U.S. patent application number 09/900350 was filed with the patent office on 2002-04-04 for pyrrolidine compounds.
Invention is credited to Aebi, Johannes, Dehmlow, Henrietta, Kitas, Eric A..
Application Number | 20020040048 09/900350 |
Document ID | / |
Family ID | 8169225 |
Filed Date | 2002-04-04 |
United States Patent
Application |
20020040048 |
Kind Code |
A1 |
Aebi, Johannes ; et
al. |
April 4, 2002 |
PYRROLIDINE COMPOUNDS
Abstract
The present invention relates to a pyrrolidine compound and
pharmaceutically acceptable esters and/or salts thereof. The
compounds are useful as inhibitors of metalloproteases, e.g. zinc
proteases, particularly zinc hydrolases, and which are effective in
treating disease states are associated with vasoconstriction of
increasing occurrences.
Inventors: |
Aebi, Johannes; (Basle,
CH) ; Dehmlow, Henrietta; (Grenzach-Wyhlen, DE)
; Kitas, Eric A.; (Arlesheim, CH) |
Correspondence
Address: |
HOFFMANN-LA ROCHE INC.
PATENT LAW DEPARTMENT
340 KINGSLAND STREET
NUTLEY
NJ
07110
|
Family ID: |
8169225 |
Appl. No.: |
09/900350 |
Filed: |
July 6, 2001 |
Current U.S.
Class: |
514/422 ;
514/423; 548/517; 548/533; 548/537 |
Current CPC
Class: |
A61P 25/06 20180101;
A61P 17/02 20180101; A61P 37/06 20180101; A61P 9/06 20180101; A61P
3/10 20180101; C07D 405/12 20130101; A61P 11/06 20180101; A61P
31/04 20180101; C07D 207/16 20130101; C07D 401/12 20130101; A61P
9/04 20180101; A61P 9/10 20180101; A61P 27/02 20180101; A61P 13/12
20180101; A61P 35/00 20180101; A61P 27/06 20180101; A61P 25/00
20180101; A61P 15/00 20180101; A61P 9/00 20180101; A61P 9/12
20180101 |
Class at
Publication: |
514/422 ;
514/423; 548/517; 548/533; 548/537 |
International
Class: |
A61K 031/401; C07D
43/02 |
Foreign Application Data
Date |
Code |
Application Number |
Jul 19, 2000 |
EP |
00114948.3 |
Claims
1. A compound of formula (I) 21wherein R.sup.1 is hydrogen,
alkylcarbonyl, or arylcarbonyl; R.sup.2 is alkyl, alkylcycloalkyl,
alkylcycloalkylalkyl, cycloalkyl, halogenalkyl, carboxyalkyl,
aminoalkyl, (mono- and dialkyl)aminoalkyl, alkoxyalkyl,
alkoxycarbonylalkyl, alkinyl, aryl, arylalkyl,
arylalkyl(alkoxycarbonyl)alkyl, arylcarbonylalkyl, aryloxyalkyl,
arylalkenyl, aryl(alkoxycarbonyl)alkyl, heteroaryl,
heteroarylalkyl, heterocyclyl or hetercycylalkyl; R.sup.3 is
hydrogen, aryl, alkyl, or arylalkyl, arylsulfonyl,
heteroarylsulfonyl; R.sup.4 is hydrogen, arylalkyl, alkyl, aryl,
cycloalkyl, cycloalkylalkyl, alkylsulfonyl, arylsulfonyl,
arylalkylsulfonyl, heteroarylsulfonyl, carboxyalkyl,
carboxyalkylsulfonyl, or alkoxycarbonylalkyl; or the groups
--NR.sup.3R.sup.4 or R.sup.5--[N--N(R.sup.4)]--R.sup.3 form a
saturated or unsaturated 5- or 6-membered aliphatic ring; R.sup.5
is hydrogen, alkylsulfonyl, arylsulfonyl, heteroarylsulfonyl,
alkoxycarbonyl, aryloxycarbonyl, heteroaryloxycarbonyl,
alkylcarbonyl, arylcarbonyl, heteroarylcarbonyl,
heteroarylalkylcarbonyl, heterocyclyl, (mono- or
di-alkylamino)-alkylcarbonyl, (mono- and dialkyl)aminosulfonyl,
arylaminocarbonyl, alkyl, alkylcarbonyl, alkoxycarbonyl, aryl,
arylalkyl, arylalkoxycarbonyl, or heteroaryl; R.sup.6 is hydrogen,
alkyl, aryl, or carboxyalkyl; X is --S(O).sub.2--,
--S(O).sub.2--NH--, --C(O)--, --C(O)NR.sup.6 or C(O)--O--or a
pharmaceutically acceptable ester, or a pharmaceutically acceptable
salt thereof.
2. A compound according to claim 1, wherein R.sup.1 is
hydrogen.
3. A compound according to claim 2, wherein R.sup.2 is alkyl,
halogenalkyl, alkylamino, alkoxy, cycloalkyl, cycloalkylamino,
aryl, arylalkyl, aryloxy, arylalkylamino, arylalkoxy, heteroaryl,
amino, or (mono- and dialkyl)amino.
4. A compound according to claim 3, wherein R.sup.2 is alkyl,
halogenalkyl, alkylamino, alkoxy, cycloalkyl, cycloalkylamino,
aryl, arylalkyl, or heteroaryl.
5. A compound according to claim 4, wherein R.sup.2 is aryl or
heteroaryl.
6. A compound according to claim 5, wherein R.sup.2 is aryl.
7. A compound according to claim 6, wherein R.sup.2 is naphthyl or
phenyl, wherein phenyl is unsubstituted or substituted by one or
more fluor or by one phenyl group.
8. A compound according to claim 7, wherein R.sup.2 is naphthyl,
2,3,4,5,6-pentafluorobenzene or biphenyl.
9. A compound according to claim 1, wherein R.sup.3 is hydrogen or
alkyl;
10. A compound according to claim 9, wherein R.sup.3 is
hydrogen.
11. A compound according to claim 1, wherein R.sup.4 is hydrogen,
arylalkyl, alkyl, arylsulfonyl, heteroarylsulfonyl,
cycloalkylalkyl, or carboxyalkyl.
12. A compound according to claim 11, wherein R.sup.4 is hydrogen,
alkyl, arylalkyl, cycloalkyl, arylsulfonyl, or carboxyalkyl.
13. A compound according to claim 12, wherein R.sup.4 is hydrogen,
alkyl, cycloalkyl, carboxyalkyl or arylalkyl.
14. A compound according to claim 13, wherein R.sup.4 is hydrogen,
alkyl or arylalkyl.
15. A compound according to claim 14, wherein R.sup.4 is hydrogen,
2,4,5-trifluorobenzyl, 2,4-difluorobenzyl, benzyl, methyl, ethyl,
isopropyl, isobutyl, benzyl or HO.sub.2C--CH.sub.2--, or
cycloalkylpropylmethyl.
16. A compound according to claim 1, wherein R.sup.5 is hydrogen,
alkylcarbonyl, alkoxycarbonyl, alkylsulfonyl, aryl, arylalkyl,
arylcarbonyl, (mono- and dialkylamino)alkylcarbonyl, (mono- and
dialkyl)aminosulfonyl, arylalkoxycarbonyl, arylaminocarbonyl,
arylsulfonyl, heteroarylcarbonyl, heteroarylalkylcarbonyl,
heteroarylsulfonyl, arylaminocarbonyl, heteroaryl, or
heterocyclyl.
17. A compound according to claim 16, wherein R.sup.5 is aryl,
arylalkyl, arylcarbonyl, arylalkoxy, arylaminocarbonyl,
arylsulfonyl, heteroarylcarbonyl, heteroarylalkylcarbonyl,
heteroarylsulfonyl, arylaminocarbonyl, heteroaryl, or
heterocyclyl.
18. A compound according to claim 17, wherein R.sup.5 is
arylsulfonyl, arylalkyl, heteroarylalkylcarbonyl,
heteroarylsulfonyl.
19. A compound according to claim 18, wherein R.sup.5 is
4-methyl-benzenesulfoonyl, benzyl, 4-methoxybenzenesulfonyl,
(1H-indol-3-yl)acetyl, thophene-2-yl, or
3,5-dimethyl-isoxyzol-4-sulfonyl- .
20. A compound according to claim 1, wherein X is --SO.sub.2--,
--C(O)--,
21. A compound according to claim 1, wherein X is --SO.sub.2--.
22. A compound according to claim 1 having the formula 22
23. A compound according to claim 22, wherein R.sup.1 is hydrogen
R.sup.2 is naphthyl or phenyl, wherein phenyl is optionally
substituted by one or more fluor or by one phenyl group; R.sup.3 is
hydrogen or alkyl R.sup.4 is hydrogen, alkyl or arylalkyl; R.sup.5
is arylsulfonyl, arylalkyl, heteroarylalkylcarbonyl,
heteroarylsulfonyl; and X is --SO.sub.2--.
24. A compound according to claim-23,
(2S,4R)-4-Mercapto-1-(naphthalene-2--
sulfonyl)-pyrrolidine-2-carboxylic acid
N'-isobutyl-N'-(4-methyl-benzenesu- lfonyl)-hydrazide.
25. A compound according to claim 23,
(2S,4R)-4-Mercapto-1-(naphthalene-2--
sulfonyl)-pyrrolidine-2-carboxylic acid
N'-(4-methyl-benzenesulfonyl)-hydr- azide
26. A compound according to claim 23,
(2S,4R)/4-Mercapto-1-(naphthalene-2--
sulfonyl)-pyrrolidine-2-carboxylic acid N'-benzyl-hydrazide
27. A compound according to claim 23,
(2S,4R)-4-Mercapto-1-(naphthalene-2--
sulfonyl)-pyrrolidine-2-carboxylic acid
N'-benzyl-N'-(4-methyl-phenylsulfo- nyl)-hydrazide.
28. A compound according to claim 23,
(2S,4R)-4-Mercapto-1-(naphthalene-2--
sulfonyl)-pyrrolidine-2-carboxylic acid
N'-methyl-N'-(4-methyl-benzenesulf- onyl)-hydrazide.
29. A compound according to claim 23,
(2S,4R)-4-Mercapto-1-(naphthalene-2--
sulfonyl)-pyrrolidine-2-carboxylic acid
N'-benzenesulfonyl-hydrazide. A compound according to claim 23,
(2S,4R)-4-Mercapto-1-(naphthalene-2-sulfo-
nyl)-pyrrolidine-2-carboxylic acid
N'-(4-methoxy-benzenesulfonyl)-hydrazid- e.
30. A compound according to claim 23,
(2S,4R)-4-Mercapto-1-(naphthalene-2--
sulfonyl)-pyrrolidine-2-carboxylic acid N'-
[(1H-indol-3-yl)-acetyl] -hydrazide.
31. A compound according to claim 23,
(2S,4R)-4-Mercapto-1-(naphthalene-2--
sulfonyl)-pyrrolidine-2-carboxylic acid
N'-thiophene-2-sulfonyl-hydrazide.
32. A compound according to claim 23,
(2S,4R)-4-Mercapto-1-(naphthalene-2--
sulfonyl)-pyrrolidine-2-carboxylic acid N'-(3,5-dimethyl
isoxazole-4-sulfonyl)-hydrazide.
33. A compound according to claim 23,
(2S,4R)-4-Mercapto-1-(naphthalene-2--
sulfonyl)-pyrrolidine-2-carboxylic acid
N'-cyclopropylmethyl-N'-(4-methyl-- benzenesulfonyl)-
hydrazide.
34. A compound according to claim 23,
(2S,4R)-4-Mercapto-1-(naphthalene-2--
sulfonyl)-pyrrolidine-2-carboxylic acid
N'-(4-methyl-benzenesulfonyl)-N'-(-
2,4,5-trifluoro-benzyl)-hydrazide.
35. A compound according to claim 23,
(2S,4R)-4-Mercapto-1-(naphthalene-2--
sulfonyl)-pyrrolidine-2-carboxylic acid
N'-(2,5-difluoro-benzyl)-N'-(4-met-
hyl-benzenesulfonyl)-hydrazide.
36. A compound according to claim 23,
(2S,4R)-4-Mercapto-1-(naphthalene-2--
sulfonyl)-pyrrolidine-2-carboxylic acid
N'-isopropyl-N'-(4-methyl-benzensu- lfonyl)-hydrazide.
37. A compound according to claim 23, (2S,4R)-
[N'-[4-Mercapto-1-(naphthal-
ene-2-sulfonyl)-pyrrolidine-2-carbonyl]
-N-(4-methyl-benzenesulfonyl)-hydr- azino]-acetic acid.
38. A compound according to claim 23,
(2S,4R)-1-(Biphenyl-4-sulfonyl)-4-me-
rcapto-pyrrolidine-2-carboxylic acid
N'-methyl-N'-(4-methyl-benzenesulfony- l)-hydrazide.
39. A compound according to claim 23,
(2S,4R)-4-Mercapto-1-(2,3,4,5,6-pent- afluoro-
benzenesulfonyl)-pyrrolidine-2-carboxylic acid
N'-(4-methyl-benzenesulfonyl)-hydrazide.
40. A compound according to claim 23,
(2S,4R)-4-Mercapto-1-(naphthalene-2--
sulfonyl)-pyrrolidine-2-carboxylic acid
N'-benzyl-N'-(4-methoxy-benzenesul- fonyl)-hydrazide.
41. A compound according to claim 23,
(2S,4R)-4-Mercapto-1-(naphthalene-2--
sulfonyl)-pyrrolidine-2-carboxylic acid
N-methyl-N'-(4-methyl-benzenesulfo- nyl)-hydrazide.43.
42. A compound according to claim 23,
(2S,4R)-4-Mercapto-1-(naphthalene-2--
sulfonyl)-pyrrolidine-2-carboxylic acid N-methyl-N'-benzyl-
N'-(4-methyl-benzenesulfonyl)-hydrazide.
Description
FIELD OF THE INVENTION
[0001] The present invention is directed to compounds which are
useful as inhibitors of metalloproteases, e.g. zinc proteases,
particularly zinc hydrolases, and which are effective in the
prophylaxis and treatment of disease states which are associated
with vasoconstriction of increasing occurrences. Examples of such
disorders are high blood pressure, coronary disorders, cardiac
insufficiency, renal and myocardial ischaemia, renal insufficiency,
dialysis, cerebral ischaemia, cardiac infarct, migraine,
subarachnoid hemorrhage, Raynaud syndrome and pulmonary high
pressure. In addition the compounds are useful as cytostatic and
cerebroprotective agents for inhibition of graft rejection, for
organ protection and for treatment of ophthalmological
diseases.
BACKGROUND OF THE INVENTION
[0002] Endothelins are peptides, that exist in three isoforms ET-1,
ET-2, and ET-3, each encoded by a distinct gene. They have been
originally discovered in the conditioned medium of porcine
endothelial cells in 1988 by Yanagisawa (Yanagisawa M; Kurihara H;
Kimura S; Tomobe Y; Kobayashi M; Mitsui Y; Yazaki Y; Goto K; Masaki
T: A novel potent vasoconstrictor peptide produced by vascular
endothelial cells [see comments]. NATURE (1988 Mar 31), 332(6163),
411-5.). The active ETs are peptides of 21 amino acids with two
intramolecular disulfide bridges. They are produced from
preproproteins of 203 to 212 amino acids which are processed by
furin like endopeptidases to the biologically inactive
big-endothelin (big-ET). The big-ETs are specifically processed to
mature ETs by a hydrolytic cleavage between amino acids 21 and 22
that are Trp.sup.21-Val.sup.22 (big-ET-1, big ET-2) and
Trp.sup.21-Ile.sup.22 in big-ET-3 respectively. Already in 1988 a
specific metalloprotease was postulated to be responsible for this
specific cleavage. In 1994 ECE-1 (endothelin converting enzyme-1)
was purified and cloned from bovine adrenal (Xu D, Emoto N, Giaid
A, Slaughter C, Kaw S, de Witt D, Yanagisawa M: ECE-1: a
membrane-bound metalloprotease that catalyzes the proteolytic
activation of big endothelin-1. Cell (1994) 78: 473-485.).
[0003] ECE-1 is a membrane bound type II zinc-endopeptidase with a
neutral pH optimum and a zinc binding motif HExxHx(>20)E. It
belongs to subfamily M13 and has a large 681 amino acid ectodomain
that comprises the active site. Other members of the M13 family are
NEP24.11 (neutral endopeptidase), PEX, a phosphate regulating
neutral endopeptidase, and Kell blood group protein that has
recently been described as a big-ET-3 processing enzyme. Members of
the M13 family of human origin are characterized by a high
molecular weight (>80 kDa) a number of conserved disulfide
bridges and a complex glycosylation pattern. The structure of NEP
has recently been solved. (Oefner et al, J. Mol. Biol. 2000, 296,
341-349). The catalytic domain of ECE and related human M13
proteinases are significantly larger (>650 amino acids) than
members of matrix metalloproteases (MMPs). Unlike the family of the
MMPs which belong to the metzincins and display a typical
HExxHxxGxxH pattern members of the M13 family are gluzincins
comprising a HExxHx(>20)E pattern. These two families are
clearly different in size of catalytic domains, structure and zinc
coordinating pattern of ligands. Active sites of the two families
show clear differences which has clear impact on type of inhibitors
and the potential selectivity.
[0004] Therefore one aspect of the present invention is to provide
compounds useful for the selective inhibition of ECE-1.
BRIEF DESCRIPTION OF THE INVENTION
[0005] The present invention relates to a compound of formula (I):
1
[0006] wherein
[0007] R.sup.1 is hydrogen, alkylcarbonyl, or arylcarbonyl;
[0008] R.sup.2 is alkyl, alkylcycloalkyl, alkylcycloalkylalkyl,
cycloalkyl, halogenalkyl, carboxyalkyl, aminoalkyl,
dialkylaminoalkyl, alkoxyalkyl, alkoxycarbonylalkyl, alkinyl, aryl,
arylalkyl, arylalkyl(alkoxycarbonyl)alkyl, arylcarbonylalkyl,
aryloxyalkyl, arylalkenyl,
[0009] aryl(alkoxycarbonyl)alkyl, heteroaryl, heteroarylalkyl,
heterocyclyl or hetercycylalkyl;
[0010] R.sup.3 is hydrogen, aryl, alkyl, or arylalkyl,
arylsulfonyl, heteroarylsulfonyl;
[0011] R.sup.4is hydrogen, arylalkyl, alkyl, aryl, cycloalkyl,
cycloalkylalkyl, alkylsulfonyl, arylsulfonyl, arylalkylsulfonyl,
heteroarylsulfonyl, carboxyalkyl, carboxyalkylsulfonyl, or
alkoxycarbonylalkyl; or
[0012] the groups--NR.sup.3R.sup.4 or
R.sup.5--[N--N(R.sup.4)]--R.sup.3 form a saturated or unsaturated
5- or 6-membered aliphatic ring;
[0013] R.sup.5 is hydrogen, alkylsulfonyl, arylsulfonyl,
heteroarylsulfonyl, alkoxycarbonyl, aryloxycarbonyl,
heteroaryloxycarbonyl, alkylcarbonyl, arylcarbonyl,
heteroarylcarbonyl, heteroarylalkylcarbonyl, heterocyclyl, (mono-
or di-alkylamino)-alkylcarb- onyl, (mono- and
dialkyl)aminosulfonyl, arylaminocarbonyl, alkyl, alkylcarbonyl,
alkoxycarbonyl, aryl, arylalkyl, arylalkoxycarbonyl, or
heteroaryl;
[0014] R.sup.6 is hydrogen, alkyl, aryl, or carboxyalkyl;
[0015] X is --S(O).sub.2--, --S(O).sub.2--NH--, --C(O)--,
--C(O)NR.sup.6 or C(O)--O--, or a pharmaceutically acceptable
esteror pharmaceutically acceptable salt thereof.
DETAILED DESCRIPTION OF THE INVENTION
[0016] The term "alkyl" as used herein, alone or in combination,
means a straight-chain or branched-chain alkyl group containing a
maximum of 7, preferably a maximum of 4, carbon atoms, e.g.,
methyl, ethyl, n-propyl, 2-methylpropyl (iso-butyl), 1-methylethyl
(iso-propyl), n-butyl, and 1,1-dimethylethyl (t-butyl).
[0017] The term "carboxy" refers to the group --C(O)OH.
[0018] The term "carbamoyl" refers to the group --C(O)NH.sub.2.
[0019] The term "carbonyl" refers to the group --C(O)--.
[0020] The term "halogen" refers to the group "fluoro, bromo,
chloro and iodo, preferably fluoro and/or chloro, most preferably
fluoro.
[0021] The term "sulfonyl" refers to the group --S(O.sub.2)--.
[0022] The term "alkenyl" refers to a hydrocarbon chain as defined
for alkyl having at least one olefinic double bond (including for
example, vinyl, allyl and butenyl).
[0023] The term "alkinyl" refers to a hydrocarbon chain as defined
for alkyl having at least one olefinic triple bond (including for
example propinyl, butin-(1)-yl, etc.
