RU2465264C2 - Deuterated hepatitis c protease inhibitors - Google Patents

Abstract

FIELD: medicine.

SUBSTANCE: invention refers to a deuterium-enriched α-ketoamide compound of formula

wherein: D means a deuterium atom; the values R¹-R⁵ are presented in cl.1 of the patent claim, and to a based pharmaceutical composition.

EFFECT: method improvement.

32 cl, 3 ex

Description

Cross reference

This application claims the priority of provisional application US serial number 60/782778, filed March 16, 2006, provisional US application serial number 60/782976, filed March 16, 2006, and provisional application US serial number 60/844771, filed September 15, 2006.

State of the art

Hepatitis C virus infection ("HCV") is an urgent medical problem for humans. HCV is considered the causative factor in most cases of hepatitis other than hepatitis A and hepatitis B, which has been determined to be present in serum in 3% of people worldwide [A. Alberti et al., "Natural History of Hepatitis C," J. Hepatology, 31., (Suppl. 1), pp. 17-24 (1999)]. In the United States alone, about four million people can be infected [MJ. Alter et al., "The Epidemiology of Viral Hepatitis in the United States, Gastroenterol. Clin. North Am., 23, pp. 437-455 (1994); MJ Alter" Hepatitis C Virus Infection in the United States, "J. Hepatology, 31. (Suppl. 1), pp. 88-91 (1999)].

Upon initial exposure to HCV, only about 20% of infected subjects develop acute clinical hepatitis, while others show spontaneous resolution of the infection. However, in almost 70% of cases, the virus stimulates the development of a chronic infection that persists for decades [S. Iwarson, "The Natural Course of Chronic Hepatitis," FEMS Microbiology Reviews, 14, pp. 201-204 (1994); D. Lavanchy, "Global Surveillance and Control of Hepatitis C," J. Viral Hepatitis, 6, pp. 35-47 (1999)]. This usually leads to relapses and a progressive complication of liver inflammation, which often leads to more serious illnesses such as cirrhosis and hepatocellular carcinoma [M.C. Kew, "Hepatitis C and Hepatocellular Carcinoma", FEMS Microbiology Reviews, 14, pp. 211-220 (1994); I. Saito et al., "Hepatitis C Virus Infection is Associated with the Development of Hepatocellular Carcinoma," Proc. Natl. Acad. Sci. USA, 87, pp. 6547-6549 (1990)]. Unfortunately, there are currently no broadly effective treatments for reducing the progression of chronic HCV.

The HCV genome encodes a polyprotein of 3010-3033 amino acids [Q.L. Choo, et al., "Genetic Organization and Diversity of Hepatitis C Virus." Proc. Natl. Acad. Sci. USA, 88, pp. 2451-2455 (1991); N. Kato et al., "Molecular Cloning of the Human Hepatitis C Virus Genome From Japanese Patients with Non-A, Non-B Hepatitis," Proc. Natl. Acad. Sci. USA, 87, pp. 9524-9528 (1990); A. Takamizawa et al., "Structure and Organization of the Hepatitis C Virus Genome Isolated From Human Carriers," J. Virol., 65, pp. 1105-1113 (1991)]. Non-structural (NS) HCV proteins are believed to provide the primary catalytic mechanism for virus replication. NS proteins are formed as a result of proteolytic cleavage of polyprotein [R. Bartenschlager et al., "Nonstructural Protein 3 of the Hepatitis C Virus Encodes a Serine-Type Proteinase Required for Cleavage at the NS3 / 4 and NS4 / 5 Junctions," J. Virol., 67, pp. 3835-3844 (1993); A. Grakoui et. al., "Characterization of the Hepatitis C Virus-Encoded Serine Proteinase: Determination of Proteinase-Dependent Polyprotein Cleavage Sites," J. Virol., 67, pp. 2832-2843 (1993); A. Grakoui et al., "Expression and Identification of Hepatitis C Virus Polyprotein Cleavage Products," J. Virol., 67, pp. 1385-1395 (1993); L. Tomei et al., "NS3 is a serine protease required for processing of hepatitis C virus polyprotein", J. Virol., 67, pp. 4017-4026 (1993)].

HCV NS 3 protein (NS3) has serine protease activity, which promotes the processing of most viral enzymes and is therefore considered essential for viral replication and infectivity. Mutations in yellow fever virus NS3 protease are known to reduce viral infectivity [Chambers, T.J. et. al., "Evidence that the N-terminal Domain of Nonstructural Protein NS3 From Yellow Fever Virus is a Serine Protease Responsible for Site-Specific Cleavages in the Viral Polyprotein", Proc. Natl. Acad. Sci. USA, 87, pp. 8898-8902 (1990)]. The first 181 amino acids in NS3 (residues 1027-1207 of the viral polyprotein) have been shown to contain the NS3 serine protease domain, which processes all four HCV polyprotein sites below [C. Lin et al., "Hepatitis C Virus NS3 Serine Proteinase: Trans-Cleavage Requirements and Processing Kinetics", J. Virol., 68, pp. 8147-8157 (1994)].

HCV NS3 serine protease and its associated cofactor, NS4A, contribute to the processing of all viral enzymes and are thus considered essential for viral replication. It turned out that this processing is similar to that carried out by the aspartyl protease of the human immunodeficiency virus, which is also involved in the processing of viral enzymes. HIV protease inhibitors that inhibit viral protein processing are potent antiviral agents in humans, and this suggests that interruption of this stage of the virus's life cycle leads to therapeutically active agents. Therefore, HCV NS3 serine protease is also an attractive target for drug development.

Currently, there are no satisfactory anti-HCV agents or treatments. Until recently, the only conventional treatment for HCV disease was interferon treatment. However, interferons have significant side effects [M. A. Wlaker et al., "Hepatitis C Virus: An Overview of Current Approaches and Progress," DDT, 4, pp. 518-29 (1999); D. Moradpour et al., "Current and Evolving Therapies for Hepatitis C," Eur. J. Gastroenterol. Hepatol., 11, pp. 1199-1202 (1999); H. L. A. Janssen et al. "Suicide Associated with Alfa-Interferon Therapy for Chronic Viral Hepatitis," J. Hepatol., 21, pp. 241-243 (1994); P.F. Renault et al., "Side Effects of Alpha Interferon," Seminars in Liver Disease, 9, pp. 273-277. (1989)] and induce long-term remission only in some (approximately 25%) cases [O. Weiland, "Interferon Therapy in Chronic Hepatitis C Virus Infection", FEMS Microbiol. Rev. 14, pp. 279-288 (1994)]. Recent pegylated forms of interferon (PEG-INTRON® and PEGASYS®) and combination therapy including ribavirin and pegylated interferon (REBETROL®) have provided only modest improvements in the rate of remission and only a partial reduction in side effects. Moreover, the prospects for effective HCV vaccines remain unclear.

