CEPHALOSPORIN DERIVATIVES

BACKGROUND OF THE INVENTION

1. Field of the Invention

The present invention is directed to new cephem derivatives represented by the general formula

in which the Acyl substituent is a group of the formula

o

II

Ar-S-CH ₂ -C-

wherein Ar is an optionally substituted lipophilic phenyl, naphthyl or pyridyl group; R ¹ represents C C ₁₀ alkyl or C ₃ -C ₆ cycloalkyl linked by a carbon atom to the quaternary nitrogen, said alkyl or cycloalkyl group having a carboxy, -S0 ₃ H or tetrazolyl substituent and said alkyl group being optionally interrupted by -S- or

R ⁷

I

N

and optionally substituted by one or more of ( -C alkylthio, hydroxy, (C C ₆ )alkylsulfinyl, (C _j -C ₆ )alkylsulfonyl, carbamoyl, ureido, C ₂ -C ₆ alkenyl, halo, oxo, hydroxyimino, heteroaryl or phenyl in which the phenyl or heteroaryl group is

optionally substituted by up to three hydroxy or ( -C alkoxy groups; R ² , R ³ , R ⁹ and R ¹⁰ are each independently hydrogen, (C ₁ -C ₁₀ )alkyl or ( -C^alkyl substituted by one or more, preferably one or two, substituents independently selected from C0 ₂ H, hydroxy and NR ^π R ¹² in which R ¹¹ and R ¹² are each independently hydrogen or (C ₁ -C ₆ )alkyl, and R ² and R ⁹ or R ³ and R ¹⁰ can optionally be joined in a ring, preferably a 5-6 membered ring; and R ⁷ is hydrogen or (C,-C ₆ )alkyl. The derivatives are gram-positive antibacterial agents especially useful in the treatment of diseases caused by methicillin-resistant Staphylococcus aureus (also referred to below as MRS A or methicillin-resistant S. aureus .

2. Description of the Prior Art

The literature discloses a vast number of cephem derivatives having a wide variety of C-3 and C-7 substituents. Applicants are not aware, however, of any literature disclosing compounds with the combination of C-3 and C-7 substituents found by applicants to give the desired activity, solubility and toxicity profile needed for a commercially viable anti-MRSA cephalosporin product.

SUMMARY OF THE INVENTION

The present invention provides a novel series of cephem derivatives of the general formula

I wherein Ar is a group of the formula

in which R ⁴ , R ⁵ and R ⁶ are each independently hydrogen, halogen, trihalomethyl, nitro, amino, hydroxy, hydroxy (Cι-C ₆ )alkyl, (Cι-C ₆ )alkyl, - (CH ₂ ) _n R ⁷ or -(CH ₂ ) _n SR ⁷ ; n is an integer of from 1 to 6; R ⁷ is hydrogen or(Cι- C ₆ )alkyl; R ¹ represents alkyl having from 1 to 10 carbons or cycloalkyl having from 3 to 6 carbons, said alkyl or cycloalkyl group being linked by a carbon atom to the quaternary nitrogen and having a carboxy, -SO ₃ H or tetrazolyl substituent, and said alkyl group being optionally interrupted by -S- or

R ⁷ -N—

and optionally substituted by one or more of (Cι-C ₆ )alkylthio, hydroxy, ( - C6)alkylsulfinyl, (Cι-C6)alkylsulfonyl, carbamoyl, ureido, C ₂ -C ₆ alkenyl, halo, oxo, hydroxyimino, heteroaryl or phenyl in which the phenyl or heteroaryl group is optionally substituted by up to three hydroxy or (Cι-C ₆ )alkoxy groups; R ⁷ is as defined above; R ² , R ³ , R ⁹ and R ¹⁰ are each independently hydrogen, (C C ₁₀ )alkyl or (C C ₁₀ )alkyl substituted by one or more, preferably one or two, substituents

independently selected from C0 ₂ H, hydroxy and NR ⁿ R ¹² in which R ¹¹ and R ¹² are each independently hydrogen or (C C ₆ )alkyl, and R ² and R ⁹ or R ³ and R ¹⁰ can optionally be joined in a ring, preferably a 5-6 membered ring; and R ⁸ is hydrogen or a protecting group; or a pharmaceutically acceptable salt or prodrug thereof.

The compounds of formula I are antibacterial agents useful in the treatment of infections in humans and other animals caused by a variety of gram-positive bacteria, particularly methicillin-resistant S. aureus.

Also included in the invention are processes for preparing the compounds of formula I and pharmaceutical compositions containing said compounds in combination with pharmaceutically acceptable carriers or diluents.

DETAILED DESCRIPTION

The present invention provides novel cephem derivatives of general formula I above which are antibacterial agents useful in the treatment of infectious diseases in humans and other animals. The compounds exhibit good activity against a variety of gram-positive microorganisms, e.g. S. pneumoniae. S_ pyogenes. S. aureus. E. faecalis. E. faecium. S. epidermidis and S. hemolyticus. and are particularly useful against strains of methicillin-resistant S. aureus.

The compounds of formula I are characterized by a substituted pyridiniumthiomethyl group of the type

at the 3-ρosition of the cephem ring and a lipophilic 7-substituent of the type

o II Ar - S - CH ₂ - C - NH -

wherin Ar is an aromatic group selected from optionally substituted phenyl, naphthyl or pyridyl.

To elaborate on the definitions for the substituents of the formula I compounds:

(a) "Halogen" includes chloro, bromo, fluoro and iodo, and is preferably chloro or bromo;

(b) "Trihalomethyl" includes trichloromethyl, trifluoromethyl, tribromomethyl and triiodomethyl, but is preferably trifluoromethyl;

(c) The aliphatic "alkyl", "alkoxy" and "alkenyl" groups may be straight or branched chains having the specified number of carbon atoms. It is preferred that such groups have up to 6 carbon atoms and most preferably up to 4 carbon atoms;

(d) "Heteroaryl" includes mono-, bi- and polycyclic aromatic heterocyclic groups containing 1-4 O, N or S atoms; preferred are 5- and 6-membered heterocyclic rings such as thienyl, furyl, thiadiazolyl, oxadiazolyl, triazolyl, isothiazolyl, thiazolyl, imidazolyl, isoxazolyl, tetrazolyl, oxazolyl, pyridyl, pyrazinyl, pyrimidinyl, pyridazinyl, pyrrolyl, pyrazolyl, etc.

(e) The alkyl or cycloalkyl R ¹ substituent is linked by a carbon atom to the quaternary nitrogen of the pyridine ring, e.g.

where C-Q represents the R ¹ substituent, and may be optionally

R ⁷ interrupted by — s — or — N — and /or optionally substituted by one or more of (C _j -C^alkylthio, hydroxy, (C C ₆ )alkylsulfinyl, ( -

C ₆ )alkylsulfonyl, carbamoyl, ureido, C ₂ -C ₆ alkenyl, halo, oxo, hydroxyimino, heteroaryl or phenyl in which the phenyl or heteroaryl group is optionally substituted by up to three hydroxy or ( -C alkoxy groups.

The term "pharmaceutically acceptable salt" as used herein is intended to include the nontoxic acid addition salts with inorganic or organic acids, e.g. salts with acids such as hydrochloric, phosphoric, sulfuric, maleic, acetic, citric, succinic, benzoic, fumaric, mandelic, p-toluenesulfonic, methanesulfonic, ascorbic, lactic, gluconic, trifluoroacetic, hydroiodic, hydrobromic, and the like.

Some of the compounds of the present invention have an acidic hydrogen and can, therefore, be converted with bases in a conventional manner into pharmaceutically acceptable salts. Such salts, e.g. ammonium, alkali metal salts, particularly sodium or potassium, alkaline earth metal salts, particularly calcium or magnesium, and salts with suitable organic bases such as lower alkylamines (methylamine, ethylamine, cyclohexylamine, and the like) or with substituted lower alkylamines (e.g. hydroxyl-substituted alkylamines such as diethanolamine, triethanolamine or tris(hydroxymethyl)aminomethane), or with bases such as piperidine or morpholine, are also intended to be encompassed by the term "pharmaceutically acceptable salt".

Compounds of formula I in the form of acid addition salts may be written as

where X represents the acid anion and R ⁸ is hydrogen or a carboxyl-protecting

Θ group. The counter anion X may be selected so as to provide pharmaceutically acceptable salts for therapeutic administration.

The carboxyl-protecting group R ⁸ is intended to include readily removable ester groups which have been conventionally employed to block a carboxyl group during the reaction steps used to prepare compounds I and which can be

removed by methods which do not result in any appreciable destruction of the remaining portion of the molecule, e.g. by chemical or enzymatic hydrolysis, treatment with chemical reducing agents under mild conditions, irradiation with ultraviolet light or catalytic hydrogenation, etc. Examples of such protecting groups include benzhydryl, p-nitrobenzyl, 2-naphthylmethyl, allyl, benzyl, p- methoxybenzyl, trichloroethyl, silyl such as trimethylsilyl, phenacyl, acetonyl, o- nitrobenzyl, 4-pyridylrnethyl and (C,-C ₆ )alkyl such as methyl, ethyl or t-butyl. Included within such protecting groups are those which are hydrolyzed under physiological conditions such as pivaloyloxymethyl, acetoxymethyl, phthalidyl,

indanyl, α-acetoxyethyl, α-pivaloyloxyethyl, and methoxy methyl. Compounds of

formula I with such physiologically hydrolyzable carboxyl protecting groups are also referred to herein and in the claims as "prodrugs". Compounds of formula I where R ⁸ is a physiologically removable protecting group are useful directly as antibacterial agents. Compounds where an R ⁸ protecting group is not physiologically removable are useful intermediates which can be easily converted to the active form by conventional deblocking procedures well-known to those skilled in the art.

Compounds of formula I wherein a hydroxyl substituent is esterified with a group hydrolyzable under physiological conditions are also included within the scope of the term "prodrug" as used herein and in the claims. Such hydroxyl protecting groups may be employed, for example, to increase the solubility of the formula I compound. Illustrative of suitable ester "prodrugs" of this type are

compounds of formula I wherein one or more hydroxy substituent groups are converted to sulfate (-OS0 ₃ H) or phosphate (-OP0 ₃ H ₂ ) groups.

