)-OH), Mirodenafil or the combination of Mirodenafil and the NO-donor at the majority of time-points of analyzed.

TABLE-US-00006 TABLE 6 IOP lowering activity following topical dosing in laser-induced ocular hypertensive non-human primates Dose IOP change (mmHg) Compound (μnoles/eye) 30 min 60 min 180 min 300 min 1440 min (1)-(2S)- 0.31 −4.0 ± 0.9 −5.3 ± 1.0 −4.9 ± 1.5 −4.4 ± 0.9 −2.6 ± 1.3 citrate salt Mirodenafil 0.31 −2.2 ± 1.4 −4.1 ± 1.5 −3.8 ± 1.0 −2.2 ± 1.1 −1.0 ± 1.1 (IIIg)-OH 0.31 −1.2 ± 0.7 −3.9 ± 1.5 −2.4 ± 1.4 −3.9 ± 1.6 −3.2 ± 1.6 Mirodenafil + 0.31 −2.2 ± 0.9 −3.0 ± 1.0 −2.9 ± 1.2 −2.1 ± 0.8 −0.6 ± 0.9 (IIIg)-OH (each compound)

EXAMPLE 28

(S)-2-(4-((4-ethoxy-3-(1-methyl-7-oxo-3-propyl-6,7-dihydro-1H-pyrazolo[4,3-d]pyrimidin-5-yl)phenyl)sulfonyl)piperazin-1-yl)ethyl-(S)-3-(2,3-bis(nitrooxy) propoxy)propanoate (Compound (15), (S) Isomer)

[0327] ##STR00068##

[0328] To a solution of Lodenafil (400 mg, 0.794 mmol) and 4-nitrophenyl 3-[(2S)-2,3-bis(nitrooxy)propoxy]propanoate (see Example 4) (226 mg, 0.951 mmol) in dry DCM (5 ml), DMAP (97 mg, 0.794 mmol) was added.

[0329] The mixture was stirred over weekend at room temperature, then it was diluted with DCM and water. The two phases were separated and the aqueous phase extracted twice with DCM. The combined organic phases were dried over MgSO.sub.4 and concentrated under reduced pressure. The residue was purified by reversed-phase chromatography (H.sub.2O/CH.sub.3CN with 0.01% FA, from 80:20 to 20:80). Dichloromethane and a saturated solution of NaHCO.sub.3 were added to the residue after purification. The two phases were separated and the aqueous phase extracted twice with DCM. The combined organic phases were dried over MgSO.sub.4 and concentrated under reduced pressure, affording 170 mg of the desired product ((Compound (15), (S) isomer) (yield: 30.0%).

[0330] .sup.1H NMR (600 MHz, dmso) δ 12.19 (s, 1H), 7.83 (dt, J=8.7, 2.4 Hz, 2H), 7.38 (d, J=8.8 Hz, 1H), 5.53 (m, 1H), 4.88 (dd, J=12.8, 3.0 Hz, 1H), 4.72 (dd, J=12.8, 6.5 Hz, 1H), 4.21 (q, 2H), 4.16 (s, 3H), 4.06 (t, 2H), 3.77-3.59 (m, 4H), 2.89 (bs, 4H), 2.77 (t, 2H), 2.55 (m, 8H), 1.74 (dd, J=14.9, 7.4 Hz, 2H), 1.33 (t, J=6.9 Hz, 3H), 0.94 (t, J=7.4 Hz, 3H).

EXAMPLE 29

(S)-(1-(4-(3-chloro-4-methoxybenzylamino)-5-(pyrimidin-2-ylmethylcarbamoyl) pyrimidin-2-yl)pyrrolidin-2-yl)methyl 6-(nitrooxy)hexanoate Fumarate (Compound (9) Fumarate Salt)

[0331] ##STR00069##

[0332] To a solution of compound (9), prepared as described in Example 1, (100 mg, 0.155 mmol) in Methanol (1.5 ml), fumaric acid (18 mg, 0.155 mmol) was added. The mixture was stirred 10 minutes at room temperature, then concentrated and the solid washed with MTBE. The solid was filtered and dried under reduced pressure, yielding 103 mg of (S)-(1-(4-(3-chloro-4-methoxybenzylamino)-5-(pyrimidin-2-ylmethylcarbamoyl) pyrimidin-2-yl)pyrrolidin-2-yl)methyl 6-(nitrooxy)hexanoate fumarate (Compound (9) fumarate, yield: 87%).

