SG185170A1

SG185170A1 - Octahydropyrrolo[1,2-a]pyrazine derivatives of carbapenem as antibacteria antibiotics

Info

Publication number: SG185170A1
Application number: SG2011032182A
Authority: SG
Inventors: Zhenhua Huang
Original assignee: Kbp Biosciences Pte Ltd
Priority date: 2011-05-05
Filing date: 2011-05-05
Publication date: 2012-11-29

Abstract

Octahydropyrrolo[1,2-alpyrazine Derivatives of Carbapenem as Antibacteria AntibioticsAbstractThe present invention belongs to the pharmaceutical field and specifically relates to Octahydropyrrolo[1,2-a]pyrazine Derivatives of Carbapenem as Antibacteria Antibiotics as shown in formula ( I ), or its pharmaceutically acceptable salts, or its readily hydrolyzed ester, or its solvate, or its stereoisomer, and intermediates as described in formula ( II ), wherein R1, R2 and R3 are as defined in specification, the present invention also relates to the processes for preparing the same, to the pharmaceutical composition containing such compounds, and to the use of these compounds in the manufacture of a medicament for the treatment and/or prophylaxis of infectious diseases. ( I ) - • •1111111111111111111111111111111111111* 6'7161*1111 11011 II *G00002*

Description

JINOLAUE

*159159*

Octahydropyrrolo[1,2-a]pyrazine Derivatives of Carbapenem as

Antibacteria Antibiotics 1. Field of The Invention

The present invention belongs to the pharmaceutical filed and specifically relates to

Octahydropyrrolo[1,2-a]pyrazine Derivatives of Carbapenem as Antibacteria Antibiotics, or its pharmaceutically acceptable salts, or its readily hydrolyzed ester, or its solvate, or its stereoisomer and intermediates, thereof, to the processes for preparing the same, to the pharmaceutical compositions containing such compounds, and to the use of these compounds in the manufacture of a medicament for the treatment and / or prophylaxis of infectious disease. 2. Background of The Invention

Carbapenems have drawn wide attention due to their broad anti-bacterial spectrum, potent anti-bacterial activity and stability towards p-lactamase. The carbapenem currently in use at clinic include Imipenem, Panipenem, Meropenem, Ertapenem, Bioapenem, Doripenem and Tebipenem.

European patent application EP0126587 published Meropenem (structure below), which is a 1B-methyl carbapenem, active to gram-positive bacteria, and gram-negative bacteria. It is especially potent against gram-negative bacteria, stable against DHP-I enzyme, can be used as a single therapy. 7 CH, Q

HiC N-CHs

TOC i,

COOH

There are more and more drug-resistant bacteria emerging due partially to antibiotics abuses, additionally, most of the carbapenems on the market have a human elimination half-life of about 1 hour, require multiple daily doses, and are less tolerable. Therefore, it is important to develop more potent, stable, and long half-life carbapenem to meet clinical needs. 3. Disclosure of The Invention

The present invention provides Octahydropyrrolo[1,2-a]pyrazine Derivatives of Carbapenem as Antibacteria Antibiotics, or its pharmaceutically acceptable salts, or its readily hydrolyzed ester, or its solvate, or its stereoisomer: Co

AR AREA

1 ee

OH rl

H;C NF

COOR? (D)

Wherein

R! is hydrogen or C,.¢ alkyl;

R? is (1) hydrogen, (2) Cy.6 alkyl, Cys alkenyl or C,.g alkynyl, which is optionally substituted by one to three x! : X! is independently selected from halogen, carboxyl, hydroxyl, amino, cyano, nitro, Cy.¢ alkoxy, halogenated Ci. alkoxy, C;.¢ alkyl amino, bis-(C,.¢ alkyl)amine, C,.¢ alkyl carbonyl,

Ci.salkyl carboxyl, C;¢ alkoxy carbonyl, carbamoyl, C,.¢ alkyl carbamoyl, bis-(Ci.¢ alkyl) carbamoyl, amino sulfonyl, C;¢ alkyl amino sulfonyl, bis-(C,s alkyl) amino sulfonyl, Ci; alkyl sulfonyl amino, Ci alkyl sulfonyl, C;¢ alkyl carbonyl amino, guanidino, Css cycloalkyl, aryl or heterocyclic,

Cs.g cycloalkyl, aryl or heterocyclic of said X' can be further substituted by the following substituents: halogen, carboxyl, hydroxyl, amino, cyano, nitro, Ci.¢ alkyl, carboxy Ci.¢ alkyl, hydroxy Ci.¢ alkyl, amino C;, alkyl, halogenated C,.¢ alkyl, Ci. alkoxy, halogenated Ci.¢ alkoxy, Cy alkoxy C.¢ alkyl, C,. alkyl amino, bis-(C,.¢ alkyl) amino, bis-(C,.¢ alkyl) amino

Cis alkyl, Ci alkyl carbonyl, C;¢alkyl carboxyl, C;s alkoxy carbonyl, carbamoyl, carbamoyl C,.¢ alkyl, C;.¢ alkyl carbamoyl, bis-(C,.¢ alkyl) carbamoyl, amino sulfonyl, amino sulfonyl Cy alkyl, Ci.¢ alkyl amino sulfonyl, bis-(C,.¢ alkyl) amino sulfonyl, C,.¢ alkyl sulfonyl amino, C,.¢ alkyl sulfonyl, C,.¢ alkyl carbonyl amino or guanidino, (3) Css cycloalkyl, aryl or heterocyclic which is optionally substituted by one to three x2,

X? is independently selected from halogen, carboxyl, hydroxyl, amino, cyano, nitro, Cy. alkyl, carboxy Ci.¢ alkyl, hydroxy Ci.¢ alkyl, amino C,, alkyl, halogenated C;. alkyl, Ci alkoxy, halogenated C; alkoxy, C;¢ alkoxy Ci.¢ alkyl, C.¢ alkyl amino, bis-(Ci.¢ alkyl) amino, bis-(C; alkyl) amino C4 alkyl, Ci-6 alkyl carbonyl, C;salkyl carboxyl, Ci alkoxy carbonyl, carbamoyl, carbamoyl Ci. alkyl, C6 alkyl carbamoyl, bis-(C,-s alkyl) carbamoyl, amino sulfonyl, amino sulfonyl Ci. alkyl, C,.¢ alkyl amino sulfonyl, bis-(C,¢ alkyl) amino sulfonyl, Ci, alkyl sulfonyl amino, C;¢ alkyl sulfonyl, Cis alkyl amino carbonyl or guanidino, :

4) -CO-Y,

Y is Cy alkyl which is optionally substituted by one to three Y!, or Cs. cycloalkyl, aryl or heterocyclic which is optionally substituted by one to three Y?,

Y! represents the above defined X! and Y? represents the above defined xX? (5) -S(O)p-Z, mis 1 or 2,

Z is amino, Cy alkyl amino, bis-(C,.¢ alkyl)amino, or C; alkyl which is optionally substituted by one to three Y!, or Cs cycloalkyl, aryl or heterocyclic which is optionally substituted by one to three Y?,

Y! represents the above defined X' and Y? represents the above defined X?%; and

R? is hydrogen or the carboxyl protecting group.

In another preferred embodiment, wherein

R'is Cys alkyl;

R% is (1) hydrogen, + (2) C14 alkyl, which is optionally substituted by one to three xX,

X! is independently selected from halogen, carboxyl, hydroxyl, amino, C4 alkoxy, halogenated C;.4 alkoxy, C;4 alkyl amino, bis-(C;4 alkyl)amine, C;4 alkyl carbonyl, carbamoyl, amino sulfonyl, guanidino, Cs.¢ cycloalkyl, phenyl, 5-6 membered heteroaryl or 3-7 membered heterocyclic,

Ci. cycloalkyl, phenyl, 5-6 membered heteroaryl or 3-7 membered heterocyclic of the said X! can be further substituted by the following substituents: halogen, Ci4 alkyl, halogenated C,4 alkyl, C,.4 alkoxy, halogenated C4 alkoxy, Ci4 alkyl amino, bis-(C;4 alkyl) amino, C4 alkyl carbonyl, carbamoyl, carbamoyl C4 alkyl, amino sulfonyl, amino sulfonyl ~~ Cj4 alkyl, or guanidino, (3) Cs. cycloalkyl, phenyl, 5-6 membered heteroaryl or 3-7 membered heterocyclic which is optionally substituted by one to three X>,

X? is independently selected from halogen, amino, C;.4 alkyl, halogenated Cj. alkyl, Ci. alkoxy, halogenated C,4 alkoxy, Ci.4 alkyl amino, bis-(C;4 alkyl) amino, bis-(C;.4 alkyl) amino Cj. alkyl, C4 alkyl carbonyl, carbamoyl, carbamoyl C4 alkyl, amino sulfonyl, amino sulfonyl C;.4 alkyl, or guanidino, 4) -CO-Y,

Y is C4 alkyl which is optionally substituted by one to three Y', or Cs cycloalkyl, phenyl, 5-6 membered heteroaryl or 3-7 membered heterocyclic which is optionally substituted by one to three Y2,

Y' represents the above defined X' and Y? represents the above defined X2, (5) -S(O)p-Z, mis 1 or 2,

Z is amino, C4 alkyl amino, bis-(C;4 alkyl)amino, or C;4 alkyl which is optionally substituted by one to three Y', or Cs cycloalkyl, phenyl, 5-6 membered heteroaryl or 3-7 membered heterocyclic which is optionally substituted by one to three Y2,

Y! represents the above defined X' and Y? represents the above defined X?; and

R? is hydrogen.

In another preferred embodiment, wherein

R! is methyl;

R? is (1) hydrogen, (2) C14 alkyl, which is optionally substituted by one to three X',

X! is independently selected from halogen, carboxyl, hydroxyl, amino, C;4 alkoxy, halogenated C4 alkoxy, C4 alkyl amino, bis-(C;.4 alkyl)amine, carbamoyl, amino sulfonyl, phenyl, pyridyl, piperidinyl, piperazinyl or morpholinyl,

The phenyl, pyridyl, piperidinyl, piperazinyl or morpholinyl of the said X' can be further substituted by the following substituents: halogen, C;4 alkyl, halogenated C;.4 alkyl, Ci4 alkoxy, halogenated C,4 alkoxy, C;.4 alkyl amino, bis-(C,.4 alkyl) amino, C;.4 alkyl carbonyl, carbamoyl, catbamoy] C14 alkyl, or amino sulfonyl, (3) phenyl, pyridyl, piperidinyl, piperazinyl, morpholinyl, tetrahydrofuranyl, thiazole, 4,5-dihydro-thiazolyl, or cyclopropyl group which is optionally substituted by one to three x2, xX? is independently selected from halogen, C4 alkyl, C,4 alkyl amino, bis-(C;4 alkyl) amino, carbamoyl, carbamoyl C;.4 alkyl, amino sulfonyl, 4) -CO-Y,

Y is Ci4 alkyl which is optionally substituted by one to three Y!, or phenyl, pyridyl, piperidinyl, piperazinyl, morpholinyl, pyrrolidinyl, furanyl, thiazolyl or 4,5-dihydro-thiazolyl which is optionally substituted by one to three Y?,

Y! is independently selected from amino, Ci4 alkyl amino, bis-(C;4 alkyl) amino,

carbamoyl, guanidino, phenyl, pyridyl, piperidinyl, morpholinyl or pyrrolidinyl, - Y? is independently selected from halogen, amino, C;.4 alkyl, C4 alkyl amino, bis-(C;4 alkyl) amino, carbamoyl, carbamoyl C;_4 alkyl or amino sulfonyl, (5) -S(O)y-Z, mis 1 or 2,

Z is amino, Ci4 alkyl amino, bis-(C;4 alkyl)amino, or C;4 alkyl which is optionally substituted by one to three Y', or phenyl, pyridyl, piperidinyl, piperazinyl, morpholinyl or pyrrolidinyl which is optionally substituted by one to three Y?,

Y! and Y? represents the above defined; and

R’ is hydrogen.

In another preferred embodiment, wherein

R! is methyl;

R? is (1) hydrogen, (2) methyl, ethyl, isopropyl, which is optionally substituted by one to three X',

X! is independently selected from halogen, hydroxyl, dimethylamine, phenyl or pyridyl,

The phenyl or pyridyl of the said X' can be further substituted by the following - substituents: halogen, methyl, carbamoyl, carbamoyl methyl, or amino sulfonyl, (3) phenyl, pyridyl, tetrahydrofuranyl, thiazole, 4,5-dihydro-thiazolyl, or cyclopropyl group which is optionally substituted by one to three X°,

X? is independently selected from halogen, methyl, carbamoyl, amino sulfonyl, (4) -CO-Y,

Y is methyl, ethyl or isopropyl which is optionally substituted by one to three Y', or phenyl, pyridyl, piperidinyl, piperazinyl, pyrrolidinyl which is optionally substituted by one to three YZ, : | Y'is independently selected from dimethylamine, carbamoyl, guanidino, phenyl, pyridyl, piperidinyl, morpholinyl or pyrrolidinyl,

Y? is independently selected from halogen, methyl, carbamoyl, carbamoyl methyl or amino sulfonyl, (5) -S(O)m-Z, mis 1 or 2,

Z is amino, dimethylamino, dimethylamino methyl, methylpiperazine or morpholine methyl; and

R? is hydrogen.

In another preferred embodiment, wherein

R! is methyl;

R? is (1) hydrogen, (2) methyl, ethyl, or isopropyl, which is optionally substituted by one to two X',

X! is independently selected from hydroxyl,or phenyl,

The said phenyl of X' can be further substituted by halogen, methyl, carbamoyl, or amino sulfonyl, (3)-CO-Y,

Y is methyl, ethyl or isopropyl, which is optionally substituted by one to two Y!, or phenyl, pyridyl, piperidinyl, which is optionally substituted by one to two Y?,

Y' is independently selected from dimethylamine, guanidino, phenyl, pyridyl, or morpholinyl,

Y? is independently selected from halogen, methyl, carbamoyl, or amino sulfonyl, 4) -S(O)m-Z, mis 1 or 2,

Z is amino, or dimethylamino; and

R? is hydrogen.