A preferred embodiment of the present invention comprises compounds of formula I wherein R ¹ is

\ CO ₂ H (D- _^ , SO_NH ₂ <S)-CH ₂ CH ₂ CH ₂ CH ₂ CH(OH)CO-H , -CH ₂ CH ₂ CH ₂ CH(0H)C0 ₂ H

H H „

N N- N

-CH ₂ CH ₂ CH ₂ S — r^ N .CH ₂ C(0)NH— ^ ^ _N -CH ₂ CH ₂ C(0)NH— r^ ^ N

N- N , N" ' _N - _N ' ,

H .

-CHjCH ₂ CH ₂ C ⁽ 0 ⁾ NH — ^ ,N -CH ₂ CH ₂ S0 ₃ H _f -(CH_)_C0NHCH_CH ₂ S0 ₃ H

N- N '

N-OH

\/ C0 ₂ H or X/^ CO,H

The most preferred Ar substituents for the compounds of the present invention are

in which R ⁴ , R ⁵ and R ⁶ are each independently hydrogen, halogen, (C _j -C ₆ )alkyl, trifluoromethyl, hydroxy, hydroxy(C ₁ -C ₆ )alkyl or amino.

Another preferred embodiment of the present invention comprises compounds of formula I in which R ¹ is ( -C alkyl substituted by an oxo o c / group and a group selected from carboxy, S0 ₃ H and tetrazolyl. Examples of such R ¹ groups include:

-C

-C

Within this embodiment the preferred Ar substituents are

in which R ⁴ , R ⁵ and R ⁶ are each independently hydrogen, halogen, ( -C alkyl, trifluoromethyl, hydroxy, hydroxy(C,-C ₆ )alkyl or amino. The most preferred Ar substituents are

In another aspect the present invention provides compounds of the formula

IA

wherein Ar is a group of the formula

in which R ⁴ , R ⁵ and R ⁶ are each independently hydrogen, halogen, trihalomethyl, nitro, amino, hydroxy, hydroxy(C _r C ₆ )alkyl, (C _r C ₆ )alkyl, -(CH ₂ ) _n OR ⁷ or -(CH ₂ ) _n SR ⁷ ;

n is an integer of from 1 to 6; R ⁷ is hydrogen or (C,-C ₆ )alkyl; and R ⁸ is hydrogen or a protecting group; and pharmaceutically acceptable salts or prodrugs thereof.

The definitions and the preparative methods discussed herein for the formula I compounds also pertain to the formula IA compounds, as do the preferred Ar substituents. The formula IA compounds are also potent gram- positive antibacterial agents especially useful for MRSA infections.

The preferred individual compounds of the present invention, all of

which have an MIC y_s a representative MRSA strain of < 8 μg/ml, are listed

below:

l-[2-carboxy-2-propen-l-yl]-4-[[(6R)-rrans-2-carboxy-8-ox o-7-[(2,5- dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0]-oct -2-en-3- yl]methylthio]pyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 1)

l-[2-carboxy-2-hydroxy ethyl]-4-[[(6R)-trans-2-carboxy-8-oxo-7-[(2,5- dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0]-oct -2-en-3- yl]methylthio]pyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 2)

l-(carboxymethyl)-4-[[(6R)-trans-2-carboxy-8-oxo-7-[(2,5- dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0]-oct -2-en-3- yl]methylthio]pyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 3)

l-[(3-carboxy-l-propyl)]-4-[[(6R)-trans-2-carboxy-8-oxo-7-[( 2,5- dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0]-oct -2-en-3- yl]methylthio]pyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 4)

l-[(5-carboxy-l-pentyl)]-4-[[(6R)-trans-2-carboxy-8-oxo-7 -[(2,5-

dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0]- oct-2-en-3- yl]methylthio]pyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 5)

l-[(R)-l-carboxy-l-ethyl]-4-[[(6R)-trans-2-carboxy-8-oxo- 7-[(2,5- dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0]-oct -2-en-3- yl]methylthio]pyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 6)

l-[(l-carboxy-l-cyclopropyl)]-4-[[(6R)-trans-2-carboxy-8- oxo-7-[(2,5- dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0]-oct -2-en-3- yl]methylthio]pyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 7)

l-[(S)-l-carboxy-2-hydroxy-l-ethyl]-4-[[(6R)-trans-2-carb oxy-8-oxo-7-[(2,5- dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0]-oct -2-en-3-

yl]methylthio]pyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 8)

1-[(1S, 2R, 3S, 4R)-l-carboxy-2,3,4,5-tetrahydroxy pent-l-yl]-4-[[(6R)-trans-2-carboxy-

8-oxo-7-[(2,5-dichlorophenylthio)acetamido]-5-thia-l-azab icyclo[4.2.0]-oct-2-en-3- yl]methylthio]pyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 9)

l-[(S)-l-carboxy-3-(methylthio)prop-l-yl]-4-[[(6R)-trans- 2-carboxy-8-oxo-7-[(2,5- dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0]-oct -2-en-3- yl]methylthio]pyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 10)

l-[(S)-l-carboxy-3-(methylsulfinyl)prop-l-yl]-4-[[(6R)-tr ans-2-carboxy-8-oxo-7-[(2,5- dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0]-oct -2-en-3- yl]methylthio]pyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 11)

l-[(S)-l-carboxy-3-(methylsulfonyl)prop-l-yl]-4-[[(6R)-tr ans-2-carboxy-8-oxo-7-[(2,5- dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0]-oct -2-en-3- yljxnethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 12)

l-[(S)-3-aminosulfonyl-l-carboxyprop-l-yl]-4-[[(6R)-trans-2- carboxy-8-oxo-7-[(2,5- dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0]-oct -2-en-3- yl]methylthio]pyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 13)

l-[(S)-4-(aminocarbonyl)amino-l-carboxybut-l-yl]-4-[[(6R) -trans-2-carboxy-8-oxo-7- [(2,5-dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2. 0]-oct-2-en-3- yl]methylthio]pyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 14)

l-[(S)-l-carboxy-(4-hydroxy phenyl)methyl]-4-[[(6R)-trans-2-carboxy-8-oxo-7-[(2,5- dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0]-oct -2-en-3- yl]methylthio]pyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 15)

l-[(S)-l-carboxy-2-(2-thienyl)ethyl]-4-[[(6R)-trans-2-car boxy-8-oxo-7-[(2,5- dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0]-oct -2-en-3- yl]methylthio]pyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 16)

l-[3-carboxy-2-hydroxy-l-propyl]-4-[[(6R)-trans-2-carboxy -8-oxo-7-[(2,5- dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0]-oct -2-en-3- yl]methylthio]pyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 17)

l-[(S)-5-carboxy-5-hydroxypent-l-yl]-4-[[(6R)-trans-2-carbox y-8-oxo-7-[(2,5- dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0]-oct -2-en-3- yl]methylthio]pyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 18)

l-[(S)-5-carboxy-5-hydroxypent-l-yl]-4-[[(6R)-trans-2-car boxy-8-oxo-7-[(2,6- dichloropyridin-4-yl)thioacetamido]-5-thia-l-azabicyclo[4.2. 0]-oct-2-en-3- yl]methylthio]pyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 19)

l-[(S)-4-carboxy-4-hydroxybut-l-yl]-4-[[(6R)-trans-2-carb oxy-8-oxo-7-[(2,5- dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0]-oct -2-en-3- yl]methylthio]pyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 20)

l-[(S)-4-carboxy-4-hydroxybut-l-yl]-4-[[(6R)-trans-2-carb oxy-8-oxo-7-[(2,6- dichloropyridin-4-yl)thioacetamido]-5-thia-l-azabicyclo[4.2. 0]-oct-2-en-3- yl]methylthio]pyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 21)

l-[(R)-4-carboxy-4-hydroxybut-l-yl]-4-[[(6R)-trans-2-carb oxy-8-oxo-7-[(2,5- dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0]-oct -2-en-3-

yl]methylthio]pyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 22)

l-[(R)-4-carboxy-4-hydroxybut-l-yl]-4-[[(6R)-trans-2-carb oxy-8-oxo-7-[(2,6- dichloropyridin-4-yl)thioacetamido]-5-thia-l-azabicyclo[4.2. 0]-oct-2-en-3- yl]methylthio]pyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 23)

l-[(S)-3-carboxy-3-hydroxyprop-l-yl]-4-[[(6R)-trans-2-car boxy-8-oxo-7-[(2,5- dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0]-oct -2-en-3- yl]methylthio]pyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 24)

l-[(S)-3-carboxy-3-hydroxyprop-l-yl]-4-[[(6R)-trans-2-car boxy-8-oxo-7-[(2,6- dichloropyridin-4-yl)thioacetamido]-5-thia-l-azabicyclo[4.2. 0]-oct-2-en-3- yl]methylthio]pyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 25)

l-[2-carboxy-2-hydroxy ethyl]-4-[[(6R)-trans-2-carboxy-8-oxo-7-[(2,6- dichloropyridin-4-yl)thioacetamido]-5-thia-l-azabicyclo[4.2. 0]-oct-2-en-3- yl]methylthio]pyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 26)

l-[(S)-3-carboxy-3-hydroxy-2,2-dimethylprop-l-yl]-4-[[(6R)-t rans-2-carboxy-8-oxo-7- [(2,5-dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2. 0]-oct-2-en-3- yl]methylthio]pyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 27)

l-[(2R,3R)-3-carboxy-2,3-dihydroxyprop-l-yl]-4-[[(6R)-tra ns-2-carboxy-8-oxo-7-[[(2,5- dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0]-oct -2-en-3- yl]methylthio]pyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 28)

l-[(2,2-dimethyl-4-carboxydioxolan-5-yl)methyl]-4-[[(6R)- trans-2-carboxy-8-oxo-7- [(2,5-dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2. 0]-oct-2-en-3- yl]methylthio]pyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 29)

l-[2-carboxy-2-oxoeth-l-yl]-4-[[(6R)-trans-2-carboxy-8-ox o-7-[(2,5- dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0]-oct -2-en-3- yl]methylthio]pyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 30)