[0024] The term "alkoxy", alone or in combination, means an alkyl
ether group in which the term `alkyl` has the significance given
earlier, such as methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy,
isobutoxy, sec.butoxy, tert.butoxy and the like.
[0025] The term "alkoxycarbonyl" refers to refers to a group of the
formula --C(O)R.sub.c wherein R.sub.c is alkoxy as defined
above.
[0026] The term "hydroxy" refers to the group --OH, the term
"cyano" to the group --CN.
[0027] The term "hydroxyalkyl" means an alkyl group as defined
earlier which is substituted by a hydroxy group.
[0028] The term "thioalkyl" refers to an alkyl group as defined
above which is substituted by a --SH group.
[0029] The term "halogenalkyl" refers to an alkyl group as defined
earlier which is substituted by one to three halogen atoms,
preferably fluoro, e.g. trifluoromethyl, 2,2,2-trifluoroethyl,
etc.
[0030] "Carboxyalkyl" means a lower-alkyl as defined above which is
substituted by a HOOC-- group.
[0031] The term "alkylcarbonyl", alone or in combination, means an
acyl group derived from an alkanecarboxylic acid, i.e. alkyl
--C(O)--, such as acetyl, propionyl, butyryl, valeryl,
4-methylvaleryl etc.
[0032] The term "cycloalkyl", alone or in combination, signifies a
saturated, cyclic hydrocarbon group with 3-8, preferably 3-6 carbon
atoms, i.e. cyclopropyl, cyclobutyl, cyclopentyl and cyclohexyl and
the like.
[0033] The term "amino" refers to the group --NH.sub.2.
[0034] The term "aryl" for R.sup.2--alone or in combination--,
refers to an aromatic carbocyclic radical, i.e. a 6 or 10 membered
aromatic or partially aromatic ring, e.g. phenyl, naphthyl or
tetrahydronaphthyl, preferably phenyl or naphthyl, and most
preferably phenyl. The aryl moiety is optionally substituted with
one or more groups, preferably 1-5, more preferably 1-3,
independently selected from halogen, preferably fluor,
alkoxycarbonyl, e.g. methylcarbonyl, carboxy, cyano, alkyl, alkoxy,
phenyl, phenoxy, trifluormethyl, trifluormethoxy, more preferably
fluor, alkoxycarbonyl, alkyl, trifluoromethyl and trifluoromethoxy
and most preferably fluor. The most preferred aromatic groups are
naphthyl or phenyl substituted with one or more fluor atoms, e.g.
naphthyl, 2,3,4,5,6-pentafluorophenyl or biphenyl.
[0035] The term "aryl" for R.sup.3 and R.sup.6--alone or in
combination--refers to an aromatic carbocyclic radical, i.e. a 6 or
10 membered aromatic or partially aromatic ring, e.g. phenyl,
naphthyl or tetrahydronaphthyl, preferably phenyl or naphthyl, and
most preferably phenyl. The aryl moiety is optionally substituted
with one or more groups, preferably 1-5, more preferably 1-3,
independently selected from halogen, preferably fluor,
alkoxycarbonyl, e.g. methylcarbonyl, carboxy, cyano, alkyl, alkoxy,
phenyl, phenoxy, trifluormethyl, trifluormethoxy, hydroxy,
alkylamido, e.g. acetamido, nitro, alkylsulfonyl, e.g.
methylsulfonyl, more preferably alkyl or alkoxy.
[0036] The term "aryl" for R.sup.4--alone or in combination--,
refers to an aromatic carbocyclic radical, i.e. a 6 or 10 membered
aromatic or partially aromatic ring, e.g. phenyl, naphthyl or
tetrahydronaphthyl, preferably phenyl or naphthyl, and most
preferably phenyl. The aryl moiety is optionally substituted with
one or more groups, preferably 1 to 3, independently selected from
halogen, preferably fluor, alkoxycarbonyl, e.g. methylcarbonyl,
carboxy, cyano, alkyl, alkoxy, phenyl, phenoxy, trifluormethyl,
trifluormethoxy, cyclohexyl, hydroxy, alkylamido, e.g. acetamido,
nitro, alkylsulfonyl, e.g. methylsulfonyl, more preferably fluor,
chlor, brom, alkoxy, carboxy, alkoxycarbonyl, and most preferably
fluor. Examples for aromatic groups are phenyl,
2,4,5-trifluorophenyl, and 2,4-difluorophenyl.
[0037] The term "aryl" for R.sup.5--alone or in combination--,
refers to an aromatic carbocyclic radical, i.e. a 6 or 10 membered
aromatic or partially aromatic ring, e.g. phenyl, naphthyl or
tetrahydronaphthyl, preferably phenyl or naphthyl, and most
preferably or phenyl. The aryl moiety is optionally substituted
with one or more groups, preferably 1-5, more preferably 1-3,
independently selected from halogen, preferably fluor,
alkoxycarbonyl, e.g. methylcarbonyl, carboxy, cyano, alkyl, alkoxy,
phenyl, phenoxy, trifluormethyl, trifluormethoxy, more preferably
alkyl or alkoxy, e.g. methyl or methoxy. Examples for these aryl
groups are 4-methyl-phenyl and 4-methoxy-phenyl.
[0038] The term "aryloxy" refers to an aryl group as defined above
attached to a parent structure via an oxy radical, i.e.,
aryl-O--.
[0039] The term "heteroaryl" for R.sup.4--alone or in
combination--refers to an aromatic monovalent mono- or bicyclic
radical having 5 to 10, preferably 5 to 6 ring atoms, containing
one to three heteroatoms, preferably one heteroatom, e.g.
independently selected from nitrogen, oxygen or sulfur. Examples of
heteroaryl groups are thiophenyl, isoxazolyl, thiazolyl, pyridinyl,
pyrrolyl, imidazolyl, tetrazolyl, preferably pyridinyl, isoxazolyl
and thiazolyl. Optionally, the heteroaryl group can be mono-, di-
or tri-substituted, independently, with phenyl, alkyl,
alkylcarbonyl, alkoxycarbonyl, hydroxy, amino, alkylamino,
dialkylamino, carboxy, alkoxycarbonylalkyl, preferably alkyl.
[0040] The term "heteroaryl" for R.sup.3 or R.sup.5--alone or in
combination--refers to an aromatic monovalent mono- or bicyclic
radical having 5 to 10, preferably 5 to 6 ring atoms, containing
one to three heteroatoms, preferably one heteroatom, e.g.
independently selected from nitrogen, oxygen or sulfur. Examples of
heteroaryl groups are pyridinyl, thiophenyl, isoxyzolyl,
isoquinolyl, quinolyl, indolyl, pyrimidine, pyridazine, and
pyrazine, preferably thiophenyl, furanyl, pyrrolidinyl, indolyl and
isoxazolyl. Optionally, the heteroaryl group can be mono-, di- or
tri-substituted, independently, with phenyl, alkyl, alkylcarbonyl,
alkoxycarbonyl, hydroxy, amino, alkylamino, dialkylamino, carboxy,
oxo, alkoxycarbonylalkyl, preferably alkyl.
[0041] The term "heterocyclyl"--alone or in combination--refers to
a non-aromatic monovalent mono- or bicyclic radical having 5 to 10,
preferably 5 to 6 ring atoms, containing one to three heteroatoms,
preferably one heteroatom, e.g. independently selected from
nitrogen, oxygen or sulfur. Optionally the heterocyclic ring can be
substituted by a group independently selected from halogen, alkyl,
alkoxy, oxocarboxy, alkoxycarbonyl, etc. and/or on a secondary
nitrogen atom (i.e. --NH--) by alkyl, arylalkoxycarbonyl,
alkylcarbonyl or on a tertiary nitrogen atom (i.e. .dbd.N--) by
oxido. Examples for heterocyclic groups are morpholinyl,
pyrrolidinyl, piperidyl, etc.
[0042] The term "dimeric form" means a compound wherein the two
R.sup.1 groups of two identical compounds of formula I have been
replaced by a common single bond or wherein R.sup.1 is
glutathione-S-- or cysteine-S-- or ester and/or alkylcarbonyl or
arylcarbonyl derivatives thereof, e.g. acetylcysteine-S-- or
benzoylcysteine-S--, preferably glutathione-S--, cysteine-S--,
acetylcysteine-S-- or benzoylcysteine-S--.
[0043] The term "pharmaceutically acceptable salt" refers to those
salts which retain the biological effectiveness and properties of
the free bases or free acids, which are not biologically or
otherwise undesirable. The salts are formed with inorganic acids
such as hydrochloric acid, hydrobromic acid, sulfuric acid, nitric
acid, phosphoric acid and the like, and organic acids such as
acetic acid, propionic acid, glycolic acid, pyruvic acid, oxylic
acid, maleic acid, malonic acid, succinic acid, fumaric acid,
tartaric acid, citric acid, benzoic acid, cinnamic acid, mandelic
acid, methanesulfonic acid, ethanesulfonic acid, p-toluenesulfonic
acid, salicylic acid, and the like. In addition these salts may be
prepared form addition of an inorganic base or an organic base to
the free acid. Salts derived from an inorganic base include, but
are not limited to, the sodium, potassium, lithium, ammonium,
calcium, magnesium salts and the like. Salts derived from organic
bases include, but are not limited to salts of primary, secondary,
and tertiary amines, substituted amines including naturally
occurring substituted amines, cyclic amines and basic ion exchange
resins, such as isopropylamine, trimethylamine, diethylamine,
triethylamine, tripropylamine, ethanolamine, lysine, arginine,
N-ethylpiperidine, piperidine, polymine resins and the like.
[0044] "Pharmaceutically acceptable esters" means that compounds of
general formula (I) may be derivatised at functional groups to
provide derivatives which are capable of conversion back to the
parent compounds in vivo. Examples of such compounds include
physiologically acceptable and metabolically labile ester
derivatives, such as methoxymethyl esters, methylthiomethyl esters
and pivaloyloxymethyl esters. Additionally, any physiologically
acceptable equivalents of the compounds of general formula (I),
similar to the metabolically labile esters, which are capable of
producing the parent compounds of general formula (I) in vivo, are
within the scope of this invention.
[0045] The compounds of formula (I) and their salts and esters are
useful in inhibiting mammalian metalloprotease activity,
particularly zinc hydrolase activity. More specifically, the
compounds of formula (I) and their salts and esters are useful as
medicaments for the treatment and prophylaxis of disorders which
are associated with diseases caused by endothelin-converting enzyme
(ECE) activity. Inhibiting of this enzyme would be useful for
treating myocardial ischaemia, congestive heart failure,
arrhythmia, hypertension, pulmonary hypertension, asthma, cerebral
vasospasm, subarachnoid haemorrhage, pre-eclampsia, kidney
diseases, atherosclerosis, Buerger's disease, Takayasu's arthritis,
diabetic complications, lung cancer, prostatic cancer,
gastrointestinal disorders, endotoxic shock and septicaemia, and
for wound healing and control of menstruation, glaucoma. In
addition the compounds are useful as cytostatic and
cerebroprotective agents, for inhibition of graft rejection, for
organ protection and for treatment of ophthalmological
diseases.
[0046] In more detail, the present invention relates to a compound
of formula (I) 2
[0047] wherein
[0048] R.sup.1 is hydrogen, alkylcarbonyl, or arylcarbonyl;
[0049] R.sup.2 is alkyl, alkylcycloalkyl, alkylcycloalkylalkyl,
cycloalkyl, halogenalkyl, carboxyalkyl, aminoalkyl,
dialkylaminoalkyl, alkoxyalkyl, alkoxycarbonylalkyl, alkinyl, aryl,
arylalkyl, arylalkyl(;alkoxycarbonyl)alkyl, arylcarbonylalkyl,
aryloxyalkyl, arylalkenyl, aryl(alkoxycarbonyl)alkyl, heteroaryl,
heteroarylalkyl, heterocyclyl or hetercycylalkyl;
[0050] R.sup.3 is hydrogen, aryl, alkyl, or arylalkyl,
arylsulfonyl, heteroarylsulfonyl;
[0051] R.sup.4 is hydrogen, arylalkyl, alkyl, aryl, cycloalkyl,
cycloalkylalkyl, alkylsulfonyl, arylsulfonyl, arylalkylsulfonyl,
heteroarylsulfonyl, carboxyalkyl, carboxyalkylsulfonyl, or
alkoxycarbonylalkyl; or the groups --NR.sup.3R.sup.4 or
R.sup.5--[N--N(R.sup.4)]--R.sup.3 form a saturated or unsaturated
5- or 6-membered aliphatic ring;
[0052] R.sup.5 is hydrogen, alkylsulfonyl, arylsulfonyl,
heteroarylsulfonyl, alkoxycarbonyl, aryloxycarbonyl,
heteroaryloxycarbonyl, alkylcarbonyl, arylcarbonyl,
heteroarylcarbonyl, heteroarylalkylcarbonyl, heterocyclyl, (mono-
or di-alkylamino)-alkylcarb- onyl, (mono- and
dialkyl)aminosulfonyl, arylaminocarbonyl, alkyl, alkylcarbonyl,
alkoxycarbonyl, aryl, arylalkyl, arylalkoxycarbonyl, or
heteroaryl;
[0053] R.sup.6 is hydrogen, alkyl, aryl, or carboxyalkyl;
[0054] X is --S(O).sub.2--, --S(O).sub.2--NH--, --C(O)--,
--C(O)NR.sup.6 or C(O)--O-- or a dimeric form, or a
pharmaceutically acceptable ester, or a pharmaceutically acceptable
salt thereof, preferably a pharmaceutically acceptable ester, or a
pharmaceutically acceptable salt thereof, and most preferably a
pharmaceutically acceptable salt thereof.
[0055] In a preferred embodiment, the invention refers to compounds
wherein R.sup.1 is hydrogen.
[0056] In a further preferred embodiment of the present invention
R.sup.2 is alkyl, halogenalkyl, alkylamino, alkoxy, cycloalkyl,
cycloalkylamino, aryl, arylalkyl, aryloxy, arylalkylamino,
arylalkoxy, heteroaryl, amino, or (mono- and dialkyl)amino; more
preferably alkyl, halogenalkyl, alkylamino, alkoxy, cycloalkyl,
cycloalkylamino, aryl, arylalkyl, or heteroaryl and even more
preferably is aryl or heteroaryl and most preferably aryl. The term
aryl in the definition for R.sup.2 especially means naphthyl or
phenyl, wherein phenyl is optionally substituted by one or more
fluor or by one phenyl group, e.g. R.sup.2 is naphthyl,
2,3,4,5,6-pentafluorophenyl or biphenyl.
[0057] According to the present invention R.sup.3 is preferably
hydrogen or alkyl, most preferably hydrogen.
[0058] In the above compounds R.sup.4 is preferably hydrogen,
arylalkyl, alkyl, arylsulfonyl, heteroarylsulfonyl,
cycloalkylalkyl, or carboxyalkyl, more preferably hydrogen, alkyl,
arylalkyl, cycloalkyl, arylsulfonyl, or carboxyalkyl, most
preferably hydrogen, alkyl, cycloalkyl, carboxyalkyl or arylalkyl,
even more preferably hydrogen, alkyl or arylalkyl, e.g. hydrogen,
2,4,5-trifluorobenzyl, 2,4-difluorobenzyl, benzyl, methyl, ethyl,
isopropyl, isobutyl, benzyl or HO.sub.2C--CH.sub.2--, or
cycloalkylpropylmethyl.
[0059] In a preferred embodiment of the present invention R.sup.5
is hydrogen, alkylcarbonyl, alkoxycarbonyl, alkylsulfonyl, aryl,
arylalkyl, arylcarbonyl, (mono- and dialkylamino)alkylcarbonyl,
(mono- and dialkyl)aminosulfonyl, arylalkoxycarbonyl,
arylaminocarbonyl, arylsulfonyl, heteroarylcarbonyl,
heteroarylalkylcarbonyl, heteroarylsulfonyl, arylaminocarbonyl,
heteroaryl, or heterocyclyl, more preferably aryl, arylalkyl,
arylcarbonyl, arylalkoxy, arylaminocarbonyl, arylsulfonyl,
heteroarylcarbonyl, heteroarylalkylcarbonyl, heteroarylsulfonyl,
arylaminocarbonyl, heteroaryl, or heterocyclyl, more preferably
arylsulfonyl, arylalkyl, heteroarylalkylcarbonyl,
heteroarylsulfonyl and most preferably 4-methyl-benzenesulfonyl,
benzyl, 4-methoxybenzenesulfonyl, (1H-indol-3-yl)acetyl,
thiophene-2-yl, or 3,5-dimethyl-isoxyzol-4-sulfonyl.
[0060] In the above described compounds X is preferably
--SO.sub.2--, --C(O)--, and most preferably --SO.sub.2--.
[0061] In the most preferred embodiment of the present invention,
the compounds may be described by the formula (II) 3
[0062] wherein R.sup.1, R.sup.2, R.sup.3, R.sup.4, R.sup.5 and X
are as defined above and pharmaceutically acceptable esters and/or
salts thereof.
[0063] In a further preferred embodiment of the present invention
R.sup.1 is hydrogen, R2 is naphthyl or phenyl, wherein phenyl is
optionally substituted by one or more fluor or by one phenyl
group,
[0064] R.sup.3 is hydrogen or alkyl, R.sup.4 is hydrogen, alkyl or
arylalkyl, R.sup.5 is arylsulfonyl, arylalkyl,
heteroarylalkylcarbonyl, heteroarylsulfonyl; and X is
--SO.sub.2--.
[0065] Preferred embodiments of the present invention are the
compounds exemplified in the examples. Especially the present
invention comprises compounds according to formula (I) or (II)
[0066]
(2S,4R)-4-Mercapto-1-(naphthalene-2-sulfonyl)-pyrrolidine-2-carboxy-
lic acid N'-isobutyl-N'-(4-methyl-benzenesulfonyl) -hydrazide;
[0067]
(2S,4R)-4-Mercapto-1-(naphthalene-2-sulfonyl)-pyrrolidine-2-carboxy-
lic acid N'-(4-methyl-benzenesulfonyl)-hydrazide;
[0068]
(2S,4R)/4-Mercapto-1-(naphthalene-2-sulfonyl)-pyrrolidine-2-carboxy-
lic acid N'-benzyl-hydrazide;
[0069] (2S,4R)-4-Mercapto-1-(naphthalene-2-sulfonyl)
-pyrrolidine-2-carboxylic acid
N'-benzyl-N'-(4-methyl-phenylsulfonyl)-hyd- razide;
[0070]
(2S,4R)-4-Mercapto-1-(naphthalene-2-sulfonyl)-pyrrolidine-2-carboxy-
lic acid N'-methyl-N'-(4-methyl-benzenesulfonyl)-hydrazide;
[0071]
(2S,4R)-4-Mercapto-1-(naphthalene-2-sulfonyl)-pyrrolidine-2-carboxy-
lic acid N'-benzenesulfonyl-hydrazide;
[0072]
(2S,4R)-4-Mercapto-1-(naphthalene-2-sulfonyl)-pyrrolidine-2-carboxy-
lic acid N'-(4-methoxy-benzenesulfonyl) -hydrazide;
[0073] (2S,4R)-4-Mercapto
1-(naphthalene-2-sulfonyl)-pyrrolidine-2-carboxy- lic acid
N'[(1H-indol-3-yl-acetyl]-hydrazide;
[0074] (2S,4R)-4-Mercapto-1-(naphthalene-2-sulfonyl)
-pyrrolidine-2-carboxylic acid
N'-thiophene-2-sulfonyl-hydrazide;
[0075] (2S,4R)-4-Mercapto-1-(naphthalene-2-sulfonyl)
-pyrrolidine-2-carboxylic acid N'-(3,5-dimethyl
isoxazole-4-sulfonyl) -hydrazide;
[0076]
(2S,4R)-4-Mercapto-1-(naphthalene-2-sulfonyl)-pyrrolidine-2-carboxy-
lic acid
N'-cyclopropylmethyl-N'-(4-methyl-benzenesulfonyl)-hydrazide;
[0077]
(2S,4R)-4-Mercapto-1-(naphthalene-2-sulfonyl)-pyrrolidine-2-carboxy-
lic acid N'-(4-methyl-benzenesulfonyl) -N'-(2,4,5-trifluoro-benzyl)
-hydrazide;
[0078] (2S,4R)-4-Mercapto-1-
(naphthalene-2-sulfonyl)-pyrrolidine-2-carbox- ylic acid
N'-(2,5-difluoro-benzyl) -N'-(4-methyl-benzenesulfonyl)
-hydrazide
[0079]
(2S,4R)-4-Mercapto-1-(naphthalene-2-sulfonyl)-pyrrolidine-2-carboxy-
lic acid N'-isopropyl-N'-(4-methyl-benzensulfonyl)-hydrazide;
[0080] (2S,4R)- [N'-
[4-Mercapto-1-(naphthalene-2-sulfonyl)-pyrrolidine-2--
carbonyl]-N-(4-methyl-benzenesulfonyl)-hydrazino]-acetic acid;
[0081]
(2S,4R)-1-(Biphenyl-4-sulfonyl)-4-mercapto-pyrrolidine-2-carboxylic
acid N'-methyl-N'-(4-methyl-benzenesulfonyl) -hydrazide;
[0082]
(2S,4R)-4-Mercapto-1-(2,3,4,5,6-pentafluoro-benzenesulfonyl)-pyrrol-
idine-2-carboxylic acid
N'-A4-methyl-benzenesulfonyl)-hydrazide;
[0083]
(2S,4R)-4-Mercapto-1-(naphthalene-2-sulfonyl)-pyrrolidine-2-carboxy-
lic acid N'-benzyl-N'-(4-methoxy-benzenesulfonyl)-hydrazide;
[0084]
(2S,4R)-4-Mercapto-1-(naphthalene-2-sulfonyl)-pyrrolidine-2-carboxy-
lic acid N-methyl-N'-(4-methyl-benzenesulfonyl)-hydrazide; and
[0085]
(2S,4R)-4-Mercapto-1-(naphthalene-2-sulfonyl)-pyrrolidine-2-carboxy-
lic acid
N-methyl-N'-benzyl-N'-(4-methyl-benzenesulfonyl)-hydrazide.