Table 1 The compounds of present invention

Structural Formula Chemical Name

OH

! CH;

HC, _ | (4R,55,65)-6-[(R)-1-hydroxyethyl]-4-methyl-3-[(7S,8aS)-2- 1 4 s— 1} methyloctahydropyrrolo[1,2-a]pyrazin-7-ylthio]-7-o0xo-1-

N NA azabicyclo[3.2.0]hept-2-ene-2-carboxylic acid

Oo

COOH

OH

H,C J, ’ 1 (4R,58,65)-6-[(R)-1-hydroxyethyl]-3-[(7S,8aS)-2-Isopropyl- 2 / s~) octahydropyrrolo[1,2-a]pyrazin-7-ylthio]-4-methyl-7-oxo- ov N N 1-azabicyclo[3.2.0]hept-2-ene-2-carboxylic acid

COOH

OH ne, I (4R,55,68)-3-[(75,8a8)-2-benzyloctahydropyrrolo{1,2-a}- 3 J ~ 1) UO pyrazin-7-ylthio]-6-[(R)-1-hydroxyethyl]-4-methyl-7-oxo- oN N 1-azabicyclo[3.2.0]hept-2-ene-2-carboxylic acid

COOH ped Fh I (4R,58,68)-3-[(7S,8a8)-2-acetyloctahydropyrrolo[1,2-a] 4 C0) pyrazin- 7-ylthio]-6-[(R)-1-hydroxyethyl]-4-methyl-7-0xo- 0 oor 1-azabicyclo[3.2.0]hept-2-ene-2-carboxylic acid

OH wet. I i (4R,55,65)-6-[(R)-1-hydroxyethyl]-3-[(7S,8aS)-2-isobutyryl-

J C1) octahydropyrrolo[ 1,2-a]pyrazin-7-ylthio]-4-methyl-7-oxo-1- oN oon N azabicyclo[3.2.0]hept-2-ene-2-carboxylic acid

OH 0

HC Fo (4R,58,65)-3-[(7S,8aS)-2-benzoyloctahydropyrrolo[1,2-a]- - C1) pyrazin-7-ylthio]-6-[(R)-1-hydroxyethyl]-4-methyl-7-0x0- 0” N N 1-azabicyclo[3.2.0]hept-2-ene-2-carboxylic acid

COOH

OH CH, 0

HC A. Ng (4R,55,65)-6-[(R)-1-hydroxyethyl]-4-methyl-7-ox0-3- 7 4 1} [(7S,8aS)-2-picolinoyloctahydropyrrolo[ 1,2-a]pyrazin- o> N N Z 7-ylthio]-1-azabicyclo[3.2.0]hept-2-ene-2-carboxylic acid

COOH

Ba CH, 0 (4R,55,68)-6-[(R)-1-hydroxyethyl]-4-methyl-7-ox0-3-

HC, CON CL [(7S,8aS)-2-(4-sulfamoylbenzoyl)octahydropyrrolo[1,2-a]

PNY Sn son. | Pyrazin-7-ylthio]-1-azabicyclo[3.2.0]hept-2-ene-2-carboxylic

COOH © | acid

Hoon 0.0

He. ’ SNH (4R,58,68)-6-[(R)-1-hydroxyethyl]-4-methyl-7-0xo-3-

J 1 2 [(7S,8aS)-2-sulfamoyloctahydropyrrolo[1,2-a]pyrazin- 0” N N 7-ylthio]-1-azabicyclo[3.2.0]hept-2-ene-2-carboxylic acid

COOH

Ba CH, (4R,58,6S5)-6-[(R)-1-hydroxyethyl]-4-methyl-7-0x0-3-

HC, <y [(7S,8aS)-2-(4-sulfamoylbenzyl)octahydropyrrolo[1,2-a]

ZN 7 NS C NH pyrazin-7-ylthio]-1-azabicyclo[3.2.0]hept-2-ene-2-carboxylic

COOH “| acid

OH wer, qu wi, | (4R,55,65)-3-[(75,8aS)-2-(2-guanidinoacetyl)octahydropyrrolo 11 ) —~ 1} hd {1,2-a)pyrazin-7-ylthio]-6-[(R)-1-hydroxyethyl]-4-methyl-7-

ZN oo N NA oxo-1-azabicyclo[3.2.0]hept-2-ene-2-carboxylic acid

H a CH; J | | (4R,58,65)-3-((7S,8aS)-2-(2-(dimethylamino)acetyl) 12 HC, N N< | octahydro-pyrrolo[1,2-a]pyrazin-7-ylthio)-6-((R)-1- #N JS N hydroxyethyl)-4-methyl-7-oxo-1-azabicyclo[3.2.0]hept-2-

S 00H ene-2-carboxylic acid ag CH; 9 (4R,58,65)-3-((7S,8aS)-2-(2-(dimethylamino)propionyl)octa- 13 HC, CY YS hydropyrrolo[1,2-a]pyrazin-7-ylthio)-6-((R)-1-hydroxyethyl)-

Nd © NA 4-methyl-7-ox0-1-azabicyclo[3.2.0]hept-2-ene-2-carboxylic © “OOH acid a CH, i 3 (4R,58,65)-6-[(R)-1-hydroxyethyl]-4-methyl-3-[(75,8a5)-2- 14 HC 2 N (2-morpholinoacetyl)octahydropyrrolo[1,2-a]pyrazin-7-

N-¥ 13 ylthio]-7-0x0-1-azabicyclo[3.2.0]hept- 2-ene-2-carboxylic ° COOH acid gn CH, 0 (4R,55,65)-6-[(R)-1-hydroxyethyl]-4-methyl-3-[(7S,8aS)-2-

HC 1) (1-methylpiperidine-4-carbonyl)octahydropyrrolo[1,2-a]

ZN JS NS No | pyrazin-7-ylthio]-7-oxo-1-azabicyclo[3.2.0]hept-2-ene-2-

COOH carboxylic acid

OH we, LT OH | (4R5S5,68)-6-[(R)-1-hydroxyethyl]-3-[(7S,825)-2-(2-hydroxy- 16 / C1) ethyl)octahydropyrrolo[ 1,2-a]pyrazin-7-ylthio]-4-methyl-7- oo“ N N oxo-1-azabicyclo[3.2.0]hept-2-ene-2-carboxylic acid : COOH

The term “Cjsalkyl”, as used hereinbefore, refers to linear, branched groups derived from removing one hydrogen from hydrocarbons containing 1 to 6 carbon(s). Examples include, but are not limited to, methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl, n-pentyl, isopentyl, 2-methylbutyl, neopentyl, 1-ethylpropyl, n-hexyl, isohesyl, 4-methylpentyl, 3-methylpentyl, 2-methylpentyl, 1-methylpentyl, 3,3-dimethylbutyl, 2,2-dimethylbutyl, 1,1-dimethylbuytl, 1,2-dimethylbutyl, 1,3-dimethylbutyl, 2,3-dimethylbutyl, 2-ethylbutyl, 1,2-dimethylpropyl, et al. the term “Cj4alkyl” refers to the above examples that contain 1 to 4 carbon(s), for example, methyl, ethyl,

n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl.

The term “Cs.3 cycloalky”, as used hereinbefore, refers to the cycloalkanes of 3 to 8 carbons atom membered size, such as cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctanyl, methylcyclopropane, dimethylcyclopropane, methylcyclobutane, dimethylcyclobutane, methylcyclopentane, dimethylcyclopentane, methylcyclohexane, dimethylcyclohexane, and so on. The term "Cj cycloalkyl” refer respectively 3 to 6 carbon atom cycloalkane in above-mentioned examples.

The term "C,.g alkenyl" , as used hereinbefore, refers straight or branched chain or cyclic alkenyl containing 2 to 8 carbon atom with double bond(s), such as vinyl, 1-propenyl, 2-propenyl, 1-butenyl, 2-butenyl, 1,3-butadienyl, 1-pentenyl, 2-pentenyl, 3-pentenyl, 1,3-pentadienyl, 1,4-pentadienyl, 1-hexenyl, 2-hexenyl, 3-hexenyl, 1,4-hexadienyl, cyclopentenyl, 1,3-cyclopentadienyl, cyclohexenyl, 1,4-cyclohexadienyl, cycloheptenyl, 1,4-cycloheptadienyl, ~ cyclooctenyl, 1,5-cyclooctadienyl and so on.

The term "C,.g alkynyl", as used hereinbefore, refers straight or branched chain containing 2 to 8 carbon atom with triple bond(s), such as acetylenyl, propynyl, 2-butynyl, 2-pentynyl, 3-pentynyl, 4-methyl-2-pentynyl, 2-hexynyl, 3-hexynyl, S-methyl-2-hexynyl, 2-heptynyl, 5-methyl-2-heptynyl, 2-octynyl, 3-octynyl and so on.

The term "C;.¢ alkoxy" , as used hereinbefore, refers to the term "Cy.¢ alkyl" structure through the oxygen atom connected with other groups such as methoxy, ethoxy, propoxy, isopropoxy, butoxy, isobutoxy, tert-butoxy, secondary butoxy, pentyloxy, isopentyloxy, hexyloxy and so on. The terms "C4 alkoxy" refer respectively to in above-mentioned specific examples 1 to 4 atoms.

The term “halo”, as used hereinbefore, include fluorine, chlorine, bromine, and iodine.

The term “halogenated C,.¢ alkyl”, “halogenated C,.¢ alkoxy” as used hereinbefore, is one or more hydrogen atoms of C;. alkyl or C;.¢ alkoxy substituted by one or more halo. The C; alkyl,

C16 alkoxy, and halo are denotes as hereinbefore.

The term“hydroxy Ci. alkyl”, “carboxy C;.¢ alkyl”, “amino C,¢ alkyl”, “C; alkoxy Cis alkyl”, as used hereinbefore, is one or more hydrogen atoms of C,.¢ alkyl independently substituted ~ by one or more hydroxy, carboxy, amino or C,.¢ alkoxy. The C,.¢ alkyl and C,.¢ alkoxy are denotes as hereinbefore.

The term “aryl”, as used hereinbefore, refers to aromatic rings, include “carbocyclic aryl” and “heteroaryl”.

The term “carbocyclic aryl” have no heteroatom, examples include but are not limited to, phenyl, naphthalenyl, anthracenyl, and phenanthrenyl.

The term “heteroaryl” refers to 5 to 20 membered cyclic group containing one or more heteroatoms except carbons, the heteroatoms means nitrogen, oxygen, sulfur, and so on. The “heteroaryl” refers to 5 to 10 ring atoms containing one or more heteroatoms (called 5-10 memebered heteroaryl) is preferred, examples include but are not limited to, furan, thiophene, pyrrole, imidazole, pyrazole, triazole, oxazole, isoxazole, thiazole, isothiazole, oxadiazole, tetrazole, pyridine, pyridazine, pyrimidine, pyrazine, triazine, benzofuran, isobenzofuran, benzothiophene, indole, isoindole, quinoline, isoquinoline, naphthyridine, acridine. The term “5-6 membered heteroaryl”, refer respectively to in above-mentioned specific examples 5 to 6 membered atoms.

The term “heterocyclic” refers to 3 to 14 membered cyclic group containing one or more heteroatoms (called 3-14 memebered heteroaryl), include saturated heterocyclic and unsaturated heterocyclic. The “heterocyclic” refers to 3 to 7 ring membered cyclic group containing one or more heteroatoms (called 3-7 memebered heterocyclic) is preferred. The examples include but are not limited to, aziridine, azetidine, pyrrolidine, 2,5-dihydro-pyrrole, 2H-pyrrole, 3H-pyrrole, piperidine, dihydropyridine, tetrahydropyridine, oxirane, oxetane, dihydropyran, pyran, azepine, oxepine, thiirane, thietane, tetrahydrothiophene, thiepane, dioxolane, dioxepane, imidazolidine, pyrazolidine, imidazoline, piperazine, tetrahydro-oxazole, dihydro-oxazole, tetrahydro-isoxazole, dihydro-isoxazole, tetrahydro-oxazine, dihydro-oxazine, oxazine, thiazoline, thiazolidine, morpholine, thiomorpholine.

The term “carboxyl protecting group”, as used hereinbefore, refers to a protecting group which can be conventionally used to substitute the acidic proton of a carboxylic acid. Examples of caboxyl protecting group include, but are not limited to, methoxymethyl, methylthiomethyl, tetrahydropyranyl, tetrahydrofuranyl, methoxyethylmethyl, benzyloxymethyl, phenacyl, allyl, p-bromophenacyl, a-methylphenacyl, p-methoxyphenacyl, diacylmethyl, N-phthalimidomethyl, methyl, ethyl, diphenyl methyl, 2,2,2-trichloroethyl, 2-haloethyl, ®-chloroalkyl, 2-(trimethylsilyl)ethyl, 2-methylthioethyl, 2-(p-nitrothiopheny)ethyl, 2-(p-methylthiophenyl)ethyl, 1-methyl-1-phenylethyl, t-butyl, cyclopentyl, cyclohexyl, di-o-nitrophenylmethyl, 9-fluorenylmethyl, 2-(9,10-dioxo)fluorenylmethyl, 5-dithiophenyl, benzyl, 2,4,6-trimethylbenzyl, p-bromobenzyl, o-nitrobenzyl, p-nitrobenzyl, p-methoxybenzyl, piperonyl, 4-pyridinylmethyl, trimethylsilyl, triethylsilyl, t-butyldimethylsilyl, isopropyldimethylsilyl, phenyldimethylsilyl,

t-butyl, phenyl, 2-pyridyl, N-hydroxypiperidinyl, N-bydroxysuccinimido, N-hydroxyphthalimido,

N-hydroxybenzotriazolyl, O-acyloxime, 2,4-dinitrothiophenyl, 2-alkyl-1,3-oxazoline, 4-alkyl-5-o0x0-1,3-oxazolidinyl, 5-alkyl-4-oxo-1,3-dioxane, triethylstannyl, tri-n-butylstannayl,

N,N’-diisopropylhydrazide, and so on.

The abbreviations in the present invention are defined as follows:

Bn: benzyl

Boc: t-butoxycarbonyl

PNB: p-nitrobenzyl : TEA : triethylamine

TFA: trifluoroacetyl

THF: tetrahydrofuranyl

DCM: CH,Cl,

DEAD: (C;Hs00CN),

DIEA: diisopropylethylamine

Pd/C: Palladium on Carbon

The present invention provides the method of preparation of compounds of formular ( I ) by nucleophilic substitution reaction of compound of formular (II), or its pharmaceutically acceptable salts, or its readily hydrolyzed ester, or its solvate, or its stereoisomer with compound of formular (IID, or its pharmaceutically acceptable salts, or its readily hydrolyzed ester, or its solvate, or its stereoisomer:

OH rl

HsC

NS (ID) © €oor® (III)

The method of preparation is generalized in the following scheme:

OH R! 0

HS HC OP(OPh

S , S N-¢ 2 0 XR 0 0 COOPNB —_— —_— i@ — »

N Compound b N N. Compound d

TFA 2 (NH Nope R

Compound a Compound ¢ Formular (II)

OH R! OH R!

CHj CH,

Ng OD th Ng S © Te PAC 0 A 2

COOPNB » ; COOH ™N N-R

Compound e Compound f

Step 1 Preparation of Compound c

In a solution of Compound a in CH,Cl, was treated with TEA and Compound b. The resulting mixture was stirred at ambient temperature for overnight. The reaction mixture was washed with saturated brine, the organic layer was dried over anhydrous Na;SOy, filtered and concentrated under reduced pressure to give crude product of Compound c.

Step 2 Preparation of compound of formular (II)

The crude Compound ¢ from Step 1 was dissolved in methanol and treated with methanolic

KOH solution. The resulting mixture was stirred at ambient temperature. Upon completion of the reaction, the reaction mixture was extracted with CH,Cl,. The combined organic layers were dried (NaxS0y), filtered and concentrated under reduced pressure to give compounds of formular (II).

Step 3 Preparation of Compound e

In a dried reaction flask, compound of formular (II) was dissolved in acetonitrile and was treated with Compound d and DIEA. The resulting mixture was stirred at ambient temperature for overnight. The reaction mixture was extracted with ethyl acetate and washed with water. The organic layer was dried over Na,SOy, filtered and concentrated under reduced pressure to give crude product of Compound e.

Step 4 Synthesis of Compound f

The crude product from Step 3 in a mixture of tetrahydrofuran and water was treated with Pd/C.

The result black suspension was stirred at ambient temperature under hydrogen atmosphere. Upon completion, the catalyst was removed by filtration and the filtered cake was washed with . tetrahydrofuran and de-ionized water. The combined filtrate was extracted with ethyl acetate. The aqueous layer was purified by Prep. HPLC and the product fractions was combined and lypholized to give Compound f.

Wherein R!, R? and R® are defined as above, L represents suitable leaving group. The synthesis of Compound a see example 1 (Step 1 to 8). X in Compound b represents halogen such as fluorine, chlorine, bromine, iodine or other replaceable group such as acid chloride.

This invention further claims compounds of formular ( I), or its pharmaceutically acceptable salts, or its readily hydrolyzed ester, or its solvate, or its stereoisomer and the preparation of intermediates as described in formular (II), or its pharmaceutically acceptable salts, or its readily hydrolyzed ester, or its solvate, or its stereoisomer:

JR?

NS (II)

Wherein, R? is defined as above.

The invention further claims the following pharmaceutically acceptable salts of formula (I) which contain alkali metal salts, such as sodium, potassium, lithium salt, etc.; alkaline earth metal salts, such as calcium, magnesium salt, etc.; other metal salts, such as aluminum, iron, zinc, copper, nickel, cobalt salt; inorganic alkali salts, such as ammonium salt; organic salts, such as tert-octyl amine, dibenzyl amine, morpholine, glucosamine, phenylglycine alkyl ester, ethylenediamine,

N-methyl glucosamine, guanidine, diethylamine, triethylamine, dicyclohexyl ammonium,

N,N'-dibenzyl ethylenediamine, chloroprocaine, procaine, diethanolamine, N-benzyl-phenylethyl amine, piperazine, tetramethyl ammonium, tris (hydroxymethyl) amino methane; halide salts, such as hydrofluoric acid salt, hydrochloric acid salt, hydrobromide, hydrogen iodate, etc.; inorganic salts, such as nitrate, perchlorate, sulfate, phosphate, etc.; lower alkyl sulfonate such as mesylate, triflate salt, ethanesulfonate, etc.; aryl sulfonates, such as benzene sulfonate, toluene sulfonate salt; organic acid, such as acetate, malate, fumarate, succinate, citrate, tartrate, oxalate, maleate, etc.; amino acid salts, such as glycine, trimethyl glycine, arginine, ornithine, glutamate, aspartate salt.

The term “readily hydrolyzed ester”, as used hereinbefore, refers to an ester that is pharmaceutically acceptable and can be hydrolyzed to the corresponding carboxyl acid in vivo. It is obvious to those skilled in the art that a hydrolysable ester of a compoud of the present invention can be formed at free carboxyl or hydroxyl of the said compound by using conventional methods.