Thus, there is a need for more effective therapeutic agents against HCV. Such inhibitors should have therapeutic potential as protease inhibitors, in particular as serine protease inhibitors, and more specifically, as HCV NS3 protease inhibitors. Specifically, such compounds may be useful as antiviral agents, in particular as anti-HCV agents.

Recently, it has been found that incorporation of deuterium into a compound reduces the rate of epimerization through the action of the deuterium isotope, thereby increasing the concentration of active isomers in vivo compared to its non-deuterated analogues.

SUMMARY OF THE INVENTION

The present invention relates to deuterated compounds of formula (I)

and their pharmaceutically acceptable salts, prodrugs and solvates. In the formula (I), D is a deuterium atom.

As for the formula (I),

D is a deuterium atom;

,R ¹ represents

,

Where

представляет собой необязательно замещенный моноциклический азагетероциклил или необязательно замещенный полициклический азагетероциклил, или необязательно замещенный полициклический азагетероцикленил, где ненасыщенность присутствует в кольце, дистальном к кольцу, содержащему группу R²¹ и к которому присоединена группа -C(O)-N(R²)-CDR³-C(O)-C(O)-NR⁴R⁵;

represents an optionally substituted monocyclic azaheterocyclyl or an optionally substituted polycyclic azaheterocyclyl, or an optionally substituted polycyclic azaheterocycle, where unsaturation is present in the ring distal to the ring containing the group R ²¹ and to which the -C (O) -N (R ² ) -CDR group is attached ³ -C (O) -C (O) -NR ⁴ R ⁵ ;

R ²¹ represents Q ³ —W ³ —Q ² —W ² —Q ¹ ; Where

each of W ² and W ³ independently represents a bond, —CO—, —CS—, —C (O) N (Q ⁴ ) -, —CO ₂ -, —O—, —N (Q ⁴ ) —C ( O) -N (Q ⁴ ) -, -N (Q ⁴ ) -C (S) -N (Q ⁴ ) -, -OC (O) NQ ⁴ -, -S-, -SO-, -SO ₂ - , —N (Q ⁴ ) -, —N (Q ⁴ ) SO ₂ -, —N (Q ⁴ ) SO ₂ N (Q ⁴ ) - and hydrogen when any of W ² and W ³ represents an end group;

each of Q ¹ , Q ² and Q ³ independently represents a bond, an optionally substituted aliphatic group, an optionally substituted heteroaliphatic group, an optionally substituted cycloaliphatic group, an optionally substituted aryl, an optionally substituted heteroaryl, an optionally substituted aralkyl, or an optionally substituted heteroaryl; or hydrogen, when any of Q ³ , Q ² or Q ¹ represents an end group, provided that Q ² cannot be a bond when both W ³ and W ² are present; and

each of R ² , R ³ and R ⁴ , independently, is H or C _1-6 alkyl;

R ⁵ represents H, alkyl, cycloalkyl, aryl optionally substituted with 1-4 alkyl groups, alkylaryl, aryl, amino, optionally substituted with 1 or 2 alkyl groups; and

R ²¹ represents Q ³ —W ³ —Q ² —W ² —Q ¹ ; where each of W ² and W ³ independently represents a bond, -CO-, -CS-, -C (O) N (Q ⁴ ) -, -CO ₂ -, -O-, -N (Q ⁴ ) -C (O) -N (Q ⁴ ) -, N (Q ⁴ ) -C (S) -N (Q ⁴ ) -, -OC (O) NQ ⁴ -, -S-, -SO-, -SO ₂ - , —N (Q ⁴ ) -, —N (Q ⁴ ) SO ₂ -, —N (Q ⁴ ) SO ₂ N (Q ⁴ ) - and hydrogen when any of W ² and W ³ represents an end group; each of Q ¹ , Q ² , and Q ³ independently represents a bond, an optionally substituted aliphatic group, an optionally substituted heteroaliphatic group, an optionally substituted cycloaliphatic group, an optionally substituted aryl, an optionally substituted heteroaryl, an optionally substituted aralkyl, or an optionally substituted heteroalkyl; or hydrogen, when any of Q ³ , Q ² or Q ¹ represents an end group, provided that Q ² cannot be a bond when both W ³ and W ² are present.

In some embodiments, implementation, R ¹ represents

Where

each of R ⁶ and R ⁸ independently represents

communication; or

optionally substituted (1,1- or 1,2-) cycloalkylene; or

optionally substituted (1,1- or 1,2-) heterocyclylene; or

methylene or ethylene substituted with one substituent selected from the group consisting of an optionally substituted aliphatic group, an optionally substituted cyclic group and an optionally substituted aromatic group, and wherein said methylene or ethylene is optionally further substituted with a substituent representing an aliphatic group;

each of R ⁷ , R ⁹ , and R ¹¹ independently represents hydrogen or an optionally substituted aliphatic group;

R ¹⁰ represents an optionally substituted aliphatic group, an optionally substituted cyclic group or an optionally substituted aromatic group;

L is —C (O) -, —OC (O) -, —NR ¹¹ C (O) -, —S (O) ₂ -, —NR ¹¹ S (O) ₂ - or a bond; and

n is 0 or 1.

In some embodiments, n is 1.

In some embodiments, R ⁶ is methylene substituted with one substituent selected from the group consisting of an optionally substituted aliphatic group, an optionally substituted cyclic group, and an optionally substituted aromatic group.

In some embodiments, R ⁶ is methylene substituted with isobutyl.

In some embodiments, R ⁷ is hydrogen.

In some embodiments, R ⁸ is methylene substituted with one substituent selected from the group consisting of an optionally substituted aliphatic group, an optionally substituted cyclic group, and an optionally substituted aromatic group. In some other embodiments, R ⁸ is methylene substituted with an optionally substituted cyclic group. In still other embodiments, R ⁸ is methylene substituted with cyclohexyl.

In some embodiments, R ⁹ is hydrogen.

In some embodiments, L is —CO—.

In some embodiments, R ¹⁰ is an optionally substituted aromatic group.

In some embodiments, the implementation of R ¹⁰ selected from the group

In some embodiments, R ¹⁰ is optionally substituted pyrazinyl (e.g., 2-pyrazinyl).