[0086] These compounds show IC.sub.50 values in the
radioimmunoassay (E on ECE-inhibition, see below) of about 50 nM to
1 .mu.M.
[0087] The invention also refers to pharmaceutical compositions
containing a compound as defined above and a pharmaceutically
acceptable excipient.
[0088] A further embodiment of the present invention refers to the
use of compounds as defined above as active ingredients in the
manufacture of medicaments comprising a compound as defined above
for the prophylaxis and treatment of disorders which are caused by
endothelin-converting enzyme (ECE) activity especially myocardial
ischaemia, congestive heart failure, arrhythmia, hypertension,
pulmonary hypertension, asthma, cerebral vasospasm, subarachnoid
haemorrhage, pre-eclampsia, kidney diseases, atherosclerosis,
Buerger's disease, Takayasu's arthritis, diabetic complications,
lung cancer, prostatic cancer, gastrointestinal disorders,
endotoxic shock and septicaemia, and for wound healing and control
of menstruation, glaucoma, diseases associated with cytostatic,
ophthalmological, and cerebroprotective indications, and organ
protection.
[0089] Further the invention refers to the use of compounds as
described above for the treatment or prophylaxis of diseases which
are associated with myocardial ischaemia, congestive heart failure,
arrhythmia, hypertension, pulmonary hypertension, asthma, cerebral
vasospasm, subarachnoid haemorrhage, pre-eclampsia, kidney
diseases, atherosclerosis, Buerger's disease, Takayasu's arthritis,
diabetic complications, lung cancer, prostatic cancer,
gastrointestinal disorders, endotoxic shock and septicaemia, and
for wound healing and control of menstruation, glaucoma, diseases
associated with cytostatic, ophthalmological, and cerebroprotective
indications, and organ protection.
[0090] In addition the invention comprises compounds as described
above for use as therapeutic active substances, in particular in
context with diseases which are associated with zinc hydrolase
activity such as myocardial ischaemia, congestive heart failure,
arrhythmia, hypertension, pulmonary hypertension, asthma, cerebral
vasospasm, subarachnoid haemorrhage, pre-eclampsia, kidney
diseases, atherosclerosis, Buerger's disease, Takayasu's arthritis,
diabetic complications, lung cancer, prostatic cancer,
gastrointestinal disorders, endotoxic shock and septicaemia, and
for wound healing and control of menstruation, glaucoma, diseases
associated with cytostatic, ophthalmological, and cerebroprotective
indications, and organ protection.
[0091] The invention also comprises a method for the therapeutic
and/or prophylactic treatment of myocardial ischaemia, congestive
heart failure, arrhythmia, hypertension, pulmonary hypertension,
asthma, cerebral vasospasm, subarachnoid haemorrhage,
pre-eclampsia, kidney diseases, atherosclerosis, Buerger's disease,
Takayasu's arthritis, diabetic complications, lung cancer,
prostatic cancer, gastrointestinal disorders, endotoxic shock and
septicaemia, and for wound healing and control of menstruation,
glaucoma, diseases associated with cytostatic, ophthalmological,
and cerebroprotective indications, and organ protection, which
method comprises administering a compound as defined above to a
human being or animal.
[0092] The invention also relates to the use of compounds as
defined above for the inhibition of zinc hydrolase activity.
[0093] The invention relates also to a process for the preparation
of a compound as defined above comprising reaction of a compound of
formula III 4
[0094] wherein R.sup.1, R.sup.2, and X are as defined above and A
is a HS-protecting group with HNR.sup.3NR.sup.4R.sup.5 for
introduction of a hydrazide: or
[0095] HNR.sup.3NR.sup.4R.sup.5 with R.sup.5 as protecting group
followed by conversion or introduction of R.sup.3 and R.sup.4;
optionally followed by conversion of a R.sup.5 and/or R.sup.2--X
group into a different R.sup.5 and/or R.sup.2--X group and/or
deprotection and or thiol liberation and wherein R.sup.3, R.sup.4
and R.sup.5 are as defined above.
[0096] The invention also refers to the above compounds whenever
manufactured by a process as described.
[0097] The compounds of formula (I) can be prepared by methods
known in the art or as described below.
[0098] Unless otherwise indicated, the substituents R.sup.1,
R.sup.2, R.sup.3R.sup.4, R.sup.5, R.sup.6, and X are as described
above. In the schemes below, all starting materials are known or
can be prepared by known methods. 5
[0099] Step a) of scheme 1 describes the persilylation of hydroxy-
and amino groups, e.g. by reaction of compound 1 with
hexamethyldisilazan/140- .degree. C. followed by reaction with
R.sup.2SO.sub.2Cl in THF or conversion to all other R.sup.2X
described later or di-t-butyldicarbonate/NaHCO.sub.3 in
dioxane/H.sub.2O (BOC protection). For inversion of the
configuration (via mesylate) the resulting alcohol 2 is treated
with MeSO.sub.3H/Ph.sub.3P/DIAD in toluene (room temperature to
80.degree. C.) or (via bromide) with LiBr/DEAD/Ph.sub.3P in THF
(4.degree. C. to room temperature) or (via chloride) with
Ph.sub.3P/CCl.sub.4 in CH.sub.2Cl.sub.2 (3.degree. C. to room
temperature). In case of retention of the configuration (via
mesylate) alcohol 2 can be transformed to a compound of formula 3
by reaction with MeSO.sub.2Cl/pyridine/DMAP (0.degree. C. to room
temperature).
[0100] For the introduction of a protected thiol moiety, compounds
of formula 3 are treated with e.g. triphenylmethanethiol or
4-methoxybenzylmercaptane and K-Ot-Bu in DMF (for Br: 0.degree. C.
to room temperature; for Cl: 0.degree. C.; for Mesylate: room
temperature to 100.degree. C.). Hydrolysis of ester 4 with aqueous
LiOH in THF (0.degree. C. to RT) gives acid 5.
[0101] The synthesis of final compounds are shown in scheme 2:
[0102] The synthesis starts with a preactivation af acid 1a
(N-hydroxy-2-pyridone, N,N-dicyclohexylcarbodiimide,
4-Ethylmorpholine in CH.sub.2Cl.sub.2 at RT) followed by reaction
with an alkyl-hydrazine (NHR.sup.3NHR.sup.4) (step a) or for
carboxylic acid hydrazide 2 (R.sup.3, R.sup.4, R.sup.5.dbd.H),
ester 1b is directly treated with hydrazine
(NH.sub.2NH.sub.2H.sub.2) in EtOH (at RT). Conversion to the free
thiol 3 is done in the following way: in case PG (protecting group)
is Tr by reaction with e.g. TFA/Et.sub.3SiH at 0.degree. C. to room
temperature or, in case PG is PMB, by reaction with e.g.
TFA/Et.sub.3SiH, at 0 to 80.degree. C. (step b).
[0103] In the case of carboxylic acid hydrazide 2 (R.sup.3,
R.sup.4, R.sup.5.dbd.H), R.sup.4 is introduced by reductive
amination: Imine formation with an aldehyde in EtOH followed by
reduction with NaBH.sub.3CN in THF gives compound 4 (step c). For
the introduction of a new R.sup.5 in case R.sup.4 is an alkyl
(R.sup.5.dbd.H), reaction with ClCOR.sup.5, ClCO.sub.2R.sup.5,
ClSO.sub.2R.sup.5 or ClSO.sub.2NR.sup.5, iPr.sub.2NEt or Huenig's
base CH.sub.2in the precence of a catalytic amount of DMAP or
DMAP-poly or R.sup.5NCO in THF at room temperature gives compound 4
which is deprotected to the final thiol 5 as described above (step
c and b).
[0104] Selective BOC deprotection of compound 2 or 4 (TFA,
CH.sub.2Cl.sub.2 at 0.degree. C.), followed by reaction with
ClCO.sub.2R.sup.2, NEM or iPr.sub.2NEt, CH.sub.2Cl.sub.2 or
R.sup.2NCO in THF at 0.degree. C. to room temperature (or
conversion to all other R.sup.2X described for
R.sup.5--introduction above) gives compound 7 (step e). Thiol
deprotection as described above gives the final thiol 8 (step b).
6
[0105] Scheme 3 shows a different way for the synthesis of
hydrazides.
[0106] Preactivation af acid 1 (N-hydroxy-2-pyridone,
N,N-dicyclohexylcarbodiimide, 4-Ethylmorpholine in CH.sub.2Cl.sub.2
at RT) followed by reaction with an alkyl-hydrazinecarboxylic acid
benzyl ester (NHR.sup.3NHR.sup.4) (step a) gives hydrazide 2 which
is converted with HBr in AcOH at 0.degree. C. to 3 (step b). A
direct conversion from 1 to 3 with preactivation and reaction with
NHR.sup.3NR.sup.4 R.sup.5is possible too (step c). Introduction of
a new R.sup.3 is done with an R.sup.3-halogenide/NaH in DMF (at
0.degree. C. to RT; .fwdarw.4, step e). Deprotection to the thiol 5
is done in the following way: in case PG is Trby reaction with e.g.
TFA/Et.sub.3SiH at 0.degree. C. to room temperature or, in case PG
is PMB, by reaction with e.g. TFA/Et.sub.3SiH, at 0 to 80.degree.
C. (step d). 7
[0107] Scheme 4 shows an other way for the synthesis of
hydrazides.
[0108] Preactivation af acid 1 (N-hydroxy-2-pyridone,
N,N-dicyclohexylcarbodiimide, 4-Ethylmorpholine in CH.sub.2Cl.sub.2
at RT) followed by reaction with a tert-butyl
2-alkyl-hydrazinecarboxylate (NHR.sup.3NHBOC) (step a) gives
hydrazide 2 which by treatment with triethylsilane in TFA at 0 to
80.degree. C. gives thiol 3 (step b). Alkylation with
alkylhalogenide (R.sup.5-halogenide) and DMF with NaH as base (at
0.degree. C. to RT) results in compound 4 which gives after
Et.sub.3SiH/TFA deprotection (as described in scheme 1) thiol 5
(step c, d).
[0109] Selective BOC-deprotection (TFA in
CH.sub.2Cl.sub.2.fwdarw.6) followed by reaction with
ClCO.sub.2R.sup.4, ClSO.sub.2R.sup.4, iPr.sub.2NEt or NEM in
CH.sub.2Cl.sub.2 in the precence of a catalytic amount of DMAP or
DMAP-poly at room temperature gives compounds 7
(R.sup.4.dbd.R.sup.5) and 8 which are separated and deprotected
(Et.sub.3SiH/TFA as described in scheme 1) to thiol 5 (step e, f
and d). The not fully substituted hydrazide 8 can be further
alkylated (R.sup.5-halogenide and DMF/NaH, 0.degree. C. to RT) and
deprotected (Et.sub.3SiH/TFA as described in scheme 1) to the thiol
5 (step g and d). 8
[0110] Scheme 5 shows further transformation of hydrazide 1.
Acylation with .gamma.-Bromo-alkanoyl chloride in the presence of
iPr.sub.2EtN in THF (0.degree. C. to RT) gives compound 2 which is
cyclised (NaH in DMF at RT). Separation of the two isomers and
deprotection of the thiol (Et.sub.3SiH/TFA as described in scheme
1) gives hydrazides 3 and 4. 9
[0111] For the preparation of compounds of formula 5 the reaction
pathway of scheme 6 can be followed: the synthesis of the starting
material 1 from hydroxyproline is described in scheme 1.
TFA/triisopropyl deprotection at reflux for 30 minutes gives thiol
2 that is attached to the resin. The final R.sup.2X is introduced
either at the beginning or after manipulations at
NR.sup.3NR.sup.4R.sup.5 (scheme 9). In the second case, R.sup.2X
(=BOC) of starting acid 1 is transformed by methods known in the
art and described for example in "The Practice of Peptide
Synthesis", M. Bodanszky and A. Bodanszky, Springer Verlag, Berlin,
1984 to a nonacid labile protecting group (e.g. R.sup.2X=FMOC, step
a: first selective BOC-deprotection with 40% TFA in
CH.sub.2Cl.sub.2 at RT followed by reaction with Fmoc-OSu in
dioxane/water and NaHCO.sub.3 as base). 10
[0112] The resin is prepared as follows (step b): The linker
4-(.alpha.,.alpha.-diphenylhydroxymethyl)benzoic acid is activated
using TPTU, DIEA in DMF and added to benzhydrylamine resin 3. The
resin is then treated with thiol 2 in CH.sub.2Cl.sub.2/TFA to give
the resin loaded starting material 5. 11
[0113] The synthesis of final compounds on resin 1 is shown in
scheme 7: The synthesis starts with a preactivation af acid 1
(TPTU, Huenig's base in DMF at RT) followed by reaction with an
alkyl-hydrazine (NH.sub.2NHR.sup.4) (step a) to give intermediates
2a, 2b or 4 (R.sup.3, R.sup.4, R.sup.5.dbd.H, srep c). Detachment
of the resin to the free thiol 3 is done with TFA/iPr.sub.3SiH in
CH.sub.2Cl.sub.2 at RT (step b). In the case of carboxylic acid
hydrazide 4, the introduction of a new R.sup.5 is done by reaction
with ClCOR.sup.5, ClCO.sub.2R.sup.5, ClSO.sub.2R.sup.5 or
ClSO.sub.2NR.sup.5, in DMF to give compound 2 which is optionally
alkylated (alkyl halgenide/DBU in DMF) to the disubstituted
hydrazide 6 (step d and f). Detachment of the resin as described
above gives the final thiol 3. In the case of reaction of hydrazide
4 with ClSO.sub.2R.sup.5, double sulfonylation to compound 2a and
2b takes place (step d), these compounds can be separated after
detachment from the resin as the corresponding thiol 3.
[0114] If R.sup.2X is FMOC: Deprotection of compound 2 (20%
piperidine/DMF then reaction with ClCO.sub.2R.sup.2, pyridine, DMF
or R.sup.2NCO in DMF at RT or conversion to all other R.sup.2X
described for R.sup.4-introduction above) gives compound 5 (step
e). Alkylation (alkyl halgenide/ DBU in DMF) and resin deprotection
as described above gives the final thiol 3 (step f and b).
[0115] The ability of the compounds of formula (I) to inhibit
metalloprotease activity, particularly zinc hydrolase activity, may
be demonstrated by a variety of in vitro and in vivo assays known
to those of ordinary skill in the art.
[0116] A) Cell Culture
[0117] A stable human umbilical vein endothelial cell line (ECV304)
was cultured in "cell factories" as described until confluency
(Schweizer et al. 1997, Biochem. J. 328: 871-878). At confluency
cells were detached with a trypsin/EDTA solution and collected by
low speed centrifugation. The cell pellet was washed once with
phosphate buffered saline pH 7.0 and stored at -80.degree. C. until
use.
[0118] B) Solubilization of ECE from ECV304 Cells
[0119] All procedures were performed at 0-4.degree. C. if not
stated otherwise. The cell pellet of 1.times.10.sup.9 was suspended
in 50 ml of buffer A (20 mM Tris/HCl, pH 7.5 containing 5 mM
MgCl.sub.2, 100 .mu.M PMSF, 20 .mu.M E64, 20 .mu.M leupeptin) and
sonicated. The resulting cell homogenate was centrifuged at 100,000
g.sub.av for 60 minutes. The supernatant was discarded and the
resulting membrane pellet was homogenized in 50 ml buffer A and
centrifugated as described. The washing of the membrane fraction in
buffer A was repeated twice. The final membrane preparation was
homogenized in 50 ml of buffer B (buffer A+0.5% Tween 20 (v/v),
0.5% CHAPS (w/v), 0.5% Digitonin (w/v)) and stirred at 4.degree. C.
for 2 hours. Thereafter the remaining membrane fragments were
sedimented as described. The resulting clear supernatant containing
the solubilized ECE was stored in 1.0 ml aliquots at -120.degree.
C. until use.
[0120] C) ECE Assay
[0121] The assay measured the production of ET-1 from human big
ET-1. To measure high numbers of samples an assay performed in 96
well plates was invented. The enzyme reaction and the
radioimmunological detection of the produced ET-1 was performed in
the same well, using a specifically developed and optimized coating
technique.
[0122] D) Coating of Plates
[0123] Fluoronunc Maxisorp White (code 437796) 96 well plates were
irradiated with 1 joule for 30 minutes in a UV Stratalinker 2400
(Stratagene). The 96 well plates were then fill with 300 .mu.l
protein A solution (2 .mu.g/ml in 0.1 M Na.sub.2CO.sub.3 pH 9.5)
per well and incubated for 48 hours at 4.degree. C. Coated plates
can be stored for up to 3 weeks at 4.degree. C. until use.
[0124] Before use the protein A solution is discarded and the
plates are blocked for 2 hours at 4.degree. C. with 0.5% BSA in
0.1M Na.sub.2CO.sub.3, pH 9.5.
[0125] Plates were washed with bidestilled water and were ready to
perform the ECE assay.
[0126] E) Screening Assay
[0127] Test compounds are solved and diluted in DMSO. 10 .mu.l of
DMSO was placed in the wells, followed by 125 .mu.l of assay buffer
(50 mM Tris/HCl, pH 7.0, 1 .mu.M Thiorphan, 0,1% NaN.sub.3, 0.1%
BSA) containing 200 ng big ET-1. The enzyme reaction was started by
the addition of 50 .mu.l of solubilized ECE (diluted in assay
buffer 1:30 to 1:60 fold (v/v)). The enzyme reaction was carried
out for 30 minutes at 37.degree. C. The enzyme reaction was stopped
by addition of 10 .mu.l 150 mM ETDA, pH 7.0.
Radioimmunoassay
[0128] The ET-1 RIA was performed principally as described earlier
(Loffler, B. -M. and Maire, J. -P. 1994, Endothelium 1: 273-286).
To plates containing the EDTA stopped enzyme reaction mixture 25
.mu.l of assay buffer containing 20000 cpm
(3-(.sup.125I)Tyr)-endothelin-1 and 25 .mu.l of the ET specific
antiserum AS-3 (dilution in assay buffer 1:1000) was added. Plates
were incubated under mixing at 4.degree. C. over night. Thereafter,
the liquid phase was sucked with a plate washer and plates were
washed once with bidestilled water. To the washed plates 200 .mu.l
scintillation cocktail (Microscint 40 LSC-Cocktail, Packard, code
6013641) was added and plates were counted for 2 minutes per well
in a Topcount.
[0129] Standard curves were prepared in plates with synthetic ET-1
with final concentrations of 0 to 3000 pg ET-1 per well. In all
plates controls for maximal ECE activity (in the presence of 10
.mu.l DMSO) and for background production of ET-1 immunoreactivity
(in the presence of 10 mM EDTA or 100 .mu.M phosphoramidon) were
performed. Assays were run in triplicate.
[0130] F) Kinetic Assay
[0131] The described assay format could be used to determine the
kinetic characteristics of the used ECE preparation as well as
different ECE inhibitors (i.e. Km, Ki) by variation of the
substrate concentration used in the assay.
[0132] G) Cell based ECE Assay
[0133] Human ECE-1c was stable expressed in MDCK cells as described
(Schweizer et al. 1997, Biochem. J. 328: 871-878). Cells were
cultured in 24 well plates to confluency in Dulbecco's modified
Eagles's medium (DMEM) supplemented with 10% (v/v) fetal bovine
serum (FBS), 0.8 mg/ml geneticin, 100 i.u./ml penicillin and 100
.mu.g/ml streptomycin in a humidified air/CO.sub.2 (19:1)
atmosphere. Before ECE assay the medium was replaced by 0.5 ml
DMEM-HBSS 1:1, 10 mM HEPES pH 7.0 supplemented with 0.1% (w/v) BSA.
The inhibitors were added in DMSO at a final concentration of 1%.
The enzyme reaction was started by the addition of 0.42 .mu.M human
big ET-1 and performed for 1.5 hours at 37.degree. C. in an
incubator. At the end of incubation, the incubation medium was
quickly removed and aliquots were analysed by radioimmunoassay for
produced ET-1 as described above.