l-[2-carboxy-2-oxoeth-l-yl]-4-[[(6R)-trans-2-carboxy-8-ox o-7-[(2,6-dichloropyridin-4- yl)thioacetamido]-5-thia-l-azabicyclo[4.2.0]-oct-2-en-3-yl]m ethylthio]pyridinium

inner salt or a pharmaceutically acceptable salt thereof (compound of Example 31)

l-[3-carbomethoxy-2-oxoprop-l-yl]-4-[[(6R)-trans-2-carboxy-8 -oxo-7-[(2,5-

dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0]- oct-2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 32)

l-[4-carboxy-2-oxoprop-l-yl]-4-[[(6R)-trans-2-carboxy-8-o xo-7-[(2,5- dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0]-oct -2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 33)

l-[2,2-dimethyl-3-carboxy-3-oxoprop-l-yl]-4-[[(6R)-trans- 2-carboxy-8-oxo-7-[(2,5- dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0]-oct -2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 34)

l-[2-carboxy-2-fluoroeth-l-yl]-4-[[(6R)-trans-2-carboxy-8 -oxo-7-[(2,5- dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0]-oct -2-en-3- yl]methylthio]pyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 35)

l-[3-carboxy-3-fluoroprop-l-yl]-4-[[(6R)-trans-2-carboxy- 8-oxo-7-[(2,5- dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0]-oct -2-en-3-

yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 36)

l-[3-carboxy-3,3-difluoroprop-l-yl]-4-[[(6R)-trans-2-carb oxy-8-oxo-7-[(2,5- dichlorophenylthio)acetamidoj-5-thia-l-azabicyclo[4.2.0]-oct -2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 37)

l-[2-(carboxymethylthio)eth-l-yl]-4-[[(6R)-trans-2-carbox y-8-oxo-7-[(2,5- dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0]-oct -2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 38)

l-[3-(2-carboxyethylthio)-l-propyl]-4-[[(6R)-trans-2-carb oxy-8-oxo-7-[(2,5- dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0j-oct -2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 39)

l-[N-(carboxymethyl)-aminocarbonylmethylj-4-[[(6R)-trans- 2-carboxy-8-oxo-7- [(2,5-dichlorophenylthio)acetamido]-5-thia-l ,abicyclo[4.2.0]-oct-2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 40)

l-[N-(carboxymethyl)-aminocarbonylmethylj-4-[[(6R)-trans-2-c arboxy-8-oxo-7-

[(2,4,5-trichlorophenylthio)acetamido]-5-thia-l-azabicycl o[4.2.0J-oct-2-en-3-

yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 41)

l-[N-(carboxymethyl)-aminocarbonylmethylj-4-[[(6R)-trans- 2-carboxy-8-oxo-7- [(2,6-dichloropyridin-4-yl)thioacetamido]-5-thia-l-azabicycl o[4.2.0j-oct-2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 42)

1 - [N-(2-carboxyethy l)-aminocarbonylmethy l]-4- [ [ (6R)-trans-2-carboxy-8-oxo-7- [ (2,5- dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0]-oct -2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 43)

l-[N-(2-carboxyethyl)-aminocarbonylmethylj-4-[[(6R)-trans -2-carboxy-8-oxo-7-[(2,6- dichloropyridin-4-yl)thioacetamido]-5-thia-l-azabicyclo[4.2. 0]-oct-2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 44)

l-[N-(2-carboxyethyl-2-hydroxy)aminocarbonylmethylj-4-[[( 6R)-trans-2-carboxy-8- oxo-7-[(2,5-dichlorophenylthio)acetamidoj-5-thia-l-azabicycl o[4.2.0j-oct-2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 45)

l-[N-(l-carboxy-2-hydroxyeth-l-yl)aminocarbonylmethyl]-4-[[( 6R)-trans-2-carboxy- 8-oxo-7-[(2,5-dichlorophenylthio)acetamidoj-5-thia-l-azabicy clo[4.2.0]-oct-2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 46)

l-[N-(l-carboxy-2-hydroxyeth-l-yl)aminocarbonylmethyl]-4- [[(6R)-trans-2-carboxy- 8-oxo-7-[(2,6-dichloropyridin-4-yl)thioacetamido]-5-thia-l-a zabicyclo[4.2.0j-oct-2- en-3-yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 47)

l-[N-(l-carboxy-3-thiomethylprop-l-yl)aminocarbonylmethyl j-4-[[(6R)-trans-2- carboxy-8-oxo-7-[(2,5-dichlorophenylthio)acetamidoj-5-thia-l -azabicyclo[4.2.0j-oct- 2-en-3-yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 48)

l-[2-((S)-N-prolyl)-2-oxoeth-l-ylj-4-[[(6R)-trans-2-carbo xy-8-oxo-7-[(2,5- dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0]-oct -2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 49)

l-[N-(l-carboxy-2-phenylethyl)aminocarbonylmethylj-4-[[(6 R)-trans-2-carboxy-8- oxo-7-[(2,5-dichlorophenylthio)acetamido]-5-thia-l-azabicycl o[4.2.0]-oct-2-en-3-

yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof

(compound of Example 50)

l-[N-(l-carboxy-2-(4-hydroxyphenyl)ethyl)aminocarbonylmet hyl]-4-[[(6R)-trans-2-

carboxy-8-oxo-7-[(2,5-dichlorophenylthio)acetamido]-5-thi a-l-azabicyclo[4.2.0]-oct- 2-en-3-yl]methylthiojρyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 51)

l-[N-(l-carboxy-2-histidylethyl)aminocarbonylmethylj-4-[[ (6R)-trans-2-carboxy-8- oxo-7-[(2,5-dichlorophenylthio)acetamidoj-5-thia-l-azabicycl o[4.2.0]-oct-2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 52)

l-[N-(carboxymethyl)-l-aminocarbonyleth-l-ylj-4-[[(6R)-tr ans-2-carboxy-8-oxo-7- [(2,5-dichlorophenylthio)acetamidoj-5-thia-l-azabicyclo[4.2. 0j-oct-2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 53)

l-[N-methyl-N-(carboxymethyl)aminocarbonylmethylj-4-[[(6R )-trans-2-carboxy-8- oxo-7-[(2,5-dichlorophenylthio)acetamido]-5-thia-l-azabicycl o[4.2.0]-oct-2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 54)

l-(sulfomethyl)-4-[[(6R)-trans-2-carboxy-8-oxo-7-[(2,5- dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0]-oct -2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 55)

l-(2-sulfoeth-l-yl)-4-[[(6R)-trans-2-carboxy-8-oxo-7-[(2, 5- dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0j-oct -2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 56)

2,6-dimethyl-l-(2-sulfoeth-l-yl)-4-[[(6R)-trans-2-carboxy -8-oxo-7-[(2,5- dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0j-oct -2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 57)

2-ethyl-l-(2-sulfoethyl)-4-[[(6R)-trans-2-carboxy-8-oxo-7 -[(2,5- dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0]-oct -2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 58)

2,3-dimethyl-l-(2-sulfoethyl)-4-[[(6R)-trans-2-carboxy-8- oxo-7-[(2,5- dichlorophenylthio)acetamidoj-5-thia-l-azabicyclo[4.2.0j-oct -2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 59)

l-[N-(2-sulfoethyl)aminocarbonylmethylj-4-[[(6R)-trans-2-car boxy-8-oxo-7-[(2,5- dichlorophenylthio)acetamidoj-5-thia-l-azabicyclo[4.2.0J-oct -2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof

(compound of Example 60)

l-[N-(2-sulfoethyl)aminocarbonylmethyl]-4-[[(6R)-trans-2- carboxy-8-oxo-7-[(2,6- dichloropyridin-4-yl)thioacetamido]-5-thia-l-azabicyclo[4.2. 0]-oct-2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 61)

l-[N-(2-sulfoethyl)-2-aminocarbonylethylJ-4-[[(6R)-trans- 2-carboxy-8-oxo-7-[(2,5- dichlorophenylthio)acetamidoJ-5-thia-l-azabicyclo[4.2.0]-oct -2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 62)

l-[N-(2-sulfoethyl)-2-aminocarbonylethylj-4-[[(6R)-trans- 2-carboxy-8-oxo-7-[(2,6- dichloropyridin-4-yl)thioacetamido]-5-thia-l-azabicyclo[4.2. 0j-oct-2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 63)

1 -[3-(5-tetrazolylthio)prop-l -yl]-4-[ [ (6R)-trans-2-carboxy-8-oxo-7- [(2,5- dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0]-oct -2-en-3-

yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 64)

l-[N-(5-tetrazolyl)aminocarbonylmethyl]-4-[[(6R)-trans-2- carboxy-8-oxo-7-[(2,5- dichlorophenylthio)acetamidoj-5-thia-l-azabicyclo[4.2.0]-oct -2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 65)

l-[N-(5-tetrazolyl)-2-aminocarbonyleth-l-ylj-4-[[(6R)-tra ns-2-carboxy-8-oxo-7-[(2,5- dichlorophenylthio)acetamidoJ-5-thia-l-azabicyclo[4.2.0]-oct -2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 66)

l-[N-(5-tetrazolyl)-3-aminocarbonylprop-l-yl]-4-[[(6R)-tr ans-2-carboxy-8-oxo-7- [(2,5-dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2. 0]-oct-2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 67)

l-[2-carboxy-2-(hydroxyimino)eth-l-yl]-4-[[(6R)-trans-2-c arboxy-8-oxo-7-[(2,5- dichlorophenylthio)acetamidoj-5-thia-l-azabicyclo[4.2.0]-oct -2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 68)

l-[N-methyl-N-(3-carboxy-l-oxoprop-l-yl)-2-aminoeth-l-yl]-4- [[(6R)-trans-2- carboxy-8-oxo-7-[(2,5-dichlorophenylthio)acetamidoj-5-thia-l -azabicyclo[4.2.0J-oct- 2-en-3-yl]methylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 69)

l-[l-carboxy-2-hydroxy-2-(2-thienyl)eth-l-ylj-4-[[(6R)-tr ans-2-carboxy-8-oxo-7-[(2,5- dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0J-oct -2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 70)

l-[N-(2-carboxy-l-oxoeth-l-yl)-2-aminoeth-l-yl]-4-[[(6R)- trans-2-carboxy-8-oxo-7- [(2,5-dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2. 0J-oct-2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 71); and

l-[2-(2-sulfoethylthio)eth-l-yl]-4-[[(6R)-trans-2-carboxy -8-oxo-7-[(2,5- dichlorophenylthio)acetamidoj-5-thia-l-azabicyclo[4.2.0J-oct -2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 72).