представляет собой замещенный моноциклический азагетероциклил.In some embodiments, implementation

represents a substituted monocyclic azaheterocyclyl.

представляет собой пирролидинил, замещенный в положении 3 атома углерода группой гетероарилокси, где гетероарил необязательно дополнительно замещен 1-4 группами галогена.In some other embodiments, implementation

represents pyrrolidinyl substituted at the 3-position of the carbon atom with a heteroaryloxy group, where heteroaryl is optionally further substituted with 1-4 halogen groups.

представляет собой

.In some embodiments, implementation

represents

.

представляет собой необязательно замещенный полициклический азагетероциклил.In some embodiments, implementation

represents an optionally substituted polycyclic azaheterocyclyl.

представляет собой

В некоторых вариантах осуществления

представляет собой

.In another embodiment

represents

In some embodiments, implementation

represents

.

или

In another embodiment, R ² is hydrogen, each of R ⁴ and R ⁵ independently is hydrogen or cyclopropyl. In another embodiment, R ³ is propyl. In another embodiment, n is 0. In another embodiment, L is —NR ¹¹ C (O) - and R ¹¹ is hydrogen. In another embodiment, R ¹⁰ is an optionally substituted aliphatic group. In another embodiment, R ¹⁰ is tert-butyl. In another embodiment, the compound is

or

,In another embodiment, R ¹ represents

,

Where

A represents - (CHX ¹ ) _a -;

B represents - (CHX ² ) _b -;

a is 0-3;

b is 0-3 provided that the sum of a + b is 2 or 3;

each of X ¹ and X ^{2 is} independently selected from hydrogen, an optionally substituted C _1-4 aliphatic group, and an optionally substituted aryl;

each of Y ¹ and Y ² independently represents hydrogen, an optionally substituted aliphatic group, an optionally substituted aryl, amino group or —OQ ⁴ ; where each Q ⁴ independently represents hydrogen or an optionally substituted aliphatic group;

R ²² represents an optionally substituted aliphatic group, an optionally substituted heteroaliphatic group, an optionally substituted cycloaliphatic group, an optionally substituted heterocycloaliphatic group, an optionally substituted aryl or optionally substituted heteroaryl. In some embodiments, R ²¹ is an optionally substituted alkylcarbonyl.

включает все стереоспецифические энантиомеры, например,

(когда A и B оба представляют собой CH₂, и Y¹ и Y² оба представляют собой H).Group

includes all stereospecific enantiomers, for example,

(when A and B both represent CH ₂ , and Y ¹ and Y ² both represent H).

In some embodiments, R ²¹ is aminoalkylcarbonyl, haloalkylcarbonyl, arylalkylcarbonyl, arylalkylcarbonyl, (cycloaliphatic group) alkylcarbonyl or (heterocycloaliphatic group) alkylcarbonyl, each of which is optionally substituted with 1-3 substituents. In some embodiments, R ²¹ is heterocycloalkyl-hydroxycarbonylamino-alkylcarbonyl, heteroaryl-carbonylamino-alkyl-carbonylamino-alkyl-carbonyl, bicycloaryl-sulfonylamino-alkylcarbonyl, aryl-alkoxy-carbonylamino-alkyl-carbonyl, alkyl-carbonyl (aliphatic group) hydroxycarbonylamino-alkyl-carbonyl, (cycloaliphatic group) alkyl-aminocarbonylamino-alkyl-carbonyl, heteroaryl-carbonylamino-alkyl-carbonylamino-alkyl-carbonyl, alkyl-aminocarbonylamino-alkyl-carbonyl or bicycloaryl -aminocarbonylamino-alkyl-carbonyl, each of which is optionally substituted with 1-3 substituents. In some embodiments, R ²² is an optionally substituted aliphatic group, an optionally substituted heteroaliphatic group, an optionally substituted cycloaliphatic group, an optionally substituted heterocycloaliphatic group, an optionally substituted aryl, or an optionally substituted heteroaryl. In some embodiments, R ²² is optionally substituted phenyl, optionally substituted naphthyl, optionally substituted anthracenyl, optionally substituted naphthalene, or optionally substituted anthracene. In some embodiments, each of X ¹ , X ² , Y ¹ and Y ² is hydrogen, each of a and b is 1.

In some embodiments, R ²¹ is an optionally substituted alkylcarbonyl.

In some embodiments, R ²¹ is aminoalkylcarbonyl, haloalkylcarbonyl, arylalkylcarbonyl, arylalkylcarbonyl, (cycloaliphatic group) alkylcarbonyl or (heterocycloaliphatic group) alkylcarbonyl, each of which is optionally substituted with 1-3 substituents.

In some embodiments, R ²¹ is heterocycloalkyl-hydroxycarbonylamino-alkylcarbonyl, heteroaryl-carbonylamino-alkyl-carbonylamino-alkyl-carbonyl, bicycloaryl-sulfonylamino-alkylcarbonyl, aryl-alkoxy-carbonylamino-alkyl-carbonyl, alkyl-carbonyl (aliphatic group) hydroxycarbonylamino-alkyl-carbonyl, (cycloaliphatic group) alkyl-aminocarbonylamino-alkyl-carbonyl, (cycloaliphatic group) alkyl-carbonylamino-alkyl-carbonyl, heteroaryl-carbonylamino-alkyl-carbonylamino-alkyl-ka Bonilla, alkyl-aminocarbonylamino-alkyl-carbonyl or bicycloaryl-aminocarbonylamino-alkyl-carbonyl, each of which is optionally substituted with 1-3 substituents.

In some embodiments, R ²² is an optionally substituted aliphatic group, an optionally substituted heteroaliphatic group, an optionally substituted cycloaliphatic group, an optionally substituted heterocycloaliphatic group, an optionally substituted aryl, or an optionally substituted heteroaryl.

In some embodiments, R ²² is optionally substituted phenyl, optionally substituted naphthyl, optionally substituted anthracenyl, optionally substituted naphthalene, or optionally substituted anthracene.

In some embodiments, each of X ¹ , X ² , Y ¹ and Y ² is hydrogen, each of a and b is 1.

In some embodiments, R ²² is an optionally substituted aliphatic group, an optionally substituted heteroaliphatic group, an optionally substituted cycloaliphatic group, an optionally substituted heterocycloaliphatic group, an optionally substituted aryl, or an optionally substituted heteroaryl.

.In one embodiment, the compound is

.

