CA2085834A1 - (1s) -hydroxyalkyl-oxapenem-3-carboxylic acids and their use as betalactamase inhibitors - Google Patents

Abstract

ABSTRACT

The invention relates to 1-oxapenem-3-carboxylic acids of the following structure:
in which R1, R3, R4 and R5 independently of one another denote pharmaceutically acceptable groups which are connected to the other part of the molecule via carbon-carbon single bonds and which contain 1 to 10 carbon atoms, and in which R2 is hydrogen, alkyl or acyl having up to 10 carbon atoms or an inorganic acid residue.
These compounds, as well as their pharmaceutically acceptable salts, esters and amide derivatives are especially useful betalactamase inhibitors. Subject of the invention are also pharmaceutical preparations containing these compounds and procedures for medication, as well as procedures for the preparation of such compounds.

Description

33~

1 Description. ((lS)-Hydroxyalkyl)oxapenem-3-carboxylic __ __ acids and their use as betalactamase lnhibitors The invention relates to l-oxapenem-3-carboxylic acids of the following structure:

.. H 3 ~ ~ -C - R
O ~ 15 COOH
in which Rl, E~3, R4 and R5 independen-tly of one another denote pharmaceutically acceptable groups connected via carbon-carbon atoms to the other part of the molecule and containing one to 10 carbon atoms and wherein R2 denote hydrogen, alkyl or acyl with up to 10 carbon atoms or an inorganic acid residue. These compounds, as well as their pharmaceutically acceptable salts, ester and amide derivatives are especially useful be-ta-lactamase inhibitorsO A matter of the invention are also pharmaceutical compositions containing these com-pounds and procedures for medication as well as proce-dures for the preparation of such compounds. -The invention relates to 2-unsubstituted oxapenem-3-car-boxylic acids substituted in their 2-position by special groups which have been described already in EP 89117433.6 and which provide the stability to the oxapenem nucleus required for the use as betalactamase inhibitors. ~n their 6-position these compounds are monosubstituted and contain a (lS)-hydroxyalkyl substituent optionally further substituted itself at oxygen or at a carbon atom. Thesè optically active compounds are useful betalactamase inhibitors and they can be described .. . . .

1 by the yeneral formu].a H O ~ R2 ~ .
S L~C ~ ~ R3 O ~ R5 C~OH

in which Rl, R3, R4 and.R5 independently of one another are selected from pharmaceutically acceptable groups connected via carbon-carbon atoms to the other part of the molecule which contain: substituted or unsubstituted alkyl, alkenyl, alkinyl, cycloalkyl, alkylcycloalkyl, alkylcycloalkenyl, cycloalkylalkyl, alkenylcycloakyl, cycloalkenylalkyl, aryl, aralkyl, aralkenyl, aralkinyl, carboxy or cyano, wherein the aforementioned alkyl, alkenyl or alkinyl molecules contain 1 to 6 carbon atoms, the cycloalkyl or the cycloalkenyl parts of the molecule contain 3 to 6 carbon atoms and the aryl part of the molecule contains 6 to 10 carbon atoms, aromatic or aliphatic heterocyclyl, heterocyclylalkyl, heterocyclylalkenyl, heterocyclylalkinyl, alkylheterocyclyl, wherein the aforementioned alkyl, alkenyl or the alkinyl parts of the molecule contain 1 to 6 carbon atoms and the heterocyclic molecular fragment is mono or bicyclic and contains 3 to 10 ring atoms of which one or more are selected from the group comprising: oxygen, sulphur and nitrogen, and where the substituents of the above-mentioned groups may be: protected or unprotected hy-droxyl, hydroxyalkyl, aminoalkyloxy, alkyloxy, acyloxy, aryloxy, heterocyclyloxy, carbamoyl, carbamoyloxy, thiocarbamoyl, thiocarbamoyloxy, alkylcarbamoyloxy, alkylthiocarbamoyloxy, mercapto, alkylthio, hydroxy-alkylthio, aminoalkylthio, amidinoalkylthio, acylthio, ~ . ~

. . .

- ~ - 2~ 3~

1 arylthio, alkylheteroarylthio, hydroxya].kylheteroaryl~
thio, heterocyclylthi.o, carbamoylthio, alkylcarbamoyl-thio, thiocarbamoyl.thio, alkylthiocarbamoylthio, protected or unprotected amino or monoalkylamino, dialkylamino, oxo, protected or unprotected oxi.mino or alkylimino, tetraalkylammonium, cycloalkylami.no, arylamino, hetero-arylamino, heterocyclamino, acylamino, ami.dino, alkyl-amidino, guanidino, alkylguanidino, carbamoylamino, alkylcarbamoylamino, thiocarbamoylamino, alkyl-thiocar-bamoylamino, nitro, chlorine, bromine, fluorine, iodine,azido, cyano, alkylsulphinyl, alkylsulphonyl, sulphon-amido, sulphamoyloxy, alkylsulphonyloxy, or pro-tected or unprotected sulpho, su].phoxy or carboxyl, where the substituents independently of one another occur one or more times and the alkyl part of the molecule thereof contains 1 to 6 carbon atoms, the aryl part of the molecule thereof contains 6 to 10 carbon atoms, and where the heteroaromatic or heterocyclic part of the molecule is mono- or bicyclic and contains 3 to 10 ring atoms, of which one or more are selected from the group comprising: oxygen, sulphur and nitrogen, and characterized in that R2 denotes hydrogen, alkyl or acyl containing up to 10 carbon atoms, sulpho, amino-sulphonyl, or phosphoroyl.

The protective groups of the abovementioned protected substituents are easily removable radicals which are known per se, such as are customarily used for this purpose in organic synthesis. Such protective groups are found, for example, in T.W. Greene, "Protective Groups in Organic Synthesis", Wiley, New York, 1981.

Furthermore, two of the groups ~3, R4 or R5 can be made together into a bridge via carbon, oxygen, nitrogen and sulphur containing parts of the molecule; they , ' .

_ 5 _ ~ ;3~

1 are then constituents of a carbocyclic or heterocyclic ring which can be three-, four-, five- or six-membered.

Furthermore, the two groups Rl and R2 can be made together into a bridge via carbon, oxygen, nitrogen and sulphur-con-taining parts of the ~nolecule; they are then constituents of a three-~ four-, five-, or six-membered carbo- or heterocyclic ring.

Examples of bridging parts of the molecule for R1 and R2 or for R3 and R4 are methylene, dimethylene, -trimethylene, tetramethylene, oxamethylene, oxadimethylene, dioxamethylene, azadimethylene, diazamethylene or the like.

Pharmaceutically acceptable groups R1, R2, R3, R4 and R5 which are bonded via a C-C single bond are groups which are, for example, customary in the B-lactam anti-biotics. Such groups are found, for example, in M.L.
Sassiver, A. Lewis in "Advances in Applied Microbiology", ed. D. Perlman, Academic Press, N.Y. (1970).

The invention furthermore relates to the pharmaceutically acceptable salts, esters and amide derivatives of the compounds according to the invention.

Pharmaceutically acceptable ester and amide derivatives are known for a long time and serve the better oral availability of many known antibiotics. Many such derivatives, e. g. pivaloyloxymethyl esters are so called prodrugs and they are described in W. DUrckheimer et al. in Adv. Drug Res. 17, 197-203 (1988~.

The invention furthermore relates to processes for the preparation of these compounds, pharmaceutical preparations containing these compounds and methods of treatment in which these compunds and preparations are administered, if an antibiotic effect is indicated.

2~58~

1 There is a continued demand of novel antibiotics. Un-fortunately -there is no static ef~icacy of any given antibiotic, because, upon con-tinuous application in large scale, resistant strains of pathogenic bacteria are created. Thus, -the search for new antibiotics ls continued.

The betalactam ~ntibiotics -take UF) the main share, i. e. two thirds of the antibiotic sales. They are appreciated because they are well tolerated.

However, in recent years, in many hospitals increased and severe infectious diseases have occurred which were caused by germs heavily producing betalactamase and attacking even third generation-cephalosporins which were relatively insensitive to betalactamases.
Such infections diseases cannot be fought back alone with these antibiotics alone.

A solution to this problem is given by betalactamase inhibitors. They are administered in combination with a betalactam antibiotic and they bind the bacterial enzymes immediately and irreversibly. The antibiotic, being protected in this manner, can develop its full efficacy.
A recent review about resistance enzymes and the use of betalactamase inhibitors is given in (H.C. Neu, The Amer. J. of Med. 79 (suppl. SB), 1 (1985).

Stable oxapenem-carboxylic acids and their use as be-talactamase inhibitors have been described in EP 8911-7433.6. Compounds having 6-hydroxyalkyl groups proved to be good inhibitors.

~ , . , ,' :, ; .