The invention further claims “solvates” of formula ( I), where "solvate" is referred to a compound crystal in which a solvent molecule is added to the crystal lattice during the crystallization process. If the solvent is water, solvates are known as hydrates; If the solvent is an organic solvent, the solvates are known as organic solvates.

The invention further claims the “stereoisomers” of formula ( I ), including all of enantiomers, diastereomeric isomers, racemic isomers, cis and trans isomers, tautomers, geometric isomers, epimers and mixtures thereof.

The present invention further provides that a said compound, or its pharmaceutically acceptable salts, or its readily hydrolyzed ester, or its solvate, or its stereoisomer, can be formulated by any known method of the field into any formulation that is pharmaceutically or clinically acceptable and given to a patient in need of treatment through a variety of administration methods, such as oral, and ex-gastrointestinal, and so on.

When administered orally, it can be formulated into conventional solid dosage forms, for example, tablets, capsules, pills, granules, and so on. When the compund is adminstered ex-gastrointestinally, it can be formulated into dosage forms for injection, for example, solution ready for injection, sterile power for injection, concentrated solution for injection, and so on. Such formulations can be manufactured with conventional methods in contemporary pharmaceutical manufacturing, depending on the properties of compouds, the formulation can include either no or certain additives.

The present invention also provides the use of compounds described hereinbefore in the manufacture of a medicament for the treatment and / or prophylaxis of infectious diseases. These compunds have potent activities against broad spectrum of bacteria, including gram-positive bacteria, gram-negative bacteria, aerobic bacteria, anaerobic bacteria, and clinical pathogens. They can be used to treat and / or prophylaxis diseases caused by pathogenic microorganisms,such as respiratory and urinary tract infections.

The compounds said hereinbefore, when compared to those disclosed in the most related prior art, have the following advantages: (1) The compounds said hereinbefore display excellent anti-bacterial activity and low toxicities, they could be used safely to treat and / or prevent various infectious diseases in mammals including human caused by pathogenic microorganisms. (2) The compounds said hereinbefore have strong activities against broad spectrum of bacteria, including gram-positive, gram-negative bacteria, and other clinical pathogens. : (3) The compounds said hereinbefore are stable against p-lactamase and renal dehydropeptidase-I (DHP-I), and could be used as a single therapy. (4) The compounds said hereinbefore have long elimination half-life, post-antibiotic effect, and long-lasting anti-bacterial activities, therefore it is more convenient to apply clinically. (5) The compounds said hereinbefore are relatively easy to synthesize, with high purity, high yield, and consistent quality, suitable for large-scale industrial production.

Example 1 _Anti-bacterial specturm and in vitro Activities

Test Bacteria:

Staphyloccocus aureus, Methicillin-resistant Staphylococcus Epidermidis(MRSE), Methicillin

Sensitive Staphylococcus Epidermidis(MSSE), Enterococcus faecali, Enterococcus faecom,

Streptococcus pneumoniae, Streptococcus agalactiae, Streptococcus pyogenes, Streptococcus viridans, Klebsiella pneumoniae (non-enzyme producing strain) , Klebsiella oxytoca(non-enzyme producing strain), Salmonella , Morganella morganii;

These bacteria are all purchased from Beijing Fuwai Hospital of Cardiovascular Diseases,

Shanghai Renji Hospital, Xijing Hospital Affiliated to the Fourth Military Medical University.

Test Compounds:

The compounds in the present invention are prepared in-house, and their chemical names and structures can be found in the preparation section of each compound;

Meropenem, Etapenem: Purchased form commercial sources;

Test Methods:

Agar dilution method, see reference Clinical and Laboratory Standards Institute. Methods for

Dilution Antimicrobial Susceptibility Tests for Bacteria that Grow Aerobically; Approved

Standard-Seventh Edition. CLSI Document M7-A7. Vol26, no.2, Wayne, PA: Clinical and

Laboratory Standards Institute, 2006.

Experimental Results and conclusions:

Table 2 Comparison of in vitro Anti-Bacterial Activities

Bacteria ~ Meropenem ____ Ftapenem __ Compoud 1 _ em Fenn Com poud 1

Enterococcus faecalis 4 8 2

MRSE 2 2 2 ono Deo ai 0.125 0.25 0.06

Salmonella 0.125 0.25 0.125 (ones proriacinn attain) 0.015 0.06 0.015

Morganella morganii 0.125 0.25 0.125

Streptococcus agalactiae 0.008 0.015 0.008

Streptococcus pneumoniae 0.25 0.25 0.125

Table 3 Comparison of in vitro Anti-Bacterial Activities . MIC (ng/mL)

Bacteria Meropenem Etapenem Compoud 2 Compoud 6 Compoud 7

Enterococcus faecali 2 8 1 1 2

Streptococcus 0.008 0.008 0.004 <0.002 0.004 pneumoniae

Table 4 Comparison of in vitro Anti-Bacterial Activities . MIC (pg/mL)

Bacteria Meropenem Etapenem Compoud4 Compoud5 Compoud 8

Streptococcus 0.03 0.06 0.06 0.06 0.06 agalactiae

Streptococcus 0.25 0.5 0.125 0.125 0.125 pneumoniae

Streptococcus 0.5 0.06 0.06 0.06 viridans

Table S Comparison of in vitro Anti-Bacterial Activities . MIC (ng/mL)

Bacteria Meropenem Etapenem Compoud 3 Compoud 9 Compoud 16

MRSE 0.5 2 0.5 —_— —_—

Streptococcus 0.03 0.25 0.03 — — agalactiae

Enterococcus 0.25 0.25 — 0.125 — : faecom

Staphyloccocus 0.125 0.125 aureus

Streptococcus 0.06 0.06 pneumoniae

Table 6 Comparison of in vitro Anti-Bacterial Activities

MIC (ng/mL)

Bacteria Meropenem Etapenem Compoud Compoud Compoud 12 13 14

Enterococcus 2 3 1 faecali

MSSE 0.06 0.25 0.125 —_—

Streptococcus 0.008 0.015 0.015 — — pyogenes

Streptococcus 0.03 0.06 — 0.03 0.06 agalactiae

As shown in Tables 2 to 6, the compounds of the invention demonstrate superior or equivalent antibacterial activities against bacteria tested, when compared with meropenem, they are more effective than ertapenem. 4. Examples

The present invention will be further illustrated with the following examples. However the scope of this invention is not limited in any way to these examples. Any variations and modifications obvious to one skilled in the art are intended to be included within the scope of the present invention.

Example 1 Preparation of (4R.55.,65)-6-[(R)-1-hvdroxyethyl]-4-methyl-3-[(7S.8a5)-2-methyl- octahydropyrrolo[1,2-a]pyrazin-7-ylthio]-7-0xo-1-azabicyclo[3.2.0]hept-2-ene-2-carboxylic acid (Compound 1)

HO, HO,

HO, HO, a [\ BNH,

N N ©

N CH3OH N° 2 THF 5 Aa CH3OH A _NBn

HO, HO, HO, Phyp

Li / / Boc,0 / DEAD

Lane pac _TEA _PhCOSH___

THF N CH,OH N HCOOH pom N DCM

L_NBn L_nH _NBoc

Q \

S TFA S CH,l S ¢ en L ° LS \

N $ TFA N NJ

L_NBoc NH LNG

OH CH

J, Se OH

H,C™™ n OH CH CH, pros OP(OPR, “I N fine <1

COOPNB S H S

CT N-¢/ 1 2 Ng N

NS Pd/C 0 ~

COOPNB COOH

Setp 1 Preparation of (25,4R)-methyl 4-hydroxypyrrolidine-2-carboxylate hydrochloride

HO, HO,

Oh mcoon 2220 0) —_— nN” COOH “cio nN” YCOMe

H H HCI

To a solution of (2S,4R)-4-hydroxypyrrolidine-2-carboxylic acid (250.4 g, 1.91 mol) in 1.5L of anhydrous methanol at 0 °C was treated dropwise with thionyl chloride (250.0 g, 2.1 mol). The resulting mixture was heated to reflux for 2 hours. The solvent and excess thionyl chloride was removed under reduced pressure to afford 312.4 g of desired product (90.0% yield).

Setp 2 Preparation of methyl (2S5,4R)-1-(2-chloroacetyl)-4-hydroxypyrrolidine- 2-carboxylate

Ho, ay © Ce { cope 0 N~ YCO,Me

H HCl THE AA

To a solution of (2S,4R)-methyl 4-hydroxypyrrolidine-2-carboxylate hydrochloride(312.4 g, 1.72 mol) in 3.5 L of anhydrous THF at 0 °C in an ice-bath was slowly added chloroacetyl chloride (213.4 g, 1.89 mol). The resulting mixture was heated to reflux for 2 hours. The solvent and excess chloroacetyl chloride was removed under reduced pressure to give desired product 350.2 g (91.9% yield).

Setp3 Preparation of (7R,8a5)-2-benzyl-7-hydroxyhexahydropyrrolo[1,2-a]pyrazine- 1,4-dione

HO, HO, { Moe TEA (A

NT COME NNO o Ac CH3OH . J Nan

To a solution of (2S,4R)-methyl 1-(2-chloroacetyl)-4-hydroxypyrrolidine-2-carboxylate (350.2 g, 1.58 mol) in 1.5 L of methanol was sequentially treated with benzylamine (338.6 g, 3.16 mol) and TEA (159.7 g, 1.58 mol). The resulting mixture was heated to reflux for 3 hours. After removing the solvent under reduced pressure, the residue was extracted with CH,Cl,. The organic extract was dried over anhydrous Na;SO4 and concentrated under reduced pressure. The residue was further recrystallized from CH,Cl; to give product 320.2 g (77.8% yield).

Setp 4 Preparation of (7R,8a5)-2-benzyloctahydropyrrolo[1,2-a]pyrazin-7-ol

HO, HO, / LiAIH, 7 (Ape ET CN

JANE L_NBn (7R,8aS)-2-benzyl-7-hydroxyhexahydropyrrolo[1,2-a]pyrazine-1,4-dione(320.2 g, 1.23 mol) was added in protions to a refluxing solution of lithium aluminum hydride (186.7 g, 4.92 mol) in 6.0 L of anhydrous THF. The resulting mixture was maintained at refluxing temperature for another 2 hours. After cooling to 0 °C, the excess lithium aluminum hydride was carefully quenched with water. The reaction mixture was filtered and the filtered cake was washed several times with THF.

The combined THF solution was concentrated under reduced pressure and the residue was recrystallized from ethyl acetate to afford desired product 143.8 g (52.8% yield).

Setp 5 Preparation of (7R,8aS)-octahydropyrrolo[1,2-a]pyrazin-7-ol formate

HO, HO

SN Pd/C (I

N Tm ON _NBn CH30H a oon "To a solution of (7R,8aS)-2-benzyloctahydropyrrolo[1,2-a]pyrazin-7-ol (143.8 g, 0.65 mol) in methanol (1.0L) was sequentially added Palladium-on carbon (35.2 g) and formic acid (33.1g, 0.72 mol). The resulting black suspension was stirred at 40 °C for 10 hours under hydrogen pressure (1 atm). The catalyst was removed by filtering through a short pad of Celite and the filtered cake was rinsed with additional methanol. The filtrate was concentrated under reduced pressure to give product 120.4 g (98.5% yield) which was used directly in next step without further purification.

Setp 6 Preparation of (7R,8aS)-tert-butyl 7-hydroxyhexahydropyrrolo[1,2-a]pyrazine- 2(1H)-carboxylate

He, Boc,0 HO,

A

Ld oo beM 1 Yipee

To a solution of (7R,8aS)-octahydropyrrolo[1,2-a]pyrazin-7-ol formate (120.4 g, 0.64 mol) in 600 mL of CH,Cl, was cooled to -10 °C in a salt-ice bath and treated with TEA (135.5 g, 1.34 mol).

After stirring for 10 min, di-tert-butyl dicarbonate (138.2 g, 0.64 mol) was added to the reaction mixture. The reaction mixture was stirred at -10 °C for 4 hours. The reaction mixture was washed twice with saturated NaHCO; solution and one time with brine. The resulting mixture was dried over anhydrous Na,SOy, filtered and concentrated under reduced pressure to give product 135.7 g (87.5% yield).

Setp 7 Preparation of (7S,8aS)-ferr-butyl 7-(benzoylthio)hexahydropyrrolo[1,2-a}- pyrazine-2(1H)-carboxylate

HO, Ph,P “2

DEAD S

(A eo, L

L_NBoe N

LNB

Triphenylphosphine (264.9 g, 1.02 mol) in 700 mL of CH,Cl, was cooled to -10 °C and stirred for 15 min. The resulting solution was treated with DEAD (175.9 g, 1.01 mol) and stirred for 1 hour followed by adding thiobenzoic acid (138.8 g, 1.01 mol). The resulting mixture was stirred for additional 1 hour and treated dropwise with (7R,8aS)-fert-butyl 7-hydroxy- hexahydropyrrolo[1,2-a]pyrazine-2(1 H)-carboxylate (135.7 g, 0.56 mol) in 300 mL of CH,Cl,. The reaction mixture was stirred at -10 °C for 6 hours. The solvent was removed under reduced pressure and the residue was extracted with ethyl acetate. The extract was concentrated and the residue was purified by flash column chromatography on silica gel to give product 137.7 g (67.8% yield).

Setp 8 Preparation of S-(75,8aS)-octahydropyrrolo[1,2-a]pyrazin-7-yl benzothioate a Q.

S TFA o

See UN __NBoe NH TFA

A solution of (7S,8aS)-tert-butyl 7-(benzoylthio)hexahydropyrrolo[1,2-a]pyrazine-2(1H)- carboxylate (137.7 g, 0.38 mol) in CH,Cl, (600 mL) was cooled to 0 °C and treated with trifluoroacetic acid (173.3 g, 1.52 mol). The resulting mixture was stirred at 0 °C for 4 hours. The reaction mixture was trituated with diethyl ether and the resulting precipitate was collected by filtration to afford product 89.8 g (86.8% yield).

Setp 9 Preparation of S-(75,8aS8)-2-methyloctahydropyrrolo[1,2-a]pyrazin-7-yl benzothioate :

CTS IN

N N

\L_NH TFA _N~

To a solution of S-(7S,8aS)-octahydropyrrolo[1,2-a]pyrazin-7-yl benzothioate (3.59 g, 10 mmol) in CH,Cl, (100 mL) at 0 °C was treated with TEA (2 g, 20 mmol) and iodomethane (1.42 g, mmol). The resulting mixture was stirred at 0 °C for overnight. The reaction mixture was washed with brine, dried over anhydrous Na,SO,4 and concentrated under reduced pressure to afford crude product which was used directly in next step without further purification.

Setp 10 Preparation of (75,8aS)-2-methyloctahydropyrrolo[1,2-a]pyrazine-7-thiol

S

LS ON

LN

The crude product of previous step was dissolved in methanol and was added to a saturated methanolic KOH solution at 0 °C. The reaction mixture was stirred at 0 °C for 2 hours. The reaction mixture was adjusted to pH=6 to 7 with IN HCI solution. Methanol was removed and the residue was extracted with CH,Cl,, dried over anhydrous Na,SO,4 and concentrated under reduced pressure to give product 0.82 g (48%).

Setp 11 Preparation of 4-nitrobenzyl (4R,5S,6S)-6-[(R)-1-hydroxyethyl]-4-methyl-3- [(75,8aS)-2-methyloctahydropyrrolo[1,2-a]pyrazin-7-ylthio]-7-0xo-1-azabicyclo[3.2.0]- hept-2-ene-2-carboxylate

OH CH

HS H,C Coho, pu CH, 0 FN CH; NT

OO \ COOPNB = ~ COOPNB : In a dried reaction flask, 0.52 g of (7S,8aS)-2-methyloctahydropyrrolo[1,2-ajpyrazine-7-thiol (3 mmol) was dissolved in acetonitrile. To this solution was added 4-nitrobenzyl (4R,5R,6S)-3-(diphenoxyphosphoryloxy)-6-[(R)-1-hydroxyethyl]-4-methyl-7-oxo-1-azabicyclo[3.2. 0]hept-2-ene-2-carboxylate (1.78 g, 3 mmol) and DIEA (0.39 g, 3 mmol). The reaction mixture was stirred at ambient temperature for overnight. The reaction mixture was extracted with ethyl acetate and washed with water. The organic layer was dried over anhydrous Na;SOy, fitered and concentrated under reduced pressure to give crude product 1.39 g (90% yield) which was used directly in next step without further purification.