The epimerization of deuterated compounds of the present invention is slower than that of similar non-deuterated compounds. As shown below, deuterated compound 1 is very slowly converted to an undeuterated intermediate, which is then converted to epimers 2 and 3 . Epimers 2 and 3 are then maintained in equilibrium, which further slows down the epimerization of deuterated compound 1.

As a result of their slow epimerization, the deuterated compounds of the present invention can increase the concentration of active isomers in vivo compared to non-deuterated analogues.

In some embodiments, the deuterium enrichment is at least 50% in the compounds of the present invention. In some embodiments, the deuterium enrichment is at least 80% in the compounds of the present invention. In some embodiments, the deuterium enrichment is at least 90% in the compounds of the present invention. In some embodiments, the deuterium enrichment is at least 99% in the compounds of the present invention.

The present invention also relates to a pharmaceutical composition comprising a pharmaceutically acceptable carrier and a compound of formula (I) or any of its embodiments described above.

The present invention also relates to a method for increasing the concentration of an active isomer of a pharmaceutical substance in vivo , comprising administering to a patient in need thereof a sufficient amount of the deuterated isomer of the pharmaceutical substance to provide a pharmaceutical effect.

The present invention also relates to a method for increasing the bioavailability of a compound, comprising replacing a hydrogen atom that is bound to a steric carbon atom in the compound with a deuterium atom. In one embodiment, the deuterated compound is a compound of formula (I) or any of its embodiments described above.

The present invention also relates to a method for inhibiting HCV protease, comprising contacting the HCV protease with a deuterated compound of formula (I) or any of its embodiments described above.

The present invention also relates to a method for treating a patient suffering from HCV infection or a condition mediated by HCV protease, comprising administering to the patient a pharmaceutically effective amount of a deuterated compound of formula (I) or any of its embodiments described above.

Also included in the scope of the present invention is a method for preparing an optically enriched compound of formula 1, wherein

carbon atoms alpha and beta relative to the carboxy group are stereo centers;

R ₁ independently represents H, an optionally substituted aliphatic group, an optionally substituted cycloaliphatic group, an optionally substituted arylaliphatic group, an optionally substituted heteroaliphatic group, or an optionally substituted heteroarylaliphatic group;

R ' ₁ represents deuterium,

R ′ ₂ represents —NHR ₂ or —OE;

R ₂ represents H, an optionally substituted aliphatic group, an optionally substituted cycloaliphatic group, an optionally substituted arylaliphatic group, an optionally substituted heteroaliphatic group or an optionally substituted heteroarylaliphatic group; and

E represents C _1-6 alkyl or benzyl.

The method includes the following steps:

a) salt formation of a compound of formula 1; and

b) crystallizing said salt to give a compound with more than 55% enantiomeric excess.

In some embodiments, R ₁ is C _1-6 alkyl, and R ′ ₂ is —NHR ₂ , where R ₂ is C _1-6 alkyl or C _1-6 cycloalkyl. In some embodiments, R ₁ is propyl and R ₂ is cyclopropyl.

In some embodiments, the method further comprises aminating a compound of formula ii

amination reagent to obtain a compound of formula iii

In some other embodiments, the amination reagent is an azide salt, and the intermediate azide compound is reduced by hydrogenation.

In some embodiments, the method further comprises oxidizing an unsaturated compound of formula i

where R ′ ₂ is —NHR ₂ or —OE, where E is C _1-5 alkyl or optionally substituted benzyl, with an oxidizing reagent to give a compound of formula ii.

In some other embodiments, the oxidizing reagent comprises tert-butyl hydroperoxide.

In some other embodiments, the oxidizing reagent further comprises a chiral reagent. In some other embodiments, the oxidizing agent is a mixture of samarium (III) isopropoxide, triphenylarsine oxide, S - (-) 1,1'-bi-2-naphthol and 4 Å molecular sieves. In some other embodiments, the oxidizing reagent comprises urea hydrogen peroxide in the presence of trifluoroacetic anhydride.

In some other embodiments, the process further comprises hydrolyzing the compound of formula ii to produce an acid, and then converting the resulting acid to an amide compound of formula ii, where R ′ ₂ is —NHR ₂ .

Also included in the scope of the present invention is a process for preparing a compound of formula 1

Where

R ₁ represents H, an optionally substituted aliphatic group, an optionally substituted cycloaliphatic group, an optionally substituted arylaliphatic group, an optionally substituted heteroaliphatic group or an optionally substituted heteroarylaliphatic group;

R ' ₁ represents deuterium,

R ₂ represents H, an optionally substituted aliphatic group, an optionally substituted cycloaliphatic group, an optionally substituted arylaliphatic group, an optionally substituted heteroaliphatic group or an optionally substituted heteroarylaliphatic group; and

the compound of formula 1 has an enantiomeric excess of more than 55%. The specified method includes the following stages:

a) oxidation of an unsaturated compound of formula i

to obtain the compounds of formula ii

b) reacting a compound of formula ii with an amination reagent to produce a compound of formula iii

c) salt formation of a compound of formula iii with an optically active organic acid; and

d) crystallizing said salt to give a compound with more than 55% enantiomeric excess.

In some embodiments, a compound of Formula 1 is (2S, 3S) -3-amino-3-deutero-N-cyclopropyl-2-hydroxyhexanamide. In some embodiments, the organic acid is L-tartaric acid or deoxycholic acid.

Also included in the scope of the present invention is a method for producing an optically enriched compound of formula 1:

Where

carbon atoms alpha and beta relative to the carboxy group are stereo centers;

R ₁ independently represents H, an optionally substituted aliphatic group, an optionally substituted cycloaliphatic group, an optionally substituted arylaliphatic group, an optionally substituted heteroaliphatic group, or an optionally substituted heteroarylaliphatic group;

R ′ ₁ represents deuterium, so that the deuterium enrichment is at least 50%;

R ′ ₂ represents —NHR ₂ or —OE;

R ₂ represents H, an optionally substituted aliphatic group, an optionally substituted cycloaliphatic group, an optionally substituted arylaliphatic group, an optionally substituted heteroaliphatic group or an optionally substituted heteroarylaliphatic group; and

E represents C _1-6 alkyl or benzyl.

The specified method includes the following stages:

a) salt formation of a compound of formula 1, and

b) crystallizing said salt to give a compound with more than 55% enantiomeric excess.

In some embodiments, R ₁ is C ₁ -C ₆ alkyl and R ′ ₂ is —NHR ₂ where R ₂ is C _1-6 alkyl or C _1-6 cycloalkyl. In some embodiments, R ₁ is propyl and R ₂ is cyclopropyl.