The more preferred individual compounds of the present invention, all of

which have a MIC < 8 μg/mL are listed below:

l-[2-carboxy-2-propen-l-yl]-4-[[(6R)-trans-2-carboxy-8-oxo-7 -[(2,5- dichlorophenylthio)acetamidoj-5-thia-l-azabicyclo[4.2.0]-oct -2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 1)

l-[2-carboxy-2-hydroxy ethylj-4-[[(6R)-trans-2-carboxy-8-oxo-7-[(2,5- dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0]-oct -2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 2)

l-[(R)-l-carboxy-l-ethylJ-4-[[(6R)-trans-2-carboxy-8-oxo- 7-[(2,5- dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0]-oct -2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 6)

l-[(S)-l-carboxy-3-(methylsulfinyl)prop-l-ylj-4-[[(6R)-tr ans-2-carboxy-8-oxo-7-[(2,5- dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0j-oct -2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 11)

l-[(S)-l-carboxy-3-(methylsulfonyl)prop-l-yl]-4-[[(6R)-tr ans-2-carboxy-8-oxo-7-[(2,5- dichlorophenylthio)acetamidoj-5-thia-l-azabicyclo[4.2.0]-oct -2-en-3-

yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof

(compound of Example 12)

l-[(S)-3-aminosulfonyl-l-carboxyprop-l-ylj-4-[[(6R)-trans -2-carboxy-8-oxo-7-[(2,5- dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0]-oct -2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 13)

l-[(S)-5-carboxy-5-hydroxypent-l-yl]-4-[[(6R)-trans-2-car boxy-8-oxo-7-[(2,5- dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0]-oct -2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 18)

l-[(S)-4-carboxy-4-hydroxybut-l-ylj-4-[[(6R)-trans-2-carb oxy-8-oxo-7-[(2,5- dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0]-oct -2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 20)

l-[(R)-4-carboxy-4-hydroxybut-l-yl]-4-[[(6R)-trans-2-carb oxy-8-oxo-7-[(2,5- dichlorophenylthio)acetamidoj-5-thia-l-azabicyclo[4.2.0J-oct -2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 22)

l-[(S)-3-carboxy-3-hydroxyprop-l-ylj-4-[[(6R)-trans-2-carbox y-8-oxo-7-[(2,6- dichloropyridin-4-yl)thioacetamidoJ-5-thia-l-azabicyclo[4.2. 0]-oct-2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 25)

l-[2-carboxy-2-hydroxy ethyl]-4-[[(6R)-trans-2-carboxy-8-oxo-7-[(2,6- dichloropyridin-4-yl)thioacetamido]-5-thia-l-azabicyclo[4.2. 0]-oct-2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 26)

l-[2-carboxy-2-oxoeth-l-ylJ-4-[[(6R)-trans-2-carboxy-8-ox o-7-[(2,5- dichlorophenylthio)acetamidoj-5-thia-l-azabicyclo[4.2.0]-oct -2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 30)

1 - [2-carboxy-2-oxoeth- 1 -y 1J-4- [ [ (6R)-trans-2-carboxy-8-oxo-7- [ (2,6-dichloropy ridin-4- yl)thioacetamido]-5-thia-l-azabicyclo[4.2.0]-oct-2-en-3-yljm ethylthio]pyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 31)

l-[3-carbomethoxy-2-oxoprop-l-ylJ-4-[[(6R)-trans-2-carbox y-8-oxo-7-[(2,5- dichlorophenylthio)acetamidoJ-5-thia-l-azabicyclo[4.2.0J-oct -2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 32)

l-[2-(carboxymethylthio)eth-l-ylj-4-[[(6R)-trans-2-carboxy-8 -oxo-7-[(2,5- dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0]-oct -2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 38)

l-[3-(2-carboxyethylthio)-l-propylj-4-[[(6R)-trans-2-carb oxy-8-oxo-7-[(2,5- dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0]-oct -2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 39)

l-[N-(carboxymethyl)-aminocarbonylmethyl]-4-[[(6R)-trans- 2-carboxy-8-oxo-7- [(2,5-dichlorophenylthio)acetamidoj-5-thia-l-azabicyclo[4.2. 0J-oct-2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 40)

l-[N-(carboxymethyl)-aminocarbonylmethylJ-4-[[(6R)-trans- 2-carboxy-8-oxo-7- [ (2,6-dichloropy ridin-4-y 1) thioacetamido] -5-thia- 1 -azabicyclo [4.2.0] -oct-2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 42)

l-[N-(2-carboxyethyl)-aminocarbonylmethylj-4-[[(6R)-trans -2-carboxy-8-oxo-7-[(2,6- dichloropyridin-4-yl)thioacetamido]-5-thia-l-azabicyclo[4.2. 0]-oct-2-en-3-

yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 44)

l-[N-(l-carboxy-2-phenylethyl)aminocarbonylmethyl]-4-[[(6 R)-trans-2-carboxy-8- oxo-7-[(2,5-dichlorophenylthio)acetamido]-5-thia-l-azabicycl o[4.2.0]-oct-2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 50)

1 -(sulf omethy l)-4- [ [ (6R)-trans-2-carboxy-8-oxo-7-[ (2,5- dichlorophenylthio)acetamidoj-5-thia-l-azabicyclo[4.2.0J-oct -2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 55)

l-(2-sulfoeth-l-yl)-4-[[(6R)-trans-2-carboxy-8-oxo-7-[(2, 5- dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0]-oct -2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 56)

2,6-dimethyl-l-(2-sulfoeth-l-yl)-4-[[(6R)-trans-2-carboxy -8-oxo-7-[(2,5- dichlorophenylthio)acetamidoJ-5-thia-l-azabicyclo[4.2.0j-oct -2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 57)

2-ethyl-l-(2-sulfoethyl)-4-[[(6R)-trans-2-carboxy-8-oxo-7-[( 2,5- dichlorophenylthio)acetamidoj-5-thia-l-azabicyclo[4.2.0]-oct -2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 58)

2,3-dimethyl-l-(2-sulfoethyl)-4-[[(6R)-trans-2-carboxy-8- oxo-7-[(2,5- dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0j-oct -2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 59)

l-[N-(2-sulfoethyl)aminocarbonylmethyl]-4-[[(6R)-trans-2- carboxy-8-oxo-7-[(2,5- dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0]-oct -2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 60)

l-[N-(2-sulfoethyl)-2-aminocarbonylethyl]-4-[[(6R)-trans- 2-carboxy-8-oxo-7-[(2,5- dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0]-oct -2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 62)

l-[N-(2-sulfoethyl)-2-aminocarbonylethylj-4-[[(6R)-trans- 2-carboxy-8-oxo-7-[(2,6- dichloropyridin-4-yl)thioacetamidoj-5-thia-l-azabicyclo[4.2. 0J-oct-2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 63)

l-[3-(5-tetrazolylthio)prop-l-yl]-4-[[(6R)-trans-2-carboxy-8 -oxo-7-[(2,5- dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0j-oct -2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 64)

l-[N-(5-tetrazolyl)aminocarbonylmethyl]-4-[[(6R)-trans-2- carboxy-8-oxo-7-[(2,5- dichlorophenylthio)acetamidoJ-5-thia-l-azabicyclo[4.2.0]-oct -2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 65)

l-[N-(5-tetrazolyl)-2-aminocarbonyleth-l-yl]-4-[[(6R)-tra ns-2-carboxy-8-oxo-7-[(2,5- dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0]-oct -2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 66)

l-[N-(5-tetrazolyl)-3-aminocarbonylprop-l-ylj-4-[[(6R)-tr ans-2-carboxy-8-oxo-7- [(2,5-dichlorophenylthio)acetamidoj-5-thia-l-azabicyclo[4.2. 0]-oct-2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 67)

l-[2-carboxy-2-(hydroxyimino)eth-l-yl]-4-[[(6R)-trans-2-c arboxy-8-oxo-7-[(2,5- dichlorophenylthio)acetamidoJ-5-thia-l-azabicyclo[4.2.0j-oct -2-en-3-

yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof

(compound of Example 68)

l-[l-carboxy-2-hydroxy-2-(2-thienyl)eth-l-yl]-4-[[(6R)-tr ans-2-carboxy-8-oxo-7-[(2,5- dichlorophenylthio)acetamidoj-5-thia-l-azabicyclo[4.2.0]-oct -2-en-3-

yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 70); and

l-[2-(2-sulfoethylthio)eth-l-yl]-4-[[(6R)-trans-2-carboxy -8-oxo-7-[(2,5- dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0j-oct -2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 72).