Setp 12 Preparation of (4R,5S,65)-6-[(R)-1-hydroxyethyl]-4-methyl-3-[(7S,8aS)-2- methyloctahydropyrrolo[1,2-a]pyrazin-7-ylthio]-7-oxe-1-azabicyclo[3.2.0]hept-2-ene-2- carboxylic acid

OH OH

CY u Ay

Oo Ny NS To o” { NS

COOPNB COOH

The crude product (3.0 g, 6 mmol) from step 11 was dissolved in a 1:1 mixture of THF and water (20 mL) and 10% Pd/C was added. The resulting black suspension was stirred at ambient temperature under hydrogen pressure for 2 hours. The catalyst was filtered off and rinsed with THF and de-ionized water. The combined filtrate was extracted with ethyl acetate. The product in aqueous layer was purified by prep. HPLC and the product fractions was freeze-dried to give solid product (21 mg, 1% yield).

Formula: C;gH27N304S Mol. Wt: 381.49 LC-MS: 382 M+H") 'H NMR (400MHz, D,0) 4.13 (t, 1H), 4.07 (dd, 1H) , 3.67 (t, 1H), 3.29 (dd, 1H), 3.21 (t, 1H), 2.88-2.94 (m, 3H), 2.76 (m, 1H), 2.57 (dd, 1H), 2.28-2.34 (m, 1H), 2.17-2.22 (m, 6H), 1.96 (t, 1H), 1.14-1.22 (m, 4H), 1.08 (d, 3H, ).

Example 2 Preparation of (4R,5S.65)-6-[(R)-1-hydroxyethyl]-3-[(7S,8a5)-2-Isopropyl- octahydropyrrolo[1,2-ajpyrazin-7-ylthio |-4-methyl-7-oxo-1-azabicyclo [3.2.0]hept-2-ene-2- carboxylic acid (Compound 2)

Cl

Dpm, 2 pom 0m 0 N_ NH TFA 0 Nn Nr ei & Q OH

Johor pn cH, He J, CH, y A © COOPNB NN coon

Setp 1-8 Procedures were same as step 1-8 in Example 1.

Setp 9 Preparation of S-(75,8aS)-2-isopropyloctahydropyrrolo[1,2-a]pyrazin-7- yl benzothioate

Cl

J A Os — o N_ NH TFA 0 NN

To a solution of S-(7S,8aS)-octahydropyrrolo[1,2-a]pyrazin-7-yl benzothioate (3.59 g, 10 mmol) in CH,Cl, (100 mL) was treated with TEA (2 g, 20 mmol) and 2-chloropropane (0.785 g, 10 mmol). The resulting mixture was stirred at 0 °C for overnight. The reaction mixture was washed with brine, dried over anhydrous Na,SQ,, filtered and concentrated under reduced pressure. The crude residue was used directly in next step without further purification.

Setp 10 Preparation of (75,8aS)-2-isopropyloctahydropyrrolo[1,2-a]pyrazine-7-thiol

Dr

Co — TO

The crude product of previous step was dissolved in methanol and was added to a saturated methanolic KOH solution at 0 °C. The reaction mixture was stirred at 0 °C for 2 hours. The reaction mixture was adjusted to pH=6 to 7 with IN HCl solution. Methanol was removed and the residue was extracted with CH,Cl,, dried over anhydrous Na;SO, and concentrated under reduced pressure to give product 0.84 g (42%).

Setp 11 Preparation of 4-nitrobenzyl (4R,5S,65)-6-[(R)-1-hydroxyethyl]-3-[(7S,8a5)-2- isopropyloctahydropyrrolo[1,2-a]pyrazin-7-ylthio]-4-methyl-7-oxo-1-azabicyclo[3.2.0]- hept-2-ene-2-carboxylate

OH CH,

Hs Wr p-ohom ai am A

TI 0 COOPNB pros —1 —/ — COOPNB

In a dried reaction flask, 0.60 g of (7S,8aS)-2-isopropyloctahydropyrrolo[1,2-a]pyrazine- 7-thiol (3 mmol) was dissolved in acetonitrile. To this solution was added 4-nitrobenzyl (4R,5R,6S)-3-(diphenoxyphosphoryloxy)-6-[(R)-1-hydroxyethyl]-4-methyl-7-oxo-1-azabicyclo- [3.2.0]hept-2-ene-2-carboxylate (1.78 g, 3 mmol) and DIEA (0.39 g, 3 mmol). The reaction mixture was stirred at ambient temperature for overnight. The reaction mixture was extracted with ethyl acetate and washed with water. The organic layer was dried over anhydrous Na;SO,, fitered and concentrated under reduced pressure to give crude product 1.30 g (80% yield) which was used directly in next step without further purification.

Setp 12 Preparation of (4R,5S,65)-6-[(R)-1-hydroxyethyl]-3-[(7S,8aS)-2-isopropyl- octahydropyrrolo[1,2-a]pyrazin-7-ylthio]-4-methyl-7-oxo-1-azabicyclo[3.2.0]hept-2-ene-2- carboxylic acid

OH CH, OH cH = UN H, eet Or © ’ COOPNB NN hic © ’ {on ne

The crude product (2.0 g, 3.68 mmol) from step 11 of Example 2 was dissolved in a 1:1 mixture of THF and water (20 mL) and 10% Pd/C (1 g) was added. The resulting black suspension was stirred at ambient temperature under hydrogen pressure for 2 hours. The catalyst was filtered off and rinsed with THF and de-ionized water. The combined filtrate was extracted with ethyl acetate. The product in aqueous layer was purified by prep. HPLC and the product fractions was freeze-dried to give solid product (70 mg, 4.6% yield).

Formula: C;oH31N304S Mol. Wt: 409.54 LC-MS: 410(M+H") 'H NMR (400MHz, D,0) 4.05-4.15 (m, 2H), 3.73 (t, 1H,), 3.54 (d, 1H), 3.37-3.47 (m, 2H), 3.29 (dd, 1H), 3.14-3.23 (m, 2H), 2.94-3.01 (m, 2H), 2.78 (t, 1H), 2.65-2.69 (m, 1H), 2.40-2.48 (m, 3H), 1.22-1.31 (m, 7H), 1.17 (d, 3H), 1.08 (d, 3H).

Example 3 Preparation of (4R,5S5,65)-3-[(7S,8a8)-2-benzyloctahydropyrrolo[1,2-a]- pyrazin-7-ylthio]-6-[(R)-1-hydroxyethyl]-4-methyl-7-0x0-1-azabicyclo[3.2.0] hept-2-ene-2- carboxylic acid (Compound 3)

Ae “J Ls NO 0 Tw TFA —— 0 Te —— TN N-Bnm a CH, " pros oko, tl Bn “He D -

COOPNB s—C N-/ N — oS {ooms Nt ? COOH

Setp 1-8 Procedures were same as step 1-8 in Example 1.

Setp 9 Preparation of S-(78,8aS5)-2-benzyloctahydropyrrolo[1,2-a]pyrazin-7-yl benzothioate

I Tw ~~ T Ten

To a solution of S-(7S,8aS)-octahydropyrrolo[1,2-a]pyrazin-7-yl benzothioate (3.59 g, 10 mmol) in CH;Cl, (100 mL) was treated with TEA (2 g, 20 mmol) and benzyl bromide (1.71 g, 10 mmol). The resulting mixture was stirred at 0 °C for overnight. The reaction mixture was washed with brine, dried over anhydrous Na,SOy, filtered and concentrated under reduced pressure. The crude residue was used directly in next step without further purification.

Setp 10 Preparation of (75,8aS)-2-benzyloctahydropyrrolo{1,2-a]pyrazine-7-thiol

Qs Hs 0 Ten Ce

The crude product of previous step was dissolved in methanol and was added to a saturated methanolic KOH solution at 0 °C. The reaction mixture was stirred at 0 °C for 2 hours. The reaction mixture was adjusted to pH=6 to 7 with IN HCl solution. Methanol was removed and the residue was extracted with CH,Cl,, dried over anhydrous Na,SO4 and concentrated under reduced pressure to give product(7S,8aS)-2-benzyloctahydropyrrolo[ 1,2-a]pyrazine-7-thiol (1.31g, 53%).

Setp 11 Preparation of 4-nitrobenzyl (4R,5S,65)-3-[(7S,8aS5)-2-benzyloctahydropyrrolo- [1,2-a]pyrazin-7-ylthio]-6-[(R)-1-hydroxyethyl]-4-methyl-7-oxo-1-azabicyclo[3.2.0]hept-2-ene- 2-carboxylate fe rd OPh OH

HS o Nf or cH. & _Bn —/ © COOPNB

In a dried reaction flask, 0.74 g of (7S,8aS)-2-benzyloctahydropyrrolo[1,2-a]pyrazine-7-thiol (3 mmol) was dissolved in acetonitrile. To this solution was added 4-nitrobenzyl (4R,5R,6S)-3-(diphenoxyphosphoryloxy)-6-[(R)-1-hydroxyethyl]-4-methyl-7-oxo-1 -azabicyclo- [3.2.0]hept-2-ene-2-carboxylate (1.78 g, 3 mmol) and DIEA (0.39 g, 3 mmol). The reaction mixture was stirred at ambient temperature for overnight. The reaction mixture was extracted with ethyl acetate and washed with water. The organic layer was dried over anhydrous Na;SOsa, fitered and concentrated under reduced pressure to give crude product 1.56 g (88% yield) which was used directly in next step without further purification.

Setp 12 Preparation of (4R,5S,65)-3-[(7S,8aS)-2-benzyloctahydropyrrolo- [1,2-a]pyrazin-7-ylthio]-6-[(R)-1-hydroxyethyl]-4-methyl-7-oxo-1-azabicyclo[3.2.0]hept-2-ene- 2-carboxylic acid

OH OH

“rs Cy” ees Oy .Bn

COOPNB © COOH

The crude product (0.59 g, 1.0 mmol) from step 11 of Example 3 was dissolved in a 1:1 mixture of THF and water (20 mL) and 10% Pd/C (1 g) was added. The resulting black suspension was stirred at ambient temperature under hydrogen pressure for 2 hours. The catalyst was filtered off and rinsed with THF and de-ionized water. The combined filtrate was extracted with ethyl acetate. The product in aqueous layer was purified by prep. HPLC and the product fractions was freeze-dried to give solid product (70 mg, 4.6% yield).

Formula: CpH3N30,S ~~ Mol. Wt: 457.59 LC-MS: 458(M+H") 'H NMR (400MHz, DLC-MSO-dg) 7.24-7.33 (m, 5H), & 5.01(s,1H), 4.11-4.12(m, 1H), 4.09-4.10 (m, 1H), 3.91-3.95 (m ,1H), 3.45-3.56 (d, J/=13.2Hz, 2H), 3.23-3.27 (m ,1H), 3.15-3.17 (m ,1H), 3.15-3.17 (m ,1H), 3.15-3.17 (m ,1H), 2.85-2.91 (m, 3H), 2.70-2.72(m ,1H), 2.50-2.52 (m ,1H), 2.11-2.15 (m ,1H), 2.07-2.11 (m ,3H) ,1.79-1.82 (m ,1H) ,1.07-1.48 (m ,6H).

Example 4 Preparation of (4R.5S,65)-3-[(7S5.8a5)-2-acetyloctahydropyrrolo[1,2-a]pyrazin- 7-ylthio]-6-[(R)-1-hydroxyethyl]-4-methyl-7-0x0-1-azabicyclo[3.2.0]hept-2-ene-2-carboxylic acid (Compound 4) oO

S Ae OH 0 HS

T= en 0 SN NAT Aa 4 oO N N

N NHTFA | J ) < ~ OH

Hye CC" Q OH OH 4 / OP(OPh), J CH, 0 cn, CH, 1 0 coors C8 WN <1 s H Ng 3 - NZ 1 Se 0 NS 0 Pd/C COOH

COOPNB

Setp 1-8 Procedures were same as step 1-8 in Example 1.

Setp 9 Preparation of S-(75,8aS)-2-acetyloctahydropyrrolo[1,2-a]pyrazin-7-yl benzothioate 0 s Aa OH ©

To < o N NHTFA n_/

To a solution of S-(7S,8aS)-octahydropyrrolo[1,2-a]pyrazin-7-yl benzothioate (3.59 g, 10 mmol) in CH,Cl, (100 mL) was treated with TEA (2 g, 20 mmol) and acetyl chloride (0.78 g, 10 mmol). The resulting mixture was stirred at 0 °C for 2 hours. The reaction mixture was washed with brine, dried over anhydrous Na,SOy, filtered and concentrated under reduced pressure. The crude residue was used directly in next step without further purification.

Setp 10 Preparation of 1-[(7S,8a5)-7-mercaptohexahydropyrrolo[1,2-a]pyrazin- 2(1H)-yl]ethanone

Dora o "NO o

O NN ~ - NK

The crude product of previous step was dissolved in methanol and was added to a saturated methanolic KOH solution at 0 °C. The reaction mixture was stirred at 0 °C for 2 hours. The reaction mixture was adjusted to pH=6 to 7 with 1N HCI solution. Methanol was removed and the residue was extracted with CH,Cl,, dried over anhydrous Na;SO, and concentrated under reduced pressure to give product 1-[(7S,8aS)-7-mercaptohexahydropyrrolo[1,2-a]pyrazin-2(1 H)-yl]ethanone (1.06 g, 53%).

Setp 11 Preparation of 4-nitrobenzyl (4R,5S,65)-3-[(7S,8aS)-2-acetyloctahydropyrrolo- [1,2-a]pyrazin-7-ylthio]-6-[(R)-1-hydroxyethyl]-4-methyl-7-oxo-1-azabicyclo[3.2.0] hept-2-ene- 2-carboxylate

OH CH

HC BR: OH tmp art BL

N NL - o N NA —/ COOPNB

In a dried reaction flask, 0.6 g of (7S,8aS)-2-acetyloctahydropyrrolo[ 1,2-a]pyrazine-7-thiol (3 mmol) was dissolved in acetonitrile. To this solution was added 4-nitrobenzyl (4R,5R,6S)-3- (diphenoxyphosphoryloxy)-6-[(R)-1-hydroxyethyl]-4-methyl-7-oxo-1-azabicyclo[3.2.0]hept-2-ene- 2-carboxylate (1.78 g, 3 mmol) and DIEA (0.39 g, 3 mmol). The reaction mixture was stirred at ambient temperature for overnight. The reaction mixture was extracted with ethyl acetate and washed with water. The organic layer was dried over anhydrous Na;SOs, fitered and concentrated under reduced pressure to give crude product 1.43 g (88% yield) which was used directly in next step without further purification.

Setp 12 Preparation of (4R,5S,65)-3-[(7S5,8a5)-2-acetyloctahydropyrrolo[1,2-a]- pyrazin-7-ylthio]-6-[(R)-1-hydroxyethyl]-4-methyl-7-oxo-1-azabicyclo[3.2.0]hept-2-ene-2- carboxylic acid go em 0 7 en 0

CH; *., J CH; 7. J 0 pic ©

COOPNB COOH

The crude product (0.7 g, 1.28 mmol) from step 11 of Example 4 was dissolved in a 1:1 mixture of THF and water (20 mL) and 10% Pd/C (0.3 g) was added. The resulting black suspension was stirred at ambient temperature under hydrogen pressure for 2 hours. The catalyst was filtered off and rinsed with THF and de-ionized water. The combined filtrate was extracted with ethyl acetate. The product in aqueous layer was purified by prep. HPLC and the product fractions was freeze-dried to give solid product (47 mg, 9.0% yield).

Due to the amide bond on the octahydropyrrolo[1,2-a]- pyrazine ring, 'H NMR exhibits two conformers.

Formula: C;9H;7N305S Mol. Wt: 409.50 LC-MS: 410(M+H") 'H NMR (400 MHz, D,0) § 4.42 (d, 0.5H), & 4.27 (d, 0.5H), 8 4.07-4.15 (m, 2H), § 3.97 (d, 0.5H), 8 3.81 (d, 0.5H), 8 3.67-3.71 (m, 1H), & 3.31 (d, 1H), 6 3.17-3.24 (m, 1.5H), 6 2.90-2.99 (m, 2.5H), 8 2.72 (t, 0.5H), 8 2.61 (t, 1H), & 2.48 (t, 0.5H), § 2.33-2.39 (m, 1H), 8 2.10-2.22 (m, 1H), 6 2.03-2.13 (m, 1H), 8 2.04 (d, 3H), 8 1.17-1.27 (m, 4H), 61.09 (d, 3H).