In some embodiments, the method further comprises the step of aminating the compound of formula ii

amination reagent to obtain a compound of formula iii

In some embodiments, the amination reagent is an azide salt, and the intermediate azide compound is reduced by hydrogenation.

In some embodiments, the method further comprises the step of oxidizing the unsaturated compound of formula i

where R ′ ₂ is —NHR ₂ or —OE, where E is C _1-5 alkyl or optionally substituted benzyl, with an oxidizing reagent to give a compound of formula ii.

In some embodiments, the oxidizing reagent comprises tert-butyl hydroperoxide. In some other embodiments, the oxidizing reagent further comprises a chiral reagent. In some embodiments, the oxidizing reagent is a mixture of samarium (III) isopropoxide, triphenylarsine oxide, S - (-) 1,1'-bi-2-naphthol and 4 Å molecular sieves. In some embodiments, the oxidizing reagent comprises urea hydrogen peroxide in the presence of trifluoroacetic anhydride.

In some embodiments, R ′ ₂ is —OE. In some embodiments, R ′ ₂ is —NHR ₂ .

In some embodiments, the method further comprises hydrolyzing the compound of Formula ii to produce an acid and then converting the resulting acid to an amide compound of Formula ii, where R ′ ₂ is —NHR ₂ .

In some embodiments, the method further comprises oxidizing a compound of formula iv

to obtain the compounds of formula ii. In some cases, the oxidation is carried out using manganese dioxide.

In some embodiments, the method further comprises reducing a compound of formula v

to obtain the compounds of formula iv. In some cases, the compound is reduced using Red-Al® and then quenched with deuterium oxide. As known in the art, "Red-Al®" refers to a compound [(CH ₃ OCH ₂ OCH ₂ ) ₂ AlH ₂ ] Na, which is commercially available, usually in the form of a solution in toluene (for example, 70% w / w .). For more information regarding Red-Al®, see, for example, Bates RW et al., Tetrahedron, 1990, 46, 4063.

Also included in the scope of the present invention is a process for preparing a compound of formula 1

Where

R ₁ represents H, an optionally substituted aliphatic group, an optionally substituted cycloaliphatic group, an optionally substituted arylaliphatic group, an optionally substituted heteroaliphatic group or an optionally substituted heteroarylaliphatic group;

R ' ₁ represents deuterium,

R ₂ represents H, an optionally substituted aliphatic group, an optionally substituted cycloaliphatic group, an optionally substituted arylaliphatic group, an optionally substituted heteroaliphatic group or an optionally substituted heteroarylaliphatic group; and

the compound of formula 1 has an enantiomeric excess of more than 55%.

The specified method includes the following stages:

a) oxidation of an unsaturated compound of formula i

to obtain the compounds of formula ii

b) reacting a compound of formula ii with an amination reagent to produce a compound of formula iii

c) salt formation of a compound of formula iii with an optically active organic acid; and

d) crystallizing said salt to give a compound with more than 55% enantiomeric excess.

In some embodiments, a compound of Formula 1 is (2S, 3S) -3-amino-3-deutero-N-cyclopropyl-2-hydroxyhexanamide. In some embodiments, the organic acid is L-tartaric acid or deoxycholic acid.

DETAILED DESCRIPTION OF THE INVENTION

I. Definitions

A. Terms

As used herein, the term “aliphatic” embraces alkyl, alkenyl and alkynyl.

As used herein, an “alkyl” group refers to a saturated aliphatic hydrocarbon group containing 1-8 (e.g., 1-6 or 1-4) carbon atoms. The alkyl group may be linear or branched. Examples of the alkyl group include, but are not limited to, methyl, ethyl, propyl, isopropyl, butyl, isobutyl, sec-butyl, tert-butyl, n-pentyl, n-heptyl and 2-ethylhexyl. The alkyl group may be optionally substituted by one or more substituents, such as alkoxy, cycloalkyloxy, heterocycloalkyloxy, aryloxy, heteroaryloxy, aralkyloxy, heteroarylalkoxy, amino, nitro, carboxy, cyano, halogen, hydroxy, sulfo, mercapto, alkylsulfonyl, alkylsulfonyl, alkylsulfonyl, amylsulfonyl, , alkylcarbonylamino, cycloalkylcarbonylamino, cycloalkylalkylcarbonylamino, arylcarbonylamino, aralkylcarbonylamino, heterocycloalkylcarbonylamino, heterocycloalkylalkylcarbonylamino, heteroarylcar bonilamino, heteroaralkylcarbonylamino, urea, thiourea, sulfamoyl, sulfamide, alkoxycarbonyl or alkylcarbonyloxy.

As used herein, an “alkenyl" group refers to an aliphatic carbon group that contains 2-8 (e.g., 2-6 or 2-4) carbon atoms and at least one double bond. Like an alkyl group, an alkenyl group may be linear or branched. Examples of the alkenyl group include, but are not limited to, allyl, isoprenyl, 2-butenyl, and 2-hexenyl. The alkenyl group may optionally be substituted with one or more substituents, such as alkoxy, cycloalkyloxy, heterocycloalkyloxy, aryloxy, heteroaryloxy, aralkyloxy, heteroarylalkoxy, amino, nitro, carboxy, cyano, halogen, hydroxy, sulfo, mercapto, alkylsulfonyl, alkylsulfonyl, alkylsulfonyl, , alkylcarbonylamino, cycloalkylcarbonylamino, cycloalkylalkylcarbonylamino, arylcarbonylamino, aralkylcarbonylamino, heterocycloalkylcarbonylamino, heterocycloalkylalkylcarbonylamino, heteroaryl arbonylamino, heteroaralkylcarbonylamino, urea, thiourea, sulfamoyl, sulfamide, alkoxycarbonyl or alkylcarbonyloxy.

As used herein, an “alkynyl" group refers to an aliphatic carbon group that contains 2-8 (e.g., 2-6 or 2-4) carbon atoms and has at least one triple bond. The alkynyl group may be linear or branched. Examples of the alkynyl group include, but are not limited to, propargyl and butynyl. The alkynyl group may be optionally substituted with one or more substituents, such as alkoxy, cycloalkyloxy, heterocycloalkyloxy, aryloxy, heteroaryloxy, aralkyloxy, heteroarylalkoxy, amino, nitro, carboxy, cyano, halogen, hydroxy, sulfo, mercapto, alkylsulfonyl, alkylsulfonyl, alkylsulfonyl, , alkylcarbonylamino, cycloalkylcarbonylamino, cycloalkylalkylcarbonylamino, arylcarbonylamino, aralkylcarbonylamino, heterocycloalkylcarbonylamino, heterocycloalkylalkylcarbonylamino, heteroaryl arbonylamino, heteroaralkylcarbonylamino, urea, thiourea, sulfamoyl, sulfamide, alkoxycarbonyl or alkylcarbonyloxy.