_ 7 _ 2~ 33~

1 2-tert~butyl-6-hydroxymethyloxapenem-3-carboxylic acid (K salt) Ia proved to be more active than clavulanic acld in its combination with me~loclllin against penicillin resistant Staph. aureus and Pseudomonas aeruginosa.
The hydroxymethyl group attached to the 6-position does not contain an additional asymmetric center. Only higher homologous side chains such as for example hydroxy-ethyl contain such a center. The investigation of various stereoisomeric 2-ter-t-butyl-6-hydroxyethyloxa-penem-3-carboxylic acids (K salts) Ib - Ie showed a surprising dependence of the betalactamase inhibiton on the configuration of the asymmetic hydroxyethyl side chain.

R ~ O~ C~3 ~ N ~ Cl -CH3 O I C~I3 COOH
I inhibition diameter mm (E.coli TEM 1) R6 Konfiguration I I+Cefaclor (lOmcg) (10+30mcg) Ia rac. hydroxymethyl trans 17 22 Ib rac. hydroxyethyl (unlike) trans 0 28 Ic rac. hydroxyethyl (unlike cis 9 26 (contains approx. 20 ~ Ib) Id ((lR)-Hydroxyethyl (like3 trans (5R, 6R~ 23 24 Ie ~(lS)-hydroxyethyl (unlike) trans (5R, 6R) 0 29 "like" means same absolute configuration of two adjacent chiral centers. As a test strain an a particularly strong betalactamase producer was used. Therefore, E. coli W 31103 R6K (TEM 1) was repeatedly incubated ..

, 3~

1 ln a nutrition medium with 20 mcg/ml of cefaclor. Cefaclor alone gave an inhibi-tion zone of 16 mm, in its combination with 10 mcg of clavulanic acid a zone of 22 mm diameter.

With an occurrence of 75 ~ TEM 1 enzymes are the most important betalactamases. They occur in a great number of enterobacteriaceae. It was found that the oxapenemcar-boxylic acids Ib and Ie showed a particularly yood activity as betalactamase inhibl-tors against E. coli TEM 1 and that they are able to protect the antibiotic ceEaclor effec-tively against the bacterial enzyme.
It is true that Ia and Id show a good activity as antibiotics.
The increase of the inhibition zone, i. e. as the protect-ing effect is rela-tively low in these cases.

Clavulanic acid is known to be a potent inhibitor of peniclllinase. However, it has been reported (Amer.
J. Med. 79 (suppl. 5B), 194 (1985) that it was not active against cephalosporinases from Enterobacter cloacae or Pseudomonas aeruginosa. Compound Ie (5R, 6R)-2-tert-butyl-((lS~-hydroxyethyloxapenem-3-carboxylic acid (K salt), on the other hand, is also effective against these germs as a betalactamase inhibitor. The only decisive matter is that the relevant antibiotic, combined with Ie, can penetrate into the xelevant bac-terium.

The compounds according to the invention are in a position to enhance the inhibition activity of all investigated penicillins, e. g. amoxycillin, piperacillin, mezlocillin or cephalosporins, e. g. cefaclor, ce operazon against betalactamase producing bacteria. Also third-generation-cephalosporins, e. g. cefotaxime or ceftriaxone (30 mcg each), generally classified as betalactamase stable, produced, for the first time, distinct inhibition zone diameters of 27 and 26 mm respectively against E. cloacae .
.

- . , :, , -~ 9 ~ 3 r~~

1 in combinations with Ie (10 mcg). The compounds according to the invention are strongly active against penicilli-nases and cephalosporinases.

The compounds Ib and Ie, outstanding as betalactamase inhibitors, contain a side chain, the configuration of which is opposite to that of thienamycin. Such oxapenemcarboxylic acids, havinq the epithienamycin side chain, are therefore preferred in their use as betalactamase inhibitors. Compounds having the (5R, 6R)-configuration and the ((lS)-hydroxyalkyl) group as a side chain in 6-position, are strongly prefered.

Hydroxyalkyloxapenemcarboxyllc acids of the (5R)-con-figuration are per se known (EP 89117433.6). They were only isolated and investigated as diastereomers.
A more exact statement about the influence of the con-figuration of the hydroxyalkyl side chain on the betalactam-ase inhibi.tory activity was not possible on this basis.
It is true, that isomerically pure (5R,6R)-oxapenemcar-boxylic acids have recently been described (M. Murakami, T. Aoki, M. Matsuura, W. Nagata, J. ~ntibiot. l9qO, 1441-9 (engl.). However, they contained exclusively hydroxyethyl side chains of the (lR)-configuration, disadvantageous for betalactamase inhibitory activity.
The compounds Ib and Ie, outstanding as betalactamase inhibitors, are not known according to the state of technology. Their betalactamase inhibitory activity against penicillinases and cephalosporinases of Gram-positive and Gram-negative bacteria is significant and surprising. They can be used as enantiomerically pure (6R,l'S)-compounds or in their racemic forms as betalactamase inhibitors.

,. ;, : , . ..

.; ' . ; - .

: ~ .

- 1 o - Z(~ 8~

l The ((lS)-hydroxyalkyl)oxapenem-3-carboxylic acids or their salts, respec-tlvely, can be prefexably administered with orally applicable antibiotics, e. g. ampicillin, amoxycillin, cefaclor, cefprozil, cefuroxime-axetil, cefixime, cefpodoxime-poxetil, cefteram-pivoxil, cefetamet-pivoxil, ceftibuten, flomoxef ancl the like. sut also combinations with perenterally aclministerable antibiotics, e. g. penicillin G, piperacillin, azlocillin, mezlocillin, ticarcllli.n, mecillinam, cefoperazone, cefalothin, cefazolin, cephaloridine, cefsudolin, cefo-taxime, cefotiam, ceftriaxone, ceEpirome, cefepime and the like are possible.

These combinations are useful in -the chemotherapy against all bacterial infectious diseases, generally Eought back with betalactam antibiotics. However, they a~e strongly preferred for diseases caused by penicillin resistant and cephalosporin resistant germs. Examples of such bacteria are Staph. aureus, H. influenzae, N. gonorrhoaeae, B. catarrhalis, Bacteroides frayilis, N. meningitidis, Citrobacter freundii, Enterobacter cloacae, Proteus mirabilis, Proteus vulgaris, Providentia rettgeri, Providencia stuarti, Pseudomonas aeruginosa, Salmonella, Serratia marescen~, E. coli and the like.

In each case, the dose of the betalactamase inhibitors according to the invention depends generally on the relevant antibiotic and on the infections disease to be fought back. This dose can, circumstances permit~ing, be equally high or similar to that of the relevant antibiotic. It can also be lower, especially with parenteral application. The preferred ratio is 1 :
10 to 1 : 1, but also other ratios are possible .

The ((lS)-hydroxyalkyl)oxapenem-3-carboxylates according to the invention, are prepared as follows:

- lL - ~ C ~ 518~3~
(Acylation) S+i - CH3 ~ ~ ~ CH
RL ~ , 3 ~ Rl ~ ~ R3 O ~ Base ~ N ~ C ~ R4 1 COOpNBz pNBzOOC R5 (Deprotection) ¦ BF3 oder CHaO OH
S~ CH EtC~N=N~CEt I S -CH
R ~ ~h3P 1~ 3 I I 3 ~ R3 0 ~ ~ 4 HCOOH ~ ~ C - R :
pNBzOOC 5 (Mitsunobu) R pNBzOOC R5 H20, HCl¦ (Hydrolysis) ~ O

OH pNBzO-CI-Cl ~ OpNBz Rl ~ 3 Base Rl ~ ~ 0 3 N ~ C - R4 0 ~ ~ cR ~ R4 (Protection) pNBzOOC R pNBzOOC R5 J~ O ¦ C12 OpNBz 3 O OpNBz ~ ~ R4 - R ~ o R
o R5 (Cyclisation) o ~ C - R4 pNBZooc ¦
8 pNBzOOC R

.

, . .

' !: ' ' ;

2~5~3.~

1 Chromatography q ~ OpNBz Q ~ OpNBz H ~ R3 H H 3 Rl r ~c R Rl ~ R
N ~ R N ~ l5 pNBzOOC pNBzOOC
8a 8b H2, Pd¦ (Kat. hydrogenation) ¦ H2, Pd (rac. series) OH VH

R ~r c R4 R~o CR3 R4 200 ~ R O ~ R
COOH COOH
9a 9~
rac.
Starting materials of type 1 have been described (J.
Antibiot. 1990, 1441 - 9). The reversal of configuration of the hydroxyalkyl side chain of 3 was effected by the Mitsunobu reaction. This procedure has been approved in the field of carbapenems (G.I. Georg, J. Kant, H.S.
Gill, J. Amer. Chem. Soc. 1987, 109, 1129 - 35). The mixture of cis, trans-isomers 8 can be separated by chromatography on silica gel. In certain cases separation is also possible by fractional crystallization.

, ; . -.
. .