Example 5 Preparation of (4R.5S,65)-6-[(R)-1-hydroxyethyl]-3-[(75.8a5)-2-isobutvryl- octahydropyrrolo[1,2-a]pyrazin-7-ylthio]-4-methyl-7-oxo-1-azabicyclo[3.2.0]hept-2-ene-2- carboxylic acid (Compound 5) 0

Oy q Nhe OO Sm 0 0 0 Tw TFA 0 OL a \_/ n—/ 0 ye ba CH, o vo OP(OPh), OH OH

N- og CH, 0 J CH, 9

S —2 / S

PNY NS paC 47 N nN

COOPNB COOH

Setp 1-8 Procedures were same as step 1-8 in. Example 1.

Step 9 Preparation of S-(7S,8aS)-2-isobutyryloctahydropyrrolo[1,2-a]pyrazin-7-yl benzothioate

0

Oppo Te Ofms 0 Ta TFA — © Ne —/

To a solution of S-(7S,8aS)-octahydropyrrolo[1,2-a]pyrazin-7-yl benzothioate (3.59 g, 10 mmol) in CH,Cl,, (100 mL) was treated with TEA (2 g, 20 mmol) and isobutyryl chloride (1.07 g, mmol). The resulting mixture was stirred at 0 °C for 2 hours. The reaction mixture was washed with brine, dried over anhydrous Na;SO,, filtered and concentrated under reduced pressure. The crude residue was used directly in next step without further purification.

Setp 10 Preparation of 1-[(7S,8aS)-7-mercaptohexahydropyrrolo[1,2-a]pyrazin- 2(1H)-yl]-2-methylpropan-1-one 0 “NN Da NH

The crude product of previous step was dissolved in methanol and was added to a saturated methanolic KOH solution at 0 °C. The reaction mixture was stirred at 0 °C for 2 hours. The reaction mixture was adjusted to pH=6 to 7 with IN HCI solution. Methanol was removed and the residue was extracted with CH,Cl, dried over anhydrous Na,SO, and concentrated under reduced pressure to give product (1.41 g, 62%).

Setp 11 Preparation of 4-nitrobenzyl (4R,5S,65)-6-[(R)-1-hydroxyethyl]-3-[(7S,8a5)-2- isobutyryloctahydropyrrolo[1,2-a]pyrazin-7-ylthio]-4-methyl-7-0xo-1-azabicyclo[3.2.0]- hept-2-ene-2-carboxylate

H;C 5 Le Q

T= o COOPNB ~~ HC, YY

Ain E —_— Pros STAN

COOPNB

In a dried reaction flask, 0.68 g of 1-[(7S,8aS)-7-mercaptohexahydropyrrolo[ 1,2-a]pyrazin- 2(1H)-yl]-2-methylpropan-1-one (3 mmol) was dissolved in acetonitrile. To this solution was added 4-nitrobenzyl (4R,5R,6S)-3-(diphenoxyphosphoryloxy)-6-[(R)-1-hydroxyethyl]-4-methyl- 7-0x0-1-azabicyclo-[3.2.0]hept-2-ene-2-carboxylate (1.78 g, 3 mmol) and DIEA (0.39 g, 3 mmol).

The reaction mixture was stirred at ambient temperature for overnight. The reaction mixture was extracted with ethyl acetate and washed with water. The organic layer was dried over anhydrous

Na,SO0,, fitered and concentrated under reduced pressure to give crude product 1.51 g (88% yield) which was used directly in next step without further purification.

Setp 12 Preparation of (4R,5S,65)-6-[(R)-1-hydroxyethyl]-3-[(7S,8aS)-2-isobutyrylocta- hydropyrrolo[1,2-a]pyrazin-7-ylthio]-4-methyl-7-oxo-1-azabicyclo[3.2.0]hept-2-ene-2- carboxylic acid gh om 0 gn a

Sma Sc oN J NN PAC ZN 7 nN

COOPNB COOH

The crude product (2.0 g, 3.5 mmol) from step 11 of Example 5 was dissolved in a 1:1 mixture of THF and water (20 mL) and 10% Pd/C (1.0 g) was added. The resulting black suspension was stirred at ambient temperature under hydrogen pressure for 2 hours. The catalyst was filtered off and rinsed with THF and de-ionized water. The combined filtrate was extracted with ethyl acetate.

The product in aqueous layer was purified by prep. HPLC and the product fractions was freeze-dried to give solid product 52 mg (3.4% yield).

Formula: C;1H3N305S Mol. Wt: 437.55 LC-MS: 438(M+H") 'H NMR (400 MHz, D,0) § 4.44 (d, 0.5H), 3 4.31 (d, 0.5H), 3 4.04-4.14 (m, 2.5H), 6 3.96 (d, 0.5H), 8 3.65-3.67 (m, 1 H), § 3.27 (dd, 1H), 5 3.16-3.21 (m, 1.5H), 3 2.86-2.96 (m, 3.5H), 6 2.67 (t, : 0.5H), 6 2.58 (t, 1H), & 2.44 (t, 0.5H), 6 2.31-2.38 (m, 1H), § 2.12-2.26 (m, 1H), 6 2.04-2.08 (m, 1H), 6 1.15-1.24 (m, 4H), 81.07 (d, 3H), 3 0.91-0.96 (m, 6H).

Example 6 Preparation of _ (4R,55,65)-3-[(75.,8aS)-2-benzoyloctahydropyrrolo[1.2-a]- pyrazin-7-ylthio]-6-[(R)-1-hydroxyethyl]-4-methyl-7-oxo-1-azabicyclo[3.2.0]hept-2-ene-2- carboxylic acid (Compound 6)

0 9S o Se HS 3 (J T= p Top 0 N N N N 0 Tr, \_ ba _/ hs /

OH

H,C Le Q - / oN NM

COOPNB

H, 7 Ho con, 9

Pd/C HC 0 /

ZN NJ

COOH

Setp 1-8 Procedures were same as step 1-8 in Example 1.

Setp 9 Preparation of S-(75,8aS)-2-benzoyloctahydropyrrolo[1,2-a]pyrazin-7-yl benzothioate

Oo

O N NHTFA —————> n_/

To a solution of S-(7S,8aS)-octahydropyrrolo[1,2-a]pyrazin-7-yl benzothioate (3.59 g, 10 mmol) in CH,Cl, (100 mL) was treated with TEA (2 g, 20 mmol) and benzoyl chloride (1.4 g, 10 mmol). The resulting mixture was stirred at 0 °C for 2 hours. The reaction mixture was washed with brine, dried over anhydrous Na,SOy, filtered and concentrated under reduced pressure. The crude residue was used directly in next step without further purification.

Setp 10 Preparation of [(7S5,8aS)-7-mercaptohexahydropyrrolo[1,2-a]pyrazin- 2(1H)-yl](phenyl)methanone

To—= p To ¢ oO : N N

N N » \ ,

The crude product of previous step was dissolved in methanol and was added to a saturated methanolic KOH solution at 0 °C. The reaction mixture was stirred at 0 °C for 2 hours. The reaction :

mixture was adjusted to pH=6 to 7 with IN HCI solution. Methanol was removed and the residue : was extracted with CH,Cl,, dried over anhydrous Na,SOj4 and concentrated under reduced pressure to give product (1.86 g, 71%).

Setp 11 Preparation of 4-nitrobenzyl (4R,5S,65)-3-[(7S5,8aS)-2-benzoyloctahydro- pyrrolo[1,2-a]pyrazin-7-ylthio]-6-[(R)-1-hydroxyethyl]-4-methyl-7-oxo-1-azabicyclo[3.2.0]hept -2-ene-2-carboxylate

HS ac CC" ?

NN oO COOPNB H,C™ ~, N bs — 0 —C 0 oO

COOPNB

In a dried reaction flask, 0.78 g of ((7S,8aS)-7-mercaptohexahydropyrrolo[1,2-a]pyrazin- 2(1H)-yl)(phenyl)methanone (3 mmol) was dissolved in acetonitrile. To this solution was added 4-nitrobenzyl (4R,5R,6S)-3-(diphenoxyphosphoryloxy)-6-[(R)-1-hydroxyethyl]-4-methyl-7-oxo-1- azabicyclo-[3.2.0Thept-2-ene-2-carboxylate (1.78 g, 3 mmol) and DIEA (0.39 g, 3 mmol). The reaction mixture was stirred at ambient temperature for overnight. The reaction mixture was extracted with ethyl acetate and washed with water. The organic layer was dried over anhydrous

Na, S04, fitered and concentrated under reduced pressure to give crude product 1.32 g (73% yield) which was used directly in next step without further purification.

Setp 12 Preparation of (4R,55,65)-3-[(7S,8aS)-2-benzoyloctahydropyrrolo[1,2-a]- pyrazin-7-ylthio]-6-[(R)-1-hydroxyethyl]-4-methyl-7-oxo-1-azabicyclo[3.2.0]hept-2-ene-2- carboxylic acid wed Eb i gem 9 uo N H HC N

COOPNB COOH

The crude product (0.8 g, 1.32 mmol) from step 11 of Example 6 was dissolved in a 1:1 mixture of THF and water (20 mL) and 10% Pd/C (0.4 g) was added. The resulting black suspension was stirred at ambient temperature under hydrogen pressure for 2 hours. The catalyst was filtered off and rinsed with THF and de-ionized water. The combined filtrate was extracted with ethyl acetate. The product in aqueous layer was purified by prep. HPLC and the product fractions was freeze-dried to give solid product 55 mg (8.7% yield).

Formula: C4H29N305S Mol. Wt: 471.57 LC-MS: 472(M+H") 'H NMR (400 MHz ,D,0) 7.39-7.45 (m, 3H), 7.31-7.34 (m, 2H), 4.58 (d, 0.5H), 4.44 (d, 0.5H), 4.13 (t, 1H), 4.07 (dd, 1H), 3.61-3.74 (m, 2H), 3.28-3.32 (m, 1H), 2.93-3.20 (m, 2.5H), 2.86 (d, 0.5H), 2.82 (d, 0.5H), 2.68 (t, 0.5H), 2.62 (t, 1H), 2.39 (m, 0.5H), 2.14-2.27 (m, 2.5H), 1.25-1.35 (m, 0.5H), 1.18 (d, 3H), 1.05-1.11 (m, 3.5H).

Example 7 Preparation of (4R.5S,65)-6-[(R)-1-hydroxyethyl]-4-methyl-7-o0x0-3-](7S.8aS)-2- picolinoyloctahydropyrrolo[1,2-alpyrazin-7-ylthio]-1-azabicyclo[3.2.0] hept-2-ene-2-carboxylic acid (Compound 7) 0 Qs HS

Ane (yo © To ? On )

OH CH, me “pom, we re N I Ny © COOPNB . IC ap

COOPNB gr CH, 0 wm HC, SSR ®

P4IC J a)

COOH

Setp 1-8 Procedures were same as step 1-8 in Example 1.

Setp 9 Preparation of S-(75,8aS)-2-picolinoyloctahydropyrrolo[1,2-a]pyrazin-7-yl benzothioate

QL a

RR CY 0 AN ¢ © N Mpa ry

To a solution of S-(7S,8aS)-octahydropyrrolo[1,2-a]pyrazin-7-yl benzothioate (3.59 g, 10 mmol) in CHCl, (100 mL) was treated with TEA (2 g, 20 mmol) and benzoyl chloride (1.4 g, 10 mmol). The resulting mixture was stirred at 0 °C for 2 hours. The reaction mixture was washed with brine, dried over anhydrous Na;SOs, filtered and concentrated under reduced pressure. The crude residue was used directly in next step without further purification.

Setp 10 Preparation of [(75,8aS)-7-mercaptohexahydropyrrolo[1,2-a]pyrazin-2(1H)- yl](pyridin-2-yl)methanone

A HS

0 TH) ? On ? om Yo

The crude product of previous step was dissolved in methanol and was added to a saturated methanolic KOH solution at 0 °C. The reaction mixture was stirred at 0 °C for 2 hours. The reaction mixture was adjusted to pH=6 to 7 with IN HCI solution. Methanol was removed and the residue was extracted with CH,Cl,, dried over anhydrous Na,SO4 and concentrated under reduced pressure to give product (2.23 g, 85%).

Setp 11 Preparation of 4-nitrobenzyl (4R,SS,65)-6-[(R)-1-hydroxyethyl}-4-methyl-7-oxo- 3-[(75,8aS)-2-picolinoyloctahydropyrrolo[1,2-a]pyrazin-7-ylthio]-1-azabicyclo[3.2.0]hept-2- ene-2-carboxylate

OH CH, / bs COOPNB . I NJ P — COOPNB

In a dried reaction flask, 0.79 g of [(7S,8aS)-7-mercaptohexahydropyrrolo[1,2-a]pyrazin- 2(1H)-yl](pyridin-2-y)methanone (3 mmol) was dissolved in acetonitrile. To this solution was added 4-nitrobenzyl (4R,5R,6S)-3-(diphenoxyphosphoryloxy)-6-[(R)-1-hydroxyethyl]-4-methyl- 7-0x0-1-azabicyclo-[3.2.0]hept-2-ene-2-carboxylate (1.78 g, 3 mmol) and DIEA (0.39 g, 3 mmol).

Na,SOy, fitered and concentrated under reduced pressure to give crude product 1.33 g (73% yield) which was used directly in next step without further purification.

Setp 12 Preparation of (4R,55,65)-6-[(R)-1-hydroxyethyl]-4-methyl-7-o0xo0-3-[(75,8a5)-2- picolinoyloctahydropyrrolo[1,2-a]pyrazin-7-ylthio]-1-azabicyclo[3.2.0]hept-2-ene-2-carboxylic acid

OH wet, 0 ng fhe = as Pd/C oF N { NS Z

COOPNB

The crude product (0.8 g, 1.32 mmol) from step 11 of Example 7 was dissolved in a 1:1 mixture of THF and water (20 mL) and 10% Pd/C (0.4 g) was added. The resulting black suspension was stirred at ambient temperature under hydrogen pressure for 2 hours. The catalyst was filtered off and rinsed with THF and de-ionized water. The combined filtrate was extracted with ethyl acetate. The product in aqueous layer was purified by prep. HPLC and the product fractions was freeze-dried to give solid product 50 mg (8.2% yield).

Formula: Co3HsN4O5S Mol. Wt: 472.56 LC-MS: 473(M+H") 'H NMR (400 MHz, D,0) 8.46 (d, 1H), 7.89 (t, 1H), 7.40-7.47 (m, 2H), 4.58 (4, 0.5H), 4.41 (4, 0.5H), 4.04-4.13 (m, 2H), 3.61-3.71 (m, 1.5H), 3.46 (d, 0.5H), 3.26-3.30 (m, 1H), 3.13-3.22 (m, 1.5H), 2.90-3.07 (m, 2.5H), 2.69 (d, 0.5H), 2.59 (t, 0.5H), 2.58-2.65 (m, 1H), 2.39-2.43 (m, 0.5H), 2.12-2.26 (m, 2.5H), 1.23-1.32 (m, 0.5H), 1.16 (d, 3H), 1.04-1.10 (m, 3.5H).

Example 8 Preparation of (4R,5S,65)-6-[(R)-1-hydroxyethyl]-4-methyl-7-0x0-3-[(75.8aS)- 2-(4-sulfamoylbenzoyl)octahydropyrrolo[1,2-glpyrazin-7-ylthio]-1-azabicyclo[3.2.0]hept-2-ene -2-carboxylic acid (Compound 8) or AA oY 0 Th TEA

HNO, ——— H,NO,S -

A | HS ba oo

I Om, ° To 0 HC [p—okom, / 0 — 0 0 COOPNB — —_—

SO,NH, SO,NH, uy Ce el , act & . 1

COOPNB COOH

Setp 1-8 Procedures were same as step 1-8 in Example 1.

Setp9 Preparation of isobutyric 4-sulfamoylbenzoic anhydride 0 0 0 0 or Ho rey

H,NO,S H,NO,S

To a solution of 4-sulfamoylbenzoic acid (11.2 g, 56 mmol) in CH,Cl, was cooled to -10 °C and treated with TEA (12.5 g, 124 mmol) followed by dropwise addition of isopropylchloroformate (11.5 g, 93 mmol). The resulting mixture was stirred for 2 hours at -10 °C. The reaction mixture was washed with water and the organic layer was dried over anhydrous Na;SO,, filtered and concentrated under reduced pressure to give product 7.28 g (48% yield).