As used herein, an “amino group” refers to —NR ^X R ^Y , where each of R ^X and R ^{Y is} independently hydrogen, alkyl, cycloalkyl, (cycloalkyl) alkyl, aryl, aralkyl, heterocycloalkyl, (heterocycloalkyl) alkyl, heteroaryl, or heteroalkyl When the term “amino” does not mean an end group (for example, alkylcarbonylamino), it is represented by the formula —NR ^X -. R ^X is as defined above.

As used herein, an “aryl” group refers to a phenyl, naphthyl or benzocondensed group containing 2-3 rings. For example, a benzofused group includes phenyl fused to one or two C _4-8 cycloaliphatic groups, for example, 1,2,3,4-tetrahydronaphthyl, indanyl, dihydroindanyl or fluorenyl. Aryl is optionally substituted with one or more substituents, such as alkyl (including carboxyalkyl, hydroxyalkyl and haloalkyl such as trifluoromethyl), alkenyl, alkynyl, cycloalkyl, (cycloalkyl) alkyl, heterocycloalkyl, (heterocycloalkyl) alkyl, aryl, heteroaryl, alkoxy, heterocycloalkyloxy, aryloxy, heteroaryloxy, aralkyloxy, heteroaralkyloxy, aroyl, heteroaroyl, amino, nitro, carboxy, alkoxycarbonyl, alkylcarbonyloxy, aminocarbonyl, alkylcarbonylamino, cycloalkylcarbonylamino, (cycloalkyl) amino, arylcarbonylamino, aralkylcarbonylamino, (heterocycloalkyl) carbonylamino, (heterocycloalkyl) alkylcarbonylamino, heteroarylcarbonylamino, heteroaralkylcarbonylamino, cyano, halogen, hydroxy, acyl, mercapto, sulfamylamidoamyloamoyl-sulfamylamino-sulfamylamino-sulfamylamido

As used herein, an “aralkyl” group refers to an alkyl group (eg, a C _1-4 alkyl group) that is substituted with an aryl group. Both “alkyl” and “aryl” are defined above. An example of an aralkyl group is benzyl.

As used herein, the term cycloaliphatic embraces cycloalkyl, cycloalkenyl and cycloalkynyl.

As used herein, a “cycloalkyl” group refers to an aliphatic carbocyclic ring of 3-10 (e.g., 4-8) carbon atoms. Examples of cycloalkyl groups include cyclopropyl, cyclopentyl, cyclohexyl, cycloheptyl, adamantyl, norbornyl, cubyl, octahydroindenyl, decahydronaphthyl, bicyclo [3.2.1] octyl, bicyclo [2.2.2] octyl, bicyclo [3.3.1] nonyl and bicyclo [3.2. 3] nonyl. A "cycloalkenyl" group as used herein refers to a non-aromatic carbocyclic ring of 3-10 (e.g., 4-8) carbon atoms having one or more double bonds. Examples of cycloalkenyl groups include cyclopentenyl, 1,4-cyclohexadienyl, cycloheptenyl, cyclooctenyl, hexahydroindenyl, octahydronaphthyl, bicyclo [2.2.2] octenyl and bicyclo [3.3.1] nonenyl. The cycloalkyl or cycloalkenyl group may optionally be substituted with one or more substituents, such as alkyl (including carboxyalkyl, hydroxyalkyl and haloalkyl such as trifluoromethyl), alkenyl, alkynyl, cycloalkyl, (cycloalkyl) alkyl, heterocycloalkyl, (heterocycloalkyl, alkyl) , alkoxy, cycloalkyloxy, heterocycloalkyloxy, aryloxy, heteroaryloxy, aralkyloxy, heteroaralkyloxy, aroyl, heteroaroyl, amino, nitro, carboxy, alkoxycarbonyl, alkylcarbonyloxy, aminocarbonyl, alkylcarbonylamino, ikloalkilkarbonilamino, (cycloalkyl) alkylcarbonylamino, arylcarbonylamino, aralkylcarbonylamino, (heterocycloalkyl) carbonylamino, (heterocycloalkyl) alkylcarbonylamino, geteroarilkarbonilamino, geteroaralkilkarbonilamino, cyano, halo, hydroxy, acyl, mercapto, alkylsulfanyl, sulfoxy, urea, thiourea, sulfamoyl, sulfamide, oxo, or carbamoyl .

Used in the present description, the term heterocycloaliphatic means heterocycloalkyl, heterocycloalkenyl and heterocycloalkynyl. As used herein, a “heterocycloalkyl” group refers to a 3-10 membered (eg, 4-8 membered) saturated ring structure in which one or more ring atoms are a heteroatom, for example, N, O or S. Examples of a heterocycloalkyl group include piperidinyl, piperazinyl, tetrahydropyranyl, tetrahydrofuryl, dioxolanyl, oxazolidinyl, isoxazolidinyl, morpholinyl, octahydrobenzofuryl, octahydrochromenyl, octahydrothiochrome, octahydrohydrodinhydrodinhydrodinhydrodinhydrodinhydride zo [b] thiophenyl, 2-oxabicyclo [2.2.2] octyl, 1-azabicyclo [2.2.2] octyl, 3-azabicyclo [3.2.1] octyl and 2,6-dioksatritsiklo [3.3.1.03,7] nonyl. A “heterocycloalkenyl” group as used herein refers to a 3-10 membered (eg, 4-8 membered) non-aromatic ring structure having one or more double bonds, and wherein one or more ring atoms are a heteroatom, for example, N, O or S. A heterocycloalkyl or heterocycloalkenyl group may be optionally substituted with one or more substituents, such as alkyl (including carboxyalkyl, hydroxyalkyl and haloalkyl, such as trifluoromethyl), alkenyl, alkynyl, cycloalkyl, (cyclic loalkyl) alkyl, heterocycloalkyl, (heterocycloalkyl) alkyl, aryl, heteroaryl, alkoxy, cycloalkyloxy, heterocycloalkyloxy, aryloxy, heteroaryloxy, aralkyloxy, heteroaralkyloxy, aroyl, heteroalkyloxy, carbonyloxy, carbonyloxy, alkoxycarbonyl cycloalkyl) alkylcarbonylamino, arylcarbonylamino, aralkylcarbonylamino, (heterocycloalkyl) carbonylamino, (heterocycloalkyl) alkylcarbonylamino, heteroarylcarbonylamino, heteroaralkylcarbonylamino, cyano, halogen, g droksi, acyl, mercapto, alkylsulfanyl, sulfoxy, urea, thiourea, sulfamoyl, sulfamide, oxo, or carbamoyl. In some cases, the substituent in the heterocycloalkyl or heterocycloalkenyl may itself be cyclic (which optionally contains one or more heteroatoms), and thus the resulting substituted heterocycloalkyl or heterocycloalkenyl is a spiro ring system, for example