, _ L3 _ 2~ 33~

1 An alternatlve procedure uses (3R,4R) 4-ace-toxy-3-((lS-tert-butyldimethylsilyloxy)ethyl)azetidin-2-one as a starting material. Accordi~y to -this procedure the reversal of configuratlon i5 superfluous; the preparation of the starting material 10 i.s Eound in J. Amer. Chem.
Soc. 1987, 109, 1129 - 35. The preparation is effected according to the Eollowlng reaction scheme:

+
S~~~ CH
3 ~, C~
1 ~ OAc NaSCH3 S- CH~

/~ H
(Su~stitution) ~ H

I Base (Alkylation) ¦ ICH2COOpNBæ

O ~ CH R3 ~Si 3 R3 _ 3~ '--CH

~ ~ - R4 ~ase `I
pNBzOOC R (ACylation) 1~ COOpNBz BF3 or OH BU4NF (Deprotection) S- CH
R ~ O 3 \~ C R 4 pNBzOOC R5 : :

1 The Eollowing transformations of 5 to 9a were described in the foregoing reactiorl scheme. Following the same reaction path, from (3S,4S)-4-Acetoxy-3-((lS) tert-butyldimethylsilyloxy)ethyl-azetidin-2-one, obtained itself from (S)-3-hydroxybu~yric acid (J. Amer. Chem.
Soc. _987, 109, 1132) was obtained (5R,65)-(lS-hydroxy-alkyl)oxapenem-3-carboxylic acid having the like-cis configuration.

Racemic hydroxyalkyloxapenemcarboxylic acids with the configuration of epithienamycin can be obtained starting from threo- or erythro-trans-3-(1-p-nitrobenzyloxycar bonyloxyethyl)-4-(2 hydroxyethylsulEonyl)azetidln-2-one ~J. Amer. Chem. Soc. 1980, 102, 2039) as a startiny 15material.
O O
O ~ OpNBz O ~ OpNBz 20Rl ~ 52-CH2cHz-H ~1 ~ S-CH3 N H NaSCH3 ~ H
(Substitution) I ICH2CH2COO~NBz t Base (Alkyl-R O ation) O OpNBzR - C ~ COCl O ~ OpNBz S-C~3 R Rl ~ S-CH3 ~ Base O ~ ~f R4 (Acylation) O
p~BzOOC R5 pNBzOOC

The following transformations of threo- or erythro-6 were effected according to the foregoing reaction scheme, whereby from threo-6 like-cis 9 and from erythro-6 unlike-trans-9 is formed _ . . .

:

- 15 - ~ ~3~ 3t~4 l The p-nitrobenyl protecting group, depicted in the reaction scheme, has been approved in the preparation of many stable oxapenem-3-carboxylic acids. This protecting group can be removed particularly easy and under mild reaction conditions using hydrogen and a hydrogenation catalyst as e. g. palladium on carbon according to method known per se. This cataly-tic hydrogenation can be carried out in a one phase mixture in a solvent like e. g. ethyl acetate but also in a two phase mixture of ethyl acetate and water in presence of an alkali or alkaline earth hydrogen carbonate. Accordinq to this method the free carboxylic acid yroup of the compounds of inventlon are transformed into their corresponding salts.

The simultaneous use of a p-nitrobenzyl protecting group for the carboxyl group (p-nitrobenzyl ester) and a p-nitrobenzyl carbonate protecting group for the alcoholic function of the compounds according to the invention is particularly preferred. By this means, both protecting groups can be removed in one working operation, by catalytic hydrogenation.

Instead of the p-nitrobenzyl protecting group, depicted in the reaction scheme, also other protecting groups, that are removable under mild conditions, can be used.
This is necessary, if the groups Rl, R3, R4 and R5 contain carbon-carbon double or triple bonds. In this case, the simultaneous use of an allyl protecting group for the carboxylate residue (allyl ester) and a allyl carbonate protecting group for the alcoholic function i9 preferred. Both protecting groups are then removed together during the last réaction step, using tetrakis-triphenylphosphine palladium and triphenylphosphine, according to the method known per se (M. Imuta, S.
Uyeo, M. Nakano, T. Yoshida, Chem. Pharm. ~ull. 33, 4371 - 81 (l9~S).

' . , :.
. " ;' ~ ~ ' ' - 16 ~ r~3~

1 In the general description of the present invention the groups are preferably selected from the pharmaceutlcally acceptable groups, connected via C-C singl.e bonds to the remaining part of the molecule which contain: sub-S stituted or unsubstituted alkyl, alkenyl, alkinyl,cycloalkyl, alkylcycloalkenyl, cyloalkylalkyl, alkenyl-cycloalkyl, cycloalkenylalkyl, aryl, aralkyl, aralkenyl, aralkinyl, carboxy, cyano in which the preceding alkyl, alkenyl or alkinyl parts of the molecule contain 1 10 to 6 carbon atoms, the cycloalkyl or the cycloalkenyl parts of the molecule contain 3 to 6 and -the aryl parts of the molecule contain 6 to 10 carbon atoms. Aromatic or aliphatic heterocyclyl, heterocyclylalkyl, hetero-cyclylalkenyl, heterocyclylalkinyl, alkylheterocyclyl, 15 in which -the preceding alkyl, alkenyl or the alkinyl parts of the molecule contain 1 to 6 carbon atoms and the heterocyclic part of the molecule is mono- or bicyclic and contains 3 to 10 ring atoms, of which one or more are selected from the group comprising: oxygen, sulphur 20 and nitrogen, and where the substituents of the above-mentioned groups may be: protected or unprotected hydroxyl, hydroxyalkyl, aminoalkyloxy, alkyloxy, acyloxy, aryloxy, heterocyclyloxy, carbamoyl, carbamoyloxy, thiocarbamoyl, thiocarbamoyloxy, alkylcarbamoyloxy, alkylthiocarba-moyloxy, mercapto, alkylthio, hydroxyalkylthio, amino-alkylthio, amidinoalkylthio, acylthio, arylthio, alkyl-heteroarylthio, hydroxyalkylheteroarylthio, hetero-cyclylthio, carbamoylthio, alkylcarbamoylthio, thio-carbamoylthio, alkylthiocarbamoylthio, protected or unprotected amino or monoalkyl.amino, dialkylamino, oxo,protected or unprotected oximino or alkylimino, tetraalkylammonium, cycloalkylamino, arylamino, hetero-arylamino, heterocyclamino, acylamino~ amidino, alkyl-amidino, guanidino, alkylguanidino, carbamoylamino, alkylcarbamoylami.no, thiocarbamoylamino, alkylthio-carbamoylamino, nitro, chlorine, bromine, fluorine, iodine, azido, cyano, alkylsulphinyl, alkylsulphonyl, - 17 - Z~ 3~

1 sulphonamido, sulphamoyloxy, alkylsulphonyloxy or protected or unprotected sulpho, sulphoxy or carboxyl, where the substituents occur one or more times and the alkyl part of the molecule thereof contains 1 to 6 carbon 5 atoms, the aryl part of the molecule thereof contains 6 to 10 carbon atoms, and where the heteroaromatic or heterocyclic part of the molecule is nono- or bicyclic and contains 3 to 10 ring atoms, of which one or more are selected from the group comprising: oxygen, sulphur 10 and nitrogen, and characterized in that R2 denotes hydrogen, alkyl ox acyl containing up to 10 carbon atoms, sulpho, aminosulphonyl and phosphoroyl.

A particularly preferred class of compound is that in which R1 is selected from the groups:

CH3 CH2-CH H2C=CH H2C=CH-CH2 HC-C HC--C-CH2 ~ ~ CH

25 3 3C~

~ ~ N N ~

CH3 CH2-CH2-'~H
and in which R2 denotes for hydrogen, alkyl or acyl with up to 10 carbon atoms, sulpho, amonosulphonyl ox phosphoroyl. R3 and R4 denote methyl and R5 is se~
lected from the groups:

CH3 CH -CH H2C=CH H2C=CH-CH2 HC-C HC-C~CH2 , ' ~' ,i ' : :

, - :

1 8 ~ 3 ~

S
s --~o~ I~o>~CH3 ~
C~I2-CH~-OH

NH2 ~N ~3 ,~ N--N ~N
2 0 ;; ~ C~2-CE12~ S~ CH2 -CH

Y~ ~--N
(:~2-CH2~, ,~ Z 2~ S~ CH3 CH2 -~2 -OH

C~2~S~ --N H3C OH

CH3 . CE3 2 -CH2-O}I

~ N--N
~7~ ~s~ 3 ~S~
C~

.
.` , ` ~ ' ., ~.
.
~, .
.. ` ., ;

8~3~
-- 19 - ~

o CH -S-c-NH CH2-S-C-NHCH3 CH2-S-CEI3 ~,NH
CH2-S-CH2-CH2-~lH2 C~I2-S-CH2-cH~ `~-c~ H

C}I -NH C~ C-NE12 C~2-21H-CHO

CH -NH~C 1l CH,-O-CEIO
H

o CH -Cl CH2-0-C~ CH3 CH -N

S S
2 5 C~2 -O-C-~H-C~I3 CE~2 -C~ CH3 CH2 -C~2 -~H2 ~ NH
3 0 H2 E~2 ~EI CH C}12-NEI-C CH-CE~ -C~ -NH2 C}IZ-C~2~cH2 ~H C\H CEI2-ClI2-COOH C~I2-COOH

j : . . : .: . : , - . , :

,~ , `:: ',' ' ~',` . ';' '' ' ' ' ' :, ' ' ' '.', ~ "'',:' .. ' ' ' ~ ' , Z~8.~3~

CE2-0-CH ~ oCOOH 2 H

ll CH2-cH2-oH CH2-cH2_o_c_~H C-N

Ol O

CH2-CH -O-C-N~IC~ CH2-CH2-C~12-0 C NH2 2 2 2 C NH CH3 CH2 CH~-NEI-CHO

15 Preferred protection groups of the protected substituents of R1, R3, R4 and R5 are benzyl, p-nitroben2yl, benzyl-oxycarbonyl, p-nitrobenzyloxycarbonyl, allyl, allyloxy-carbonyl, trimethylsilyl, tert-butyldimethylsilyl, benzylidene and oxomethylene.