[0134] The ECE screening assay was validated by the measurement of
the characteristic inhibitor constants of phosphoramidon (IC.sub.50
0.8.+-.0.2 .mu.M) and CGS 314447 (IC.sub.50 20.+-.4 nM) [De
Lombaert, Stephane; Stamford, Lisa B.; Blanchard, Louis; Tan,
Jenny; Hoyer, Denton; Diefenbacher, Clive G.; Wei, Dongchu;
Wallace, Eli M.; Moskal, Michael A.; et al. Potent non-peptidic
dual inhibitors of endothelin-converting enzyme and neutral
endopeptidase 24.11. Bioorg. Med. Chem. Lett. (1997), 7(8),
1059-1064]. The two inhibitors were measured with IC.sub.50 values
not significantly different from those described in the literature
but measured with different assay protocols. In the cell based
assay phosphoramidon showed an IC.sub.50 of 4 .mu.M. This assay
gave additional information about the inhibitory potency of
inhibitors under much more physiologic conditions, as e.g. the ECE
was embedded in a normal plasma membrane environment. It is
important to state, that the screening assay was performed in the
presence of 1 .mu.M Thiorphan to block any potential big ET-1
degradation due to the action of NEP24.11. No NEP activity was
present in MDCK-ECE-1c transfected cells in preliminary experiments
when ET-1 production was measured in presence or absence of
thiorphan. In subsequent experiments no thiorphan was added in the
incubation medium.
[0135] According to the above methods, the compounds of the present
invention show IC.sub.50 values in the radioimmunoassay (E on
ECE-inhibition) of about 50 nM to about 1000 .mu.M. The preferred
compounds show values of 50 nM to 1 .mu.M.
[0136] As mentioned earlier, medicaments containing a compound of
formula I are also an object of the present invention as is a
process for the manufacture of such medicaments, which process
comprises bringing one or more compounds of formula I and, if
desired, one or more other therapeutically valuable substances into
a galenical administration form.
[0137] The pharmaceutical compositions may be administered orally,
for example in the form of tablets, coated tablets, dragees, hard
or soft gelatin capsules, solutions, emulsions or suspensions.
Administration can also be carried out rectally, for example using
suppositories; locally or percutaneously, for example using
ointments, creams, gels or solutions; or parenterally, for example
using injectable solutions.
[0138] For the preparation of tablets, coated tablets, dragees or
hard gelatin capsules the compounds of the present invention may be
admixed with pharmaceutically inert, inorganic or organic
excipients. Examples of suitable excipients for tablets, dragees or
hard gelatin capsules include lactose, maize starch or derivatives
thereof, talc or stearic acid or salts thereof.
[0139] Suitable excipients for use with soft gelatin capsules
include for example vegetable oils, waxes, fats, semi-solid or
liquid polyols etc.; according to the nature of the active
ingredients it may however be the case that no excipient is needed
at all for soft gelatin capsules.
[0140] For the preparation of solutions and syrups, excipients
which may be used include for example water, polyols, saccharose,
invert sugar and glucose.
[0141] For injectable solutions, excipients which may be used
include for example water, alcohols, polyols, glycerin, and
vegetable oils.
[0142] For suppositories, and local or percutaneous application,
excipients which may be used include for example natural or
hardened oils, waxes, fats and semi-solid or liquid polyols.
[0143] The pharmaceutical compositions may also contain preserving
agents antioxidants, solubilising agents, stabilizing agents,
wetting agents, emulsifiers, sweeteners, colorants, odorants, salts
for the variation of osmotic pressure, buffers, coating agents or
antioxidants. They may also contain other therapeutically valuable
agents.
[0144] The dosages in which the compounds of formula I are
administered in effective amounts depend on the nature of the
specific active ingredient, the age and the requirements of the
patient and the mode of application. In general, dosages of 0.1-100
mg/kg body weight per day come into consideration, although the
upper limit quoted can be exceeded when this is shown to be
indicated.
The following specific examples are provided as a guide to assist
in the practice of the invention, and are not intended as a
limitation on the scope of the invention.
EXAMPLES
General Remarks
[0145] All reactions were done under argon.
[0146] Abbreviations: EtOH Ethanol, THF Tetrahydrofuran, Et.sub.2O
Diethylether, MeOH Methanol, CH.sub.2Cl.sub.2, EDCI
N-(3-Dimethylaminopropyl)-N'-ethylcarbodiimide hydrochloride, HOBT
1-Hydroxybenzotriazole, DBU
1,8-Diazabicyclo[5.4.0]undec-7-ene(1,5-5), LAH Lithium aluminium
hydride, LDA lithium diisopropylamide, DEAD Diethyl
azodicarboxylate, DIAD Diisopropyl azodicarboxylate, DMAP
4-Dimethylaminopyridine, DMAP-poly
4-(N-Benzyl-N-methylamino)pyridine, polymer-supported (polystyrol
based 2% DVB, ca 1,6 mmol "DMAP"/g resin), NEM N-ethylmorpholine,
NMM N-methylmorpholine, TBAF tetrabutylammonium fluoride, DIEA
diethylamine, DMF dimethylformamide, TFA trifluoroacetic acid, TPTU
2-(2-pyridon-1-yl)-1,1,3,3-tetramethyl uronium tetrafluorobrate,
iPr.sub.2NEt Huenigs base or N-ethyldiisopropylamine, FMOC-OSu
9-Fluorenylmethyloxycarbonyl-N-hydroxysuccinimide ester.
Example 1
Starting Materials (Esters--Scheme 1)
[0147] 40 g (220 mmol) of L-hydroxyproline
methylester-hydrochloride (twice suspended in toluene and
evaporated under reduced pressure to remove water) was suspended in
600 ml hexamethyldisilazane and refluxed for 2 h. The solution was
evaporated under reduced pressure and dissolved in 100 ml THF. 49.9
g (220 mmol) of 2-naphthalene-sulfonyl chloride in 200 ml of THF
were added slowly and stirred for 16 h at RT. 150 ml H.sub.2O were
added and after 1 h the solvents were evaporated. The residue was
partitioned between water/ethyl acetate (3.times.), the organic
phases were washed with 10% NaCl and dried over Na.sub.2SO.sub.4 to
give 60.4 g (82%) of
(2S,4R)-4-Hydroxy-1-(naphthalene-2-sulfonyl)-pyrr-
olidine-2-carboxylic acid methyl ester, MS: 335 (M.sup.+). 12
[0148] In analogy
[0149] L-hydroxyproline benzylester-hydrochloride and
1-naphthalenesulfonyl chloride gave
(2S,4R)-4-Hydroxy-1-(naphthalene-2-su-
lfonyl)-pyrrolidine-2-carboxylic acid benzyl ester, MS: 411
(MH.sup.+);
[0150] L-hydroxyproline benzylester-hydrochloride and
methanesulfonyl chloride gave
(2S,4R)-4-Hydroxy-1-methanesulfonyl-pyrrolidine-2-carboxyli- c acid
benzyl ester, mp 132-133.degree. C., MS: 300 (MH.sup.+);
[0151] L-hydroxyproline methylester-hydrochloride and
methanesulfonyl chloride gave after extraction with
CH.sub.2Cl.sub.2
(2S,4R)-4-Hydroxy-1-methanesulfonyl-pyrrolidine-2-carboxylic acid
methyl ester, mp 115.5-117.degree. C., MS: 164 (M-COOMe).
[0152] Via Mesylate: A biphasic solution of 13.9 ml (215 mmol)
methanesulfonic acid, 29.8 ml (215 mmol) triethylamine and 58.7 g
(224 mmol) triphenylphosphine in 150 ml toluene was added to a
suspension of 60 g (179 mmol)
(2S,4R)-4-Hydroxy-1-(naphthalene-2-sulfonyl)-pyrrolidine--
2-carboxylic acid methyl ester in 300 ml toluene which was stirred
mechanically. After adding 44.9 ml (233 mmol) of diisopropyl
azodicaboxylate (exothermic!) the solution was heated for 2.5 h at
80.degree. C. 300 ml water was added at RT and extracted with
ethylacetate (3.times.300 ml). The organic phase was washed with
aqueous 10% KHSO.sub.4 (2.times.100 ml), 10% NaCl (2.times.150 ml),
dried over Na.sub.2SO.sub.4 and evaporated to give 180 g of crude
product. Flash chromatography (ethyl acetate/hexane 1:1) gave 63.7
g (86%) of (4S,
2S)-4-Methanesulfonyloxy-1-(naphthalene-2-sulfonyl)-pyrrolidine-2-carboxy-
lic acid methylester.
[0153] 64.2 g (167 mmol) of triphenylmethanthiol was slowly added
at RT to a solution of 17.9 g (160 mmol) of potassium tert-butylate
in 300 ml DMF and stirred mechanically for 30 min. Then 63 g (152
mmol) of
(4S,2S)-4-Methanesulfonyloxy-1-(naphthalene-2-sulfonyl)-pyrrolidine-2-car-
boxylic acid methylester in 300 ml DMF were added at 20.degree. C.
by cooling at the end with an ice bath. The reaction was heated for
1.3 h at 100.degree. C., cooled, evaporated to 400 ml and extracted
with 250 ml aqueous saturated NH.sub.4Cl/ethyl acetate
(3.times.300). The organic phases were washed with aq. 10% NaCl,
dried (Na.sub.2SO.sub.4) and evaporated. Flash chromatography
(CH.sub.2Cl.sub.2/MeOH 99:1) gave 58.6 g (65%,
(2S,4R)/(2R,4R)-isomer ca 4:1, .sup.1H-NMR) and 9.2 g (10%,
(2S,4R)/(2R,4R)-isomer ca 1:1, .sup.1H-NMR) of
(2S,4R)-1-(Naphthalene-2-s-
ulfonyl)-4tritylsulfanyl-pyrrolidine-2-carboxylic acid methyl
ester, MS: 594 (MH.sup.+).
[0154] In analogy:
[0155] (2S,4R)-4-Hydroxy-1-methanesulfonyl-pyrrolidine-2-carboxylic
acid methyl ester gave after 3.75 h at 80.degree. C.
(4S,2S)-4-Methanesulfonyl-
oxy-1-(methylsulfonyl)-pyrrolidine-2-carboxylic acid methylester
which was heated for 45 min at 100.degree. C. with
triphenylmethanthiolate to give
(2S,4R)-1-Methanesulfonyl-4-tritylsulfanyl-pyrrolidine-2-carboxylic
acid methyl ester ((2S,4R)/(2R,4R)-isomer ca 9:1, .sup.1H-NMR), MS:
482 (MH.sup.+);
(2S,4R)-4-Hydroxy-1-methanesulfonyl-pyrrolidine-2-carboxylic acid
benzyl ester gave after 5 h at 80.degree. C.
(2S,4S)-1-Methanesulfon-
yl-4-methanesulfonyloxy-pyrrolidine-2-carboxylic acid benzyl ester
which was heated for 30 min with 4-methoxybenzylthiol/potassium
tert-butylate to give
(2S,4S)-1-Methanesulfonyl-4-methanesulfonyloxy-pyrrolidine-2-carb-
oxylic acid benzyl ester, mp 91-92.degree. C., MS: 453
(M+NH4.sup.+).
[0156] Via bromide: To a solution of 76.5 g (291.6 mmol, 6 eq)
triphenylphosphine in 650 ml THF were added 44.6 ml (286.8 mmol,
5.9 eq) DEAD in 70 ml THF at a temperature between 1.5-4.5.degree.
C. over a period of 0.5 h. The solution was stirred for 0.5 h
before 42.2 g (486.1 mmol, 10 eq) LiBr were added, and the reaction
mixture was recooled to 4.degree. C. for the addition of 20 g (48.6
mmol) (2S,4R)-4-Hydroxy-1-(na-
phthalene-2-sulfonyl)-pyrrolidine-2-carboxylic acid benzyl ester in
75 ml THF. After stirring at RT for 3 h, water was added and the
suspension concentrated and redissolved in 700 ml ethyl acetate and
water. The layers were separated, the inorganic one was extracted
with 100 ml of ethyl acetate (3.times.), and the combined organic
layers were washed with brine, dried over MgSO.sub.4 and
evaporated. Triphenylphosphine oxide was removed by crystallization
from ethyl acetate/hexane and the mother liquid was purified by
column chromatography on silica gel with hexane:ethyl acetate 3:1
yielding 13.4 g (62%) of (2S,4S)-4-Bromo-1-(naph-
thalene-2-sulfonyl)-pyrrolidine-2-carboxylic acid benzyl ester as
colorless solid, mp 97-98.degree. C., MS: 473 (MH.sup.+).
[0157] 3.38 g (30.1 mmol, 1.1 eq) potassium tert. butylate in 150
ml DMF were treated with 4.4 ml (31.5 mmol, 1.15 eq)
4-methoxybenzyl mercaptane at 0.degree. C. The solution was stirred
at RT for 1 h before 12.99 g (27.4 mmol)
(2S,4S)-4-Bromo-1-(naphthalene-2-sulfonyl)-pyrrolidine-2-carb-
oxylic acid benzyl ester in 100 ml DMF were added. The reaction was
stirred at RT overnight, DMF was removed under vacuum, and the
residue redissolved in ethyl acetate and 1M aq. KHSO.sub.4. The
layers were separated, and the organic one washed with brine, dried
over Na.sub.2SO.sub.4 and evaporated. The crude oil was purified by
flash chromatography on silica gel with hexane/ethyl acetate
(3:1-2:1) as eluent yielding 7.23 g (48%)
(2S,4R)-4-(4-Methoxy-benzylsulfanyl)-1-(naph-
thalene-2-sulfonyl)-pyrrolidine-2-carboxylic acid benzyl ester as
light yellow solid, mp 90-91.degree. C., MS: 547 (M.sup.+).
[0158] In analogy:
[0159] (2S,4R)-4-Hydroxy-pyrrolidine-1,2-dicarboxylic acid
1-tert-butyl ester 2-methyl ester with
4-methoxybenzylthiol/potassium tert-butylate gave
(2S,4R)-4-(4-Methoxy-benzylsulfanyl)-pyrrolidine-1,2-dicarboxylic
acid 1-tert-butyl ester 2-methyl ester, MS: 382 (MH.sup.+).
[0160]
(2S,4R)-4-Hydroxy-1-(naphthalene-2-sulfonyl)-pyrrolidine-2-carboxyl-
ic acid methyl ester with 4-methoxybenzylthiol/potassium
tert-butylate gave
(2S,4R)-4-(4-Methoxy-benzylsulfanyl)-1-(naphthalene-2-sulfonyl)-pyrr-
olidine-2-carboxylic acid methyl ester as colorless oil, MS: 472
(MH.sup.+);
[0161] Via chloride:
((2S,4R)-4-Tritylsulfanyl-pyrrolidine-1,2-dicarboxyli- c acid
1-tert-butyl ester 2-methyl ester: the synthesis of the
intermediate of the present invention is known in the art and
described for example in International Patent Application WO
9820001 and European Patent Application Publication No.
EP-A-696593.)
[0162] A solution of 374 g (1.48 mol)
(2S,4R)-4-Hydroxy-pyrrolidine-1,2-di- carboxylic acid 1-tert-butyl
ester 2-methyl ester in 1.6 l CH.sub.2Cl.sub.2 was treated with 680
g (2.6 mol) triphenylphosphine, cooled to 3-5.degree. C. and
treated in 10 min with 1.24 l (12.8 mol) CCl.sub.4, after 2 h at
this temperature cooling was stopped, the reaction temperature
raised during 2 h to 35.degree. C. It was cooled down to 20.degree.
C. and stirred for further 45 min. After addition of 4 l of
n-heptane, the reaction was evaporated to 2.9 l, cooled to
0.degree. C., filtered, the residue was treated twice the same way,
the third time by dissolving the residue again in 2 l of
CH.sub.2Cl.sub.2. The solvents were evaporated and filtered through
silica gel with hexane/tert.-butyl-methylether 9:1 as eluent.
Evaporation of the solvents gave 347 g (89%) of
(2S,4S)-4-Chloro-pyrrolidine-1,2-dicarboxylic acid 1-tert-butyl
ester 2-methyl ester, MS: 246 (MH.sup.+).
[0163] A solution of 76 g (0.68 mol) potassium-tert.-butylate in
1.5 l DMF was cooled (-3.degree. C.) and treated slowly (1.5 h)
with 202 g (0.73 mol) triphenylmethanethiol in 0.8 l DMF (at max
1.degree. C.). After 2.5 h at 0.degree. C., a solution of 161 g
(0.61 mol) of (2S,4S)-4-Chloro-pyrrolidine-1,2-dicarboxylic acid
1-tert-butyl ester 2-methyl ester in 0.35 1 DMF was added. The
reaction was stirred over night at 2.degree. C., evaporated,
dissolved in 1.5 l ethyl acetate, poured into 2.7 l aqueous
saturated NH.sub.4Cl solution and extracted with ethyl acetate
(2.times.). The organic phase was washed with aqueous saturated
NaHCO.sub.3, dried over Na.sub.2SO.sub.4 and evaporated. HPLC on
silica gel with hexane/ethyl acetate (95:5 to 7:3) gave 268 g (87%)
(2S,4R)-4-Tritylsulfanyl-pyrrolidine-1,2-dicarboxylic acid
1-tert-butyl ester 2-methyl ester, MS: 504 (MH.sup.+).
Example 2
Hydrolysis (Scheme1)
[0164] To a solution of 14.8 g (31.6 mmol)
(2S,4R)-4-(4-Methoxy-benzylsulf-
anyl)-1-(naphthalene-2-sulfonyl)-pyrrolidine-2-carboxylic acid
methyl ester in 950 ml THF were added 950 ml 0.1M LiOH (95 mmol) at
0.degree. C. The solution was stirred at RT for 2 h, diluted with
ice water, acidified by the addition of 1M KHSO.sub.4 (pH 2), and
extracted with ethyl acetate. The combined organic phases were
washed with brine, dried over Na.sub.2SO.sub.4 and were evaporated.
The product was crystallized from ethyl acetate/hexane yielding
13.15 g (90%) (2S,4R)-4-(4-Methoxy-benzylsu-
lfanyl)-1-(naphthalene-2-sulfonyl)-pyrrolidine-2-carboxylic acid as
colorless solid, MS: 456 (MH.sup.+).
[0165] Analogously the following compounds were prepared:
[0166]
(2S,4R)-1-Methanesulfonyl-4-tritylsulfanyl-pyrrolidine-2-carboxylic
acid as light brown foam, MS: 466 (M-H.sup.-);
[0167]
(2S,4R)-4-(4-Methoxy-benzylsulfanyl)-pyrrolidine-1,2-dicarboxylic
acid 1-tert-butyl ester MS: 366 (M-H.sup.-);
[0168] Additional compounds were prepared according to the
following references:
[0169] tert-Butyl 2-(isobutyl)hydrazinecarboxylate (Faessler,
Alexander; Bold, Guido; Capraro, Hans-Georg; Cozens, Robert;
Mestan, Juergen; Poncioni, Bernard; Roesel, Johannes;
Tintelnot-Blomley,
[0170] Marina; Lang, Marc. Aza-Peptide Analogs as Potent Human
Immunodeficiency Virus Type-1 Protease Inhibitors with Oral
Bioavailability. J. Med. Chem. (1996), 39(16), 3203-3216).
[0171] tert-Butyl 2-(methyl)hydrazinecarboxylate (Lenman, Morag M.;
Lewis, Arwel; Gani, David. Synthesis of fused
1,2,5-triazepine-1,5-diones and some N2- and N3-substituted
derivatives: potential conformational mimetics for cis-peptidyl
prolinamides. J. Chem. Soc., Perkin Trans. 1 (1997), Issue 16,
2297-2311).
[0172] N-Methyl-hydrazinecarboxylic acid benzyl ester (Lenman,
Morag M.; Lewis, Arwel; Gani, David. Synthesis of fused
1,2,5-triazepine-1,5-diones and some N2- and N3-substituted
derivatives: potential conformational mimetics for cis-peptidyl
prolinamides. J. Chem. Soc., Perkin Trans. 1 (1997), Issue 16,
2297-2311).
Example 3
Synthesis of Hydrazides
Example 3a
Sequence A (Scheme 2)
[0173] (step1) To a solution of 4.4 g (9.6 mmol)
(2S,4R)-4-(4-Methoxy-benz-
ylsulfanyl)-1-(naphthalene-2-sulfonyl)-pyrrolidine-2-carboxylic
acid in 200 ml CH.sub.2Cl.sub.2 were added 1.2 g (10.8 mmol,1.1 eq)
N-hydroxy-2-pyridone, followed by 2.2 g (10.7 mmol, 1.1 eq)
N,N-dicyclohexylcarbodiimide in 25 ml CH.sub.2Cl.sub.2 at 0.degree.
C. over a period of 30 min. The suspension was stirred for
additional 4 h at that temperature before 4.2 ml (33.0 mmol, 3.4
eq) NEM and 1.9 g (mmol, 1.05 eq) isobutylhydrazine-sulfate were
added. The reaction mixture was stirred at RT over night. The
suspension was treated with 0.55 ml (9.6 mmol, 1.0 eq) glacial
acetic acid in 10 ml water and stirred for 1.5 h, diluted with aq.