The most preferred individual compounds of the present invention are listed below:

l-[2-carboxy-2-propen-l-yl]-4-[[(6R)-trans-2-carboxy-8-ox o-7-[(2,5- dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0]-oct -2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 1)

l-[2-carboxy-2-hydroxy ethyl]-4-[[(6R)-trans-2-carboxy-8-oxo-7-[(2,5- dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0J-oct -2-en-3-

yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 2)

l-[(S)-3-aminosulfonyl-l-carboxyprop-l-ylj-4-[[(6R)-trans -2-carboxy-8-oxo-7-[(2,5- dichlorophenylthio)acetamidoJ-5-thia-l-azabicyclo[4.2.0]-oct -2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 13)

l-[(S)-5-carboxy-5-hydroxypent-l-yl]-4-[[(6R)-trans-2-car boxy-8-oxo-7-[(2,5- dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0J-oct -2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 18)

l-[(S)-4-carboxy-4-hydroxybut-l-yl]-4-[[(6R)-trans-2-carb oxy-8-oxo-7-[(2,5- dichlorophenylthio)acetamidoj-5-thia-l-azabicyclo[4.2.0]-oct -2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 20)

l-[(R)-4-carboxy-4-hydroxybut-l-yl]-4-[[(6R)-trans-2-carb oxy-8-oxo-7-[(2,5- dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0j-oct -2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 22)

l-[(S)-3-carboxy-3-hydroxyprop-l-ylj-4-[[(6R)-trans-2-carbox y-8-oxo-7-[(2,6- dichloropyridin-4-yl)thioacetamidoj-5-thia-l-azabicyclo[4.2. 0J-oct-2-en-3-

yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 25)

l-[2-carboxy-2-oxoeth-l-yl]-4-[[(6R)-trans-2-carboxy-8-ox o-7-[(2,5-

dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0J- oct-2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 30)

l-[2-carboxy-2-oxoeth-l-yl]-4-[[(6R)-trans-2-carboxy-8-ox o-7-[(2,6-dichloropyridin-4- yl)thioacetamidoj-5-thia-l-azabicyclo[4.2.0J-oct-2-en-3-yl]m ethylthioJpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 31)

l-[2-(carboxymethylthio)eth-l-yl]-4-[[(6R)-trans-2-carbox y-8-oxo-7-[(2,5- dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0J-oct -2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 38)

l-[3-(2-carboxyethylthio)-l-propylJ-4-[[(6R)-trans-2-carb oxy-8-oxo-7-[(2,5- dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0]-oct -2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 39)

l-[N-(2-carboxyethyl)-aminocarbonylmethyl]-4-[[(6R)-trans-2- carboxy-8-oxo-7-[(2,6- dichloropyridin-4-yl)thioacetamido]-5-thia-l-azabicyclo[4.2. 0]-oct-2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 44)

l-(2-sulfoeth-l-yl)-4-[[(6R)-trans-2-carboxy-8-oxo-7-[(2, 5- dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0j-oct -2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 56)

2,6-dimethyl-l-(2-sulfoeth-l-yl)-4-[[(6R)-trans-2-carboxy -8-oxo-7-[(2,5- dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0J-oct -2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 57)

2,3-dimethyl-l-(2-sulfoethyl)-4-[[(6R)-trans-2-carboxy-8- oxo-7-[(2,5- dichlorophenylthio)acetamidoJ-5-thia-l-azabicyclo[4.2.0J-oct -2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 59)

l-[N-(2-sulfoethyl)-2-aminocarbonylethyl]-4-[[(6R)-trans- 2-carboxy-8-oxo-7-[(2,5- dichlorophenylthio)acetamidoj-5-thia-l-azabicyclo[4.2.0J-oct -2-en-3-

yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 62)

l-[N-(2-sulfoethyl)-2-aminocarbonylethyl]-4-[[(6R)-trans- 2-carboxy-8-oxo-7-[(2,6- dichloropyridin-4-yl)thioacetamidoj-5-thia-l-azabicyclo[4.2. 0]-oct-2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 63)

l-[3-(5-tetrazolylthio)prop-l-yIJ-4-[[(6R)-trans-2-carbox y-8-oxo-7-[(2,5- dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0J-oct -2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 64)

l-[N-(5-tetrazolyl)aminocarbonylmethyl]-4-[[(6R)-trans-2- carboxy-8-oxo-7-[(2,5- dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0]-oct -2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 65)

l-[N-(5-tetrazolyl)-2-aminocarbonyleth-l-yl]-4-[[(6R)-tra ns-2-carboxy-8-oxo-7-[(2,5- dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0J-oct -2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 66)

l-[N-(5-tetrazolyl)-3-aminocarbonylprop-l-yl]-4-[[(6R)-trans -2-carboxy-8-oxo-7- [(2,5-dichlorophenylthio)acetamidoj-5-thia-l-azabicyclo[4.2. 0J-oct-2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 67); and

l-[2-carboxy-2-(hydroxyimino)eth-l-ylj-4-[[(6R)-trans-2-c arboxy-8-oxo-7-[(2,5- dichlorophenylthio)acetamido]-5-thia-l-azabicyclo[4.2.0]-oct -2-en-3- yljmethylthiojpyridinium inner salt or a pharmaceutically acceptable salt thereof (compound of Example 68).

The compounds of the present invention can be made by conventional methods. Two suitable procedures are summarized in the following reaction scheme:

Ar . OH

ArSH

Y

R = ester protecting group such as diphenylmethyl (DP ) or para-methoxybenzyl (PMB)

Method 1

Method 2

UI

To elaborate on the above process, thiol VII is converted into the arylthioacetic acid derivative VI by treatment with bromoacetic acid under basic conditions (e.g. aqueous sodium or potassium hydroxide). The reaction temperature for this step is typically between 20 °C and 100 °C. Starting thiol YJI is commercially available or can be prepared according to known literature procedures. Following acidification of the reaction mixture, the product VI is

typically isolated by crystallization or, if necessary, it can be purified by chromatography.

Arylthioacetic acid VI is then coupled with a suitable cephem intermediate having a 3-substituent leaving group. For example, the leaving group may be acetoxy or halo. In the preferred embodiment illustrated by the reaction scheme, the cephem intermediate is the 3-chloro cephem V, but other suitable cephem intermediates with equivalent leaving groups at the 3-position could also be employed. The cephem intermediate V may be acylated with VJ or a reactive derivative thereof by conventional acylation procedures well-known in the cephalosporin art to give N-acylated intermediate IV. In addition to using the free arylthioacetic acid, e.g. with a suitable condensing agent such as dicyclohexylcarbodiimide, acylating agent VI may also be employed in the form of equivalent acylating derivatives such as an acid anhydride, mixed anhydride, activated ester, or acid halide. The cephem intermediate preferably has the carboxyl group protected by a conventional carboxyl-protecting group which can be readily removed. Examples of such protecting groups are discussed above and include benzyl, 4-nitrobenzyl, 4-methoxybenzyl, diphenylmethyl, allyl, and the like. Other examples of suitable protecting groups are disclosed in Protective Groups in Organic Synthesis. Theodora W. Greene (John Wiley & Sons, 1981), Chapter 5. In one embodiment, intermediate V may be acylated with acid Yl in the presence of dicyclohexylcarbodiimide and in an inert solvent such as tetrahydrofuran or dichloromethane. The reaction temperature is typically between -20 °C and 50 °C. Upon completion of the reaction, insoluble material is

removed by filtration, the filtrate is concentrated, and the residue is treated with a relatively non-polar solvent such as diethyl ether or ethyl acetate resulting in precipitation of the desired product. Alternatively, acid VJ may be converted to the corresponding acid chloride, for example by treatment with thionyl chloride with or without a solvent such as dichloromethane, followed by coupling with cephem amine V in the presence of a base such as triethylamine or N- methylmorpholine to give intermediate IV. Cephem IV is typically isolated after aqueous work-up and evaporation of volatile solvents followed by trituration of the compound with a relatively non-polar solvent such as diethyl ether or ethyl acetate. This intermediate may be used in the next reaction step as the X = chloride derivative, or can be converted to the X = bromide or X = iodide derivative by treatment with the appropriate metal halide in a solvent such as acetone.

Conversion of cephem IV to the target quaternary cephems I is accomplished by two different methods. One method for preparation of I entails displacement of an appropriate 3-substituent leaving group with 4- mercaptopyridine followed by quaternization of the pyridyl nitrogen, and then deprotection of the cephem carboxylate ester. For example, cephem IY (X = Cl) is treated with optionally substituted 4-mercaptopyridine and sodium iodide in a one-pot reaction to give intermediate fl. Reaction of thiopyridyl derivative H with a reactive alkylating agent provides the quaternary cephem intermediate T. Examples of alkylating agents are α-halocarbonyl derivatives such as N- substituted haloacetamides. The alkylation reaction is carried out in an inert

solvent such as acetone, dimethylformamide, or tetrahydrofuran and is run at temperatures between -20 °C and 100 °C Removal of the cephem carboxylate ester protecting group to give I is then accomplished under acidic conditions. For example, when R is diphenylmethyl or 4-methoxybenzyl, I is obtained upon treatment of i_ with trifluoroacetic acid neat or in an inert solvent such as methylene chloride. A reagent such as anisole may also be employed to scavenge the liberated ester protecting group. The reaction temperature is usually at or below room temperature. The deprotection may also be carried out by treatment with other protic acids such as hydrochloric acid in a solvent such as methanol. The final product is typically isolated by precipitation or crystallization. In some cases, cephem I is purified by column chromatography, for example on reversed- phase adsorbent.

In a second method of preparing quaternary cephems I, intermediate IY is deprotected under acidic conditions, followed by reaction of the resulting intermediate IV with a thiopyridone derivative ffl. For example, when R is diphenylmethyl or 4-methoxybenzyl, cephem acid IV is obtained upon treatment of IY with trifluoroacetic acid neat or in an inert solvent such as methylene chloride. A reagent such as anisole may also be employed to scavenge the liberated ester protecting group. The reaction temperature is usually at or below room temperature. The deprotection may also be carried out by treatment with other protic acids such as hydrochloric acid in a solvent such as methanol. The final product is typically isolated by precipitation or crystallization. Reaction of IV with a thiopyridone derivative III in a solvent such as

dimethylformamide, dimethyl sulfoxide, ethanol, methanol, or other appropriate solvents at a temperature between -20 °C and 100 °C affords target quaternary cephem I. The final product is isolated as described above.

Thiopyridones fll are typically prepared according to a method analogous to that described in T. Takahashi et al., European Patent Application No. 209751 and in I.E. El-Kholy et al., J. Heterocyclic Chem. Vol. 11, p. 487 (1974). This procedure entails reaction of 4-thiopyrone (European Patent No. 209751) with an appropriate primary amine in a solvent such as aqueous methanol or ethanol at a temperature ranging between 0 °C and 78 ° C. The primary amine may be in the form of a zwitterion in examples where there is a free acid group present in the molecule. In these cases, a base such as sodium hydroxide, sodium bicarbonate, or pyridine is added to form the free amine in situ. The product may be isolated as its sodium salt by evaporation of volatile solvents, followed by trituration with a solvent such as diethyl ether or ethyl acetate. Alternatively, the reaction mixture may be acidified and extracted with an organic solvent to afford the product as the free carboxylic acid. If the carboxylate group is protected as an ester, the amine may be free or present as an acid salt. In the latter case, a base such as sodium hydroxide, sodium bicarbonate, or pyridine is added to form the free amine in situ. The product is typically isolated by precipitation or by reverse phase column chromatography following removal of volatile solvents.