The products according to the invention form a large plurality of pharmacologically acceptable salts with inorganic and organic bases. These include, for example, metal salts which are derived from alkali metal or 25 alkaline earth metal hydroxides, carbonates or bicarbonates, and salts which are derived from primary, secondary or tertiary amines, such as monoalkylamines, dialkylamines, trialkylamines, lower alkanolamines, di-lower alkanolamines, lower alkkylenediamines, N,N-diaralkyl lower alkylenedi-30 amines, aralkylamines, amino-substituted lower alkanols, N,N-di-lower alkylamino-substituted lower alkanols, amino-, polyamino- and guanidino-substituted lower alkanoic acids and nitrogen con~aining heterocyclic amines. Examples of salts are those which are derived from sodium hydroxide, sodium carbonate, sodium bicarbonate, potassium carbonate, potassium hydroxide, calcium carbonate, trimethylamine, triethylamine, piperidine, morpholine, - - . :, .

., , . . .

;

- 21 - ~ 83~

1 quinlne, lysine, protamine, ~rginine, procaine, ethanolamine, morphine, benzylamine, ethylenediamine, N,N'-dibenzyl~
ethylenediamine, diethanolamir1e, piperazine, dimethyl-aminoe-thanol, 2~amino-2-methyl-1-propanol, theophylline, N-methylglucamlne and the like.

The invention furthermore relates to salts of amino groups which are contained on the side chains or Rl, R3, R4 and R5 in certain species of the compounds accordiny to the invention. Such pharmaceutically acceptable acid addition sal-ts are derived from organic and inorganic acids, such ~s HCl, HBr, citric acid, tartaric acid and the like.

The salts can be monosalts, such as -the monosodium salt, which is Eormed by treat~ent of 1 equivalent of sodium hydroxide with 1 equivalent of the products according to the invention, and also disalts. Such salts can be obtained by treatment of 1 equivalent of a base having a divalent cation, such as calcium hydroxide, with 1 equivalent of the compounds according to the invention. The salts according to the invention are pharmacologically acceptable, non-toxic derivatives which can be used as active constituents in suitable pharmaceutical dosis unit forms. They can also be combined with other medicines with the formation of preparations having a wide spectrum of activity.

The new compounds according to the invention are useful belalactamase inhibitors which are effective against various gram-positive and gram-negative pathogens.
The free acid and in particular its salts, such as the amine and metal salts, in particular the alkali metal and alkaline earth metal salts, can be employed for the removal of sensitive pathogens from dental and medicinal instruments, for the destruction of micro-organisms and for therapeutic use in humans and animals.

. . : ~ . : .

. -~'. , , ' ; . .;' ,, . : .

-' ~ ` : ~ "~, , , - . ~ ;

3~

1 For this latter purpose, pharmacologically accepta~le salts with inorganic and oryanic bases, such as are known per se and used in the adminis-tration of penicillins and cephalosporins, are used. For example, salts, such as alkali metal and alkaline earth metal salts, and primary, secondary and tertiary amine salts may be used for this purpose. These salts may be used for pharmaceutically acceptable liquid and solid excipients with the suitable dose unit forms such as pills, tablets, capsules, suppositories, syrups, elixirs and the llke, which may be prepared by processes which are known per se.

The new compounds are useful betalactamase inhibitors against various gram-positive and gram-negative bacteria and are accordingly used in human and veterinary medicine.
The compounds according to the invention may be used as antibacterial medicines ~or the -treatment of infections which are caused by gram-positive or gram-negative bacteria, for example against Staphylococcus aureus, Escherichia coli, Klebsiella pneumoniae, sacillus subtilis, Salmonella typhosa, Pseudomonas and Bacterium proteus.

The betalactamase inhibitors according to the invention may furthermore be used as additives for animal feeds, for the preservation of foodstuffs or feeds and as disinfectants. For example, they can be used in aqueous preparations in concentration in the range from 0.1 -to lO0 parts of antibiotic/million parts of solution for the destruction and inhibition of the growth of harmful bacteria on medicinal and dental instruments and as bactericides in industrial applications, for example in water-based pai~ts and in soft water for paper mills, and for the inhibition of the growth of harmful bacteria.

-.

, ; :

~ - 23 ~ 3~A

1 The products according to -the lnvention can be used alone or together as active constituents in any Large number of pharmaceutical preparations. These products and their correspondiny salts may be used in capsule 5 form or as tablets, powder or liquid solutions or as suspensions or elixirs. They may be administered orally, intravenously or intramuscularly.

Moreover, pharmaceutical preparations are preferred which contain, besides the betalactamase inhibitors and additives known per se, also a betalactam antibiotic.
The mixture ratio between inhibitor and antibiotic amoun-ts preferably from 1 : 10 to 1 : 2, but also other doses are possible.
Moreover, pharmaceutical preparations are particularly preferred which, besides the hetalactamase inhibitors according to the invention, contain a penicillin or a cephalosporin.
The preparations are preferably administered in a form suitable for absorption thrcugh the gastrointestinal tract. Tablets and capsules for oral administratlon may be present in dose unit form and may contain customary pharmaceutical excipients, such as binders, for example syrup, acacia gum, gelatin, sorbitol, traganth or poly-vinylpyrrolidone; fillers, for example lactose, sugar, maize starch, calcium phosphate, sorbi-tol or glycerol;
lubricants, for example magnesium stearate, talc, poly-ethylene glycol, silica; disintegrants, for examplepotato starch, or acceptable wetting agents, such . .

:-- '. ' ~ ~
, - ~4 - ~ 3~

1 as sodium lauryl sulphate. The tablets may be coated by processes which are well known per se. Oral and liquid preparations may be present in the form of aqueous or oily suspensions, solutions, emulsions, syrups, elixirs etc., or they may be present as a dry product, for example for reconstitution wi-th water or o-ther suitable excipients before use. Such liquid preparations may contain additives which are known per se, such as suspending agents, for example sorbitol syrup, methyl-cellulose, glucose/sugar syrup, gelatin, hydroxyethyl-cellulose, carboxymethylcellulose, alumlnium stearate gel, or hydrogenated edible oils, for example almond oil, fractionated coconut oil, oily esters, propylene glycol or ethyl alcohol; preservatives, for example methyl or propyl p-hydroxybenzoate or sorbic acid.
Suppositories will contain suppository foundations which are known per se, for example cocoa butter or other glycerides.

Preparations for injection may be present in dose unit form in ampoules or in containers having a number of doses together with an added preservative. The prepa-rations may be present in the form of suspensions, solutions or emulsions in oily or aqueous excipients, and they may contain formulating agents, such as suspend-ing agents, stabilizers and/or dispersants. Alternatively, the active constituent may be present in powder form for reconstitution with a suitable excipient, for example sterile, pyrogen-free water, before use.

: . . : , .. -:

, - 25 - 2~ 3~

l The preparations rnay also be present in a sui-table form for absorption by the mucous membranes of the nose and -the throat or the bronchial tissue, and they may usefully be present in the form of powders or liquld sprays or inhalants, pastilles, as painting agents for the throat, etc. For medication of the eyes and ears, the preparations may be u!3ed in liquid or semi-solid form in the form of individual capsules, or -they may be used as drops or the like. Topical applications may be present or formulated in hydrophobic or hydrophilic foundations such as ointments, creams, lotlons, painting agents, powders and the like.

The preparations according to the invention may additional-ly contain another constituent, such as stabilizers, binders, antioxidan-ts, preservatives, lubricants, suspending agents, viscosity enhancers or flavour enhancers and the like additionally to the excipient. Other active constituents may additionally ~e contained in the prepa-rations to that a wider spectrum of antibiotic activityis obtained.

For veterinary medicine, the preparations may, for example, be formulated as an intramammary preparation in either long-acting or rapid-release foundations.