[0333] .sup.1H NMR (600 MHz, DMSO-d6) δ 13.13 (m, 2H), 9.17 (m, 1H), 8.76 (m, 3H), 8.54 (m, 1H), 7.38 (m, 1H), 7.24 (m, 2H), 7.04 (m, 1H), 6.63 (s, 2H), 4.52 (m, 5H), 4.20 (m, 2H), 3.81 (s, 3H), 3.52 (m, 2H), 3.32 (m, 2H), 2.31 (m, 2H), 1.92 (m, 4H), 1.64 (m, 2H), 1.50 (m, 2H), 1.32 (m, 2H).

EXAMPLE 30

(S)-(1-(4-(3-chloro-4-methoxybenzylamino)-5-(pyrimidin-2-ylmethylcarbamoyl) pyrimidin-2-yl)pyrrolidin-2-yl)methyl 6-(nitrooxy)hexanoate Maleate (Compound (9) Maleate Salt)

[0334] ##STR00070##

[0335] To a solution of compound (9), prepared as described in Example 1, (100 mg, 0.155 mmol) in Methanol (1.5 ml), maleic acid (18 mg, 0.155 mmol) was added. The mixture was stirred 10 minutes at room temperature, then concentrated and the solid washed with MTBE. The solid was filtered and dried under reduced pressure, giving 113 mg of (S)-(1-(4-(3-chloro-4-methoxybenzylamino)-5-(pyrimidin-2-ylmethylcarbamoyl) pyrimidin-2-yl)pyrrolidin-2-yl)methyl 6-(nitrooxy)hexanoate 2-maleate (Compound (9) maleate, yield: 96%).

[0336] .sup.1H NMR (600 MHz, DMSO-d6) δ 12.35 (m, 3H), 9.18 (m, 1H), 8.77 (m, 3H), 8.54 (s, 1H), 7.38 (m, 2H), 7.22 (m, 1H), 7.04 (m, 1H), 4.55 (m, 6H), 4.20 (m, 3H), 3.98 (m, 1H), 3.80 (s, 3H), 3.49 (m, 2H), 2.75 (m, 2H), 2.65 (m, 2H), 2.31 (m, 2H), 1.92 (m, 4H), 1.56 (m, 4H), 1.32 (m, 2H).

EXAMPLE 31

Intraocular Pressure (IOP) Lowering Effects in Ocular Normotensive New Zealand White Rabbits

[0337] In this study the efficacies in lowering intraocular pressure of the compound of the invention (12)-(2S)-citrate salt (disclosed in Example 8) was assessed in ocular normotensive New Zealand white rabbits.

Experimental Procedure

[0338] Male New Zealand white (NZW) rabbits were topically administered (50 μL) with vehicle (phosphate buffer pH 6.0, cremophor EL 5% w/v, DMSO 0.3% w/v, benzalkonium chloride 0.02% w/v), or (12)-(2S)-citrate salt at 1% concentration.

[0339] Intraocular pressure was determined using a pneumatonometer (Model 30™ Reichert, Depew, N.Y., USA) prior to (basal) and at selected time point following ocular dosing. One topical drop of NOVESINA 0.4% (purchased from Thea Pharma) was applied to the eye prior to each intraocular pressure measurement.

[0340] Results

[0341] Data are expressed as mean±SEM of n=9. Efficacy (E) as intraocular pressure (TOP) change was calculated versus basal and vehicle values as follow: (IOP.sub.drug−IOP.sub.basal drug)−(IOP.sub.veh−IOP.sub.basal veh). The TOP values are taken at 30, 60 and 120 minutes post-dosing. The results reported in Table 7 demonstrated that the nitric oxide releasing phosphodiesterase type 5 inhibitor of the invention (compound (12)-(2S)-citrate salt) efficiently lowered TOP at several time points post dosing.

TABLE-US-00007 TABLE 7 IOP lowering activity following topical dosing in ocular normotensive New Zealand white rabbits IOP change (mmHg) Compound 30 min 60 min 120 min (12)-(2S)-citrate salt −5.6 ± 0.9 −3.9 ± 1.0 −2.4 ± 0.8