The thiopyridone derivatives of formula III are another aspect of the present invention. The preferred R ¹ , R ² , R ³ , R ⁹ and R ¹⁰ substituents of

derivatives III are as disclosed above in connection with the end-products of formula I.

It will be understood that where the substituent groups used in the above

reactions contain certain reaction-sensitive functional groups such as carboxylate groups which might result in undesirable side-reactions, such groups may be protected by conventional protecting groups known to those skilled in the art. Suitable protecting groups and methods for their removal are illustrated, for example, in Protective Groups in Organic Synthesis. Theodora W. Greene (John Wiley & Sons, 1991). It is intended that such "protected" intermediates and end- products are included within the scope of the present disclosure and claims.

The desired end-product of formula I may be recovered either as the zwitterion or in the form of a pharmaceutically acceptable acid addition salt, e.g. by addition of the appropriate acid such as HCl, HI or methanesulfonic acid to the

zwitterion. Compounds of formula I where Rβ is hydrogen, an anionic charge or a pharmaceutically acceptable salt thereof may be converted by conventional

procedures to a corresponding compound where R^ is a physiologically hydrolyzable ester group.

It will be appreciated that certain products within the scope of formula I may have a C-3 substituent group which can result in formation of optical isomers. It is intended that the present invention include within its scope all

such optical isomers as well as epimeric mixtures thereof, i.e. R- or S- or racemic forms.

The novel cephalosporin derivatives of general formula I (including the

IA compounds) wherein R^ is hydrogen, an anionic charge or a physiologically hydrolyzable carboxyl-protecting group, or the pharmaceutically acceptable salts or prodrugs thereof, are potent antibiotics active against many gram-positive bacteria. While they may be used, for example, as animal feed additives for promotion of growth, as preservatives for food, as bactericides in industrial applications, for example in waterbased paint and in the white water of paper

mills to inhibit the growth of harmful bacteria, and as disinfectants for destroying or inhibiting the growth of harmful bacteria on medical and dental equipment, they are especially useful in the treatment of infectious disease in humans and other animals caused by the gram-positive bacteria sensitive to the new derivatives. Because of their excellent activity against MRSA organisms, they are particularly useful in the treatment of infections resulting from such bacteria.

The pharmaceutically active compounds of this invention may be used alone or formulated as pharmaceutical compositions comprising, in addition to the active cephem ingredient, a pharmaceutically acceptable carrier or diluent. The compounds may be administered by a variety of means, for example, orally, topically or parenterally (intravenous or intramuscular injection). The pharmaceutical compositions may be in solid form such as capsules, tablets, powders, etc. or in liquid form such as solutions, suspensions or emulsions.

Compositions for injection, the preferred route of delivery, may be prepared in unit dose form in ampules or in multidose containers and may contain additives such as suspending, stabilizing and dispersing agents. The compositions may be in ready-to-use form or in powder form for reconstitution at the time of delivery with a suitable vehicle such as sterile water.

The dosage to be administered depends, to a large extent, on the particular compound being used, the particular composition formulated, the route of administration, the nature and condition of the host and the particular situs and organism being treated. Selection of the particular preferred dosage and route of application, then, is left to the discretion of the physician or veterinarian. In general, however, the compounds may be administered parenterally or orally to mammalian hosts in an amount of from about 50 mg/day to about 20 g/day. Administration is generally carried out in divided doses, e.g., three to four times a day, analogous to dosing with a cephalosporin such as cefotaxime.

IN VITRO ACTIVITY

Samples of the compounds prepared below in Examples 1 - 72 after solution in water and dilution with Nutrient Broth were found to exhibit the following ranges of Minimum Inhibitory Concentrations (MIC) versus the indicated microorganisms as determined by tube dilution. The MICs were determined using a broth micro dilution assay in accordance with that

recommended by the National Committee for Clinical Laboratory Standards (NCCLS). Mueller-Hinton medium was used except for Streptococci which was tested in Todd Hewitt broth. The final bacterial inoculate contained

approximately 5 x 10^ cfu/ml and the plates were incubated at 35°C for 18 hours

in ambient air (Streptococci in 5% CO2). The MIC was defined as the lowest drug

concentration that prevented visible growth.

Microorganism MIC range in ug/ml

S. aureus methicillin resistant A27223 1 - 8

S. pneumoniae A9585 0.0005 - 2

S. pyogenes A9604 0.0005 - 2

E. faecalis A20688 0.25 - 8

E. faecium A24885 0.5 - 16 5. aureus A9537, penicillinase negative 0.0005 - 0.125

S. aureus A15090, penicillinase positive 0.03 - 1

S. epidermidis A24548 0.001 - 1

S. epidermidis A25783, methicillin resistant 0.015 - 2

S. hemolyticus A21638 0.015 - 8 5. hemolyticus A27235, methicillin resistant 0.25 - 32

IN VIVO ACTIVITY

The in vivo therapeutic efficacy of the compounds prepared in Examples 1 - 72 below after intramuscular injection to mice experimentally infected with the representative MRSA strain A27223 was also measured.

The determination of the effectiveness of antimicrobial agents in Staphylococcus aureus systemic infection in mice

Organisms: The test organism, MRSA strain A27223 used to generate systemic infection in mice, is grown on two large Brain Heart Infusion Agar plates. On each plate, 0.5 ml of frozen stock culture is plated out. Plates are then incubated

for 18 hours at 30°C. The next day each plate is washed with 20 ml of Brain Heart

Infusion Broth and then pooled together. A microscopic direct count of microorganism is done using a 1:1000 dilution of plate wash. After a direct count is obtained, the number of organisms per milliliter is calculated. The count is adjusted to the desired amount of inoculum by diluting in 4% hog mucin. The

desired challenge (amount of organisms given to mice) is 2.4 x 10^ cfu/0.5 ml /mouse for MRSA strain A27223. The mice are infected intraperitoneally with 0.5 ml of challenge. Ten non-treated infected mice are used as controls.

Mice: Mice used are male ICR mice. The average weight of the animals is from 20 to 26 grams.

Drug preparation and treatment: Compounds are tested at 4 dose levels, (25, 6.25, 1.56, and 0.39 mg/kg) and prepared in 5% cremophor, unless otherwise specified. Vancomycin is used as the control compound, and is dosed at 6.25, 1.56, 0.39, and 0.098 mg/kg. It is prepared in 0.1M phosphate buffer. There are five infected mice per dose level, and they are treated with 0.2 ml of the test compound, preferably by intramuscular injection. Treatment begins 15 minutes and 2 hours

post-infection.

Test duration: A PD50 (the dose of drug given which protects 50% of mice from

mortality) runs for 5 days. During this time, mortality of mice are checked every day and deaths are recorded. The cumulative mortality at each dose level is used to calculate a PD50 value for each compound. Surviving mice are sacrificed at

the end of day 5 by CO2 inhalation.

Calculation: Actual calculation of PD50 is performed with a computer program

using the Spearman-Karber procedure.

Results: The in vivo efficacy, expressed as the PD50 value, ranged from about 0.8

to 22 mg/kg (for certain compounds, more than one test was carried out; the indicated range is for at least one test result when multiple tests were done).

ILLUSTRATIVE EXAMPLES

The following examples illustrate the invention, but are not intended as a limitation thereof. The abbreviations used in the examples are conventional abbreviations well-known to those skilled in the art. Some of the abbreviations used are as follows:

In the following examples, all temperatures are given in degrees

Centigrade. Melting points were determined on an electrothermal apparatus and

are not corrected. Proton and carbon-13 nuclear magnetic resonance (*H and ¹³ C NMR) spectra were recorded on a Bruker AM-300 or a Varian Gemini 300 spectrometer. All spectra were determined in CDCI ₃ , DMSO-d ₆ , CD ₃ OD, or D ₂ O

unless otherwise indicated. Chemical shifts are reported in δ units relative to

tetramethylsilane (TMS) or a reference solvent peak and interproton coupling constants are reported in Hertz (Hz). Splitting patterns are designated as follows: s, singlet; d, doublet; t, triplet; q, quartet; m, multiple.; br, broad peak; dd, doublet of doublets; dt, doublet of triplets; and app d, apparent doublet, etc. Infrared spectra were determined on a Perkin-Elmer 1800 FT-IR spectrometer from 4(

cm ^-1 to 400 cm ^-1 , calibrated to 1601 cm ^-1 absorption of a polystyrene film, and are

reported in reciprocal centimeters (cm ^-1 ). Mass spectra were recorded on a Kratos MS-50 or a Finnegan 4500 instrument utilizing direct chemical ionization (DCI,

isobutene), fast atom bombardment (FAB), or electron ion spray (ESI). Ultraviolet spectra were determined on a Hewlett Packard 8452 diode array spectrophotometer in the solvent indicated.

Analytical thin-layer chromatography (TLC) was carried out on precoated silica gel plates (60F-254) and visualized using UV light, iodine vapors, and /or staining by heating with methanolic phosphomolybdic acid. Column chromatography, also referred to as flash chromatography, was performed in a

glass column using finely divided silica gel at pressures somewhat above atmospheric pressure with the indicated solvents. Reversed-phase analytical thin-layer chromatography was carried out on precoated reverse phase plates and visualized using UV light or iodine vapors. Reversed-phase column chromatography was performed in a glass column using Baker Octadecyl (Ciβ), 40

μm.

Example 1

l-f2-carboxy-2-propen-l-ylj-4-fr(6R -trans-2-carboxy-8-oxo-7-r(2.5- dichlorophenylthio)acetamidoj-5-thia-l-azabicyclo[4.2.01-oct -2-en-3- yljmethylthiojpyridinium bromide

A. 2.5-Dichlorophenylthioacetic acid

A mixture of 2,5-dichlorothiophenol (10.3 g, 57.5 mmol) and bromoacetic acid (8.03 g, 57.8 mmol) in water (225 mL) was treated with 10 N NaOH (13 mL, 130 mmol) and the mixture was heated at 100 °C for 1 h. The reaction mixture was then cooled to 0 °C and acidified to pH 1 with 6N HCl. The product precipitated and was collected by filtration to give 13.0 g (95% yield) of 2,5-

dichlorophenylthioacetic acid as white crystals, m.p. 118 °C. H NMR (300 MHz, CDC1 ₃ ) δ 3.74 (s, 2 H), 7.15 (dd, J=2, 9 Hz, 1 H), 7.32 (d, J=9 Hz, 1 H), 7.36 (d, J=2 Hz, 1

H). Anal. Calcd. for C ₈ H ₆ 0 ₂ SCl ₂ : C, 40.53; H, 2.55. Found: C, 40.46; H, 2.64.