A “heteroaryl” group as used herein refers to a monocyclic, bicyclic or tricyclic ring structure containing 5-15 ring atoms, where one or more ring atoms are heteroatoms, for example, N, O or S, and where one or more rings of a bicyclic or tricyclic ring structure are aromatic. Some examples of heteroaryl are pyridyl, furyl, pyrrolyl, thienyl, thiazolyl, oxazolyl, imidazolyl, indolyl, 2,3-dihydroindolyl, quinolyl, 1,2-dihydroquinolyl, 1,2,3,4-tetrahydroquinolyl, tetrazolyl, benzofuryl, 2, 3-dihydrobenzofuranyl, benzthiazolyl, xanthene, thioxanthene, phenothiazine, dihydroindole and benzo [1,3] dioxol. Heteroaryl is optionally substituted with one or more substituents, such as alkyl (including carboxyalkyl, hydroxyalkyl and haloalkyl such as trifluoromethyl), alkenyl, alkynyl, cycloalkyl, (cycloalkyl) alkyl, heterocycloalkyl, (heterocycloalkyl) alkyl, aryl, heteroaryloalkyl, alk heterocycloalkyloxy, aryloxy, heteroaryloxy, aralkyloxy, heteroaralkyloxy, aroyl, heteroaroyl, amino, nitro, carboxy, alkoxycarbonyl, alkylcarbonyloxy, aminocarbonyl, alkylcarbonylamino, cycloalkylcarbonylamino, (cycloalkyl) rbonylamino, arylcarbonylamino, aralkylcarbonylamino, (heterocycloalkyl) carbonylamino, (heterocycloalkyl) alkylcarbonylamino, heteroarylcarbonylamino, heteroaralkylcarbonylamino, cyano, halogen, hydroxy, acyl, mercaptoamino, sulfo, carbamyl, sulfo, carbamocyl, alkoxy, sulfo, carbamyl, carbamide, sulfocam, carbamide, alkyl

A “heteroalkyl” group as used herein refers to an alkyl group (eg, a C _1-4 alkyl group) that is substituted with a heteroaryl group. Both “alkyl” and “heteroaryl” are defined above.

As used herein, a “cyclic group” includes cycloalkyl, heterocycloalkyl, cycloalkenyl, heterocycloalkenyl, aryl or heteroaryl, each of which is defined above.

As used herein, an “acyl” group refers to a formyl group or an alkyl-C (= O) - group, wherein “alkyl” is defined above. Acetyl and pivaloyl are examples of acyl groups.

As used herein, a “carbamoyl" group refers to a group having the structure —O — CO — NR ^X R ^Y or —NR ^X —CO — OR ^Z , wherein R ^X and R ^{Y are as} defined above and R ^Z may be alkyl, aryl , aralkyl, heterocycloalkyl, heteroaryl or heteroaralkyl.

As used herein, the carboxy and sulfo groups refer to —COOH and —SO ₃ H, respectively.

Used in the present description, the group "alkoxy" refers to the group alkyl-O-, where "alkyl" is defined above.

As used herein, the group “sulfoxy” refers to —O — SO — R ^X or —SO — OR ^X , wherein R ^{X is as} defined above.

As used herein, a sulfanyl group refers to —SR ^X , where R ^{X is as} defined above.

As used herein, a sulfinyl group refers to —S (O) —R ^X , where R ^{X is as} defined above.

Used in the present description, the sulfonyl group refers to -S (O) ₂ -R ^X , where R ^{X is} defined above.

Used in the present description, "halogen" or the group "halo" refers to fluorine, chlorine, bromine or iodine.

As used herein, a “sulfamoyl” group refers to the structure —S (O) ₂ —NR ^X R ^Y or —NR ^X —S (O) ₂ —R ^X , where R ^X , R ^Y, and R ^{Z are as} defined above.

As used herein, a “sulfamide” group refers to the structure —NR ^X —S (O) ₂ —NR ^Y R ^Z wherein R ^X , R ^Y, and R ^{Z are as} defined above.

As used herein, the urea group refers to the —NR ^X —CO — NR ^Y R ^Z structure and the thiourea group refers to the —NR ^X —CS-NR ^Y R ^Z structure. R ^X , R ^Y and R ^{Z are as} defined above.

Used in the present description, the group "guanidino" refers to the structure-N = C (NR ^X R ^Y ) N (R ^X R ^Y ), where R ^X and R ^{Z are} defined above.

As used herein, the term “amidino group” refers to the structure —C═ (NR ^X ) N (R ^X R ^Y ), wherein R ^X and R ^{Y are as} defined above.

As used herein, the term “oximino group” refers to the structure —C═N — OR ^X , where R ^{X is as} defined above.

The effective amount used in the present description is defined as the amount needed to provide a therapeutic effect in the patient being treated, and is usually determined based on the age, surface area, body weight and condition of the patient. The dose ratio for animals and humans (calculated in milligrams per square meter of body surface area) is described by Freireich et al., Cancer Chemother. Rep., 50: 219 (1966). The surface area of the body can be approximately determined based on the height and weight of the patient. See, for example, Scientific Tables, Geigy Pharmaceuticals, Ardsley, New York, 537 (1970).

As used herein, the term “patient” refers to mammals, including humans.

The antagonist used in the present description is a molecule that binds to the receptor without activating the receptor. It competes with endogenous ligand (s) or substrate (s) for the receptor binding site (s), thus inhibiting the receptor's ability to transduce the intracellular signal in response to binding to the endogenous ligand.