The dose to be administered depends to a large extent on the condition of the subject to be treated and the weight of the host, and the route and frequency of administration. The parenteral route is preferred for generalized infections and the oral route for in-testinal infections~ In general, a daily oral dose contains about 1 to about 200 mg of active constituent/kg .. . ..
, ' ' ` ', ` ,. ~

- 26 - ~ ¢~ 83~

1 of body weight of the subject in one or more admlnist-rations per day. A preferred daily dose for adult humans is in the range of about 5 to lO0 mg of active constituent/kg of body weight.

The preparations according to the invent:ion may be administered in various unit dose forms, for example in solid or liquid, orally absorbable dose forms. The preparations per unit dose may contain 0.1 to 99 6 of active material either in solid or liquid form.
The preferred range is about lO to 60 ~. The preparations will in general contain 15 to about 1500 mg of active constituent, however in general it is preferred to use a dose in the range from about 50 to 500 mg. With parenteral administration, the unit dose is normally the pure compound in a sterile water solution or may be present in the form of a soluble powder which can be dissolved.

~, ~' ~: . ' ' ' :

- 27 ~ 3~

1 The following examples explaln the products, procedures, preparations and methods of therapy.

Example 1 Pre~aration of (5R,6R)-trans-2-t:ert-but~l_ -((15)-hydroxyethy].)oxapenem-3-carboxy]ic acid a_d its potassium salt H O--H
~ H ~I
H C / ~ ~ CH3 R = H, K
~ C - CH

COOR

Acylation: Intermediate 2 (~1, R3, R4, R5 = CH3) In a three-necked flask fitted with a mechanical stirrer, rubber septum, starting material 1 (Rl, R3, R4, R5 = CH3) (35.5 g) is dissolved in 980 ml of dry tetra-hydrofuran under n.itrogen and the solution cooled to -78 C. Then, with stirring, 9.8 ml of pivaloyl chloride is added with a syringe and subsequently 165 ml of a 1 molar solution of lithium-bis-trimethylsilylamide was added. The mixture is stirred for additional 30 minutes at -78 C and then it is added to 5 l of toluene.
The obtained solution is then washed subsequently with 3 l lN HCl, 3 l saturated NaCl and once again with 1 lt of saturated NaCl solution and the organic layer dried over magnesium sulfate. Removal of the solvent in vacuo and chromatography of the residue on 1 kg of silica gel using toluene-ethyl acetate 9 : 1 affords pure 2 as a viscous oil (36.2 g, 86 %). IR-spectrum in CH2Cl2 : 2920, 2850, 1760, 1750, 1705, 1605, 1525, 1350, 840 cm~1.

- , ~ ' ' :
, .

- 28 - ~ 3~

l Deprotection: Intermediate 3 (Rl, R3, R4, R5 = CH3) OH
¦ S -CH.
H3C ~ O
O ~ C ~ C~

pNBzO~C CH3 In a two-necked round-bottmed flask fitted with rubber septum, reflux condenser and magnetic stirrer to a solution of 2 (R1, R3, R4, R5 = CH3) (5.53 g) in 48 ml of dry tetrahydrofuran, was added 5.71 ml of acetic acid and 26.7 ml of a 1.1 molar solution of tetrabutyl-ammonium fluoride in THF with a syringe. The septumwas then replaced by a stopper and the mixture stirred magnetically under nitrogen for approximately 22 hours.
Subsequently, it .is refluxed for 90 minutes. The reaction mixture is diluted with 1 lt of toluene and waished subsequently with 400 ml saturated NaCl, 150 ml 2N
RHCO3-solution and once again with 150 ml of saturated NaCl. The organic layer is dried over magnesium sulfate and the solvent removed in vacuo. The crystalline residue is chromatographed on 140 g silica gel using toluene-ethyl acetate (3 : 1), yielding 3.57 g 3 (82 ~) of a crystalline product. IR-spectrum in CH2Cl2:
3600, 2960, 1765, 1750, 1710, 1~10, 1525, 1350 cm~l.

3~

. . : , 2~S~

l Mitsunobu-Reaction: Interme_iate 4 (R1, R3, R4, R5 CH3) ~ CH=O
.
, S~ ~3 3 ~ () ~ ~ ~ CH3 pNBzOOC ~3 I'o a solution of 3 (4.2 g) and triphenylphosphine (5.03 g) in 72 ml of dry tetrahydrofuran, subsequently is added 1.09 ml of formic acid and 3.78 ml of diisopropyl azodicarboxylate at 0 C under argon and -the reaction mixtu.re is stirred at 0 C for 45 minutes and room temperature for an additional 90 min. The reaction solution is dissolved with 720 ml of toluene and then subsequently washed with 720 ml potions of water, dilute NaHCO3 solution and 10 % NaCl solution. The organic layer is dried over magnesium sulfate and the solvent removed in vacuo. Chromatography of the crude product on 200 g of silica gel (0.040 - 0.063 mm) using toluene-ethyl acetate (97 : 3) and with toluene-ethyl acetate (95 : 5) yields 1.95 g (44 %) of a pale yellow oil.
IR spectrum in CH2Cl2: 2930, 1765, 1750, 1720, 1605, 1525, 1350, 1175 cm~l.

Hydrolysis: Intermediate 5 (R1, R3, R4, R5 - CH3 OH
/S C~3 H3C ~ ~ CH3 O ~ C CH
pNBzOOC CH3 .

, - 30 - 2~ 3~

1 To a solutlon of 4 ~Rl, R3, R4, R5, = CH3) 11.81 g) in 39 ml of methanol under stirring at 0 C is added 3.83 ml of 1.0 N aqueous HC1 dropwise and the mixture stirred at room temperature for 9 hours. The reaction mixture is diluted with 200 ml of toluene and subsequently washed with 100 ml portions of dilute NaHCO3 solution and 10 % NaCl solution. The organic layer is dried over magnesium sulfate and the solvent removed in vacuo.
After drying in high vacuum 1.70 g of a colourless noncrystalline solid is obtained (100 %). The crude product 4 is used without urther purification. IR-spectrum in CH2C12: 3600, 2970, 1765, 1755, 1715, 1610, 1525, L350 cm~l.

Protection: Intermed_ate 6 (Rl, R3, R4, R5_= CH3) O ~ OpNBz S ~ C~
o '~f ~;~ c~
pNBzOOC CH3 To a solution of 5 (R1, R3, R4, R5 = CH3~ (1.63 g) in 8 ml of methylene chloride at -8 C with stirring is added a solution of p-nitrobenzyl chloroformate (1.10 g) in 3 ml of methylene chloride and subsequently, within 15 minutes, solid N,N-dimethylaminopyridine (0.62 g) is added. After 5 minutes of stirring, a white precipitate is formed. The reaction mixture is stirred for 2 hours at -5 C and subsequently is diluted with 75 ml of methylene chloride and washed with 50 ml of 10 % NaCl solution. The organic layer is dried over magnesium sulfate and the solvent removed `

. ~ ,.,, -. - .
, ;
. .

- 31 - 2~

1 ln vacuo. The crude mixture of 6 is -then chrornatographed using toluene-ethyl acetate (9 : 1), where upon 1.87 g (81 ~) of pure 6 as a noncrystalline so]id is obtained.
IR spectrum in CH2C12: 2970, 1765, 1755, 1715, 1610, 5 1525, 1350, 1245, 850 cm~l.

Chlorinatlon: Intermediate 7 (Rl, R3, R4, R5 = CH3) o OpNBz O ~ N ~ ~ I 3 pNBZOOC c~3 In a 250 ml two-necked flask with rubber septum, a balloon filled with nitrogen and a mechanical stirrer, at -40 C with stirring to a solution of 6 (Rl, R3, R4, R5 = CH3) in 115 ml of methylene chloride was intro-duced chlorine gas into the reaction solution with a syringe and the mixture stirred for 10 minutes sub-sequently. The cold reaction mixture is then poured on an aqueous solution of 5.74 g anhydrous sodium hydrogen sulfite and 4.57 g of anhydrous sodium carbonate and the mixture shaken. The aqueous phase is extracted with a small portion of methylene chloride and the extraction solutions combined. Drying over magnesium sulfate and removal of the solvent in vacuo affords a colourless crude product 7. Chromatography on 4.3 g of silica gel at 0 C using toluene (10 ml fractions) and using to-luene-ethyl acetate (10 ml fractions) affords 1.49 g pure product 7 (85 ~) as a noncrystalline white solid.