NaHCO.sub.3 (5%) and extracted with CH.sub.2Cl.sub.2. The combined
organic phases were washed with 1M KHSO.sub.4 solution, water and
brine, dried over Na.sub.2SO.sub.4 and evaporated. Tituration with
hexane yields 5.02 g (quant)
(2S,4R)-4-(4-Methoxy-benzylsulfanyl)-1--
(naphthalene-2-sulfonyl)-pyrrolidine-2-carboxylic acid
N'-isobutyl-hydrazide, which was directly subjected to the
following reaction. 13
[0174] (step2) 222 mg (0.42 mmol)
(2S,4R)-4-(4-Methoxy-benzylsulfanyl)-1-(-
naphthalene-2-sulfonyl)-pyrrolidine-2-carboxylic acid
N'-isobutyl-hydrazide in 10 ml TFA were treated with 0.68 ml (4.2
mmol, 10 eq) triethylsilane at 80.degree. C. for 90 min. The
solvent was evaporated in vacuo and the crude product was purified
by flash chromatography yielding 148 mg (86%)
(2S,4R)-4-Mercapto-1-(naphthalene-2--
sulfonyl)-pyrrolidine-2-carboxylic acid N'-isobutyl-hydrazide as
light yellow crystalline, MS: 408 (MH.sup.+).
[0175] Analogously the following compounds were prepared:
[0176] From
(2S,4R)-4-(4-Methoxy-benzylsulfanyl)-1-(naphthalene-2-sulfonyl-
)-pyrrolidine-2-carboxylic acid and
[0177] b) benzylhydrazine followed by deprotection:
(2S,4R)-4-Mercapto-1-(naphthalene-2-sulfonyl)-pyrrolidine-2-carboxylic
acid N'-benzyl-hydrazide as white solid, MS: 442 (MH.sup.+);
[0178] c)p-toluenesulfonylhydrazine followed by deprotection:
(2S,4R)-4-Mercapto-1-(naphthalene-2-sulfonyl)-pyrrolidine-2-carboxylic
acid N'-(4-methyl-benzenesulfonyl)-hydrazide as white solid, MS:
506 (MH.sup.+);
[0179] d) methylhydrazine followed by deprotection:
(2S,4R)-4-Mercapto-1-(naphthalene-2-sulfonyl)-pyrrolidine-2-carboxylic
acid N'-methyl-hydrazide as white crystalline, MS: 366
(MH.sup.+);
[0180] From
(2S,4R)-1-Methanesulfonyl-4-tritylsulfanyl-pyrrolidine-2-carbo-
xylic acid and p-toluenesulfonylhydrazine followed by deprotection:
(2S,4R)-4-Mercapto-1-methanesulfonyl-pyrrolidine-2-carboxylic acid
N'-(4-methyl-benzenesulfonyl)-hydrazide as white crystalline, MS:
394 (MH.sup.+);
Example 3b
Sequence B (Step1 from Sequence A followed by step 3,4-Scheme2)
[0181] (step 3) 4.3 g (3.15 mmol)
(2S,4R)-4-(4-Methoxy-benzylsulfanyl)-1-(-
naphthalene-2-sulfonyl)-pyrrolidine-2-carboxylic acid
N'-isobutyl-hydrazide in 450 ml CH.sub.2Cl.sub.2 were treated with
5.6 ml (32.6 mmol, 4 eq) N-ethyldiisopropylamine, 3.1 g (16.3 mmol,
2 eq) p-toluene sulfonyl chloride and 100 mg (0.8 mmol, 0.1 eq)
DMAP at 0.degree. C. and was stirred at RT over night. 2.05 g (16.1
mmol, 2 eq) MeNHCH.sub.2CO.sub.2K were added and, the solution was
stirred at RT for 1 h, 1M KHSO.sub.4 solution was added and, the
phases were separated. The organic layer was extracted with sat.
NaHCO.sub.3 and, the inorganic layers were washed with
CH.sub.2Cl.sub.2. The combined organic phases were washed with
brine, dried over Na.sub.2SO.sub.4 and evaporated. Purification of
the crude residue by flash chromatography with hexane:ethyl acetate
2:1 as eluent yields 2.68 g (48%)
(2S,4R)-4-(4-Methoxy-benzylsulfanyl)-1-(naphthalene-2-sulfonyl)-pyrrolidi-
ne-2-carboxylic acid
N'-isobutyl-N'-(4-methyl-benzenesulfonyl)-hydrazide, which was
directly subjected to the following reaction.
[0182] (step 4) To 2.68 g (3.9 mmol, 1 eq)
(2S,4R)-4-(4-Methoxy-benzylsulf-
anyl)-1-(naphthalene-2-sulfonyl)-pyrrolidine-2-carboxylic acid
N'-isobutyl-N'-(4-methyl-benzenesulfonyl)-hydrazide in 100 ml TFA
were added 6.2 ml (39 mmol, 10 eq) triethylsilane and the mixture
was heated to 80.degree. C. for 1.5 h, concentrated in vacuo and
redissolved in toluene and evaporated. Tituration with hexane
yields the crude product which was further purified by flash
chromatography with hexane:ethyl acetate 1:1 yielding 1.65 g (74%)
(2S,4R)-4-Mercapto-1-(naphthalene-2-sul-
fonyl)-pyrrolidine-2-carboxylic acid
N'-isobutyl-N'-(4-methyl-benzenesulfo- nyl)-hydrazide as white
crystalline, MS: 562 (MH.sup.+).
[0183] Analogously, the following compounds were prepared (steps
1,3,4):
[0184] From
(2S,4R)-4-(4-Methoxy-benzylsulfanyl)-1-(naphthalene-2-sulfonyl-
)-pyrrolidine-2-carboxylic acid
[0185] b) and methylhydrazine, followed by reaction with
p-toluenesulfonyl chloride and deprotection:
(2S,4R)-4-Mercapto-1-(naphthalene-2-sulfonyl)--
pyrrolidine-2-carboxylic acid
N'-methyl-N'-(4-methyl-phenylsulfonyl)-hydra- zide as white
crystalline, MS: 520 (MH.sup.+);
[0186] c) and isobutylhydrazine.sulfate, followed by reaction with
4-t-butyl-phenylsulfonyl chloride and deprotection:
(2S,4R)-4-Mercapto-1-(naphthalene-2-sulfonyl)-pyrrolidine-2-carboxylic
acid N'-(4-tert-butyl-benzenesulfonyl)-N'-isobutyl-hydrazide as
white crystalline, MS: 604 (MH.sup.+);
[0187] d) and methylhydrazine, followed by reaction with
methanesulfonyl chloride and deprotection:
(2S,4R)-4-Mercapto-1-(naphthalene-2-sulfonyl)--
pyrrolidine-2-carboxylic acid
N'-methanesulfonyl-N'-methyl-hydrazide as white solid, MS: 444
(MH.sup.+);
[0188] e) and isobutylhydrazine.sulfate, followed by reaction with
methanesulfonyl chloride and deprotection:
(2S,4R)-4-Mercapto-1-(naphthal-
ene-2-sulfonyl)-pyrrolidine-2-carboxylic acid
N'-isobutyl-N'-methanesulfon- yl-hydrazide as white crystalline,
MS: 486 (MH.sup.+);
[0189] f) and benzylhydrazine, followed by reaction with
methanesulfonyl chloride and deprotection:
(2S,4R)-4-Mercapto-1-(naphthalene-2-sulfonyl)--
pyrrolidine-2-carboxylic acid
N'-benzyl-N'-methanesulfonyl-hydrazide as white crystalline, MS:
520 (MH.sup.+);
[0190] g) and benzylhydrazine, followed by reaction with
p-toluenesulfonyl chloride and deprotection:
(2S,4R)-4-Mercapto-1-(naphthalene-2-sulfonyl)--
pyrrolidine-2-carboxylic acid
N'-benzyl-N'-(4-methyl-phenylsulfonyl)-hydra- zide as white
crystalline, MS: 596 (MH.sup.+); From
(2S,4R)-1-Methanesulfonyl-4-tritylsulfanyl-pyrrolidine-2-carboxylic
acid
[0191] a2) and benzylhydrazine, followed by reaction with
methanesulfonyl chloride and deprotection:
(2S,4R)-4-Mercapto-1-methanesulfonyl-pyrrolidi- ne-2-carboxylic
acid N'-benzyl-N'-methanesulfonyl-hydrazide as white solid, MS: 408
(MH.sup.+);
[0192] b2) and isobutylhydrazine.sulfate, followed by reaction with
methanesulfonyl chloride and deprotection:
(2S,4R)-4-Mercapto-1-methanesu- lfonyl-pyrrolidine-2-carboxylic
acid N'-isobutyl-N'-methanesulfonyl-hydraz- ide as colorless solid,
MS: 450 (MH.sup.+);
[0193] c2) and isobutylhydrazine.sulfate, followed by reaction with
p-toluenesulfonyl chloride and deprotection:
(2S,4R)-4-Mercapto-1-methane- sulfonyl-pyrrolidine-2-carboxylic
acid N'-isobutyl-N'-(4-methyl-benzenesul- fonyl)-hydrazide as
colorless solid, MS: 374 (MH.sup.+);
[0194] d2) and isobutylhydrazine.sulfate, followed by reaction with
4-tert-butyl-benzenesulfonyl chloride and deprotection:
(2S,4R)-4-Mercapto-1-methanesulfonyl-pyrrolidine-2-carboxylic acid
N'-(4-tert-butyl-benzenesulfonyl)-N'-isobutyl-hydrazide as white
crystalline, MS: 492 (MH.sup.+);
Example 3c
Sequence C (Scheme 2)
[0195] Analogously to sequence A,B (steps 1,3), from
(2S,4R)-4-(4-Methoxy-benzylsulfanyl)-pyrrolidine-1,2-dicarboxylic
acid 1-tert-butyl ester and methylhydrazine, followed by treatment
with p-toluenesulfonyl chloride
(2S,4R)-2-[N'-Methyl-N'-(4-methyl-phenylsulfon-
yl)-hydrazinocarbonyl]-4-(4-methoxy-benzylsulfanyl)-pyrrolidine-1-carboxyl-
ic acid 1-tert-butyl ester, which was directly subjected to the
following reaction.
[0196] (step5) 2.37 g (4.3 mmol)
(2S,4R)-2-[N'-Methyl-N'-(4-methyl-phenyls-
ulfonyl)-hydrazinocarbonyl]-4-(4-methoxy-benzylsulfanyl)-pyrrolidine-1-car-
boxylic acid 1-tert-butyl ester in 10 ml CH.sub.2Cl.sub.2 were
treated with 4 ml TFA at 0.degree. C. and kept in the freezer over
night. The solvent was evaporated, the crude material was dissolved
and evaporated with toluene (2.times.) and hexane (3.times.)
yielding
(2S,4R)-2-[N'-Methyl-N'-(4-methyl-phenylsulfonyl)-hydrazinocarbonyl]-4-(4-
-methoxy-benzylsulfanyl)-pyrrolidine as TFA salt as crude product,
which was subjected to the following reaction without further
purification.
[0197] (step 6) To 250 mg (0.44 mmmol, 1.0 eq)
(2S,4R)-2-[N'-Methyl-N'-(4--
methyl-phenylsulfonyl)-hydrazinocarbonyl]-4-(4-methoxy-benzylsulfanyl)-pyr-
rolidine-TFA in 5 ml CH.sub.2Cl.sub.2 were added 360 .mu.l (2.1
mmol, 4.8 eq) N-ethyldiisopropylamine and 70 .mu.l (0.56 mmol, 1.2
eq) phenyl chloroformate at 0.degree. C. The solution was stirred
at RT over night, 1N KHSO.sub.4 was added, the phases were
separated and the inorganic one was extracted with
CH.sub.2Cl.sub.2, the organic layer was washed with 1M KHSO.sub.4
and brine, dried over Na.sub.2SO.sub.4 and evaporated.
[0198] (step 7) The crude material was redissolved in 10 ml TFA
and, 700 .mu.l (4.4 mmol, 10 eq) triethylsilane were added at RT
and the solution was stirred at 80.degree. C. for 70 min.
Evaporation and flash chromatography with hexane:ethyl acetate 1:1
followed by lyophilisation yields 149.2 mg (75%)
(2S,4R)-2-[N'-Methyl-N'-(4-methyl-phenylsulfonyl)-h-
ydrazinocarbonyl]-4-mercapto-pyrrolidine-1-carboxylic acid phenyl
ester as white solid, MS: 450 (MH.sup.+).
[0199] In a similar manner (step 6,7) the following compounds were
prepared: from
(2S,4R)-2-[N'-Methyl-N'-(4-methyl-phenylsulfonyl)-hydrazin-
ocarbonyl]-4-(4-methoxy-benzylsulfanyl)-pyrrolidine-TFA with
n-butyl chloroformate, i-propyl chloroformate, butylsulfamoyl
chloride, cyclopropylsulfamoylchloride,
benzylsulfamoylchloride:
[0200]
(2S,4R)-2-[N'-Methyl-N'-(4-methyl-phenylsulfonyl)-hydrazinocarbonyl-
]-4-mercapto-pyrrolidine-1-carboxylic acid butyl ester as colorless
gum, MS: 430 (MH.sup.+);
[0201]
(2S,4R)-2-[N'-Methyl-N'-(4-methyl-phenylsulfonyl)-hydrazinocarbonyl-
]-4-mercapto-pyrrolidine-1-carboxylic acid isopropyl ester as white
solid, MS: 416 (MH.sup.+);
[0202]
(2S,4R)-2-[N'-Methyl-N'-(4-methyl-phenylsulfonyl)-hydrazinocarbonyl-
]-4-mercapto-pyrrolidine-1-sulfonic acid butylamide as white lyoph
solid, MS: 463 (M-H).sup.-;
[0203]
(2S,4R)-2-[N'-Methyl-N'-(4-methyl-phenylsulfonyl)-hydrazinocarbonyl-
]-4-mercapto-pyrrolidine-1-sulfonic acid cyclopropylamide as white
lyoph solid, MS: 447 (M-H).sup.-;
[0204]
(2S,4R)-2-[N'-Methyl-N'-(4-methyl-phenylsulfonyl)-hydrazinocarbonyl-
]-4-mercapto-pyrrolidine-1-sulfonic acid benzylamide as white lyoph
solid, MS: 497 (M-H).sup.-;
[0205] From
(2S,4R)-2-[N'-Isobutyl-N'-(4-methyl-phenylsulfonyl)-hydrazinoc-
arbonyl]-4-(4-methoxy-benzylsulfanyl)-pyrrolidine-1-carboxylic acid
1-tert-butyl ester was treated with i-propyl chloroformate, n-butyl
chloroformate, benzyl chloroformate, phenyl chloroformate,
quinoline-8-sulfonyl chloride, thiophene-2-sulfonyl chloride,
benzylsulfamoylchloride, butylsulfamoyl chloride or
cyclopropylsulfamoylchloride according to protocols (steps 5-7) to
give the following compounds:
[0206]
(2S,4R)-2-[N'-Isobutyl-N'-(4-methyl-phenylsulfonyl)-hydrazinocarbon-
yl]-4-mercapto-pyrrolidine-1-carboxylic acid isopropyl ester as
white solid, MS: 458 (MH.sup.+);
[0207]
(2S,4R)-2-[N'-Isobutyl-N'-(4-methyl-phenylsulfonyl)-hydrazinocarbon-
yl]-4-mercapto-pyrrolidine-1-carboxylic acid butyl ester as white
solid, MS: 472 (MH.sup.30 );
[0208]
(2S,4R)-2-[N'-Isobutyl-N'-(4-methyl-phenylsulfonyl)-hydrazinocarbon-
yl]-4-mercapto-pyrrolidine-1-carboxylic acid benzyl ester as white
solid, MS: 506 (MH.sup.+);
[0209]
(2S,4R)-2-[N'-Isobutyl-N'-(4-methyl-phenylsulfonyl)-hydrazinocarbon-
yl]-4-mercapto-pyrrolidine-1-carboxylic acid phenyl ester as white
solid, MS: 492 (MH.sup.+);
[0210]
(2S,4R)-4-Mercapto-1-(quinoline-8-sulfonyl)-pyrrolidine-2-carboxyli-
c acid N'-isobutyl-N'-(4-methyl-benzyl)-hydrazide as white solid,
MS: 563 (MH.sup.+);
[0211]
(2S,4R)-4-Mercapto-1-(thiophen-2-sulfonyl)-pyrrolidine-2-carboxylic
acid N'-isobutyl-N'-(4-methyl-benzyl)-hydrazide as white solid, MS:
518 (MH.sup.+);
[0212]
(2S,4R)-2-[N'-Isobutyl-N'-(4-methyl-phenylsulfonyl)-hydrazinocarbon-
yl]-4-mercapto-pyrrolidine-1-sulfonic acid benzylamide as white
lyoph solid, mp 67.degree. C. MS: 539 (M-H).sup.-;
[0213]
(2S,4R)-2-[N'-Isobutyl-N'-(4-methyl-phenylsulfonyl)-hydrazinocarbon-
yl]-4-mercapto-pyrrolidine-1-sulfonic acid butylamide as white
lyoph solid, MS: 467 (M-H).sup.-;
[0214]
(2S,4R)-2-[N'-Isobutyl-N'-(4-methyl-phenylsulfonyl)-hydrazinocarbon-
yl]-4-mercapto-pyrrolidine-1-sulfonic acid cyclopropylamide as
white lyoph solid, MS: 489 (M-H).sup.-.
Example 3d
Sequence D--Direct Formation from the Ester (Scheme 3)
[0215] Analogously to Ruye Xing and Robert P. Hanzlik, J. Med.
Chem. 1998, 41, 1344-1351 the following reactions were carried
out:
[0216] (step 8) To a solution of 5 g (10.6 mmol, 1 eq)
(2S,4R)-4-(4-Methoxy-benzylsulfanyl)-1-(naphthalene-2-sulfonyl)-pyrrolidi-
ne-2-carboxylic acid methyl ester in 20 ml methanol were added 6.45
ml (110 mmol, 10 eq) hydrazine-hydrate and the solution was stirred
at RT for 3 days. The solvent was evaporated, followed by solving
and evaporating with EtOH, ether and hexane. The light yellow solid
was dried in vacuo giving
(2S,4R)-4-(4-Methoxy-benzylsulfanyl)-1-(naphthalene-2-sul-
fonyl)-pyrrolidine-2-carboxylic acid hydrazide, mp 130.degree. C.,
MS: 472 (MH.sup.+).
[0217] (step 9) 150 mg (0.3 mmol, 1.0 eq)
(2S,4R)-4-(4-Methoxy-benzylsulfa-
nyl)-1-(naphthalene-2-sulfonyl)-pyrrolidine-2-carboxylic acid
hydrazide in 5 ml TFA and 0.65 ml triisopropylsilane were stirred
for 3 d at RT. The solvent was evaporated and the residue
redissolved in sat. NaHCO.sub.3 solution:ethyl acetate, the phases
were separated and the inorganic one was extracted with ethyl
acetate. The combined organic phases were washed with water and
brine. Column chromatography yields 68 mg ( 61%)
(2S,4R)-4-Mercapto-1-(naphthalene-2-sulfonyl)-pyrrolidine-2-carboxylic
acid hydrazide as white foam, MS: 352 (MH.sup.+). 14
Example 3e
Sequence E--intermediates (Scheme 3)
[0218] Analogously to Sequence D step 8, the following compound was
prepared from
(2S,4R)-1-(Naphthalene-2-sulfonyl)-4-tritylsulfanyl-pyrroli-
dine-2-carboxylic acid methyl ester and hydrazine hydrate:
[0219]
(2S,4R)-1-(Naphthalene-2-sulfonyl)-4-tritylsulfanyl-pyrrolidine-2-c-
arboxylic acid hydrazide as off-white solid, mp 172.degree. C., MS:
594 (MH.sup.+).
[0220] (step 10) To a suspension of 3.0 g (5.05 mmol)
(2S,4R)-1-(Naphthalene-2-sulfonyl)
tritylsulfanyl-pyrrolidine-2-carboxyli- c acid hydrazide in ethanol
were added 0.56 ml (5.6 mmol, 1.1 eq) benzaldehyde at RT, and the
reaction mixture was heated to 80.degree. C. for 3 h. The solvent
was evaporated and the residue was purified by flash chromatography
on silica gel with ethyl acetate:hexane 1:1 as eluent yielding 2.75
g (80%) (2S,4R)-1-(Naphthalene-2-sulfonyl)-4-tritylsulfanyl-
-pyrrolidine-2-carboxylic acid benzylidene-hydrazide as white foam,
MS: 682 (MH.sup.+).
[0221] (step 11) To 2.48 g (3.64 mmol)
(2S,4R)-1-(Naphthalene-2-sulfonyl)--
4-tritylsulfanyl-pyrrolidine-2-carboxylic acid
benzylidene-hydrazide were added 228.6 mg (3.64 mmol, 1.0 eq)
NaBH.sub.3CN in 12.4 ml THF, followed by 691.8 mg (3.64 mmol, 1.0
eq) toluene-4-sulfonic acid in 8.7 ml THF. The solution was stirred
at RT for 2 h, additional mmol,0.3 eq) NaBH.sub.3CN were added, and
the reaction was stirred overnight at RT. The mixture was diluted
with ethyl acetate and washed with brine, sat. NaHCO.sub.3 solution
and brine, dried over Na.sub.2SO.sub.4 and the solvent was
evaporated.