B. (6R)-trans-3-ChIoromethyl-7-f(2,5-dichlorophenyl)thioacet-am idol-8-oxo-g- thia-l-azabicyclol4.2.01oct-2-ene-2-carboxylate. diphenylmethyl ester

Method a: A solution of 2,5-dichlorophenylthioacetic acid (13.0 g, 54.9 mmol) in methylene chloride (55 mL) and thionyl chloride (10 mL, 137 mmol) was heated at reflux for 3 h. The reaction mixture was allowed to cool to room temperature and was concentrated in vacuo. The residue was evaporated two times from toluene to give 14 g of 2,5-dichlorophenylthioacetyl chloride (100% yield) as a

slightly colored product which was used in the next step without purification. *H NMR (300 MHz, CDCI ₃ ) δ 4.13 (s, 2 H), 7.22 (dd, J=2, 9 Hz, 1 H), 7.35 (d, J=9 Hz, 1 H),

7.39 (d, J=2 Hz, 1 H).

(6R)-trans-3-Chloromethyl-7-amino-8-oxo-5-thia-l-azabicyc lo-[4.2.0Joct-2-

ene-2-carboxylate, diphenylmethyl ester, HCl salt was stirred in a biphasic mixture of EtOAc and saturated NaHCθ ₃ for 0.5 h. The layers were separated,

and the organic solution was dried over anhydrous MgSO_ι, filtered, and

concentrated to dryness. The free base (9.15 g, 22.0 mmol) was dissolved in THF (200 mL), cooled to 0 °C, and treated with N-methylmorpholine (3.34 g, 33.0 mmol) and 2,5-dichlorophenylthioacetyl chloride (6.75 g, 26.4 mmol). The reaction mixture was stirred for 1 h at 0 °C, diluted with EtOAc (1000 mL) and washed with water (1000 mL) and brine (100 mL). The organic solution was then dried (MgSO-i) and the solvents were evaporated in vacuo. The residue was

stirred with ether (100 mL). The product solidified and was collected by filtration to give 12.0 g (86% yield) of (6R)-trans-3-chloromethyl-7-[(2,5-dichlorophenyl)- thioacetamidoj-8-oxo-5-thia-l-azabicyclo[4.2.0]oct-2-ene-2-c arboxylate,

diphenylmethyl ester, m.p. 120 °C. H NMR (300 MHz, CDC1 ₃ ) δ 3.43 (d, J=18 Hz,

1 H), 3.59 (d, J=18 Hz, 1 H), 3.69 (d, J=17 Hz, 1 H), 3.79 (d, J=17 Hz, 1 H), 4.36 (d, J=12 Hz, 1 H), 4.41 (d, J=12 Hz, 1 H), 4.98 (d, J=5 Hz, 1 H), 5.81 (dd, J=5, 9 Hz, 1 H), 6.98 (s, 1 H), 7.14-7.44 (m, 14 H). Anal. Calcd for C ₂₉ H ₂₃ N ₂ O ₄ S ₂ CI ₃ : C, 54.94; H, 3.66; N, 4.42. Found: C, 55.18; H, 3.84; N, 4.62.

Method b: (6R)-trans-3-Chloromethyl-7-amino-8-oxo-5-thia-l-azabicyclo[ 4.2.0joct- 2-ene-2-carboxylate, diphenylmethyl ester, HCl salt (Otsuka, 248 g, 0.55 mol) was treated with NaHCθ ₃ (56 g, 0.66 mol) in water (1.6 L) at 0 °C The mixture was

stirred at 0 °C for 0.5 h and then CH2CI2 (1.5 L) was added. The biphasic mixture

was filtered through Celite and the Celite pad was washed with CH ₂ CI ₂ (2 L total).

The layers were separated and the organic solution was dried over anhydrous MgSθ ₄ , filtered, and concentrated to a volume of ca. 2 L. The free amine solution

was then added to a mixture of 2,5-dichlorothio-phenylacetic acid (130 g, 0.55

mol) and dicyclohexylcarbodiimide (144 g, 0.70 mol) in THF (1 L) at room temperature. The reaction mixture was stirred for 2.5 h and then was filtered through Celite, washing the Celite pad with several portions of acetone. The filtrate was concentrated in vacuo to give a solid mass. The solid was slurried in Et ₂ θ and then collected by filtration, washing the solid with several portions of

Et ₂ θ. The solid was dried under high vacuum over P2O5 to give 268 g (77% yield)

of (6R)-trans-3-chloromethyl-7-[(2,5-dichlorophenyl)thioacetami do]-8-oxo-5-thia- l-azabicyclo[4.2.0]oct-2-ene-2-carboxylate, diphenylmethyl ester (see above for analytical data).

C (6R)-tτans-3-(4-Pyridylthiomethyl)-7-K2,5-dichlorophenvl)-t hioacetamidol- 8-oxo-5-thia-l-azabicyclo[4.2.01oct-2-ene-2-carboxylate, diphenylmethyl ester

A solution of (6R)-trans-3-chloromethyl-7-[(2,5-dichlorophenyl)- thioacetamidoj-8-oxo-5-thia-l-azabicyclo[4.2.0]oct-2-ene-2-c arboxylate, diphenylmethyl ester (6.00 g, 9.46 mmol) in acetone (100 mL) was treated with sodium iodide (4.26 g, 28.4 mmol). The mixture was stirred at 20 °C for 3 h and then condensed under reduced pressure to a volume of 50 mL. The concentrated solution was diluted with EtOAc (200 mL) and washed with ice water (100 mL). The organic solution was washed with saturated NaHSθ ₄ (20 mL), dried

(MgSθ ₄ ), and evaporated under reduced pressure. The residue was stirred with

ether (30 mL). The product solidified and was collected by filtration to give 6.40 g of (6R)-trans-3-iodomethyl-7-[(2,5-dichlorophenyl)thio-acetamid o]-8-oxo-5-thia-l- azabicyclo[4.2.0]oct-2-ene-2-carboxylate, diphenylmethyl ester (93% yield) as a tan

solid, m.p. 124 °C. Η NMR (300 MHz, CDCI ₃ ) δ 3.43 (d, J=18 Hz, 1 H), 3.69 (d, J=17

Hz, 1 H), 3.70 (d, J=18 Hz, 1 H), 3.78 (d, J=17 Hz, 1 H), 4.27 (d, J=9 Hz, 1 H), 4.33 (d, J=9 Hz, 1 H), 4.96 (d, J=5 Hz, 1 H), 5.75 (dd, J=5, 9 Hz, 1 H), 7.00 (s, 1 H), 7.20-7.46 (m, 14 H). Anal. Calcd. for C ₂ 9H ₂₃ N ₂ O ₄ S ₂ CI ₂ I: C, 48.01; H, 3.20; N, 3.86. Found: C,

48.00; H, 3.14; N, 3.76.

(6R)-trans-3-Iodomethyl-7-[(2,5-dichlorophenyl)thioacetam ido]-8-oxo-5- thia-l-azabicyclo[4.2.0]oct-2-ene-2-carboxylate, diphenylmethyl ester (3.00 g, 4.14 mmol) was dissolved in THF (50 L) at 0 °C and treated with 4-mercaptopyridine (0.504 g, 4.54 mmol). A solution of 2,6-lutidine (0.576 g, 5.38 mmol) in THF (1 mL) was added next, and the reaction mixture was stirred at 0 °C for 0.5 h and then at 20 °C for 1 h. The mixture was diluted with ethyl acetate (500 mL) and the organic solution was washed with water (2 x 500 mL) and brine (100 mL). The solution was then dried (MgSθ ₄ ) and evaporated under reduced pressure to give an oil which was treated with E. ₂ O (50 mL) to give a solid. The solid was

collected by filtration and purified by column chromatography on silica gel (CH ₂ CI ₂ to 30% EtOAc/CH ₂ Cl ₂ ) to give 1.50 g of (6R)-trans-3-[(4-

pyridylthiomethyl]-7-[(2,5-dichlorophenyl)-thioacetamidoJ -8-oxo-5-thia-l- azabicyclo[4.2.0Joct-2-ene-2-carboxylate, diphenylmethyl ester as a tan solid (49%

yield), m.p. 122 °C *H NMR (300 MHz, CDCI3) δ 3.37 (d, J=18 Hz, 1 H), 3.52 (d, J=18

Hz, 1 H), 3.68 (d, J=17 Hz, 1 H), 3.76 (d, J=17 Hz, 1 H), 3.96 (d, J=13 Hz, 1 H), 4.16 (d, J=13 Hz, 1 H), 4.93 (d, J=5 Hz, 1 H), 5.76 (dd, J=5, 9 Hz, 1 H), 6.95-7.42 (m, 16 H), 7.49 (d, J=9 Hz, 1 H), 8.29 (d, J=6 Hz, 2 H). Anal. Calcd. for C34H27N ₃ O ₄ S ₃ CI ₂ : C, 57.62; H, 3.84; N, 5.93. Found: C, 57.27; H, 3.68; N, 5.79.