' . . ' ' . ' , ' .~ , 33~

1 IR-spectrum in Cfl2C12: 2970, 1790, 1755, 1720, 1610, 1525, 1350, 1245, 850 cm~l.

Cyclisation: ~-Nitrobenzyl (5R,6R) and (5S,6R)-2-.
tert-Butyl-6-((lS~-p-nitrobenzyl~xycarbony-loxyet-hx~) oxapenem-3-carboxylate 8 (R1, R3, R4, R5 = CH3) ~ ~ OpNBZ

3 ~ ~ ~ CH3 O C~
pN~zOOC' 3 In a two-necked flask fit-ted with a rubber septum, a balloon filled with nitrogen and a magne-tic stirrer, with stirring at -30 C to a solution of 7 (R1, R3, R4, R5 = CH3) (1.35 g) in 45 ml of dry tetrahydrofuran is slowly added by a syringe 2.55 ml of a 0.92 molar solution of potassium tert-butoxide in tert-butanol and the reactiorl mixture stirred subsequently for 90 minutes at -30 C. The reaction solution is diluted with 240 ml of ethyl acetate and subsequently washed one after the other with 220 ml of 10 % NaC1, 100 ml of 10 % NaCl and 100 ml of saturated NaCl solution.
Drying over magnesium sulfate and removal of the solvent in vacuo affords a cis, trans-isomeric mixture of 8.
IR spectrum in CH2C12: 2970, 1800, 1750, 1715, 1605, 1580, 1525, 1350, 1315, 1240, 1090, 850 cm~l.

:' ~ -:

3 3 ~$~31~

1 Chromatographic Separation of (5R,6R) and_(5S,6R) p-Nitrobenzy~l 2-tert-Butyl-6-((lS)-p-nitrobenzyloxycarbon oxyethyl)-oxa~enem-3-carb~y~ s 8a and 8b (Rl, R3 R4, R5 = CH3) O O
OpNBz ~ ~ OpNBZ

' ~ H3C ~ ~
~ 3 ~ N ~ 1 3 O I CH O ¦ C~
pNBZooc 3 PN~Zooc 3 8a ab The cis, trans-isomeric mixtllre of 8 (Rl, R3, R4, R5 = CH3) (1.25 g) is chromatographed on 70 g silica gel (5 - 20 ~m) at -12 C using hexane-ethyl acetate (1 : 1), where upon 705 mg (56 ~) pure 8a and 352 (28 %) 8b, which still contains some 8a, was obtained. The respective fractio~s are evaporated at 0 C in high cacuo at 0.5 Torr. 8a:
~l25 = +50 (c = 1, ethyl acetate).
8a: NMR in CD3CN: o = 1.3 (s, 9H), 1.43 (d, 3H, J = 6.5 Hz), 3.99 (dd, lH, J = 1 Hz, J = 3.5 Hz), 5.19 (m, lH), 5.26 (s, 2H), 5.33 (ABq, 2H, J = 14 Hz, J = 85 Hz), 5.78 (d, lH, J = 1 Hz), 7.67 to 7.58 (2d, 4H, J = 8.4 Hz), 8.19 (2d, 4H, J = 8.4 H7).

8b: NMR in CD3CN: ~ = 1.23 (s, 9H), 1.42 (d, 3H, J =
6.5 Hz) 4.19 (m, lH), 5.19 (m, lH), 5.24 (s, lH), 5.33 (ABq, 2H, J = 14 Hz, J = 82.6 Hz), 5.89 (d, lH, J =
3.1 Hz), 7.59 to 7.66 (2d, 4H, J = 9 Hz), 8.21 (2d, 4H, J = 9 Hz).

'` ' ' ~

- 34 _ ~ ~t~3 1 SeparatLon of (5R,6R) and (5S,6R) p-Nl~t _b~y~
tert-Butyl-6-(~lS)-~-nitrobenzyloxycarbony-oxyethyl-) oxapenem-3-carboxylates 8a and 8b (R1, R3, R4, R5 = CH3) by Fractional Crystalli~ation --_ The cis, trans-isomeric mixture 8 (R1, R3, R4, R5) (1.30 g) is dissolved in 5 ml of ethyl acetate, hexane is added until it becomes turbid and it is left for two days at -20 C, where upon the isomer 8b, containing small amounts of 8b, crystallizes. The crystals are collected on a filter and the mother liquor is immedia-tely evaporated in high vacuum at 0.5 Torr, leaving pure 8a as a white noncrystalline solid (755 mg, 61 ~).

Isomerisation of (5S,6R) to (5R,6R) p-Nitrobenzyl 2-tert-Butyl-6 ((lS)-p-nitrobenzyloxycarbonyloxyethyl) oxapenem-3-carboxylates 8b to 8a (R1, R3, R4, R5 = CH ) _ _ _ _ _ 3_ 8b (R1, R3, R4, R5 = CH3) (510 mg) is dissolved in ethyl acetate (10 ml) and left for three days a-t room temperature where upon the thermodynamic equilibrium of 69 % 8a and 31 ~ 8b is establlshed. According to the afsrementioned procedures more of the pure isomer 8a (255 mg, 50 %) is obtained by chromatography at -15 C or by fractional crystallization.

Catalytic Hydrogenation: (5R,6R)-2 tert-Butyl-((lS)-6-hydroxyethyl) oxapenem-3-carboxylic Acid- Potassium Salt 9a (R1, R3, R4, R5 = CH3) _ _ _ 0~
H ~ CH

H3C ~ ~ ~ C ~ CH3 O C~3 COOH
K

' :~ '.".; ' ' ' ' ,. .

:, S~ fl 1 In a hydrogen apparatus fitted with a rubber septum, 620 my of Pd on charcoal (10 %) in 48 ml of ethyl acetate is prehydrogenated at 0 C Eor 7 mlnutes and subsequently a freshly prepared solution of 8a (R1, R3, R4, R5 = CH3) (618 mg) in 6 ml of ethyl acetate is added wl-th a syringe and hydrogenated for 30 mlnutes at 0 C. Within 20 minutes 210 ml of hydrogen is adsorbed. The catalyst is removed by filtration and the filtrate is carefully extracted at 0 C with a cold solution of 81 mg cf potassium hydrogen carbonate in bidestilled water (20 ml~. The aqueous layer is extracted twice with por-tlons of ethyl acetate (10 ml), the remaining ethyl acetate removed shortly by evacuation and lyophilized at 0.001 Torr, where upon 188 mg (59 %) of potassium salt 9a as a white voluminous powder is obtained. UV-spectrum in H2O: ~ max = 262 nm. NMR spectrum in D2O : ~= 1.25 (s, 9H), 1.39 (d, 3H, J = 603 Hz), 3.84 (d, lH, 4.6 Hz), 4.3 (m, lH), 5.73 (s, lH).

Example 2 Pre~aration of Racemic cis- and trans-2 tert-Butyl-6-(unlike-hydroxyethyl~ oxapenem-3-carboxylic Acid . _ and its Potassium_Salt H O - H
~ .- ~ ~
H3C ~ \ ~ O C~I3 R - H, K

O '~~~~ ~ rac. trans COO~, K

.
" , : . :
`
: , : .
., . , ~ .. ' , .

.

~ 36 ~

1 Substitution: Intermediate 15 (Rl = CH3) O ~ OpNBz ,5--CE~
H C " "r~ 3 ~ N H
o To a suspension of (erythro-trans) 14 (Rl = CH3) (3.56 g) in acetonitrile (9.7 ml) under viqorous stirring is added a solution of sodium methanethiolate (0.93 g) in water (9.7 ml) at 0C within 15 minutes and the mixture stirred for additional. 30 minutes at 0 C.
The yellow, turbid reaction solution is then diluted with 75 ml of methylene chloride and washed with 25 ml of water. The aqueous phase is extracted with three portions of methylene chloride, 20 ml each, and the combined organic phases are washed with 30 ml of saturated NaCl solution. The organic phase is dried over magnesium sulfate and the solvent removed in vacuo. The obtained crude product 15 is subsequently chromatographed on 200 g of silica gel using toluene-ethyl acetate (9 : l) and toluene ethyl acetate (4 : 1), where upon 2.45 g (82 ~ of crystalline product 15 (erythro, trans) is obtained. IR spectrum in C~2Cl2 3400, 3060, 2990, 1775, 1755, 1610, 1525, 1350, 1245, 850.

': ` ' . ~,, , -,. . . . .

.

33~

1 Alkylation: Intermediate 16 (R1 = CH3) o O OpNBz H3C~'-` ~ S -CE~3 \
`I
pN~zOOC
To a solution of 15 (erythro-trans) (R1 = CH3) (2.30 g) in acetonitrile (23 ml) at 0 C, were added within 5 minutes p-nitrobenzyl iodoacetate (3.26 g) with stirring and subsequently, within 15 minutes solid caesium carbonate (3.30 g) with vigorous stirring at 0 C and subsequently the reaction mixture was stirred for 1 additional hour. To the mixture 230 ml of toluene is added and it is subsequently washed with portions of 80 ml of saturated sodium hydrogen carbonate and saturated sodium chloride solution. The organic phase is dried over magnesium sulfate and the solvent removed in vacuo. The obtained crude product is then chromatographed on 220 g of silica gel using toluene-ethyl acetate (9 . 1), where upon 2.21 g of a colourless solid is 25 obtained (62 %). IR spectrum in CH2C12: 3060, 1770, 1750, 1610, 1525, 1350, 1245, 1190, cm~1.