[0222] The residue was dissolved in 11 ml 1M NaOH and 15 ml THF and
stirred for 1 h, diluted with ethyl acetate and 9 ml 1M KHSO.sub.4
were added, followed by 5% NaHCO.sub.3 solution. The slightly basic
solution was extracted with ethyl acetate. The organic phase was
washed with water and brine, dried over Na.sub.2SO.sub.4 and
evaporated yielding 2.45 g (98%)
(2S,4R)-1-(Naphthalene-2-sulfonyl)-4-tritylsulfanyl-pyrrolidine-2-c-
arboxylic acid N'-benzyl-hydrazide as white foam, MS: 684
(MH.sup.+). (analogously to Alexander Faissler, Guido Bold,
Hans-Georg Capraro, Robert Cozens, Jurgen Mestan, Bernard Poncioni,
Johannes Rosel, Marina Tintelnot-Blomley, and Marc Lang, J. Med.
Chem. 1996, 39, 16, 3203-3216.)
[0223] Analogously, the following compound was prepared from
(2S,4R)-1-(Naphthalene-2-sulfonyl)-4-tritylsulfanyl-pyrrolidine-2-carboxy-
lic acid hydrazide and 2,5-difluoro-benzaldehyde:
[0224]
(2S,4R)-1-(Naphthalene-2-sulfonyl)-4-tritylsulfanyl-pyrrolidine-2-c-
arboxylic acid N'-(2,5-difluoro-benzyl)-hydrazide as white foam,
MS: 720 (MH.sup.+).
Example 3e
Sequence E-final Products (Scheme 3)
[0225] (step 12) To 200 mg (0.3 mmol)
(2S,4R)-1-(Naphthalene-2-sulfonyl)-4-
-tritylsulfanyl-pyrrolidine-2-carboxylic acid N'-benzyl-hydrazide
in 3 ml CH.sub.2Cl.sub.2 were added 60 .mu.l (0.35 mmol, 1.2 eq)
N,N-diisopropylethylamine, 72.5 mg (0.35 mmol, 1.2 eq)
4-methoxybenzene sulfonylchloride and 22.5 mg (0.12 eq) DMAP-poly
at 0.degree. C. The suspension was shaken at RT for 3 days,
additional 25 .mu.l (0.15 mmol, 0.5 eq) N,N-diisopropylethylamine,
31 mg (0.15 mmol, 0.5 eq) 4-methoxybenzene sulfonylchloride and
22.5 mg (0.12 eq) DMAP-poly were added and the reaction was
continued for a day. After filtration and washing of the resin with
CH.sub.2Cl.sub.2, the organic phase was concentrated and the crude
material purified by flash chromatography yielding 152 mg (61%)
(2S,4R)-1-(Naphthalene-2-sulfonyl)-4-tritylsulfanyl-
-pyrrolidine-2-carboxylic acid
N'-benzyl-N'-(4-methoxy-benzenesulfonyl)-hy- drazide which was
directly subjected to the following reaction.
[0226] (step 13) To 148 mg (0.17 mmol) )
(2S,4R)-1-(Naphthalene-2-sulfonyl-
)-4-tritylsulfanyl-pyrrolidine-2-carboxylic acid
N'-benzyl-N'-(4-methoxy-b- enzenesulfonyl)-hydrazide in 3.0 ml TFA
were added 276 .mu.l (1.73 mmol, 10 eq) triethylsilane at 0.degree.
C. The mixture was stirred at RT for 30 min, the solvent was
evaporated and the residue was redissolved in ethyl acetate, sat.
aq. NaHCO.sub.3 solution, the layers were separated and the
inorganic one extracted with ethyl acetate. The combined organic
layers were washed with brine, dried over Na.sub.2SO.sub.4 and
evaporated. The residue was purified by flash chromatography with a
gradient of ethyl acetate:hexane (1:1.5 to 1:1) yielding 76 mg
(72%)
(2S,4R)-4-Mercapto-1-(naphthalene-2-sulfonyl)-pyrrolidine-2-carboxylic
acid N'-benzyl-N'-(4-methoxy-benzenesulfonyl)-hydrazide as white
foam, MS: 612 (MH.sup.+).
[0227] Analogously, the following compounds were prepared from
(2S,4R)-1-(Naphthalene-2-sulfonyl)-4-tritylsulfanyl-pyrrolidine-2-carboxy-
lic acid N'-benzyl-hydrazide and p-toluoyl chloride, acetyl
chloride, p-anisoyl chloride followed by deprotection:
[0228] (b) (2S,4R)-4-Methyl-benzoic acid
N-benzyl-N'-[4-mercapto-1-(naphth-
alene-2-sulfonyl)-pyrrolidine-2-carbonyl]-hydrazide as white foam,
MS: 560 (MH.sup.+).
[0229] (c)
(2S,4R)-4-Mercapto-1-(naphthalene-2-sulfonyl)-pyrrolidine-2-car-
boxylic acid N'-acetyl-N'-benzyl-hydrazide as white solid, mp
64.degree. C., MS: 484 (MH.sup.+).
[0230] (d) (2S,4R)-4-Methoxy-benzoic acid
N-benzyl-N'-[4-mercapto-1-(napht-
halene-2-sulfonyl)-pyrrolidine-2-carbonyl]-hydrazide as white foam,
MS: 576 (MH.sup.+).
[0231] From
(2S,4R)-1-(Naphthalene-2-sulfonyl)-4-tritylsulfanyl-pyrrolidin-
e-2-carboxylic acid N'-(2,5-difluoro-benzyl)-hydrazide and
[0232] 2,4-difluorobenzoyl chloride, 2-thiophene carbonyl chloride,
methanesulfonyl chloride, methoxybenzenesulfonyl chloride,
2-thiophenesulfonyl chloride, benzenesulfonyl chloride,
4-fluorobenzenesulfonyl chloride:
[0233] (e) (2S,4R)-2,4-Difluoro-benzoic acid
N-(2,5-difluoro-benzyl)-N'-[4-
-mercapto-1-(naphthalene-2-sulfonyl)-pyrrolidine-2-carbonyl]-hydrazide
as white foam, MS: 618 (MH.sup.+).
[0234] (f) (2S,4R)-Thiophene-2-carboxylic acid
N-(2,5-difluoro-benzyl)-N'--
[4-mercapto-1-(naphthalene-2-sulfonyl)-pyrrolidine-2-carbonyl]-hydrazide
as white foam, MS: 588 (MH.sup.+).
[0235] (g)
(2S,4R)-1-(Naphthalene-2-sulfonyl)-4-tritylsulfanyl-pyrrolidine-
-2-carboxylic acid
N'-(2,5-difluoro-benzyl)-N'-methanesulfonyl-hydrazide as white
foam, MS: 556 (MH.sup.+).
[0236] (h)
(2S,4R)-1-(Naphthalene-2-sulfonyl)-4-tritylsulfanyl-pyrrolidine-
-2-carboxylic acid
N'-(2,5-difluoro-benzyl)-N'-(4-methoxy-benzenesulfonyl)- -hydrazide
as white foam, MS: 648 (MH.sup.+).
[0237] (i)
(2S,4R)-1-(Naphthalene-2-sulfonyl)-4-tritylsulfanyl-pyrrolidine-
-2-carboxylic acid
N'-(2,5-difluoro-benzyl)-N'-(thiophene-2-sulfonyl)-hydr- azide as
white foam, MS: 624 (MH.sup.+).
[0238] (j)
(2S,4R)-1-(Naphthalene-2-sulfonyl)-4-tritylsulfanyl-pyrrolidine-
-2-carboxylic acid
N'-(2,5-difluoro-benzyl)-N'-benzenesulfonyl-hydrazide as white
foam, MS: 618 (MH.sup.+).
[0239] (k)
(2S,4R)-1-(Naphthalene-2-sulfonyl)-4-tritylsulfanyl-pyrrolidine-
-2-carboxylic acid
N'-(2,5-difluoro-benzyl)-N'-(4-fluoro-benzenesulfonyl)-- hydrazide
as white foam, MS: 636 (MH.sup.+).
Example 3f
Synthesis According to Scheme 4
[0240] Analogously to Sequence A--step 1, from
(2S,4R)-4-(4-Methoxy-benzyl-
sulfanyl)-1-(naphthalene-2-sulfonyl)-pyrrolidine-2-carboxylic acid
and N-Methyl-hydrazinecarboxylic acid benzyl ester was prepared
intermediate
[0241]
(2S,4R)-N'-[4-(4-Methoxy-benzylsulfanyl)-1-(naphthalene-2-sulfonyl)-
-pyrrolidine-2-carbonyl]-N-methyl-hydrazinecarboxylic acid benzyl
ester as off-white crystalline, MS: 620 (MH.sup.+).
[0242] (step 14) 2.45 g (1.52 mmol)
(2S,4R)-N'-[4-(4-Methoxy-benzylsulfany-
l)-1-(naphthalene-2-sulfonyl)-pyrrolidine-2-carbonyl]-N-methyl-hydrazineca-
rboxylic acid benzyl ester were dissolved in 9 ml acetic acid and 9
ml HBr (33% in acetic acid) were added at 0.degree. C. stirred for
4 h at 0.degree. C. The solution was concentrated and the residue
was dissolved in CH.sub.2Cl.sub.2 and sat. aq NaHCO.sub.3 solution,
the layers were separated and the inorganic one was extracted with
CH.sub.2Cl.sub.2 and the combined organic phases were washed with
brine and dried over Na.sub.2SO.sub.4. The crude product was
purified by flash chromatography with ethyl acetate:hexane 1:1
yielding 380 mg (44%)
(2S,4R)-4-(4-Methoxy-benzylsulfanyl)-1-(naphthalene-2-sulfonyl)-pyrrolidi-
ne-2-carboxylic acid N-benzyl-N-methyl-hydrazide as white solid, mp
83.2.degree. C., MS: 576 (MH.sup.+).
[0243] Deprotection according to Scheme 2 (step 4) gave:
[0244]
(2S,4R)4-Mercapto-1-(naphthalene-2-sulfonyl)-pyrrolidine-2-carboxyl-
ic acid N-benzyl-N-methyl-hydrazide as white solid, mp 69.degree.
C., MS: 456 (MH.sup.+).
[0245] (step16) 180 mg (0.31 mmol)
(2S,4R)-4-(4-Methoxy-benzylsulfanyl)-1--
(naphthalene-2-sulfonyl)-pyrrolidine-2-carboxylic acid
N-benzyl-N-methyl-hydrazide in 7.5 ml DMF were treated with 16.5 mg
(0.34 mmol, 1.1 eq) NaH and 21 .mu.l (0.34 mmol, 1.1 eq) methyl
iodide at 0.degree. C. and, the solution was stirred for 3 h at RT.
The solution was diluted with water and CH.sub.2Cl.sub.2. The
phases were separated and the inorganic layer was extracted with
CH.sub.2Cl.sub.2, the combined organic layers were washed with
brine, dried over Na.sub.2SO.sub.4 and were evaporated. The crude
material was purified by flash chromatography with a gradient of
ethyl acetate:hexane 2:1 to ethyl acetate yielding 110 mg (60%)
(2S,4R)-4-(4-Methoxy-benzylsulfanyl)-1-(naphthalene-2-sulfonyl)--
pyrrolidine-2-carboxylic acid N'-benzyl-N,N-dimethyl-hydrazide
which was deprotected using the protocol described 15
[0246] for Scheme 2 (step 4) to give
(2S,4R)-4-Mercapto-1-(naphthalene-2-s-
ulfonyl)-pyrrolidine-2-carboxylic acid
N'-benzyl-N,N-dimethyl-hydrazide as white solid, mp 133.2.degree.
C., MS: 470 (MH.sup.+).
Example 3g
Synthesis According to Scheme 5
[0247] Analogously to Sequence A--step 1, from
(2S,4R)-4-(4-Methoxy-benzyl-
sulfanyl)-1-(naphthalene-2-sulfonyl)-pyrrolidine-2-carboxylic acid
and tert-Butyl 2-(methyl)hydrazinecarboxylate was prepared
intermediate (2S,4R)-N'-[4-(4-Methoxy-benzylsulfanyl)-1-(
naphthalene-2-sulfonyl)-pyrr-
olidine-2-carbonyl]-N'-methyl-hydrazinecarboxylic acid tert-butyl
ester as white crystalline, MS: 586 (MH.sup.+).
[0248] (step 19) To 200 mg (0.34 mmol)
(2S,4R)-N'-[4-(4-Methoxy-benzylsulf-
anyl)-1-(naphthalene-2-sulfonyl)-pyrrolidine-2-carbonyl]-N'-methyl-hydrazi-
necarboxylic acid tert-butyl ester in 10 ml TFA were added 0.54 ml
(3.4 mmol, 10 eq) triethylsilane at RT and the solution was stirred
at 80.degree. C. for 1 h. The solvent was evaporated in vacuo and
the crude material was purified by flash chromatography with ethyl
acetate/hexane yielding 61.6 mg (40%)
(2S,4R)-4-Mercapto-1-(naphthalene-2-sulfonyl)-pyrr-
olidine-2-carboxylic acid N-methyl-N'-trifluoroacetyl-hydrazide as
white solid, MS: 461 (MH.sup.+).
[0249] (step 20, 21) According to the procedure (Scheme 4, step 16)
was prepared from (2S,4R)-N'-[4-(4-Methoxy-benzylsulfanyl)-1-(
naphthalene-2-sulfonyl)-pyrrolidine-2-carbonyl]-N'-methyl-hydrazinecarbox-
ylic acid tert-butyl ester and benzyl bromide
(2S,4R)-N'-[4-(4-Methoxy-ben- zylsulfanyl)-1-(
naphthalene-2-sulfonyl)-pyrrolidine-2-carbonyl]-N'-methyl-
-N-benzyl-hydrazinecarboxylic acid tert-butyl ester which was
directly BOC and PMB-deprotected according to Scheme 2 (step 4) to
give
(2S,4R)-4-Mercapto-1-(naphthalene-2-sulfonyl)-pyrrolidine-2-carboxylic
acid N-benzyl-N-methyl-hydrazide as colorless oil, mp, MS: 456
(MH.sup.+).
[0250] (Step 22) 2 g (3.4 mmol)
(2S,4R)-N'-[4-(4-Methoxy-benzylsulfanyl)-1-
-(naphthalene-2-sulfonyl)-pyrrolidine-2-carbonyl]-N'-methyl-hydrazinecarbo-
xylic acid tert-butyl ester in 9 ml CH.sub.2Cl.sub.2 were treated
with 3.4 ml TFA for 2 h. The solution was concentrated, redissolved
in toluene and evaporated. The residue was dissolved in ethyl
acetate and sat. aq. NaHCO.sub.3, the layers were separated and the
inorganic one was extracted with ethyl acetate, the combined
organic ones were washed wit brine, dried over Na.sub.2SO.sub.4 and
concentrated yielding 1.67 g (quant)
(2S,4R)-4-(4-Methoxy-benzylsulfanyl)-1-(naphthalene-2-sulfonyl)-p-
yrrolidine-2-carboxylic acid N-methyl-hydrazide as white solid, MS:
486 (MH.sup.+).
[0251] (Step 23) To 1.67 g (3.4 mmol)
(2S,4R)-4-(4-Methoxy-benzylsulfanyl)-
-1-(naphthalene-2-sulfonyl)-pyrrolidine-2-carboxylic acid
N-methyl-hydrazide were added 0.78 g (4.08 mmol, 1.2 eq)
toluenesulfonyl chloride, 0.7 ml (4.08 mmol, 1.2 eq) N-ethyl
diisopropylamine and 106 mg (0.17 mmol, 0.05 eq) DMAP-resin,
followed by additional 324 mg (1.7 mmol, 0.5 eq) toluenesulfonyl
chloride after 3 d. The reaction was filtered, 1M HCl was added and
the inorganic phase was extracted with CH.sub.2Cl.sub.2. The
combined organic phases were washed with brine, dried over
Na.sub.2SO.sub.4 and were concentrated. The crude product was
purified by flash chromatography using ethyl acetate:hexane 1:2
yielding 350 mg (13%)
(2S,4R)-4-(4-Methoxy-benzylsulfanyl)-1-(naphthalene-2-sulfon-
yl)-pyrrolidine-2-carboxylic acid
N-methyl-N'N'-bis-(4-methyl-benzenesulfo- nyl)-hydrazide as white
solid, MS: 794 (MH.sup.+) and 630 mg (29%)
(2S,4R)-4-(4-Methoxy-benzylsulfanyl)-1-(naphthalene-2-sulfonyl)-pyrrolidi-
ne-2-carboxylic acid
N-methyl-N'-(4-methyl-benzenesulfonyl)-hydrazide as white solid,
MS: 640 (MH.sup.+).
[0252] These compounds were deprotected according to Scheme 2 (step
4) giving:
[0253]
(2S,4R)-4-Mercapto-1-(naphthalene-2-sulfonyl)-pyrrolidine-2-carboxy-
lic acid N-methyl-N'N'-bis-(4-methyl-benzenesulfonyl)-hydrazide as
white crystalline, mp 110.degree. C., MS: 674(MH.sup.+).
[0254]
(2S,4R)-4-Mercapto-1-(naphthalene-2-sulfonyl)-pyrrolidine-2-carboxy-
lic acid N-methyl-N'-(4-methyl-benzenesulfonyl)-hydrazide as white
crystalline, mp 103.5.degree. C., MS: 520 (MH.sup.+).
[0255] (step 25,26) According to procedure (Scheme 4, step 16) was
prepared from
(2S,4R)-4-(4-Methoxy-benzylsulfanyl)-1-(naphthalene-2-sulfo-
nyl)-pyrrolidine-2-carboxylic acid
N-methyl-N'-(4-methyl-benzenesulfonyl)-- hydrazide and benzyl
bromide (2S,4R)-4-(4-Methoxy-benzylsulfanyl)-1-(napht-
halene-2-sulfonyl)-pyrrolidine-2-carboxylic acid
N-methyl-N'-benzyl- N'-(4-methyl-benzenesulfonyl)-hydrazide which
was directly deprotected according to Scheme 2 (step 4) to give
(2S,4R)-4-Mercapto-1-(naphthalene--
2-sulfonyl)-pyrrolidine-2-carboxylic acid N-methyl-N'-benzyl-
N'-(4-methyl-benzenesulfonyl)-hydrazide as white solid, mp
82.5.degree. C., MS: 610 (MH.sup.+). 16
Example 3 h
Synthesis of Cyclic Compounds (Schema 6)
[0256] (step 27) To 150 mg (0.25 mmol)
(2S,4R)-1-(Naphthalene-2-sulfonyl)--
4-tritylsulfanyl-pyrrolidine-2-carboxylic acid hydrazide in 65 ml
THF were added 52 .mu.l (0.3 mmol, 1.2 eq) iPr.sub.2EtN and 29
.mu.l (0.25 mmol) 4-bromobutyryl chloride at 0.degree. C. The
solution was stirred at RT for 2 h, the solution was concentrated
and redissolved in ethyl acetate/H.sub.2O. The inorganic phase was
extracted with ethyl acetate and, the organic phase was washed with
brine and dried over Na.sub.2SO.sub.4. Column chromatography with
ethyl acetate:hexane 1:1 yields 170 mg (92%)
(2S,4R)-1-(Naphthalene-2-sulfonyl)-4-tritylsulfanyl-p-
yrrolidine-2-carboxylic acid N'-(4-bromo-butyryl)-hydrazide which
was dissolved in 120 ml DMF and treated with 17 mg (0.38 mmol, 55%
in mineral oil) NaH and the solution stirred for 2 h, was
concentrated and dissolved in ethyl acetate/H.sub.2O. The inorganic
phase was extracted with ethyl acetate, the organic phase washed
with brine and dried over Na.sub.2SO.sub.4. The crude product was
purified by column chromatography yielding 75 mg (45%)
(2S,4R)-4-tritylsulfanyl-1-(naphthalene-2-sulfonyl)--
pyrrolidine-2-carboxylic acid (2-oxo-pyrrolidin-1-yl)-amide as
white foam and 40 mg (25%) (2S,4R)-1-[4- tritylsulfanyl
-1-(naphthalene-2-sulfonyl)--
pyrrolidine-2-carbonyl]-tetrahydro-pyridazin-3-one as white foam.
The two compounds were treated separately in TFA (2 ml/mmol
tritylsulfanyl) with 10 eq triethylsilane at 0.degree. C. to RT,
until no educt could be detected, to yield
(2S,4R)-4-Mercapto-1-(naphthalene-2-sulfonyl)-pyrrolid-
ine-2-carboxylic acid (2-oxo-pyrrolidin-1-yl)-amide as white foam,
MS: 420 (MH.sup.+) and
(2S,4R)-1-[4-Mercapto-1-(naphthalene-2-sulfonyl)-pyrrolidi-
ne-2-carbonyl]-tetrahydro-pyridazin-3-one as white foam, MS: 420
(MH.sup.+), respectively. 17
Example 4
Solid Phase Synthesis of Hydrazide Derivatives
Building Block synthesis (Scheme 7)
[0257]
(2S,4R)-4-(4-Methoxy-benzylsulfanyl)-pyrrolidine-1,2-dicarboxylic
acid 1-tert-butyl ester (19.48 g, 53 mmol) was treated with TFA (80
ml) in CH.sub.2Cl.sub.2 (120 ml) for 15 min. The reaction mixture
was concentrated under reduced pressure and the resultant dark red
oil was triturated in diethyl ether/n-hexane (1:4 v/v, 860 ml). The
precipitated salt was collected and dried under reduced pressure
(18.9 g) and directly used in the next step.