Setp 10 Preparation of S-(7S5,8aS$)-2-(4-sulfamoylbenzoyl)octahydropyrrolo[1,2-a]- pyrazin-7-yl benzothioate

Ls mr, Ope TO 0 To ~~ 0 0 N nu TFA 0.0 —/ 1 - SO,NH,

To a solution of S-(7S,8aS)-octahydropyrrolo[1,2-a]pyrazin-7-yl benzothioate (3.59 g, 10 mmol) in CHCl, (100 mL) was treated with TEA (2 g, 20 mmol) and isobutyric 4-sulfamoylbenzoic anhydride (2.71 g, 10 mmol). The resulting mixture was stirred at 0 °C for 2 hours. The reaction mixture was washed with brine, dried over anhydrous Na;SO,, filtered and concentrated under reduced pressure. The crude residue was used directly in next step without further purification.

Setp 11 Preparation of 4-[(7S,8aS)-7-mercaptooctahydropyrrolo[1,2-a]pyrazine- 2-carbonyl]benzenesulfonamide

A,» "Ces 0 To, 0 : 0 \_/ _

SO,NH, SO,NH,

The crude product of previous step was dissolved in methanol and was added to a saturated methanolic KOH solution at 0 °C. The reaction mixture was stirred at 0 °C for 2 hours. The reaction mixture was adjusted to pH=6 to 7 with 1N HCI solution. Methanol was removed and the residue was extracted with CH,Cl,, dried over anhydrous Na,SO, and concentrated under reduced pressure to give product (2.45 g, 72%).

Setp 12 Preparation of 4-nitrobenzyl (4R,5S,6S5)-6-[(R)-1-hydroxyethyl]-4-methyl-7-o0xo- 3-[(7S,8aS)-2-(4-sulfamoylbenzoyl)octahydropyrrolo[1,2-a]pyrazin-7-ylthio]-1-azabicyclo- [3.2.0]hept-2-ene-2-carboxylate

OH

Om 0 rd OPh 0 Va JO es x CH, a

COOPNB < CC OL

Pros NA SO,NH,

SO,NH, COOPNB

In a dried reaction flask, 0.79 g of 4-[(7S,8aS)-7-mercaptooctahydropyrrolo[1,2-a]pyrazine-2- carbonyl]benzenesulfonamide (3 mmol) was dissolved in acetonitrile. To this solution was added 4-nitrobenzyl (4R,5R,6S)-3-(diphenoxyphosphoryloxy)-6-[(R)-1-hydroxyethyl]-4-methyl-7-oxo-1- : azabicyclo[3.2.0]hept-2-ene-2-carboxylate (1.78 g, 3 mmol) and DIEA (0.39 g, 3 mmol). The reaction mixture was stirred at ambient temperature for overnight. The reaction mixture was extracted with ethyl acetate and washed with water. The organic layer was dried over anhydrous

Na,SOq, fitered and concentrated under reduced pressure to give crude product 1.54 g (75% yield) which was used directly in next step without further purification.

Setp 13 Preparation of (4R,5S,65)-6-[(R)-1-hydroxyethyl]-4-methyl-7-o0x0-3-[(7S,8aS)- 2-(4-sulfamoylbenzoyl)octahydropyrrolo[1,2-a]pyrazin-7-ylthio]-1-azabicyclo[3.2.0]hept-2-ene -2-carboxylic acid a re AOL o wed & N I

COOPNB COOH

The crude product (0.9 g, 1.4 mmol) from step 12 of Example 8 was dissolved in a 1:1 mixture of THF and water (20 mL) and 10% Pd/C (0.4 g) was added. The resulting black suspension was stirred at ambient temperature under hydrogen pressure for 2 hours. The catalyst was filtered off and rinsed with THF and de-ionized water. The combined filtrate was extracted with ethyl acetate.

The product in aqueous layer was purified by prep. HPLC and the product fractions was freeze-dried to give solid product 50 mg (8.0% yield). “Due to the amide bond on the octahydropyrrolo[1,2-a]- pyrazine ring, 'H NMR exhibits two conformers.

Formula: C4H30N407S; Mol. Wt: 550.65 LC-MS: 551(M+H") 'H NMR (400 MHz, D,0) & 7.88 (d, 2H), § 7.51 (dd, 2H), & 4.58 (d, 0.5H), & 4.42 (d, 0.5H), & 4.03-4.12 (m, 2H), 6 3.62-3.68 (m, 1.5H), 6 3.48 (d, 0.5H), 6 3.26-3.30 (m, 1H), § 3.14-3.22 (m, 1.5H), 8 2.91-3.08 (m, 2.5H), 4 2.68 (d, 0.5H), 8 2.59 (t, 0.5H), 6 2.58-2.65 (m, 1H), § 2.37-2.41 (m, 0.5H), 6 2.14-2.28 (m, 2.5H), § 1.23-1.32 (m, 0.5H), 51.18 (d, 3H), 8 1.04-1.09 (m, 3.5H). . Example 9 Preparation of (4R,5S,68)-6-[(R)-1-hydroxyethyl]-4-methyl-7-ox0-3- [(7S,8aS)-2-sulfamoyloctahydropyrrolo[1,2-a]pyrazin-7-ylthio]-1-azabicyclo[3.2.0]hept-2-ene- 2-carboxylic acid (Compound 9)

Setp 1-8 Procedures were same as step 1-8 in Example 1.

Setp 9 Preparation of S-(75,8aS)-2-[N-[(4-nitrobenzyloxy)carbonyl]sulfamoyl]octa- hydropyrrolo[1,2-a]pyrazin-7-yl benzothioate ps

Om SE 0 N_ NH TFA — >» HN~¢

MV» ~ To a solution of 4-nitrobenzyl alcohol (2.73 g, 17.9 mmol) in acetonitrile was cooled to 0 °C and dropwise treated with a CHCl; solution of chlorosulfonyl isocyanate (2.5 g, 17.9 mmol). The resulting mixture was stirred at 0 °C for 1 hour and used directly in next reaction with purification.

The above solution was slowly added to a solution of S-(7S,8aS)-octahydropyrrolo- [1,2-a]pyrazin-7-yl benzothioate (4.27 g, 11.9 mmol) and TEA (5 mL, 35.7 mmol) in CH,Cl, (100 mL). The resulting mixture was stirred at 0 °C for 2 hours and at ambient temperarure for additional hours. The reaction mixture was washed with brine, dried over anhydrous Na,SO,, filtered and concentrated under reduced pressure. The crude residue was purified by flash column chromatography on silica gel to afford product 2.18 g (35.2% yield). - Setp 10 Preparation of 4-nitrobenzyl (75,8aS)-7-mercaptohexahydropyrrolo[1,2-a]- pyrazin-2(1H)-ylsulfonylcarbamate

On On 0 hg 2 ) N_N~¢%o .

O.N yf 0 on yb 2 0 0

S-(7S,8aS)-2-[N-[(4-nitrobenzyloxy)carbonyl]sulfamoyl]octahydropyrrolo- [1,2-a]pyrazin-7-yl benzothioate (2.18 g, 4.19 mmol) was dissolved in methanol and was added to a saturated methanolic KOH solution at 0 °C. The reaction mixture was stirred at 0 °C for 2 hours. The reaction mixture was adjusted to pH=6 to 7 with IN HCI solution. Methanol was removed and the residue was extracted with CH,Cl, dried over anhydrous Na,SO; and concentrated under reduced pressure to give product (1.25 g, 72%).

Setp 11 Preparation of 4-nitrobenzyl (4R,5S,6S5)-6-[(R)-1-hydroxyethyl]-4-methyl-3- [(7S,8a8)-2-[N-[(4-nitrobenzyloxy)carbonyl]sulfamoyl]octahydropyrrolo[1,2-a]pyrazin-7-ylthi 0]-7-oxo-1-azabicyclo[3.2.0]hept-2-ene-2-carboxylate

HS : OH CH

T= 0 ST dhom OH go o { nc, rs NP x ~ COOPNB q C1} N “CL

COOPNB

In a dried reaction flask, 2.725 g of 4-nitrobenzyl (7S,8aS)-7-mercaptohexahydropyrrolo- [1,2-a]pyrazin-2(1H)-ylsulfonylcarbamate (6.55 mmol) and DIEA (2.17 g, 13 mmol) was dissolved in acetonitrile. To this solution was added 4-nitrobenzyl (4R,5R,6S)-3- (diphenoxyphosphoryloxy)-6-[(R)-1-hydroxyethyl]-4-methyl-7-oxo-1-azabicyclo[3.2.0]hept-2-ene- 2-carboxylate (4.28 g, 7.21 mmol). The reaction mixture was stirred at ambient temperature for 72 hours under nitrogen atmosphere. The reaction mixture was extracted with ethyl acetate and washed with water. The organic layer was dried over anhydrous Na,SOy, fitered and concentrated under reduced pressure to give crude product 1.4 g (28% yield) which was used directly in next step without further purification.

Setp 12 Preparation of (4R,5S,65)-6-[(R)-1-hydroxyethyl]-4-methyl-7-0x0-3-[(7S,8aS)- 2-sulfamoyloctahydropyrrolo[1,2-a]pyrazin-7-ylthio]-1-azabicyclo[3.2.0]hept-2-ene-2-carboxyl ic acid

OH net. 0 rp we), os 5

COOPNB Pd/C COOH

The product (1.4 g, 1.84 mmol) from step 11 of Example 9 was dissolved in a 1:1 mixture of

THF and water (20 mL) and 10% Pd/C (0.4 g) was added. The resulting black suspension was stirred at ambient temperature under hydrogen pressure for 2 hours. The catalyst was filtered off and rinsed with THF and de-ionized water. The combined filtrate was extracted with ethyl acetate.

The product in aqueous layer was purified by prep. HPLC and the product fractions was freeze-dried to give solid product 280 mg (35% yield).

Formula: C;7H2¢N4O6S; Mol. Wt: 446.54 LC-MS: 447(M+H") 'HMR: (400 MHz, DLC-MSO _d6) &: 6.74 (s, 2H), 4.92 (s, 1H), 3.84-3.91 (m, 2H), 3.46 (d, 2H), 2.87-2.97 (m, 3H), 2.62-2.67 (m, 1H), 2.23-2.33 (m, 2H), 2.03-2.09 (m, 2H), 1.1 (d, 3H), 1.0 (d, 3H).

Example 10 Preparation of (4R,55.,65)-3-[(7S.8a8)-2-[2-(dimethylamino)acetyl]octahydro- pyrrolo[1.2-alpyrazin-7-ylthio]-6-[(R)-1-hydroxyethyl]-4-methyl-7-0x0-1-azabicyclo[3.2.0}hept -2-ene-2-carboxylic acid (Compound 12)

Step 1-8 Procedures were same as step 1-8 in Example 1.

Step 9 Preparation of S-(75,8aS5)-2-[2-(dimethylamino)acetyl]octahydropyrrolo[1,2-a]- pyrazin-7-yl benzothioate oO

Hom a! Apron 0

N NH TFA 0 N — “Nucl A ps

To a solution of S-(7S,8aS)-octahydropyrrolo[1,2-a]pyrazin-7-yl benzothioate (4.27 g, 11.9 mmol) in CHCl, (200 mL) was treated with TEA (5 mL, 35.7 mmol) and chloroacetyl chloride (1.35 g, 11.9 mmol). The resulting mixture was stirred at ambient temperature for overnight. To the reaction mixture was added dimethylamine hydrochloride salt (0.97 g, 11.9 mmol) and TEA (1.7 mL, 11.9 mmol). The resulting mixture was heated to reflux and stirred until the reaction completed.

The reaction mixture was washed with saturated aqueous NaHCO; solution and brine, dried over anhydrous Na,SOy, filtered and concentrated under reduced pressure. The residue was purified by flash column chromatography on silica gel to give product 1.70 g (41.2%).

Step 10 Preparation of 2-(dimethylamino)-1-[(7S,8aS)-7-mercaptohexahydropyrrolo-

[1,2-a]pyrazin-2(1H)-yl]ethanone 0 NN </ ARN \

To a solution of S-(7S,8aS)-2-[2-(dimethylamino)acetyl}octahydropyrrolo[1,2-a]pyrazin-7-yl benzothioate (1.70 g, 4.9 mmol) in methanol was added to a saturated methanolic KOH solution at 0 °C. The reaction mixture was stirred at 0 °C for 2 hours. The reaction mixture was adjusted to pH=6 to 7 with IN HCI solution. Methanol was removed and the residue was extracted with CHCl, dried over anhydrous Na,SO4 and concentrated under reduced pressure to give product (0.99 g, 83%).

Step 11 Preparation of 4-nitrobenzyl (4R,5S,65)-3-[(7S,8aS)-2-[2-(dimethylamino)acetyl- octahydropyrrolo[1,2-a]pyrazin-7-ylthio]-6-[(R)-1-hydroxyethyl]-4-methyl-7-oxo0-1-azabicyclo -[3.2.0]hept-2-ene-2-carboxylate

In a dried reaction flask, 0.97 g of 2-(dimethylamino)-1-[(7S,8aS)-7-mercaptohexahydro- pyrrolo[1,2-a]pyrazin-2(1 H)-yl]ethanone (4.0 mmol) and DIEA (1.03 g, 8 mmol) was dissolved in acetonitrile (100 mL). To this solution was added 4-nitrobenzyl (4R,5R,6S)- 3-(diphenoxyphosphoryloxy)-6-[(R)-1-hydroxyethyl]-4-methyl-7-oxo-1-azabicyclo-[3.2.0]hept-2- ene-2-carboxylate (2.38 g, 4 mmol). The reaction mixture was stirred at ambient temperature for 24 hours under nitrogen atmosphere. The reaction mixture was extracted with ethyl acetate and washed with water. The organic layer was dried over anhydrous Na,SOy, fitered and concentrated under reduced pressure to give crude product 1.62 g (69% yield) which was used directly in next step without further purification.

Step 12 Preparation of (4R,5S,6S)-3-[(7S,8aS)-2-[2-(dimethylamino)acetyl]octahydro- pyrrolo[1,2-a]pyrazin-7-ylthio]-6-[(R)-1-hydroxyethyl]-4-methyl-7-0xo-1-azabicyclo[3.2.0]hept -2-ene-2-carboxylic acid

OH cut & JA _ HC 2. & a

COOPNB COOH

The product (1.62 g, 2.76 mmol) from step 11 of Example 10 was dissolved in a 1:1 mixture of

The product in aqueous layer was purified by prep. HPLC and the product fractions was freeze-dried to give solid product 287 mg (23% yield).

Due to the amide bond on the octahydropyrrolo[1,2-a]- pyrazine ring, "H NMR exhibits two conformers.

Formula: CpHiN4OsS Mol.Wt.: 452.57 LC-MS: 453(M+H") "HNMR: (400 MHz, D,0) 8: 4.39(d, 0.5H), 4.25(d, 0.5H), 4.09 (t, 1H), 4.04 (d, 1H), 3.84(d, 0.5H), 3.60-3.69 (m, 1.5H), 3.55(d, 2H), 3.27(d, 1H), 3.09-3.19 (m, 1.5H), 2.83-2.95(m, 2.5H), 2.69-2.75 (m, 0.5H) 2.57(t, 1H), 2.48 (t, 0.5H), 2.33-2.41(m, 7H), 2.06-2.31 (m, 2H), 1.14-1.18(m, 4H), 1.06 (d, 3H).

Example 11 Preparation of (4R.55,65)-3-[(7S.8a85)-2-[2-(dimethylamino)propionyl]octa- hydropvyrrolo[1,2-a]pyrazin-7-ylthio]-6-[(R)-1-hydroxyethyl]-4-methyl-7-0x0-1-azabicyclo[3.2.

O]hept-2-ene-2-carboxylic acid (Compound 13)

Step 1-8 Procedures were same as step 1-8 in Example 1.

Step 9 Preparation of S-(75,8a8)-2-(2-(dimethylamino)propionyl)Octahydropyrrolo [1,2-a]-pyrazin-7-yl benzothioate

A

Cm He Dn 0 N_ NH TFA — 0 NN pe

H HCI \

To a solution of S-(7S,8aS)-octahydropyrrolo[1,2-a]pyrazin-7-yl benzothioate (4.27 g, 11.9 mmol) in CHCl, (200 mL) was treated with TEA (5 mL, 35.7 mmol) and 2-chloropropionyl chloride (1.51 g, 11.9 mmol). The resulting mixture was stirred at ambient temperature for overnight. To the reaction mixture was added dimethylamine hydrochloride salt (0.97 g, 11.9 mmol) in 20 mL of CH,Cl, and TEA (1.7 mL, 11.9 mmol). The resulting mixture was heated to reflux and

: stirred until the reaction completed. The reaction mixture was washed with saturated aqueous

NaHCO3; solution and brine, dried over anhydrous Na;SO, filtered and concentrated under reduced pressure. The residue was purified by flash column chromatography on silica gel to give product 1.59 g (37.2% yield).