Acylation: Intermediate 6 (R1, R3, R4, R5 - CH3) ll O OpNBz ~3C/ ~ ~ ~ C~3 pNBzOOC CH3 :- .

, ' - 38 ~ 33~

l To a solu-tion of 16 (erythro) (2.2.L y) and pivaloyl chloride (0.54 ml) in 53 ml of tetrahydro~uran is added dropwise at -70 C under nitrogen and with vigorous stirring a lM solution (8.28 ml) of lithium-bis-trimethyl-silylamlde in THF within 15 minutes and the reactionmixture stirred for additional 30 minutes at -70 C.
The reaction solution is diluted with 350 ml o~ toluene, washed with 300 ml 2N HCl and twice with portions, 300 ml each, of saturated NaCl solution. Drying of the organic phase over MgSO4 and removal of -the solvent affords the crude product 6. Chromatography on silica gel (100 g) using toluene-ethyl acetate (9 : 1) a~fords 1.98 g (77 %) pure 6 as a colourless noncrystalline solid. IR spectrum in CH2Cl2: 2980, 1770, 1755, 1715, 1610, 1525, 1350, 1245, 1190 cm~1.

Further transformations.

According to the procedure for the preparation of (5R,6R)-trans-2-tert-butyl-6-(lS)-hydroxyethyl)oxapenem-3-carboxylic acid (example 1) starting from racemic 6 (Rl, R3, R4, R5 = CH3), via steps chlorination, cyclisation and catalytic hydrogenation the racemic compounds 9a (trans) and _ (cis) are obtained (as potassium salts).
Compound 9b contains approx. 20 % 9a.

9a: NMR spectrum in D2O : ~ - 1.25 (s, 9H), 1.34 (d, 3H, J = 6 Hz), 3.84 (d, lH, J = 4.5 Hz), 4.27 to 4.30 (m, lH), 5.73 (s, lH).
9b: NMR spectrum in D2O : ~ = 1.26 (s, 9H), 1.34 (d, 3H, J = 6.4 Hz), 3.88 to 3.90 (dd, lH, J = 3.2 Hz, J = 8.8 Hz), 4.26 to 4.30 (m, l H), 5.79 (d, lH, J
= 3.2 Hz).

.. ..

,.
' .: ,..
:.

39 ~ t~33~

1 Example 3 Preparation of (like)-cis-2-tert-butyl-6-(1-hydroxyethyl)-oxapenem-3-carboxylic acid and its potassium sal-t -~ O - H
\ . H H
~ IEl3 ~ ~ ~ CH
CO(: ~

Starting from threo-trans~3 (1-p-nitrobenzyloxycarbonyl-oxyethyl)-4-(2-hydroxyethylsulfonyl)azetidin 2-one 14 (Rl, R3, R4, R5 = CH3) according to the aforementioned procedure racemic (like)-cis-2--tert-butyl-6-(1-hydroxy-ethyl)oxapenem-3-carboxylic acid sodium salt is obtained.
UV spectrum in H2O: ~ max 263 nm ( ~ = 4500).

Example 4 Alternative pre~aration _f (5R,6R)-trans-2-tert-butyl-6-((lS~-hydroxyethyl)oxapenem-3-carboxylic acid potassium salt without reversal of configuration ~H

3 ~ 1 3 o CR3 COOR

- .: . ., :,.

.
-- . . . .

, ~

- 40 ~ 3~

l Substitution: Intermedia-te ll (R1 = Ci-13) ~Sl~ 3 ~ CH~
H3C ~ S- CH3 ~H

In a three-neclced flask, Eitted with a stirrer and dropping funnel, to a suspension of (3R,4R)-4-acetoxy-3-((lS)-tert-butyldimethylsilyloxyethyl)azetldin-2-one (28.74 g) in 110 ml of acetonitril at 0 C with lS vigorous stirring was added dropwise a solution of sodium methanethiolate (10.52 g~ in 110 ml of water within 15 minutes. The colourless reaction mixture is subsequently stirred at room temperature for 75 minutes. The mixture is diluted with 500 ml of methylene chloride and washed with 250 ml of water. The aqueous phase is further extracted with three portions of methy-lene chloridej 200 ml each, and the combined organic phases dried over magnesium sulfate. Evaporation of the solution in vacuo affords 11 as a noncrystalline solid. Yield 23.9 g (87 ~). IR spectrum in CH2Cl2:
3400, 2960, 2930, 2860, 1775, 1365, 1145, 1070 cm~1.

Alkylation: Intermediate 12 (R1 = CH3 S+i - CH3 . CH3 H3C ~ S- CH3 O
COOpNBz , . "~

- 41 ~ 5~

1 In a four-necked flask, fitted with a stirrer, rubber septum and a balloon filled with nitrogen, at -70 C
to a solution of intermediate 11 ~Rl = CH3) (24.~ g) in 196 ml of dry tetrahydrofuran is slowly added 36 ml of a 2.5 M solution of butyl lithium in hexane. The obtained solution is further stirred for 20 rninutes at -20 C and this solution is added wlth a syrinye to a solution of p nitrobenzyl iodoacetate (34.7 g) in 106 ml of dry N,N-dimethylformamide within 5 minutes and with vigorous stirring. The reaction mixture is further stirred for 50 minutes at 0 C and then diluted with 2 lt of ether, and washed with two portions, 400 ml each, of 2 % NaCl solution. The combined aqueous phases are again extracted with 300 ml oE ether and lS the combined organic extracts dried over magnesium sulfate. Evaporation of the solution in vacuo affords the crude product 12. Chromatography on 1.6 kg of silica gel using toluene-ethyl acetate (97 : 3) and toluene-ethyl acetate (9 : 1) affords 31.7 g (75 %) 12 as a highly viscous, colourless oil. IR spectrum in CH2Cl2: 3050, 2955, 2925, 2855, 1760, 1610, 1530, 1380, 1350, 1185 cm~l.

Acylation: Intermediate 13 (R1, R3, R4, R5 = CH3) +

,,CH;
/S~ .
O \ CH3 3 ~ ~ CR3 ~NBzOOC CH3 ":
:

2~ ~ S~3 - ~2 -1 According to the given procedure for the preparation of 2 (Rl, R3, R4, R5 = CH3) starting from 12 (R1, R3, R4, R5, = CH3) -the intermediate 13 as a noncrystalline viscous oil in 90 % yi~ld is obtained. IR spectrum in CH2C12: 2920, 2850, 1760, 1750, 1605, 1350 c~-l.

Deprotection: Intermediate 5 ~_1, R3, R4, R5 = CH3) OH
/S- CE~3 H3C ~ O
~ CH3 pWBzOOC C~3 According to the described procedure (example 1) for the preparation of 3 ~Rl, ~3, R4, R5 = CH3~ starting from 13 the intermediate S as a colourless noncrystalline solid was obtained in 80 ~ yield. .TR spectrum in CH2C12:
3600, 2970, 1765, 1755, 1715, 1610, :i525, 1350 cm~l.

Example 5 Transformation of the hydroxyalkyl side chain: (5R,6R)-2-tert-butyl-6-((lS)-sulfoxyethyl)oxapenem-3~carboxylic acid disodium salt (Rl, R3, R4, R5 = CH~: R2 = SO3H) H O~S-OR
~-" O
H3C / ~ I CH3 C - CH3 R = H, Na COOR

, . .. .

.. ..

~ .
~' ' :
.~

43 ~ 58~4 1 To a suspension of (5R,6R)-2-tert-butyl-6-((lS)-h~clroxy-ethyl)oxapenem-3-carboxylic acld potassium salt (14 mg) in 3 ml of dry -tetrahydrofuran is added 20 mcl of trimethylsil~l chlorosulfonate with stirring at 0 C
and the reaction mlxture stirred for additional 15 minutes.
To the reaction mixture subsequently is added 2.4 ml precooled phosphate buffer pH 7.~ and it ls stirred for 30 minutes. Chromatography on reversed phase silica gel (RP-18) in acetonitrile-water (1 : 4) at 0 C affords 7.5 mg of pure sulfate as a disodium salt. UV spectrum in H2O: ~ max = 275 nm (~ = 5000). NMR spectrum in D2O: 1.26 (s, 9H), 1.48 (d, J = 6.5 Hz, 3H), 4.0~ (d, = 3.5 Hz, lH), 4.89 (m, lH), 5.84 (s, lH).

Example 6 Preparation of pharmaceutical preparations A unit dose form is prepared by mixing 60 mg (5R,6R)-tert-butyl-6-((lS) (hydroxyethyl)oxapenem-3-carboxylic acid K-salt with 120 mg of ampicillin, 20 mg of lactose and 5 mg of magnesium stearate and the 205 mg of mixture added to a No. 3 gelatine capsule. Similarly, if more active constituents and less lactose are used, other dose forms may be prepared and filled into No. 3 gelatin capsules: and should it be necessary to mix more than 205 mg of constituents together, larger capsules, and also compressed tablets and pills, may also be produced.
The following examples illustrate the preparation of pharmaceutical preparations.