[0258] The TFA salt (18.9 g, 53 mmol) of (2S,
4R)-4-(4-methoxy-benzylsulfa- nyl)-pyrrolidine-2-carboxylic acid in
1,4-dioxane/H.sub.2O (300 ml) containing NaHCO.sub.3 (17.8 g, 212
mmol) was treated with Fmoc-OSu (19.7 g, 58.3 mmol) and
magnetically stirred for 16 h. The reaction mixture was diluted
with water (400 ml) and washed with diethyl ether (2.times.). Ethyl
acetate (400 ml) and HCl (25%, 50 ml) were added. The organic phase
was extracted and washed H.sub.2O, NaCl sat. and dried MgSO.sub.4.
Filtration and concentration under reduced pressure yielded a foam
(22.5 g). 18
[0259] The above foam (20.7 g, 42.3 mmol) was dissolved in TFA (350
ml) and triisopropylsilane (43.5 ml) was added. The mixture was
refluxed for 0.5 h and concentrated under reduced pressure. Diethyl
ether (100 ml) and n-hexanes (300 ml) were added yielding a
precipitate. The supernatant was decanted and the precipitate was
dried under reduced pressure and high vacuum to yield a white foam
(2S, 4R)-4-sulfanyl-1-(fluorenylmethoxycarbo-
nyl)-pyrrolidine-2-carboxylic acid (9.6 g, MS: 370 MH.sup.+)
Resin Derivatization (Scheme 8)
[0260] The linker 4-(.alpha.,.alpha.-diphenylhydroxymethyl)benzoic
acid (18.3 g, 60 mmol) was activated using TPTU (17.8 g, 60 mmol),
DIEA (30.8 ml, 180 mmol) in DMF(abs., 250 ml) for 3 min. The
mixture was added to a flask containing benzhydrylamine resin
(loading-NH.sub.2 0.9 mmol/g, 44.4 g) and the flask was shaken for
1 h. The resin was collected on a filter and washed
(3.times.alternating DMF/isopropanol), CH.sub.2Cl.sub.2, ether and
dried: 54.65 g, 0.65 mmol/g (loading based on mass increase).
[0261] To the CH.sub.2Cl.sub.2 washed resin above (46.9 g, 30
mmol), was added a mixture of (2S,
4R)-4-sulfanyl-1-(naphthalene-2-sulfonyl)-pyrroli-
dine-2-carboxylic acid (12.2 g, 36 mmol) in CH.sub.2Cl.sub.2 (abs.
550 ml), TFA (80 ml). The red colored mixture was shaken for 1.5 h
and the resin was then filtered, washed (3.times.alternating
CH.sub.2Cl.sub.2/isopropanol), CH.sub.2Cl.sub.2, ether and dried:
42 g, 0.65 mmol/g (loading based on mass increase).
[0262] To the CH.sub.2Cl.sub.2 washed resin above (33.5 g, 22
mmol), was added a mixture of (2S,
4R)-4-sulfanyl-1-(fluorenylmethoxycarbonyl)-pyrro- lidine-2-
carboxylic acid (9.7 g, 26 mmol) in CH.sub.2Cl.sub.2 (abs. 450 ml),
TFA (67 ml). The red colored mixture was shaken for 1.5 h and the
resin was then filtered, washed (3.times.alternating
CH.sub.2Cl.sub.2/isopropanol), CH.sub.2Cl.sub.2, ether and dried:
42 g, 0.59 mmol/g (loading based on mass increase). 19
Example 5
Parallel Chemistry on Solid Phase (Scheme 9)
Typical Procedure: steps 1,2
[0263] Resin derivatized with (2S,
4R)-4-sulfanyl-1-(naphthalene-2-sulfony-
l)-pyrrolidine-2-carboxylic acid (0.4 g, 0.26 mmol) above, was
treated with DMF (abs. 5 ml), TPTU (0.18 g, 0.61 mmol), DIEA (0.21
ml, 1.21 mmol) for 10 min. The DMF solution was removed under
vacuum and the reaction flask was charged with benzyloxycarbonyl
hydrazide (0.13 g, 0.77 mmol) in DMF (abs. 3 ml). The reaction
mixture was shaken for 0.5 h and the resin was collected at the
filter, washed (3.times.alternating DMF/isopropanol),
CH.sub.2Cl.sub.2, ether and dried.
[0264] The resin (440 mg ) was treated with 40%
TFA/CH.sub.2Cl.sub.2 (10 ml), triisopropylsilane (0.5 ml) for 10
min and the filtrate was collected and concentrated under reduced
pressure and the residue was freeze-dried from acetic acid (10 ml)
yielding 41 mg (2S,
4R)-N'-[4-mercapto-1-(naphthalene-2-sulfonyl)-pyrrolidine-2-carbonyl]-hyd-
razinecarboxylic acid benzyl ester as a white lyophilisate, MS:
508.3 (MNa.sup.+).
[0265] Other compounds prepared in parallel, via the above
procedure, were indicated in Table 1. 20
Typical Procedure: steps 3,2
[0266] Resin derivatized with (2S,
4R)-4-sulfanyl-1-(naphthalene-2-sulfony-
l)-pyrrolidine-2-carboxylic acid (0.5 g, 0.33 mmol) above, was
treated with DMF (abs. 5 ml), TPTU (0.19 g, 0.65 mmol), DIEA (0.22
ml, 1.3 mmol) for 10 min. The DMF solution was removed under vacuum
and the reaction flask was charged with 4-methoxybenzenesulfonyl
hydrazide (0.20 g, 1.00 mmol) in DMF (abs. 5 ml). The reaction
mixture was shaken for 1 h and the resin was collected at the
filter, washed (3.times.alternating DMF/isopropanol),
CH.sub.2Cl.sub.2, ether and dried.
[0267] The resin (540 mg ) was treated with 40%
TFA/CH.sub.2Cl.sub.2, triisopropylsilane (0.5 ml) for 15 min and
the filtrate was collected and concentrated under reduced pressure
and the residue was purified by prep.RP-HPLC and the desired
fractions were pooled and freeze-dried from acetic acid (10 ml)
yielding (2S, 4R)-4-mercapto-1-(naphthalene-2-sulfony-
l)-pyrrolidine-2-carboxylic acid
N'-(4-methoxy-benzenesulfonyl)-hydrazide as a white lyophilisate,
MS: 520.1 (MH.sup.-)
[0268] Other compounds prepared in parallel, via the above
procedure, were indicated in Table 1.
Typical Procedure: steps 3,6,2
[0269] Resin derivatized with (2S,
4R)-4-sulfanyl-1-(naphthalene-2-sulfony-
l)-pyrrolidine-2-carboxylic acid (4.3 g, 2.75 mmol) above, was
treated with DMF (abs. 30 ml), TPTU (1.63 g, 5.50 mmol), DIEA (1.41
ml, 8.25 mmol) for 15 min. The DMF solution was removed under
vacuum and the reaction flask was charged with
toluene-4-sulfonhydrazide (1.54 g, 8.25 mmol) in DMF (abs. 30 ml).
The reaction mixture was shaken for 16 h and the resin was
collected at the filter, washed (3.times.alternating
DMF/isopropanol), CH.sub.2Cl.sub.2, ether and dried.
A typical Alkylation step e.g.,
[0270] To the resin (0.8 g, 0.45 mmol) was added DMF (abs. 10 ml),
diazabicyclo[5.4.0]undec-7-ene (0.08 ml, 0.54 mmol),
2,5-difluorobenzylbromide (0.11 g, 0.54 mmol) and the mixture
shaken for 16 h and the resin was collected at the filter, washed
(3.times.alternating DMF/isopropanol), CH.sub.2Cl.sub.2, ether and
dried.
[0271] The resin (0.16 g ) was treated with 40%
TFA/CH.sub.2Cl.sub.2 (10 ml), triisopropylsilane (0.5 ml) for 15
min and the filtrate was collected and concentrated under reduced
pressure and the residue was purified by prep. RP-HPLC and the
desired fractions were pooled and freeze-dried from acetic acid
yielding (2S, 4R)-4-mercapto-1-(naphthalene-
-2-sulfonyl)-pyrrolidine-2-carboxylic acid
N'-(2,5-difluorobenzyl)-N'-(4-m- ethyl-benzenesulfonyl)-hydrazide
as a white lyophilisate, MS: 632.0 (MH.sup.+).
[0272] Other compounds prepared in parallel, via the above
procedure, were indicated in Table 1. Disubstituted products were
also obtained.
Typical Procedure: steps 4,5,2
[0273] Resin derivatized with (2S,
4R)-4-sulfanyl-1-(naphthalene-2-sulfony-
l)-pyrrolidine-2-carboxylic acid (1.0 g, 0.65 mmol) above, was
treated with DMF (abs. 10 ml), TPTU (0.39 g, 1.3 mmol), DIEA (0.45
ml, 2.6 mmol) for 10 min. The DMF solution was removed under vacuum
and the reaction flask was charged with hydrazine hydrate (25%,
0.42 ml, 3.25 mmol) in DMF (abs. 8 ml). The reaction mixture was
shaken for 1 h and the resin was collected at the filter, washed
(3.times.alternating DMF/isopropanol), CH.sub.2Cl.sub.2, ether and
dried.
[0274] To this resin (0.22 g, 0.14 mmol) was added DMF (abs. 3 ml),
DIEA (0.10 ml, 0.60 mmol), 4-fluorobenzenesulfonyl chloride (0.11
g, 0.56 mmol) and the mixture shaken for 3.5 h and the resin was
collected at the filter, washed (3.times.alternating
DMF/isopropanol), CH.sub.2Cl.sub.2, ether and dried.
[0275] The resin (0.25 mg) was treated with 40%
TFA/CH.sub.2Cl.sub.2 (10 ml), triisopropylsilane (0.5 ml) for 15
min and the filtrate was collected and concentrated under reduced
pressure and the residue was purified by prep.RP-HPLC and the
desired fractions were pooled and freeze-dried from acetic acid (10
ml) yielding (2S,
4R)-4-mercapto-1-(naphthalene-2-sulfonyl)-pyrrolidine-2-carboxylic
acid N'-(4-fluoro-benzenesulfonyl)-hydrazide as a white
lyophilisate, MS: 510.2 (MH.sup.-).
[0276] Other compounds prepared in parallel, via the above
procedure, were indicated in Table 1.
Typical Procedure: steps 3,7,2
[0277] Resin derivatized with ((2S,
4R)-4-sulfanyl-1-(fluorenylmethoxycarb-
onyl)-pyrrolidine-2-carboxylic acid (20.1 g, 11.9 mmol) above, was
treated with DMF (abs. 150 ml), TPTU (7.1 g, 23.8 mmol), DIEA (6.1
ml, 35.7 mmol) for 10 min. The DMF solution was removed under
vacuum and the reaction flask was charged with toluene-4-sulfon
hydrazide ( 6.65 g, 35.7 mmol) in DMF (abs. 100 ml). The reaction
mixture was shaken for 16 h and the resin was collected at the
filter, washed (3.times.alternating DMF/isopropanol), DMF.
[0278] Fmoc group removal was achieved with 20% piperidine/DMF
(2.times.5 min).
[0279] Pyrrolidine substitution reactions followed e.g.,
[0280] To this resin (0.60 g, 0.30 mmol) was added DMF (abs. 6 ml),
pyridine (0.12 ml, 1.50 mmol), 8-quinolinesulfonyl chloride (0.08
g, 0.36 mmol) and the mixture shaken for 16 h and the resin was
collected at the filter, washed (3.times.alternating
DMF/isopropanol), CH.sub.2Cl.sub.2, ether and dried.
[0281] The resin (0.25 mg) was treated with 40%
TFA/CH.sub.2Cl.sub.2 (10 ml), triisopropylsilane (0.5 ml) for 15
min and the filtrate was collected and concentrated under reduced
pressure and the residue was purified by prep.RP-HPLC and the
desired fractions were pooled and freeze-dried from acetic acid (10
ml) yielding (2S,
4R)-4-mercapto-1-(naphthalene-2-sulfonyl)-pyrrolidine-2-carboxylic
acid N'-(4-fluoro-benzenesulfonyl)-hydrazide as a white
lyophilisate, MS: 505.3(MH.sup.-)
[0282] Other compounds prepared in parallel, via the above
procedure, were indicated in Table 1.
1TABLE 1 Steps: Name Ion Spray MS Educt Scheme 9
(2S,4R)-4-Mercapto-1-(naphthalene-2- M + H 428.4 phenylhydrazide
3,2 sulfonyl)-pyrrolidine-2-carboxylic acid N'- phenyl-hydrazide
Benzoic acid (2S,4R)-N'-[4-mercapto-1- M + H 456.3 Benzoic acid
hydrazide 1,2 (naphthalene-2-sulfonyl)-pyrrolid- ine-2-
carbonyl]-hydrazide (2S,4R)-N'-[4-Mercapto-1-(naphtha- lene-2- M +
H 410.3 carboxylic acid methylester 1,2
sulfonyl)-pyrrolidine-2-carbonyl]- hydrazide hydrazinecarboxylic
acid methyl ester (2S,4R)-N'-[4-Mercapto-1-(naphthalene-2- M + Na
508.3 benzyloxycarbonyl 1,2 sulfonyl)-pyrrolidine-2-carbonyl]-
hydrazide hydrazinecarboxylic acid benzyl ester Furan-2-carboxylic
acid (2S,4R)-N'-[4- M + H 446.3 2-furoic acid hydrazide 1,2
mercapto-1-(naphthalene-2-sulfonyl)-
pyrrolidine-2-carbonyl]-hydrazide Nicotinic acid
(2S,4R)-N'-[4-mercapto-1- M + H 457.3 Nicotinic acid hydrazide 1,2
(naphthalene-2-sulfonyl)-pyrrolidine-2- carbonyl]-hydrazide
trifluoro-acetate (1:1) (2S,4R)-4-Mercapto-1-(naphthalene-2- M +
NH4 523.2 toluene-4-sulfon hydrazide 3,2 sulfonyl)-pyrrolidine-2-c-
arboxylic acid N'-(4- methyl-benzenesulfonyl)-hydrazide
(2S,4R)-4-Mercapto-1-(naphthalene-2- M + H 352.2 hydrazine hydrate
4,2 sulfonyl)-pyrrolidine-2-carboxylic acid hydrazide
trifluoro-acetate (1:1) (2S,4R)-4-Mercapto-1-(naphthalene-2- M - H
520.1 4-methoxybenzenesulfonyl 3,2 sulfonyl)-pyrrolidine-2-carboxy-
lic acid N'-(4- hydrazide methoxy-benzenesulfonyl)-hydrazide
(2S,4R)-4-Mercapto-1-(naphthalene-2- M + H 470.2 4-methyl-benzoyl-
1,2 sulfonyl)-pyrrolidine-2-carboxylic acid N'-(4- hydrazide
methyl-benzoyl)-hydrazide (2S,4R)-4-Mercapto-1-(naphthalene-2- M +
H 476.1 thiophen-3-yl-acetyl- 1,2 sulfonyl)-pyrrolidine-2-carboxyl-
ic acid N'- hydrazide (thiophen-3-yl-acetyl)-hydrazide
(2S,4R)-4-Mercapto-1-(naphthalene-2- M + H 509.3
1H-indol-3-yl-acetyl- 1,2 sulfonyl)-pyrrolidine-2-carboxylic acid
N'-(1H- hydrazide indol-3-yl-acetyl)-hydrazide
(2S,4R)-4-Mercapto-1-(naphthalene-- 2- M + H 462.2
thiophene-2-carbonyl- 1,2 sulfonyl)-pyrrolidine-2-ca- rboxylic acid
N'- hydrazide (thiophene-2-carbonyl)-hydrazide
(2S,4R)-N'-[4-Mercapto-1-(naphthalene-2- M + H 489.2
3-fluoro-phenylamide 1,2 sulfonyl)-pyrrolidine-2-carbonyl]-
hydrazide hydrazinecarboxylic acid (3-fluoro-phenyl)- amide
(2S,4R)-4-Mercapto-1-(naphthalene-2- M - H 490.2 benzenesulfonyl
hydrazide 3,2 sulfonyl)-pyrrolidine-2-carboxylic acid N'-
benzenesulfonyl-hydrazide (2S,4R)-4-Mercapto-1-(naphthalene-2- M +
H 394.3 acetyl hydrazide 1,2 sulfonyl)-pyrrolidine-2-carboxylic
acid N'- acetyl-hydrazide Isonicotinic acid
(2S,4R)-N'-[4-mercapto-1- M + H 457.3 Isonicotinic acid hydrazide
1,2 (naphthalene-2-sulfonyl)-pyrrolidine-2- carbonyl]-hydrazide
trifluoro-acetate (1:1) (2S,4R)-N'-[4-Mercapto-1-(naphthalene-2- M
+ H 471.2 phenylamide carboxylic 1,2 sulfonyl)-pyrrolidine-2-carbo-
nyl]- acid hydrazinecarboxylic acid phenylamide
(2S,4R)-4-Mercapto-1-(naphthalene-2- M + H 437.3
dimethylaminoacetyl 1,2 sulfonyl)-pyrrolidine-2-carboxylic acid N'-
hydrazide dimethylaminoacetyl-hydrazide trifluoro-acetate (1:1)
(2S,4R)-4-Mercapto-1-(naphthalene-2- M + NH4 677.3 toluene-4-sulfon
hydrazide 3,2 sulfonyl)-pyrrolidine-2-carboxylic acid N',N'-
bis-(4-methyl-benzenesulfonyl)-hydrazide (2S,4R)-4-Mercapto-1-(na-
phthalene-2- M + NH4 537.2 1.toluene-4-sulfon 3,6,2
sulfonyl)-pyrrolidine-2-carboxylic acid N- hydrazide
methyl-N'-(4-methyl-benzenesulfonyl)- 2.iodomethane hydrazide
(2S,4R)-4-Mercapto-1-(naphthalene-2- M + H 534.2 1.toluene-4-sulfon
3,6,2 sulfonyl)-pyrrolidine-2-carboxylic acid N,N'- hydrazide;
2.iodomethane dimethyl-N'-(4-methyl-benzenesulfonyl)- hydrazide
(2S,4R)-4-Mercapto-1-(naphthalene-2- M + NH4 545.1
2,4-difluorosulfonyl 4,5,2 sulfonyl)-pyrrolidine-2-carboxylic acid
N'-(2,4- chloride difluoro-benzenesulfonyl)-hydrazide
(2S,4R)-4-Mercapto-1-(naphthalene-2- M + H 510.2
2-fluoro-benzenesulfonyl 4,5,2 sulfonyl)-pyrrolidine-2-carboxylic
acid N'-(2- chloride fluoro-benzenesulfonyl)-hydrazide
(2S,4R)-4-Mercapto-1-(naphtha- lene-2- M + H 510.2
4-fluoro-benzenesulfonyl 4,5,2 sulfonyl)-pyrrolidine-2-carboxylic
acid N'-(4- chloride fluoro-benzenesulfonyl)-hydrazide
(2S,4R)-4-Mercapto-1-(naphthalen- e-2- M + NH4 685.2
4-fluoro-benzenesulfonyl 4,5,2 sulfonyl)-pyrrolidine-2-carboxylic
acid N,N'-bis- chloride (4-fluoro-benzenesulfonyl)-hydrazide
(2S,4R)-4-Mercapto-1-(naphtha- lene-2- M + NH4 447.2
methanesulfonyl chloride 4,5,2 sulfonyl)-pyrrolidine-2-carboxylic
acid N'- methanesulfonyl-hydrazide
(2S,4R)-4-Mercapto-1-(naphthalene-2- M + H 498.0
2-thiophenesulfonyl 4,5,2 sulfonyl)-pyrrolidine-2-carboxyl- ic acid
N'- chloride thiophene-2-sulfonyl-hydrazide
(2S,4R)-N-[4-[N'-[4-Mercapto-1-(naphthalene- M + NH4 566.1
N-acetylsulfanilyl chloride 4,5,2 2-sulfonyl)-pyrrolidine-2-carbon-
yl]- hydrazinosulfonyl]-phenyl]-acetamide
(2S,4R)-N'-[4-Mercapto-1-(naphthalene-2- M + H 459.3
N,N-dimethylsulfamoyl 4,5,2 sulfonyl)-pyrrolidine-2-carbonyl]-
chloride hydrazinecarboxylic acid dimethylamide
(2S,4R)-4-Mercapto-1-(naphthalene-2- M + NH4 569.7 4- 4,5,2
sulfonyl)-pyrrolidine-2-carboxylic acid N'-(4-
methylsulfonylbenzenesulf
methanesulfonyl-benzenesulfonyl)-hydrazide onyl chloride
(2S,4R)-4-Mercapto-1-(naphthalene-2- M + H 511.2
3,5-dimethylisoxazole-4- 4,5,2 sulfonyl)-pyrrolidine-2-carboxylic
acid N'-(3,5- sulfonyl chloride
dimethyl-isoxazole-4-sulfonyl)-hydrazide
(2S,4R)-4-Mercapto-1-(naphthalene-2- M + H 670.1
3,5-dimethylisoxazole-4- 4,5,2 sulfonyl)-pyrrolidine-2-carboxylic
acid N,N'-bis- sulfonyl chloride
(3,5-dimethyl-isoxazole-4-sulfonyl)-hydrazide
(2S,4R)-4-Mercapto-1-(naphthalene-2- M + NH4 475.2
isopropylsulfonyl chloride 4,5,2 sulfonyl)-pyrrolidine-2-carboxylic
acid N'- isopropanesulfonyl-hydrazide
(2S,4R)-4-Mercapto-1-(naphthalene-2- M + H 510.3
3-fluoro-benzenesulfonyl 4,5,2 sulfonyl)-pyrrolidine-2- -carboxylic
acid N'-(3- chloride fluoro-benzenesulfonyl)-hydrazide
(2S,4R)-4-Mercapto-1-(naphthalene-2- M + NH4 684.9
3-fluoro-benzenesulfonyl 4,5,2 sulfonyl)-pyrrolidine-2-carboxylic
acid N'N'-bis- chloride (3-fluoro-benzenesulfonyl)-hydrazide
(2S,4R)-4-Mercapto-1-(naphthalene-2- M - H 558.1 1.