Step 10 Preparation of 2-(dimethylamino)-1-((75,8a5)-7-mercaptohexahydropyrrolo [1,2-a]- pyrazin-2(1H)-yl)propan-1-one gy = o 3 AON 0 , —_— AE / ~ “I \

S-(78,8a8)-2-[2-(dimethylamino)propionyl]octahydropyrrolo[1,2-a]pyrazin-7-yl benzothioate (1.59 g, 4.4 mmol) dissolved in methanol was added to a saturated methanolic KOH solution at 0 °C.

The reaction mixture was stirred at 0 °C for 2 hours. The reaction mixture was adjusted to pH=6 to 7 with IN HCI solution. Methanol was removed and the residue was extracted with CH,Cl,, dried over anhydrous Na,SO4 and concentrated under reduced pressure to give product (0.91 g, 81%).

Step 11 Preparation of 4-nitrobenzyl (4R,5S,6S5)-3-[(7S5,8aS)-2-[2-(dimethylamino)- propionyl]octahydropyrrolo[1,2-a]pyrazin-7-ylthio]-6-[(R)-1-hydroxyethyl]-4-methyl-7-0x0-1- azabicyclo[3.2.0]hept-2-ene-2-carboxylate bs CH, °

In a dried reaction flask, 0.91 g of 2-(dimethylamino)-1-[(7S,8aS)-7-mercaptohexahydro- pyrrolo[1,2-a]pyrazin-2(1 H)-yl]ethanone (3.56 mmol) and DIEA (1.03 g, 8 mmol) was dissolved in acetonitrile (100 mL). To this solution was added 4-nitrobenzyl (4R,5R,65)-3-(diphenoxyphosphoryloxy)-6-[(R)-1-hydroxyethyl]-4-methyl-7-oxo-1-azabicyclo- [3.2.0]hept-2-ene-2-carboxylate (2.11 g, 3.56 mmol). The reaction mixture was stirred at ambient temperature for 24 hours under nitrogen atmosphere. The reaction mixture was extracted with ethyl acetate and washed with water. The organic layer was dried over anhydrous Na;SO,, fitered and concentrated under reduced pressure to give crude product 1.51 g (71% yield) which was used directly in next step without further purification.

Step 12 Preparation of (4R,58,65)-3-[(7S5,8aS)-2-[2-(dimethylamino)propanoyl]

octahydro-pyrrolo[1,2-a]pyrazin-7-ylthio]-6-[(R)-1-hydroxyethyl]-4-methyl-7-0xo-1-azabi cyclo[3.2.0]hept-2-ene-2-carboxylic acid nt fH JA oct JA

COOPNB COOH

The product (1 51 g, 2.52 mmol) from step 11 of Example 11 was dissolved in a 1:1 mixture of

The product in aqueous layer was purified by prep. HPLC and the product fractions was freeze-dried to give solid product 140 mg (12% yield).

Formula: C2oH34N405S Mol. Wt: 466.59 LC-MS: 467(M+H") "HNMR: (400 MHz, D,0) 8: 4.40-4.43(m, 0.5H), 4.29-4.36(m, 1.5H), 4.06-4.14(m, 2H), 3.85-3.95 (m, 0.5H), 3.67-3.82(m, 1.5H), 3.29(d, 1H), 3.18-3.25(m, 1.5H), 2.96-3.02(m, 2.5H), 2.73-2.85(m, 6.5H), 2.59-2.63(m, 1.5H) 2.35-2.38(m, 1H) 2.12-2.21(m, 2H) 1.37-1.42(m, 3H) 1.13-1.26(m, 4H) 1.08(d, 3H)

Example 12: Peparation of (4R.5S5,65)-6-[(R)-1-hydroxyethyl]-4-methyl-3-[(7S5.,8aS5)-2- (2-morpholinoacetyl)octahydropyrrolo[1,2-alpyrazin-7-ylthio]-7-oxe-1-azabicyclo[3.2.0] hept- 2-ene-2-carboxylic acid (compound 14)

Stepl-8 Procedures were same as step 1-8 in Example 1.

Step 9 Preparation of S-(7S,8aS)-2-(2-morpholinoacetyl)octahydropyrrolo [1,2-a]pyrazin-7-yl benzothioate 0

To a solution of S-(7S,8aS)-octahydropyrrolo[1,2-a]pyrazin-7-yl benzothioate (4.27 g, 11.9 mmol) in CH,Cl, (200 mL) was treated with TEA (5 mL, 35.7 mmol) and 2-chloroacetyl chloride (1.35 g, 11.9 mmol) in 20 mL of CH,Cl,. The resulting mixture was stirred at ambient temperature for overnight. To the reaction mixture was added morpholine (1.03 g, 11.9 mmol) in 20 mL of

CH,Cl. The resulting mixture was heated to reflux and stirred until the reaction completed. The reaction mixture was washed with saturated aqueous NaHCOs solution and brine, dried over anhydrous Na;SOy, filtered and concentrated under reduced pressure. The residue was purified by flash column chromatography on silica gel to give product 2.40 g (56% yield).

Step 10 Preparation of 1-[(78,8aS)-7-mercaptohexahydropyrrolo[1,2-a] pyrazin-2(1H)-yl]-2-morpholinoethanone

S-(78,8aS)-2-(2-morpholinoacetyl)octahydropyrrolo[ 1,2-a]pyrazin-7-yl benzothioate (2.60 g, 6.67 mmol) dissolved in methanol was added to a saturated methanolic KOH solution at 0 °C. The reaction mixture was stirred at 0 °C for 2 hours. The reaction mixture was adjusted to pH=6 to 7 with IN HCI solution. Methanol was removed and the residue was extracted with CH,Cl,, dried over anhydrous Na;SO, and concentrated under reduced pressure to give product (1.29 g, 68%).

Step 11 Preparation of 4-nitrobenzyl (4R,5S,65)-6-[(R)-1-hydroxyethyl]-4-methyl-3- [(75,8aS5)-2-(2-morpholinoacetyl)octahydropyrrolo[1,2-a]pyrazin-7-ylthio]-7-oxo-1-azabicyclo- © [3.2.0]hept-2-ene-2-carboxylate

HC A 4 8 ae 0

Oy Aa 7 {coms NN

In a dried reaction flask, 1.29 g of 1-[(7S,8aS)-7-mercaptohexahydropyrrolo[1,2-a]pyrazin- 2(1H)-yl]- 2-morpholinoethanone (4.52 mmol) and DIEA (1.03 g, 8 mmol) was dissolved in acetonitrile (100 mL). To this solution was added 4-nitrobenzyl (4R,5R,6S)-3-(diphenoxyphosphoryloxy)-6-[(R)-1-hydroxyethyl]-4-methyl-7-oxo-1-azabicyclo- [3.2.0]hept-2-ene-2-carboxylate (2.69 g, 4.52 mmol). The reaction mixture was stirred at ambient temperature for 24 hours under nitrogen atmosphere. The reaction mixture was extracted with ethyl acetate and washed with water. The organic layer was dried over anhydrous Na,SO,, fitered and concentrated under reduced pressure to give crude product 1.36 g (48% yield) which was used directly in next step without further purification. :

Step 12 Peparation of (4R,5S,65)-6-[(R)-1-hydroxyethyl]-4-methyl-3-[(7S,8a8)-2-(2- morpholinoacetyloctahydropyrrolo[1,2-a]pyrazin-7-ylthio]-7-oxo-1-azabicyclo[3.2.0] hept-2- ene-2-carboxylic acid

COOPNB

The product (1.36 g, 2.16 mmol) from step 11 of Example 12 was dissolved in a 1:1 mixture of

The product in aqueous layer was purified by prep. HPLC and the product fractions was freeze-dried to give solid product 171 mg (16% yield).

Formula: C;3H34N406S Mol. Wt: 494.60 LC-MS: 495(M+H") 'HNMR: (400 MHz, D,0) 3: 4.40(d, 0.5H,), 4.25(d, 0.5H), 4.03-4.11 (m, 2H), 3.93 (d, 0.5H), 3.75(d, 0.5H), 3.55-3.65 (m, 5H), 3.35-3.91(m, 2.5H), 3.17-3.26 (m, 1H), 3.05-3.15(m, 0.5H), 2.86-2.94 (m, 2.5H) 2.59-2.62 (m, 0.5H) 2.46-2.59(m, 6H) 2.34-2.45 (m, 1H) 2.11-2.21(m, 2H) 1.11-1.20 (m, 4H) 1.05 (d, 3H). 'HNMR: (400 MHz, D,0O) &: 4.40(d, 0.5H), 4.25(d, 0.5H), 4.03-4.11(m, 2H), 3.93(d, 0.5H), 3.75(d, 0.5H), 3.55-3.65(m, 5H), 3.35-3.91(m, 2.5H), 3.17-3.26(m, 1H), 3.05-3.15(m, 0.5H), 2.86-2.94(m, 2.5H), 2.59-2.62(m, 0.5H), 2.46-2.59(m, 6H), 2.34-2.45(m, 1H), 2.11-2.21(m, 2H), 1.11-1.20 (m, 4H), 1.05 (d, 3H).

Example 13 Preparation of (4R.5S5.65)-6-[(R)-1-hvdroxyethyl]-3-[(7S.8aS)-2-(2-hydroxy- ethyl)octahydropyrrolo[1.2-a]pyrazin-7-ylthio]-4-methyl-7-o0xo-1-azabicyclo[3.2.0]hept-2-ene- 2-carboxylic acid (Compund 16)

Setp 1-8 Procedures were same as step 1-8 in Example 1.

Step 9 Preparation of S-(75,8aS)-2-(2-hydroxyethyl)octahydropyrrolo[1,2-a]pyrazin- 7-yl benzothioate

S Br Ae = HO J AY —_— N 0 N NH TFA a; on

To a solution of S-(7S,8aS)-octahydropyrrolo[1,2-alpyrazin-7-yl benzothioate (4.27 g, 11.9 mmol) in CH,Cl, (200 mL) was treated with TEA (5 mL, 35.7 mmol) and 2-bromoethanol (1.35 g, 11.9 mmol). The resulting mixture was heated to reflux and stirred until the reaction completed. The reaction mixture was washed with saturated aqueous NaHCO; solution and brine, dried over anhydrous Na;SO,, filtered and concentrated under reduced pressure. The residue was purified by flash column chromatography on silica gel to give product 2.26 g (62% yield).

Step 10 Preparation of 2-[(7S,8aS)-7-mercaptohexahydropyrrolo[1,2-a]pyrazin- 2(1H)-yl] ethanol 0 N —_— N N

S-(75,8aS)-2-(2-hydroxyethyl)octahydropyrrolo[1,2-a]pyrazin-7-yl benzothioate (2.26 g, 7.37 mmol) dissolved in methanol was added to a saturated methanolic KOH solution at 0 °C. The reaction mixture was stirred at 0 °C for 2 hours. The reaction mixture was adjusted to pH=6 to 7 with IN HCI solution. Methanol was removed and the residue was extracted with CH,Cl,, dried over anhydrous Na,SO, and concentrated under reduced pressure to give product (0.77 g, 52%).

Step 11 Preparation of 4-nitrobenzyl (4R,5S,65)-6-[(R)-1-hydroxyethyl]-3-[(7S,8aS5)-2- (2-hydroxyethyl)octahydropyrrolo[1,2-a]pyrazin-7-ylthio]-4-methyl-7-oxo-1-azabicyclo[3.2.0]- hept-2-ene-2-carboxylate

J 0 OH

HC 1 J CH,

HS \_ ORO, ch NAOH

Ala o N-/ s—1)

N NTL COOPNB o ~~ OH > COOPNB

In a dried reaction flask, 0.77 g of 2-[(7S,8aS)-7-mercaptohexahydropyrrolo[1,2-a]pyrazin- (1H)-yl]ethanol (3.83 mmol) and DIEA (1.03 g, 8 mmol) was dissolved in acetonitrile (100 mL). To this solution was added 4-nitrobenzyl (4R,5R,6S5)-3-(diphenoxyphos- phoryloxy)-6-[(R)-1-hydroxyethyl]-4-methyl-7-oxo-1-azabicyclo[3.2.0]hept-2-ene-2-carboxylate (MAP) (2.27 g, 4.83 mmol). The reaction mixture was stirred at ambient temperature for 24 hours under nitrogen atmosphere. The reaction mixture was extracted with ethyl acetate and washed with water. The organic layer was dried over anhydrous Na;SOs, fitered and concentrated under reduced pressure to give crude product 1.23 g (59% yield) which was used directly in next step without further purification.

Stepl2 Preparation of (4R,5S,65)-6-[(R)-1-hydroxyethyl]-3-[(7S,8aS)-2- (2-hydroxyethyl)-octahydropyrrolo[1,2-a]pyrazin-7-ylthio]-4-methyl-7-o0xo-1-azabicyclo[3 .2.0]hept-2-ene-2- carboxylic acid

The product (1.23 g, 2.25 mmol) from step 11 of Example 13 was dissolved in a 1:1 mixture of

The product in aqueous layer was purified by prep. HPLC and the product fractions was freeze-dried to give solid product 101 mg (11% yield).

Formula: C;gH2oN305S Mol. Wt: 411.52 LC-MS: 412(M+H") 'HNMR: (400 MHz, D,0) &: 3.98-4.12 (m, 2H),3.52-3.68 (m, 3H), 3.21-3.27 (m, 1H), 3.09-3.10 (m, 1H), 2.98 (d, 1H), 2.76-2.90 (m, 3H), 2.48-2.57 (m, 3H), 2.11-2.34 (m, 4H), 1.92-2.01 (m, 1H), 1.05-1.19 (m, 4H), 1.03 (d, 3H).

The following compounds can also be prepared according to the method described above:

OH CH

He, 3 gn CH 0, 19 s— 7 3 5 | A XZ

FN / N A PN —13 i

COOH ) _ « COOH 0 He A. 3 . gt CH, 0 9

N cc.

T A 1) ; 34 3 . TY 0 F Nn nn

COOH ° re COOH

H

COOH ? a COOH , Ho J, 3 pH CH, 0 g } T > S CL 36 HC ‘, S Cy

COOH ’ o { "

COOH wot i Hs a" oo P 23 3L7 N CONH, HC, A.