_ l Tabl_ tng (5R,6R)-tert-sutyl-6-((ls)-hydroxyethyl)-oxapenem-3-carboxylic acid, K-salt 60 ~mpicillin 120 Maize starch V.S.P. 6 Magnesium stearate 232 Dicalcium phosphate 192 Lactose, V.S.P. l90 The active constituents are mixed with the calcium phosphate, lactose and about half the maize starch. The mlxture is then granulated with 6 mg of maize starch and coarse-sieved. It is dried in high vacuum and again sieved through sieves having internal mesh widths of 1.00 mm (No. 16 screens). The rest of the maize starch and the magnesium stearate is added and the mixture is pressed to give tablets which each weigh 800 mg and have a diameter of about 1.27 cm (0.5 in.).

Parenteral solution (5R,6R)-tert-Butyl-6-~(lS)-hydroxyethyl)-100 mg oxapenem-3-carboxylic acid, K-salt Ampicillin 500 mg Sterile water (is added from a separate ampoule using a syringe immediately before use) 2 ml 3~

1 Oph-thalmic solution -(5R,6R)-tert-Butyl-6-((lS)-hydroxyethyl)-oxapenem-3-carboxylate, K-salt 20 mg 5 Ampicillin 100 mg Hydroxypropylmethylcellulose 5 mg Sterile water to 1 ml (is added from a separate ampoule using a syringe immediately before use) Otical solution (5R,6R)-2-tert-Butyl-6-((lS)-hydroxyethyl)-oxapenem-3-carboxylate, K-salt 20 mg 15 Ampicillin 100 mg Benzalkonium chloride 0.1 mg Sterile water to (is added from a separate ampoule immediately before use with a syringe) 1 ml 20 Topical cream or ointment (5R,6R)-2-tert-Butyl-6-((lS)-hydroxyethyl)-oxapenem-3-carboxylate, K-salt 20 mg Ampicillin 100 mg 25 Polyethylene glycol 4000 V.S.P. 400 mg Polyethylene glycol 400 V.S.P. 100 mg End of description .
, '

Claims (13)

1. Compounds of the structural formula their pharmaceutically acceptable salts, esters and amide derivatives, in which R1, R3, R4 and R5 independently of one another denote pharmaceutically acceptable groups having 1 to 10 carbon atoms which are connected with the other part of the molecule via carbon-carbon single bonds, and in which R2 denotes hydrogen, alkyl or acyl having 1 to 10 carbon atoms or an inorganic acid residue.

2. Compounds according to Claim 1, characterized in that R1, R3, R4 and R5 independently of one another denote substituted or unsubstituted alkyl, alkenyl, alkinyl, cycloalkyl, alkylcycloalkyl, alkylcycloalkenyl, cycloalkylalkyl, alkenylcycloakyl, cycloalkenylalkyl, aryl, aralkyl, aralkenyl, aralkinyl, carboxy or cyano, in which the aforementioned alkyl, alkenyl or alkinyl parts of the molecule contain 1 to 6 carbon atoms, the cycloalkyl or the cycloalkenyl parts of the molecule contain 3 to 6 carbon atoms and the aryl parts of the molecule contain 6 to 10 carbon atoms, aromatic or aliphatic heterocyclyl, heterocyclylalkyl, hetero-cyclylalkenyl, heterocyclylalkinyl, alkylheterocyclyl, in which the aforementioned alkyl, alkenyl or the alkinyl parts of the molecule contain 1 to 6 carbon atoms and the heterocyclic part of the molecule is mono or bicyclic and contains 3 to 10 ring atoms of which one or more are selected from the group comprising: oxygen, sulphur and nitrogen, and where the substituents of the above-mentioned groups may be: protected or unprotected hy-droxyl, hydroxyalkyl, aminoalkyloxy, alkyloxy, acyloxy, aryloxy, heterocyclyloxy, carbamoyl, carbamoyloxy, thiocarbamoyl, thiocarbamoyloxy, alkylcarbamoyloxy, alkylthiocarbamoyloxy, mercapto, alkylthio, hydroxy-alkylthio, aminoalkylthio, amidinoalkylthio, acylthio, arylthio, alkylheteroarylthio, hydroxyalkylhetero-arylthio, heterocyclylthio, carbamoylthio, alkylcarba-moylthio, thiocarbamoylthio, alkylthiocarbamoylthio, protected or unprotected amino or monoalkylamino, di-alkylamino, oxo, protected or unprotected oximino or alkylimino, tetraalkylammonium, cycloalkylamino, aryl-amino, heteroarylamino, heterocyclamino, acylamino, amidino, alkylamidino, guanidino, alkylguanidino, car bamoylamino, alkylcarbamoylamino, thiocarbamoylamino, alkylthiocarbamoylamino, nitro, chlorine, bromine, fluorine, iodine, azido, cyano, alkylsulphinyl, alkyl-sulphonyl, sulphonamido, sulphamoyloxy, alkylsulphonyl-oxy, or protected or unprotected sulpho, sulphoxy or carboxyl, where the substituents independently of one another occur one or more times and the alkyl part of the molecule thereof econtains 1 to 6 carbon atoms, the aryl part of the molecule thereof contains 6 to 10 carbon atoms, and where the heterocyclic part of the molecule is mono- or bicyclic and contains 3 to 10 ring atoms, of which one or more are selected from the group comprising: oxygen, sulphur and nitrogen, and characterized in that R2 denotes hydrogen, alkyl or acyl containing 1 to 10 carbon atoms, sulpho, ami.no-sulphonyl, or phosphoroyl.

3. Compounds according to Claim 2, characterized in that R1 is selected from the groups:
CH3 CH2-CH H2C=CH H2C=CH-CH2 HC?C HC?C-CH2 and in which R2 denotes hydrogen, alkyl or acyl having 1 to 10 carbon atoms, sulpho, aminosulphonyl or phosphoroyl and in which R3 and R4 denote methyl and R5 is selected from the groups:

CH3 CH2-CH3 H2C=CH H2C=CH-CH2 HC?C HC?C-CH2 CH2-S-?-NH2 CH2-S-?-NHCH3 CH2-S-CH3 CH2-S-CH2-CH2 -NH2 CH2-NH2 CH2-NH-CHO

CH2-O-CHO

CH2-C1 CH2-N3 CH-2CH2-NH

CH2-CH2-CH2-NH2- CH2-CH2-COOH CH2-COOH

CH2-O-CH3 CHOOH CH2-OH

CH2-CH2-OH CH2-CH2-O-?-NH2 C?N

CH2-CH2-O-?-NHCH3

4. Compounds according to Claim 3, characterized in that R1 and R2 have the above mentioned definitions and R3, R4 and R5 denote methyl.

5. Pharmaceutical preparation, characterized in that it contains a compound according to at least one of Claims 1 to 4 and a pharmaceutical excipient therefor.

6. Pharmaceutical preparation, characterized in that it exists in a dose unit form and in that it contains a therapeutically effective amount of a compound according to at least one of Claims 1 to 4 and a pharmaceutical excipient therefor.

7. Pharmaceutical preparation, characterized in that it contains a compound according to at least one of Claims 1 to 4, a betalactam antibiotic and a pharmaceutical excipient therefor.

8. Pharmaceutical preparation, characterized in that it exists in a dose unit form, and in that it contains a therapeutically effective amount of a compound according to at least one of Claims 1 to 4, a betalactam antibiotic and a pharmaceutical excipient therefor.

9. Use of compounds of the structural formula their pharmaceutically acceptable salts, esters and amide derivatives, in which R1, R3, R4 and R5 independently of one another denote pharmaceutically acceptable groups having 1 to 10 carbon atoms which are connected to the other part of the molecule via carbon-carbon single bonds, and in which R2 denotes hydrogen, alkyl or acyl having 1 to 10 carbon atoms or an inorganic acid residue, as betalactamase inhibitors.

10. Procedure for the preparation of compounds according to Claim 1, characterized in that optically active, protected or unprotected 4-acetoxy-3-((1S)-hydroxyalkyl)azetidin-2-one is used as a starting material or as intermediate.

11. Procedure for the preparation of compounds according to Claim 1, characterized in that optically active, protected or unprotected 4-acetoxy-3-((1R)-hydroxyalkyl)azetidin-2-one is used as a starting material or intermediate.

12. Racemic (1',6-unlike)-trans-6-(1'-hydroxyalkyl)oxa-penem-3-carboxylic acids according to at least one of claims 1 -4, including their enantiomeric components, and their use as betalactamase inhibitors.

13. Racemic (1',6-unlike)-cis-6-(1'-hydroxyalkyl)oxa-penem-3-carboxylic acids according to at least one of claims 1 -4, including their enantiomeric components, and their use as betalactamase inhibitors.