Toluene-4-sulfon 3,6,2 sulfonyl)-pyrrolidine-2-carboxylic acid N'-
hydrazide cyclopropylmethyl-N'-(4-methyl- 2. Bromomethyl
benzenesulfonyl)-hydrazide cyclopropane (2S,4R)-4-Mercapto-1-(nap-
hthalene-2- M + H 478.2 alpha-bromo-2,5- 4,5,2
sulfonyl)-pyrrolidine-2-carboxylic acid N'-(2,5- difluorotoluene
difluoro-benzyl)-hydrazide (2S,4R)-4-Mercapto-1-(naphthalene-2- M +
NH4 667.1 1. Toluene-4-sulfon 3,6,2 sulfonyl)-pyrrolidine-2-carb-
oxylic acid N'-(4- hydrazide 2. methyl-benzensulfonyl)-N'-(2,4,5-t-
rifluoro- 2,4,5-trifluorobenzyl benzyl)-hydrazide bromide
(2S,4R)-4-Mercapto-1-(naphthalene-2- M + H 632.0 1.
Toluene-4-sulfon 3,6,2 sulfonyl)-pyrrolidine-2-carboxylic acid
N'-(2,5- chloride 2. difluoro-benzyl)-N'-(4-methyl-benzensulfonyl)-
2,5-trifluorobenzyl hydrazide bromide
(2S,4R)-4-Mercapto-1-(naphthalene-2- M + H 548.1 1.
Toluene-4-sulfon 3,6,2 sulfonyl)-pyrrolidine-2-carboxylic acid N'-
hydrazide isopropyl-N'-(4-methyl-benzensulfonyl)- 2. Isopropyl
iodide hydrazide (2S,4R)-[N'-[4-Mercapto-1-(naphthalene-2- M + H
564.1 1. Toluene-4-sulfon 3,6,2
sulfonyl)-pyrrolidine-2-carbonyl]-N-(4-meth- yl- hydrazide
benzenesulfonyl)-hydrazino]-acetic acid 2. Tert.-butyl bromoacetate
(2S,4R)-4-Mercapto-2-[N'-(4-methyl- M - H 448.2 1. Toluene-4-sulfon
3,7,2 benzenesulfonyl)-hydrazinocarbonyl- ]- hydrazide
pyrrolidine-1-carboxylic acid benzyl ester 2. Benzyloxycarbonyl
chloride (2S,4R)-1-(2,6-Difluoro-benzoy- l)-4-mercapto- M - H 454.3
1. Toluene-4-sulfon 3,7,2 pyrrolidine-2-carboxylic acid
N'-(4-methyl- hydrazide benzenesulfonyl)-hydrazide 2.
2,6-Difluoro-benzoyl chloride
(2S,4R)-4-Mercapto-1-(quinoline-8-sulfonyl)- M - H 505.3 1.
Toluene-4-sulfon 3,7,2 pyrrolidine-2-carboxylic acid N'-(4-methyl-
hydrazide benzenesulfonyl)-hydrazide 2. 8-quinolinesulfonyl
chloride (2S,4R)-4-Mercapto-2-[N'-(4-methyl- M - H 414.3 1.
Toluene-4-sulfon 3,7,2 benzenesulfonoyl)-hydrazinocarbonyl]-
hydrazide pyrrolidine-1-carboxylic acid butyl ester 2. Butyl
chloroformate (2S,4R)-4-Mercapto-2-[N'-(4-methyl- M - H 393.0 1.
Toluene-4-sulfon 3,7,2 benzenesulfonyl)-hydrazinocarbonyl]-
hydrazide pyrrolidine-1-sulfonic acid amide 2. Chloro-sulfonic acid
amide (2S,4R)-1-(Biphenyl-4-sulfonyl)-4-mercapto- M + NH4 549.4 1.
Toluene-4-sulfon 3,7,2 pyrrolidine-2-carboxylic acid N'-(4-methyl-
hydrazide benzenesulfonyl)-hydrazide 2. Biphenyl-4-sulfonyl
chloride (2S,4R)-1-(3-Fluoro-benzenes- ulfonyl)-4- M + NH4 491.4 1.
Toluene-4-sulfon 3,7,2 mercapto-pyrrolidine-2-carboxylic acid
N'-(4- hydrazide methyl-benzenesulfonyl)-hydrazide 2. 3-Fluoro-
benzenesulfonyl chloride (2S,4R)-1-(2-Fluoro-benzenesulfonyl)-4- M
+ NH4 491.4 1. Toluene-4-sulfon 3,7,2
mercapto-pyrrolidine-2-carboxylic acid N'-(4- hydrazide
methyl-benzenesulfonyl)-hydrazide 2. 2-Fluoro- benzenesulfonyl
chloride (2S,4R)-4-Mercapto-1-(2,4,5-trif- luoro- M + NH4 527.3 1.
Toluene-4-sulfon 3,7,2 benzenesulfonyl)-pyrrolidine-2-carboxylic
acid hydrazide N'-(4-methyl-benzenesulfonyl)-hydrazide 2.
2,4,5-Fluoro- benzenesulfonyl chloride
(2S,4R)-4-Mercapto-1-(4-phenoxy- M + NH4 565.4 1. Toluene-4-sulfon
3,7,2 benzenesulfonyl)-pyrrolidine-2-car- boxylic acid hydrazide
N'-(4-methyl-benzenesulfonyl)-hydrazide 2. 4-phenoxy-
benzenesulfonyl chloride
(2S,4R)-1-(Biphenyl-4-sulfonyl)-4-mercapto- M + H 546.4 1.
Toluene-4-sulfon 3,7,6,2 pyrrolidine-2-carboxylic acid
N'-methyl-N'-(4- hydrazide methyl-benzenesulfonyl)-hydrazide 2.
Biphenyl-4-sulfonyl chloride 3. iodomethane
(2S,4R)-1-(3-Fluoro-benzenesulfonyl)-4- M + H 488.4 1.
Toluene-4-sulfon 3,7,6,2 mercapto-pyrrolidine-2-carboxylic acid N'-
hydrazide methyl-N'-(4-methyl-benzenesulfonyl)- 2. 3-Fluoro-
hydrazide benzenesulfonyl chloride 3. iodomethane
(2S,4R)-1-(2-Fluoro-benzenesulfonyl)-4- M + H 488 1.
Toluene-4-sulfon 3,7,6,2 mercapto-pyrrolidine-2-carboxylic acid N'-
hydrazide methyl-N-(4-methyl-benzenesulfonyl)- 2. 2-Fluoro-
hydrazide benzenesulfonyl chloride 3. iodomethane
(2S,4R)-4-Mercapto-1-(2,4,5-trifluoro- M + NH4 541.3 1.
Toluene-4-sulfon 3,7,6,2 benzenesulfonyl)-pyrrolidine-2-carboxylic
acid hydrazide N'-methyl-N'-(4-methyl-benzenesulfonyl)- 2.
2-Fluoro- hydrazide benzenesulfonyl chloride 3. iodomethane
(2S,4R)-4-Mercapto-1-(4-phenoxy- M + H 562 1. Toluene-4-sulfon
3,7,6,2 benzenesulfonyl)-pyrrolidine-2-carboxylic acid hydrazide
N'-methyl-N'-(4-methyl-benzenesulfonyl)- 2. 4-phenoxy- hydrazide
benzenesulfonyl chloride 3. iodomethane
(2S,4R)-4-Mercapto-1-(4-propyl- M - H 496.1 1. Toluene-4-sulfon
3,7,2 benzenesulfonyl)-pyrrolidine-2-carboxylic acid hydrazide
N'-(4-methyl-benzenesulfonyl)-hydrazide 2. 4-propyl-
benzenesulfonyl chloride (2S,4R)-1-(3,5-Dimethyl-isoxazole-4-sulfo-
nyl)- M - H 473.0 1. Toluene-4-sulfon 3,7,2
4-mercapto-pyrrolidine-- 2-carboxylic acid N'-(4- hydrazide
methyl-benzenesulfonyl)-hydrazi- de 2. 3,5-Dimethyl-isoxazole-
4-sulfonyl chloride (2S,4R)-4-Mercapto-1-(4-methanesulfonyl- M - H
532.0 1. Toluene-4-sulfon 3,7,2
benzenesulfonyl)-pyrrolidine-2-carboxylic acid hydrazide
N'-(4-methyl-benzenesulfonyl)-hydrazide 2. 4-methanesulfonyl-
benzenesulfonyl chloride (2S,4R)-4-Mercapto-1-thiophene-2-sulfon-
yl- M - H 460.2 1. Toluene-4-sulfon 3,7,2 pyrrolidine-2-carboxylic
acid N'-(4-methyl- hydrazide benzenesulfonyl)-hydrazide 2.
thiophene-2-sulfonyl chloride (2S,4R)-4-Mercapto-2-[N'-(4- -methyl-
M - H 421.3 1. Toluene-4-sulfon 3,7,2
benzenesulfonyl)-hydrazinocarbonyl]- hydrazide
pyrrolidine-1-sulfonic acid dimethylamide 2. Chloro sulfonic acid
dimethylamide (2S,4R)-1-(4-Fluoro-benzenesulfonyl)-4- M - H 472.1
1. Toluene-4-sulfon 3,7,2 mercapto-pyrrolidine-2-carboxylic acid
N'-(4- hydrazide methyl-benzenesulfonyl)-hydrazide 2. 4-Fluoro-
benzenesulfonyl chloride (2S,4R)-1-Isopropanesulfonyl-4-mercapto- M
- H 420.3 1. Toluene-4-sulfon 3,7,2 pyrrolidine-2-carboxylic acid
N'-(4-methyl- hydrazide benzenesulfonyl)-hydrazide 2.
Isopropanesulfonyl chloride
(2S,4R)-1-(2,4-Difluoro-benzenesulfonyl)-4- M - H 490.2 1.
Toluene-4-sulfon 3,7,2 mercapto-pyrrolidine-2-carboxylic acid
N'-(4- hydrazide methyl-benzenesulfonyl)-hydrazide 2. 2,4-Difluoro-
benzenesulfonyl chloride
(2S,4R)-4-Mercapto-1-(2,3,4,5,6-pentafluoro- M - H 544.0 1.
Toluene-4-sulfon 3,7,2 benzenesulfonyl)-pyrrolidine-2-carboxylic
acid hydrazide N'-(4-methyl-benzenesulfonyl)-hydrazide 2.
2,3,4,5,6-pentafluoro- benzenesulfonyl chloride
(2S,4R)-4-Mercapto-1-methanesulfonyl- M - H 392.1 1.
Toluene-4-sulfon 3,7,2 pyrrolidine-2-carboxylic acid N'-(4-methyl-
hydrazide benzenesulfonyl)-hydrazide 2. methanesulfonyl chloride
(2S,4R)-1-(2,5-Difluoro-benzenesulfonyl)-4- M - H 490.2 1.
Toluene-4-sulfon 3,7,2 mercapto-pyrrolidine-2-carboxylic acid
N'-(4- hydrazide methyl-benzenesulfonyl)-hydrazide 2. 2,5-Difluoro-
benzenesulfonyl chloride 2S,4R)-4-Mercapto-1-(4-methyl- M - H 468.1
1. Toluene-4-sulfon 3,7,2 benzenesulfonyl)-pyrrolidine-2-carboxylic
acid hydrazide N'-(4-methyl-benzenesulfonyl)-hydrazide 2. 4-methyl-
benzenesulfonyl chloride (2S,4R)-N-[4-[4-Mercapto-2-]N'-(4-methyl-
M - H 511.2 1. Toluene-4-sulfon 3,7,2 benzenesulfonyl)-hydrazinoca-
rbonyl]- hydrazide pyrrolidine-1-sulfonyl]-phenyl)-acetamide 2.
4-acetamide phenylsulfonyl chloride
(2S,4R)-1-(4-Butoxy-benzenesulfonyl)-4- M - H 526.0 1.
Toluene-4-sulfon 3,7,2 mercapto-pyrrolidine-2-carboxylic acid
N'-(4- hydrazide methyl-benzenesulfonyl)-hydrazide 2. 4-Butoxy-
benzenesulfonyl chloride (2S,4R)-4-Mercapto-1-(2-naphthalen-1-yl- M
- H 526.0 1. Toluene-4-sulfon 3,7,2 ethylsulfonyl)-pyrrolidine-2-
-carboxylic acid N'- hydrazide (4-methyl-benzenesulfonyl)-hydrazid-
e 2. 2-naphthalen-1-yl- ethylsulfonyl chloride
(2S,4R)-1-(4-Chloro-benzenesulfonyl)-4- M + NH4 507.3 1.
Toluene-4-sulfon 3,7,2 mercapto-pyrrolidine-2-carboxylic acid
N'-(4- hydrazide methyl-benzenesulfonyl)-hydrazide 2. 4-Chloro-
benzenesulfonyl chloride (2S,4R)-4-Mercapto-1-(4-trifluoromethyl- M
+ NH4 541.4 1. Toluene-4-sulfon 3,7,2 benzenesulfonyl)-pyrrolidi-
ne-2-carboxylic acid hydrazide N'-(4-methyl-benzenesulfonyl)-hydra-
zide 2. 4-trifluoromethyl- benzenesulfonyl chloride
(2S,4R)-1-(2-Methyl-benzenesulfonyl)-4- M + NH4 487.4 1.
Toluene-4-sulfon 3,7,2 mercapto-pyrrolidine-2-carboxylic acid
N'-(4- hydrazide methyl-benzenesulfonyl)-hydrazide 2. 2-Methyl-
benzenesulfonyl chloride (2S,4R)-1-(Naphthalene-1-sulfonyl)-4- M +
H 506.3 1. Toluene-4-sulfon 3,7,2 mercapto-pyrrolidine-2-carboxyli-
c acid N'-(4- hydrazide methyl-benzenesulfonyl)-hydrazide 2.
Naphthalene-1-sulfonyl chloride (2S,4R)-4-Mercapto-1-phen-
ylmethanesulfonyl- M + NH4 487.4 1. Toluene-4-sulfon 3,7,2
pyrrolidine-2-carboxylic acid N'-(4-methyl- hydrazide
benzenesulfonyl)-hydrazide 2. Phenylmethanesulfonyl chloride
(2S,4R)-1-(2,6-Dichloro-benzenesulfonyl)-4- M + NH4 541.3 1.
Toluene-4-sulfon 3,7,2 mercapto-pyrrolidine-2-carboxylic acid
N'-(4- hydrazide methyl-benzenesulfonyl)-hydrazide 2. 2,6-Dichloro-
benzenesulfonyl chloride (2S,4R)-4-Mercapto-1-(2-trifluoromethyl- M
+ NH4 541.4 1. Toluene-4-sulfon 3,7,2
benzenesulfonyl)-pyrrolidine-2-carboxylic acid hydrazide
N'-(4-methyl-benzenesulfonyl)-hydrazide 2. 2-trifluoromethyl-
benzenesulfonyl chloride
(2S,4R)-1-(3-Methyl-benzenesulfonyl)-4- M + NH4 487.4 1.
Toluene-4-sulfon 3,7,2 mercapto-pyrrolidine-2-carboxylic acid
N'-(4- hydrazide methyl-benzenesulfonyl)-hydrazide 2. 3-Methyl-
benzenesulfonyl chloride (2S,4R)-1-(3-Chloro-benzenesulfonyl)-4- M
+ NH4 507.3 1. Toluene-4-sulfon 3,7,2 mercapto-pyrrolidine-2-carbo-
xylic acid N'-(4- hydrazide methyl-benzenesulfonyl)-hydrazide 2.
3-Chloro- benzenesulfonyl chloride
(2S,4R)-1-(2-Chloro-benzenesulfonyl)-4- M + NH4 507.3 1.
Toluene-4-sulfon 3,7,2 mercapto-pyrrolidine-2-carboxylic acid
N'-(4- hydrazide methyl-benzenesulfonyl)-hydrazide 2. 2-Chloro-
benzenesulfonyl chloride (2S,4R)-1-(3,4-Dichloro-benzenesulfonyl)--
4- M + NH4 541.3 1. Toluene-4-sulfon 3,7,2
mercapto-pyrrolidine-2-c- arboxylic acid N'-(4- hydrazide
methyl-benzenesulfonyl)-hydrazide 2. 3,4-Dichloro- benzenesulfonyl
chloride (2S,4R)-4-Mercapto-1-(4-methoxy- M + NH4 503.4 1.
Toluene-4-sulfon 3,7,2 benzenesulfonyl)-pyrrolidine-2-carboxylic
acid hydrazide N'-(4-methyl-benzenesulfonyl)-hydrazide 2.
4-methoxy- benzenesulfonyl chloride
(2S,4R)-4-Mercapto-1-(3-trifluoromethyl- M + NH4 541.3 1.
Toluene-4-sulfon 3,7,2 benzenesulfonyl)-pyrrolidi- ne-2-carboxylic
acid hydrazide N'-(4-methyl-benzenesulfonyl)-hydra- zide 2.
3-trifluoromethyl- benzenesulfonyl chloride
(2S,4R)-1-Benzenesulfonyl-4-mercapto- M + NH4 473.4 1.
Toluene-4-sulfon 3,7,2 pyrrolidine-2-carboxylic acid N'-(4-methyl-
hydrazide benzenesulfonyl)-hydrazide 2. benzenesulfonyl chloride
(2S,4R)-1-(3,4-Difluoro-benzenesulfonyl)-4- M + NH4 509.3 1.
Toluene-4-sulfon 3,7,2 mercapto-pyrrolidine-2-carboxylic acid
N'-(4- hydrazide methyl-benzenesulfonyl)-hydrazide 2. 3,4-Difluoro-
benzenesulfonyl chloride
(2S,4R)-1-(2,6-Difluoro-benzenesulfonyl)-4- M + NH4 509.3 1.
Toluene-4-sulfon 3,7,2 mercapto-pyrrolidine-2-carboxylic acid
N'-(4- hydrazide methyl-benzenesulfonyl)-hydrazide 2. 2,6-Difluoro-
benzenesulfonyl chloride (2S,4R)-4-Mercapto-1-(2,3,4-trifluoro- M +
NH4 527.3 1. Toluene-4-sulfon 3,7,2
benzenesulfonyl)-pyrrolidine-2-carboxylic acid hydrazide
N'-(4-methyl-benzenesulfonyl)-hydrazide 2. 2,3,4-trifluoro-
benzenesulfonyl chloride
Example A
[0283] Tablets containing the following ingredients can be
manufactured in a conventional manner:
2 Ingredients Per tablet Compound of formula I 10.0-100.0 mg
Lactose 125.0 mg Maize starch 75.0 mg Talc 4.0 mg Magnesium
stearate 1.0 mg
Example B
[0284] Capsules containing the following ingredients can be
manufactured in a conventional manner:
3 Ingredients Per capsule Compound of formula I 25.0 mg Lactose
150.0 mg Maize starch 20.0 mg Talc 5.0 mg
Example C
[0285] Injection solutions can have the following composition:
4 Compound of formula I 3.0 mg Gelatine 150.0 mg Phenol 4.7 mg
Water for injection solutions ad 1.0 ml
Example D
[0286] 500 mg of compound of formula I are suspended in 3.5 ml of
Myglyol 812 and 0.08 g of benzyl alcohol. This suspension is filled
into a container having a dosage valve. 5.0 g of Freon 12 under
pressure are filled into the container through the valve. The Freon
is dissolved in the Myglyol-benzyl alcohol mixture by shaking. This
spray container contains about 100 single dosages which can be
applied individually.
* * * * *