TOOT 7 |” =O

COOH o” { nS 7 — COOH 7 ] OH 4 HC, N N me, re A 3 vg aS Ln 38 yt 0 ZN Ng NM

COOH ° ~ COOH

Jp a 25 Hy C7, N XN H c, ? 0

NH. 7 5

COCH ’ 0 N nN 7 — 5 5 COOH \ H

H;C 7%. rH 1 26 To 4 OT 40 HC, ’ 1

N

FN NA Nt ~ 3

COOH °

I - - om COOH 3 of CH S 27 HC . Py HC, ’ pp

N J $ NJ ! H S } "

N-/ NS

COOH © — COOH

HC" i L 28 BCT N HC ’ LY

Ng AN +2 s— 1) 5 . Ng NJ

HC", ’ 29 ’ 5 NSN He. A 7) SR 43 N 0 Z Ng

COOH 0 NS — — . COOH 5 “, Cy + NH,

Presto SRSA

COOH

Claims

1. A compound of formula (I), or its pharmaceutically acceptable salts, or its readily hydrolyzed ester, or its solvate, or its stereoisomer thereof: OH rl H;C NE © COOR? (I) Wherein R! is hydrogen or Cy.¢ alkyl; R? is . (1) hydrogen, (2) C1. alkyl, Cp. alkenyl or C,.5 alkynyl, which is optionally substituted by one to three xX! X! is independently selected from halogen, carboxyl, hydroxyl, amino, cyano, nitro, Cy. alkoxy, halogenated Ci. alkoxy, Ci. alkyl amino, bis-(C.¢ alkyl)amine, C,. alkyl carbonyl, Cicalkyl carboxyl, Ci. alkoxy carbonyl, carbamoyl, Cy alkyl carbamoyl, bis-(Ci alkyl) carbamoyl, amino sulfonyl, C.¢ alkyl amino sulfonyl, bis-(C alkyl) amino sulfonyl, Ci alkyl sulfonyl amino, Ci. alkyl sulfonyl, Ci alkyl carbonyl amino, guanidino, Cs.g cycloalkyl, aryl or heterocyclic,

Cs.g cycloalkyl, aryl or heterocyclic of said X! can be further substituted by the following substituents: halogen, carboxyl, hydroxyl, amino, cyano, nitro, Ci.¢ alkyl, carboxy Ci. alkyl, hydroxy Ci.¢ alkyl, amino Cj alkyl, halogenated C,.¢ alkyl, Ci alkoxy, halogenated Ci. alkoxy, C1. alkoxy C.¢ alkyl, Ci alkyl amino, bis-(C1.¢ alkyl) amino, bis-(C. alkyl) amino Cis alkyl, Ci alkyl carbonyl, Ciealkyl carboxyl, Ci. alkoxy carbonyl, carbamoyl, carbamoyl Cp. alkyl, Cy. alkyl carbamoyl, bis-(Ci.¢ alkyl) carbamoyl, amino sulfonyl, amino sulfonyl Ci. alkyl, Ci alkyl amino sulfonyl, bis-(Ci¢ alkyl) amino sulfonyl, Ci.¢ alkyl ~ sulfonyl amino, C.¢ alkyl sulfonyl, C;.¢ alkyl carbonyl amino or guanidino, (3) Cs.3 cycloalkyl, aryl or heterocyclic which is optionally substituted by one to three x2, xX? is independently selected from halogen, carboxyl, hydroxyl, amino, cyano, nitro, Ci. alkyl, carboxy Ci. alkyl, hydroxy Ci. alkyl, amino Ci alkyl, halogenated Ci. alkyl, Ci. alkoxy, halogenated Cj. alkoxy, Ci alkoxy Ci alkyl, Ci alkyl amino, bis-(Ci alkyl) amino, bis-(Ci.¢ alkyl) amino Cy alkyl, Ci. alkyl carbonyl, Cy.salkyl carboxyl, Cis alkoxy carbonyl, carbamoyl, carbamoyl Cs alkyl, Ci. alkyl carbamoyl, bis-(Ci. alkyl) carbamoyl, amino sulfonyl, amino sulfonyl Ci. alkyl, Ci alkyl amino sulfonyl, bis-(Ci.s alkyl) amino sulfonyl, Cis alkyl sulfonyl amino, Ci. alkyl sulfonyl, Ci.¢ alkyl amino carbonyl or guanidino, 4) -CO-Y, Y is Ci alkyl which is optionally substituted by one to three Y!, or Cig cycloalkyl, aryl or heterocyclic which is optionally substituted by one to three Y?, Y! represents the above defined X! and Y? represents the above defined xX? (5) -S(O)m-Z, mis 1 or 2, Z is amino, Ci alkyl amino, bis-(Ci.s alkyl)amino, or Ci alkyl which is optionally substituted by one to three Y', or Cs cycloalkyl, aryl or heterocyclic which is optionally substituted by one to three Y?, Y' represents the above defined X! and Y? represents the above defined X%; and R? is hydrogen or the carboxyl protecting group.

2. A compound claimed in claim 1, or its pharmaceutically acceptable salts, or its readily hydrolyzed ester, or its solvate, or its stereoisomer: Wherein R'is C4 alkyl; Ris (1) hydrogen, (2) C1.4 alkyl, which is optionally substituted by one to three X!, X' is independently selected from halogen, carboxyl, hydroxyl, amino, C4 alkoxy, halogenated C14 alkoxy, Ci4 alkyl amino, bis-(Ci4 alkyl)amine, C4 alkyl carbonyl, carbamoyl, amino sulfonyl, guanidino, Cs cycloalkyl, phenyl, 5-6 membered heteroaryl or 3-7 membered heterocyclic,

Cs. cycloalkyl, phenyl, 5-6 membered heteroaryl or 3-7 membered heterocyclic of the said X' can be further substituted by the following substituents: halogen, Ci4 alkyl, halogenated Cy alkyl, C14 alkoxy, halogenated Cy.4 alkoxy, Ci. alkyl amino, bis-(Ci4 alkyl) amino, C14 alkyl carbonyl, carbamoyl, carbamoyl C4 alkyl, amino sulfonyl, amino sulfonyl Ci alkyl, or guanidino, (3) Cs. cycloalkyl, phenyl, 5-6 membered heteroaryl or 3-7 membered heterocyclic which is optionally substituted by one to three x2, X? is independently selected from halogen, amino, C;.4 alkyl, halogenated C14 alkyl, C14 alkoxy, halogenated Ci4 alkoxy, C4 alkyl amino, bis-(Ci4 alkyl) amino, bis-(C;4 alkyl) amino Ci. alkyl, Cy alkyl carbonyl, carbamoyl, carbamoyl C;.4 alkyl, amino sulfonyl, amino sulfonyl C4 alkyl, or guanidino, 4) -CO-Y, Y is C4 alkyl which is optionally substituted by one to three Y!, or Cs. cycloalkyl, phenyl, 5-6 membered heteroaryl or 3-7 membered heterocyclic which is optionally substituted by one to three Y?, Y! represents the above defined X' and Y? represents the above defined X2, (5) -S(O)n-Z, mis 1 or2, Z is amino, C;4 alkyl amino, bis-(C14 alkyl)amino, or C;4 alkyl which is optionally substituted by one to three Y!, or Cs cycloalkyl, phenyl, 5-6 membered heteroaryl or 3-7 membered heterocyclic which is optionally substituted by one to three Y?, Y! represents the above defined X! and Y? represents the above defined X?; and R? is hydrogen.

3. A compound claimed in claim 2, or its pharmaceutically acceptable salts, or its readily hydrolyzed ester, or its solvate, or its stereoisomer: Wherein R' is methyl; R%is (1) hydrogen, (2) C14 alkyl, which is optionally substituted by one to three xX, X'! is independently selected from halogen, carboxyl, hydroxyl, amino, Ci4 alkoxy, halogenated Cj alkoxy, C14 alkyl amino, bis-(C4 alkyl)amine, carbamoyl, amino sulfonyl, phenyl, pyridyl, piperidinyl, piperazinyl or morpholinyl, The phenyl, pyridyl, piperidinyl, piperazinyl or morpholinyl of the said X! can be further substituted by the following substituents: halogen, C4 alkyl, halogenated C4 alkyl, Ci4 alkoxy, halogenated C,.4 alkoxy, C4 alkyl amino, bis-(C;.4 alkyl) amino, C;.4 alkyl carbonyl, carbamoyl, carbamoyl C;.4 alkyl, or amino sulfonyl, (3) phenyl, pyridyl, piperidinyl, piperazinyl, morpholinyl, tetrahydrofuranyl, thiazole,

4,5-dihydro-thiazolyl, or cyclopropyl group which is optionally substituted by one to three x2,

X? is independently selected from halogen, C;.4 alkyl, C14 alkyl amino, bis-(Ci.4 alkyl) amino, carbamoyl, carbamoyl C4 alkyl, amino sulfonyl, 4) -CO-Y,

Y is Ci4 alkyl which is optionally substituted by one to three Y', or phenyl, pyridyl, piperidinyl, piperazinyl, morpholinyl, pyrrolidinyl, furanyl, thiazolyl or 4,5-dihydro-thiazolyl which is optionally substituted by one to three Y?,

Y! is independently selected from amino, Ci4 alkyl amino, bis-(Ci4 alkyl) amino, carbamoyl, guanidino, phenyl, pyridyl, piperidinyl, morpholinyl or pyrrolidinyl,

Y? is independently selected from halogen, amino, Ci. alkyl, C14 alkyl amino, bis-(Ci4 alkyl) amino, carbamoyl, carbamoyl C;4 alkyl or amino sulfonyl,

(5) -S(O)y-Z, mis 1 or 2,

Z is amino, C4 alkyl amino, bis-(C;4 alkyl)amino, or Ci4 alkyl which is optionally substituted by one to three Y', or phenyl, pyridyl, piperidinyl, piperazinyl, morpholinyl or pyrrolidinyl which is optionally substituted by one to three Y?,

Y! and Y? represents the above defined; and R® is hydrogen.

4, A compound claimed in claim 3, or its pharmaceutically acceptable salts, or its readily hydrolyzed ester, or its solvate, or its stereoisomer: Wherein

R! is methyl;

R?is

(1) hydrogen,

(2) methyl, ethyl, isopropyl, which is optionally substituted by one to three X!,

X! is independently selected from halogen, hydroxyl, dimethylamine, phenyl or pyridyl, The phenyl or pyridyl of the said X! can be further substituted by the following substituents: halogen, methyl, carbamoyl, carbamoyl methyl, or amino sulfonyl, (3) phenyl, pyridyl, tetrahydrofuranyl, thiazole, 4,5-dihydro-thiazolyl, or cyclopropyl group which is optionally substituted by one to three X>, X? is independently selected from halogen, methyl, carbamoyl, amino sulfonyl, 4) -CO-Y,

Y is methyl, ethyl or isopropyl which is optionally substituted by one to three Y!, or phenyl, pyridyl, piperidinyl, piperazinyl, pyrrolidinyl which is optionally substituted by one to three Y?, Y'is independently selected from dimethylamine, carbamoyl, guanidino, phenyl, pyridyl, piperidinyl, morpholinyl or pyrrolidinyl, Y? is independently selected from halogen, methyl, carbamoyl, carbamoyl methyl or amino sulfonyl, 5) -S(O)n-Z, mis 1 or 2, Z is amino, dimethylamino, dimethylamino methyl, methylpiperazine or morpholine methyl; and R’is hydrogen.

5. A compound claimed in claim 4, or its pharmaceutically acceptable salts, or its readily hydrolyzed ester, or its solvate, or its stereoisomer: Wherein R' is methyl; R? is (1) hydrogen, (2) methyl, ethyl, or isopropyl, which is optionally substituted by one to two X', X'is independently selected from hydroxyl,or phenyl, The said phenyl of X' can be further substituted by halogen, methyl, carbamoyl, or amino sulfonyl, 3) -CO-Y, Y is methyl, ethyl or isopropyl, which is optionally substituted by one to two Y!, or phenyl, pyridyl, piperidinyl, which is optionally substituted by one to two Y>, Y' is independently selected from dimethylamine, guanidino, phenyl, pyridyl, or morpholinyl, Y? is independently selected from halogen, methyl, carbamoyl, or amino sulfonyl, 4) -S(O)p-Z, mis 1 or 2, Z is amino, or dimethylamino; and R? is hydrogen.

6. A compound claimed in claim 5, or its pharmaceutically acceptable salts, or its readily hydrolyzed ester, or its solvate, or its stereoisomer, and the said compound is selected from the followings: (4R,58,65)-6-[(R)-1-hydroxyethyl]-4-methyl-3-[(7S,8aS)-2-methyloctahydropyrrolo[ 1,2-a]pyra zin-7-ylthio]-7-oxo-1-azabicyclo[3.2.0]hept-2-ene-2-carboxylic acid, (4R,5S,6S5)-6-[(R)-1-hydroxyethyl]-3-[(7S,8aS)-2-Isopropyl-octahydropyrrolo[ 1,2-a]pyrazin-7- ylthio]-4-methyl-7-oxo-1-azabicyclo[3.2.0]hept-2-ene-2-carboxylic acid, (4R,58,6S5)-3-[(7S,8aS)-2-benzyloctahydropyrrolo[ 1,2-a]-pyrazin-7-ylthio]-6-[(R)-1-hydroxyet hyl]-4-methyl-7-ox0-1-azabicyclo[3.2.0]hept-2-ene-2-carboxylic acid, (4R,58,6S5)-3-[(7S,8aS)-2-acetyloctahydropyrrolo[1,2-a]pyrazin-7-ylthio]-6-[(R)-1-hydroxyeth yl]-4-methyl-7-o0x0-1-azabicyclo[3.2.0]hept-2-ene-2-carboxylic acid, - (4R,58,6S5)-6-[(R)-1-hydroxyethyl]-3-[(7S,8aS)-2-isobutyryl-octahydropyrrolo[1,2-a]pyrazin-7 -ylthio]-4-methyl-7-oxo0-1-azabicyclo[3.2.0]hept-2-ene-2-carboxylic acid, (4R,58,685)-3-[(7S,8aS)-2-benzoyloctahydropyrrolo[ 1,2-a]-pyrazin-7-ylthio]-6-[ (R)-1-hydroxye thyl]-4-methyl-7-oxo0-1-azabicyclo[3.2.0]hept-2-ene-2-carboxylic acid, (4R,5S,65)-6-[(R)-1-hydroxyethyl]-4-methyl-7-0x0-3-[(7S,8aS)-2-picolinoyloctahydropyrrolo[ 1,2-a]pyrazin-7-ylthio]-1-azabicyclo[3.2.0]hept-2-ene-2-carboxylic acid, (4R,5S,65)-6-[(R)-1-hydroxyethyl]-4-methyl-7-0x0-3-[(7S,8aS)-2-(4-sulfamoylbenzoyl)octahy dropyrrolo[1,2-a]pyrazin-7-ylthio]-1-azabicyclo[3.2.0]hept-2-ene-2-carboxylic acid, (4R,58,6S5)-6-[(R)-1-hydroxyethyl]-4-methyl-7-0x0-3-[(7S,8aS)-2-sulfamoyloctahydropyrrolo[ 1,2-a]pyrazin-7-ylthio]-1-azabicyclo[3.2.0]hept-2-ene-2-carboxylic acid, (4R,5S,65)-6-[(R)-1-hydroxyethyl]-4-methyl-7-0x0-3-[(7S,8aS)-2-(4-sulfamoylbenzyl)octahyd ropyrrolo[1,2-a]pyrazin-7-ylthio]-1-azabicyclo[3.2.0]hept-2-ene-2-carboxylic acid, (4R,5S,65)-3-[(7S,8aS)-2-(2-guanidinoacetyl)octahydropyrrolo[1,2-a]pyrazin-7-ylthio]-6-[ (R)- 1-hydroxyethyl]-4-methyl-7-oxo-1-azabicyclo[3.2.0]hept-2-ene-2-carboxylic acid, (4R,58,6S)-3-((7S,8aS)-2-(2-(dimethylamino)acetyl)octahydro-pyrrolo[ 1,2-a]pyrazin-7-ylthio)- 6-((R)-1-hydroxyethyl)-4-methyl-7-oxo-1-azabicyclo[3.2.0]hept-2-ene-2-carboxylic acid, (4R,58,6S5)-3-((7S,8aS)-2-(2-(dimethylamino)propionyl)octa-hydropyrrolo[ 1,2-a]pyrazin-7-ylt hio)-6-((R)-1-hydroxyethyl)-4-methyl-7-oxo0-1-azabicyclo[3.2.0]hept-2-ene-2-carboxylic acid, (4R,5S,65)-6-[(R)-1-hydroxyethyl]-4-methyl-3-[(7S,8aS)-2-(2-morpholinoacetyl)octahydropyrr olo[1,2-a]pyrazin-7-ylthio]-7-oxo-1-azabicyclo[3.2.0]hept- 2-ene-2-carboxylic acid, (4R,58,65)-6-[(R)-1-hydroxyethyl]-4-methyl-3-[(7S,8aS)-2-(1-methylpiperidine-4-carbonyl)oct ahydropyrrolo[1,2-a]pyrazin-7-ylthio]-7-oxo0-1-azabicyclo[3.2.0]hept-2-ene-2-carboxylic acid, and (4R,585,65)-6-[(R)-1-hydroxyethyl]-3-[(7S,8aS)-2-(2-hydroxy-ethyl)octahydropyrrolo[ 1,2-a]lpyr azin-7-ylthio]-4-methyl-7-oxo0-1-azabicyclo[3.2.0]hept-2-ene-2-carboxylic acid.

7. A pharmaceutical dosage form comprising a compound claimed in claims 1-6, or its pharmaceutically acceptable salts, or its readily hydrolyzed ester, or its solvate, or its stereoisomer, characterized in that the said dosage form contains one or more pharmaceutical carriers.

8. A method of the treatment or prevention of infectious diseases comprising a compound claimed in claims 1-6, or its pharmaceutically acceptable salts, or its readily hydrolyzed ester, or its solvate, or its stereoisomer.

9. A method of preparation of a compound of formula (I) comprising a nucleophilic substitution reactionin between an intermediate of formula (II) and an intermediate of formula (III) OH rl! HsC NS a CooR* (II) Wherein, R',R* and R® are defined as in claim 1 ,and Lis a leaving group.

10. A compound of formula (II), or its pharmaceutically acceptable salts, or its readily hydrolyzed ester, or its solvate, or its stereoisomer: R2 an Wherein, R? is defined as in claim 1.