D. l-f2-Carboxy-2-propen-l-yll-4-ff(6R)-trans-2-(diphenylmethyl -carboxy)-8- oxo-7-f(2.5-dichlorophenylthio)acetamidol-5-thia-l-azabicycl of4.2.01-oct-2-en-3- yllmethylthiolpyridinium bromide

A solution of (6R)-trans-3-(4-pyridylthiomethyl)-7-[(2,5-dichlorophenyl)- thioacetamido]-8-oxo-5-thia-l-azabicyclo[4.2.0joct-2-ene-2-c arboxylate, diphenylmethyl ester (600 mg, 0.845 mmol) in 5 mL of DMF was treated with 2-

(bromomethyl) acrylic acid (Aldrich) (279 mg, 1.69 mmol). This mixture was stirred at rt for 4 h, and then 50 mL of ether was added, resulting in the precipitation of the desired product. The solid was collected by filtration washing with ether to give 667 mg (85%) of l-[2-carboxy-2-propen-l-yl]-4-[[(6R)-trans-2- (diphenylmethyl carboxy )-8-oxo-7-[(2,5-dichlorophenylthio)acetamido]-5-thia-l-

azabicyclo[4.2.0]-oct-2-en-3-yljmethylthio]pyridinium bromide, as a tan solid. ¹ H

NMR (DMSO-d6) δ 9.35 (d, J=8 Hz, IH), 8.73 (d, J=7 Hz, 2H), 7.88 (d, J=7 Hz, 2H),

7.18-7.49 (m, 13 H), 6.97 (s, IH), 6.48 (s, IH), 6.18 (s, IH), 5.81 (dd, J=5,8 Hz, IH), 5.35 (s, 2H), 5.23 (d, J=5 Hz, IH), 4.31 (d, J=12 Hz, IH), 7.23 (d, J=12 Hz, IH), 3.95 (s, 2H),

3.83 (d, J=18 Hz, IH), 3.59 (d, J=18 Hz, IH). IR (KBr) 1780, 1714, 1626, 1450, 1106 cnr

^! ; MS (ESI) 792 (MH)+; Anal, calcd. for C ₃ 8H ₃₁ N3O6S3 ₂ • HBr • 0.35 HC(O)

N(CH ₃ )2 • 1.5 H ₂ 0: C, 50.64; H, 4.08; N, 5.07. Found: C, 50.68, H, 3.93; N, 5.05.

E. l-r2-Carboxy-2-propen-l-yll-4-rr(6R)-trans-2-carboxy-8-oxo-7 -r(2.5- dichlorophenyIthio)acetamidol-5-thia-l-azabicyclof4.2.01-oct -2-en-3- yllmethylthiolpyridinium bromide

A slurry of l-[2-carboxy-2-propen-l-yl]-4-[[(6R)-trans-2-(diphenylmethyl carboxy)-8-oxo-7-[(2,5-dichlorophenylthio)acetamidoj-5-thia- l-azabicyclo[4.2.0J-oct-

2-en-3-yl]methylthio]pyridinium bromide (580 mg, 0.665 mmol) in 22 mL of

CH2CI2 was cooled to 0°C (ice bath) under an atomosphere of N2. Anisole (2.8

mL) was added followed by trifluoroacetic acid (8 mL). The cooling bath was removed, and the reaction was allowed to stir at rt for 1 h. The solution was concentrated in vacuo to remove the CH2CI2 and most of the trifluoroacetic acid.

The residue was treated with ether to precipitate a yellow solid. This solid was collected by filtration washing with ether. The solid was stirred with ethyl acetate for 30 min. stirred with acetone for 30 min., collected by filtration, and dried in vacuo to give 322 mg (66%) of l-[2-carboxy-2-propen-l-yl]-4-[[(6R)-trans-2-carboxy- 8-oxo- 7-[(2,5-dichlorophenylthio)acetamidoJ-5-thia-l-azabicyclo[4. 2.0J-oct-2-en-3-

yljmethylthiojpyridinium bromide as a tan solid. IH NMR (DMSO-d6) δ 9.27 (d,

J=8 Hz, IH), 8.73 (d, J=6 Hz, 2H), 8.06 (d, J=6 Hz, 2H), 7.45-8.48 (m, 2H), 7.24 (dd, J=2,9 Hz, 2 H), 6.41 (s, IH), 6.08 (s, IH), 5.63 (dd, J=5,8 Hz, IH), 5.19 (s, 2H), 5.09 (d, J=5 Hz, IH), 4.43 (s, 2H), 3.92 (s, 2H), 3.70 (d, J=18 Hz, IH), 3.48 (d, J=18 Hz, IH). IR (KBr) 1774, 1702, 1680, 1628, 1106 cm-1; MS (ESI) 626 (MH)+, Anal: Calcd for

C25H2iN3θ ₆ S ₃ Cl2xHBr x0.5Me CO; C, 42.44; H, 3.13; N, 5.94. Found: C, 43.25; H,

3.42, N, 5.71.

Example 2

l-r2-Carboxy-2-hydroxyethyll-4-r[(6Rl-trans-2-carboxy-8-o xo-7-[(2.5- dichlorophenylthio)acetamidoJ-5-thia-l-azabicyclof4.2.0j-oct -2-en-3- yljmethylthiojpyridinium chloride

A. (6R)-trans-3-Chloromethyl-7-[ ⁽ 2.5-dichlorophenyl ⁾ -thioacetamidol-8-oxo-5- thia-l-azabicycIo[4.2.01oct-2-ene-2-carboxylic acid

A slurry of (6R)-trans-3-chloromethyl-7-[(2,5-dichlorophenyl)- thioacetamido]-8-oxo-5-thia-l-azabicyclo[4.2.0joct-2-ene-2-c arboxylate, diphenylmethyl ester (10.0 g, 15.8 mmol) in CH2CI2 (200 mL) at 0 °C was treated

with anisole (24 mL) and then trifluoroacetic acid (80 mL). The solution was stirred for 1 h at 0 °C and then concentrated under reduced pressure. The residue was stirred with Et2θ, and the resulting solid was collected by filtration to give

5.20 g of (6R)-trans-3-chloro-methyl-7-[(2,5-dichlorophenyl)-thioaceta midoj-8-oxo- 5-thia-l-azabicyclo[4.2.0joct-2-ene-2-carboxylic acid as a white solid (70% yield),

m.p. 125 °C. *H NMR (300 MHz, DMSO-d ₆ ) δ9.28 (d, J=8 Hz, 1 H), 7.47 (dd, J=2, 8

Hz, 2 H), 7.24 (dd, J=2, 8 Hz, 1 H), 5.70 (dd, J=5, 8 Hz, 1 H), 5.13 (d, J=5 Hz, 1 H), 4.58 (d, J=ll Hz, 1 H), 4.52 (d, J=ll Hz, 1 H), 3.91 (s, 2 H), 3.70 (d, J=18 Hz, 1 H) 3.51 (d, J=18 Hz, 1 H).

B. l-f2-carboxy-2-hydroxyethy11-4-rr(6R)-trans-2-carboxv-8-oxo- 7-r(2.5- dichlorophenylthio)acetamidol-5-thia-l-azabicycIof4.2.01-oct -2-en-3- yllmethylthiolpyridinium chloride

1. Preparation of l-[N-(2-carboxy-2-hydroxyethyl)-4-thiopyridone sodium salt

A solution of D,L-isoserine (1.00 g, 9.52 mmol) in a mixture of water (10 mL) and ethanol (10 mL) was treated with 9.52 mL of IN NaHCθ3 followed by

thiopyrone (1.07 g, 9.52 mmol). The mixture was stirred at reflux for 2 h, and concentrated in vacuo. The residue was triturated with ether to give 1.78 g (85%) of l-[N-(2-carboxy-2-hydroxyethyl)-4-thiopyridone sodium salt as a brown solid.

H NMR (DMSO-d ⁶ ) δ 7.50 (d, J=7 Hz, 2H), 7.08 (d, J=7 Hz, 2H), 4.17 (dd, J=3,4 Hz,

IH), 4.03 (dd, J=6,14 Hz, IH), 3.83 (dd, J=3,6 Hz, IH). ^C (75 MHz) δ 188.62, 172.79,

137.09, 129.44, 70.78, 60.40; IR (KBr) 3384 (br), 1622, 1114 cur ¹ ; MS (ESI) 198 (M)\

2. l-[2-Carboxy-2-hydroxyethyl]-4-[[(6R)-trans-2-carboxy-8-oxo- 7-[(2,5- dichlorophenylthio)acetamidoJ-5-thia-l-azabicyclo[4.2.0]-oct -2-en-3- yljmethylthiojpyridinium chloride

A solution of cephem 6(R)-trans-3-chloromethyl-7-[(2,5-dichlorophenyl)- thioacetamido]-8-oxo-5-thia-l-azabicyclo[4.2.0]oct-2-ene-2-c arboxylic acid (60 Omg, 1.28 mmol) in 5 mL of DMF was treated with thiopyridone l-[N-(2-carboxy-2- hydroxyethyl)-4-thiopyridone Na salt(255 mg, 1.28 mmol). After stirring the mixture for 3h at rt under a nitrogen atmosphere, it was concentrated in vacuo. The residue was stirred with ether until a solid formed. This solid was collected by filtration and washed with ethyl acetate followed by acetone to give 564 mg (63%) of l-[2-carboxy-2-hydroxyethyl]-4-[[(6R)-trans-2-carboxy-8-oxo- 7-[(2,5- dichlorophenylthio)acetamidoJ-5-thia-l-azabicyclo[4.2.0J-oct -2-en-3-

yljmethylthiojpyridinium chloride as a tan solid. H NMR (DMSO-d ⁶ ) δ 9.36 (d,

J=6 Hz, IH), 8.67 (d, J=6 Hz, 2H), 8.07 (d, J=6 Hz, 2H), 7.42-7.48 (m, 2H), 7.21 (dd,

J=2,9 Hz IH), 5.57 (dd, J=5,8 Hz, IH), 5.05 (d, J=5 Hz, IH), 4.80 (d, J=ll Hz, IH), 4.39- 4.60 (m, 4H), 3.93 (s, 2H), 3.64 (d, J=18 Hz, IH), 3.45 (d, J=18 Hz, IH). IR (KBr) 3422

(br), 1774 , 1703, 1675, 1628, 1450, 1108 cm" ¹ . MS (ESI) 630 (MH) ⁺ ; Anal. Calcd. for C ₂₄ H ₂₁ O ₇ N ₃ S ₃ CI ₂ • HCl • 2.0 H ₂ 0: C, 40.97; H, 3.69; N, 5.97. Found: C, 40.95; H,

3.43; N, 5.83

The following additional compounds were prepared according to the general procedures of Examples 1 and 2 by varying the thiol starting material and the pyridine or thiopyridone derivative:

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