The phrase "optionally substituted" is used interchangeably with the phrase "substituted or unsubstituted." As described herein, the compounds of the present invention may optionally be substituted with one or more substituents, such as generally illustrated above or exemplified by specific classes, subclasses, and species of the present invention. Unless otherwise indicated, each of the specific groups for the variables R ¹ , R ² , R ³ , R ⁴ and R ⁵ in the formula (I) may optionally be substituted with one or more substituents described in this application. Each substituent of a particular group is additionally optionally substituted with one to three substituents selected from halogen, cyano, alkoxy, hydroxyl, nitro, haloalkyl and alkyl. For example, an alkyl group may be substituted with alkylsulfanyl, and alkylsulfanyl may optionally be substituted with one to three substituents selected from halogen, oxo, cyano, alkoxy, hydroxyl, nitro, haloalkyl and alkyl. As a further example, alkyl may be substituted with (cycloalkyl) carbonylamino and the cycloalkyl portion of (cycloalkyl) carbonylamino may optionally be substituted with one to three substituents selected from halogen, cyano, oxo, alkoxy, hydroxyl, nitro, haloalkyl and alkyl.

As a rule, the term “substituted”, regardless of whether the term “optionally” is present before it or not, refers to the replacement of hydrogen radicals in a given structure with a radical of said substituent. Specific substituents are described above in the definitions and below in the description of the compounds and their examples. Unless otherwise indicated, an optionally substituted group may contain a substituent at each substitutable position of the group, and when in any particular structure more than one position may be substituted by more than one substituent selected from the specified group, and such substituents may be either the same, either excellent in every position. A ring substituent, such as heterocycloalkyl, may be bonded to another ring, such as cycloalkyl, to form a spiro-bicyclic ring system, for example, both rings contain one atom common to these rings. As should be understood by those skilled in the art, combinations of substituents contemplated by the present invention are those combinations that provide stable or chemically achievable compounds.

The phrase “stable or chemically achievable” as used herein refers to compounds that are not substantially altered, subject to conditions that make it possible to obtain, determine and, preferably, isolate, purify and use them for one or more purposes, which are disclosed in this application. In some embodiments, a stable compound or chemically achievable compound is a compound that is substantially unchanged when it is maintained at a temperature of 40 ° C. or lower, in the absence of moisture, or in other chemically reactive conditions, for at least a week.

Unless otherwise specified, the structures provided herein also include all isomeric (eg, enantiomeric, diastereomeric, and geometric (or conformational)) forms of these structures; for example, the R- and S-configurations for each asymmetric center, (Z) and (E) double bond isomers and (Z) and (E) conformational isomers. Therefore, individual stereochemical isomers, as well as enantiomeric, diastereomeric and geometric (or conformational) mixtures of the compounds of the present invention are included in the scope of the present invention. Unless otherwise indicated, all tautomeric forms of the compounds of the present invention are included in the scope of the present invention.

In addition, unless otherwise indicated, the structures presented in this application also include compounds that differ only in the presence of one or more isotopically enriched atoms. For example, compounds having the structures described in the present invention, in addition to replacing hydrogen with deuterium or tritium, or replacing carbon with ¹³ C- or ^l4 C-enriched carbon, are included in the scope of the present invention. Such compounds are useful, for example, as analytical tools or probes in biological assays.

An N-oxide derivative or pharmaceutically acceptable salt of each of the compounds of formula (I) is also included within the scope of the present invention. For example, the ring nitrogen atom of the imidazole or pyrazole core ring or a nitrogen-containing heterocyclyl substituent can form an oxide in the presence of a suitable oxidizing agent, such as m-chloroperbenzoic acid or H ₂ O ₂ .

A compound of formula (I) that is inherently acidic (for example, containing a carboxyl or phenolic hydroxyl group) can form a pharmaceutically acceptable salt, such as a sodium, potassium, calcium or gold salt. Also included within the scope of the present invention are salts formed with pharmaceutically acceptable amines, such as ammonia, alkylamines, hydroxyalkylamines and N-methylglycamine. The compound of formula (I) can be treated with an acid to give acid addition salts. Examples of such acids include hydrochloric acid, hydrobromic acid, hydroiodic acid, sulfuric acid, methanesulfonic acid, phosphoric acid, p-bromophenyl sulfonic acid, carbonic acid, succinic acid, citric acid, benzoic acid, oxalic acid, malic acid, malic acid , fumaric acid, ascorbic acid, maleic acid, acetic acid and other mineral and organic acids, well known to specialists in this field. Acid addition salts can be obtained by treating a compound of formula (I) in the form of its free base with a sufficient amount of acid (e.g., hydrochloric acid) to obtain an acid addition salt (e.g., hydrochloride salt). The acid addition salt can again be converted to the free base by treating the salt with a suitable dilute aqueous alkaline solution (for example, sodium hydroxide, sodium bicarbonate, potassium carbonate or ammonia). The compounds of formula (I) can also be, for example, in the form of achiral compounds, racemic mixtures, optically active compounds, pure diastereomers or a mixture of diastereomers.

B. Abbreviations

The abbreviations below have the following meanings. If an abbreviation is not defined, it has generally accepted meaning.

Bemp = 2-tert-butylimino-2-diethylamino-1,3-dimethylperhydro-1,3,2-diazaphosphorin Boc = tert-butoxycarbonyl Bop = benzotriazol-1-yloxy-tris (dimethylamino) phosphonium hexafluorophosphate usd = widened doublet ear.s = widened singlet Cdi = carbonyldiimidazole d = doublet dd = doublet of doublets Dic = diisopropylcarbodiimide DMF = dimethylformamide DMAP = dimethylaminopyridine DMSO = dimethyl sulfoxide EDCI = ethyl 1- (3-dimethylaminopropyl) carbodiimide eq. = equivalents EtOAc = ethyl acetate g = grams Hobt = 1-hydroxybenzotriazole Dipa = Hunig base = diisopropylethylamine l = liter m = multiplet M = molar max = maximum meq = milliequivalent ml = milliliter mm = millimeter mmol = millimole Moc = methoxycarbonyl n = normal N / a = Not available ng = nanograms nm = nanometers OD = optical density PEPC = 1- (3- (1-pyrrolidinyl) propyl) -3-ethylcarbodiimide PP-HOBT = piperidine-piperidin-1-hydroxybenzotriazole psi = pounds per square inch Ph = phenyl quart = quartet quint = quintet rpm = rpm from = singlet t = triplet TFUK = trifluoroacetic acid TLC = thin layer chromatography μl = microliter UV = ultraviolet

II. The compounds of the present invention

Typically, the deuterated compounds of the present invention can be synthesized by methods known in the art as methods for synthesizing their non-deuterated forms, except that the deuterated starting material or reactant is used in the synthesis method. Examples of suitable methods include the methods described in US application No. 60/711530; WO 02/18369; WO 07/022459; Advanced Organic Chemistry, 2 ^nd Ed., P. 204, J. March, McGraw Hill, New York, NY, 1997; and Synthesis A: Elemes and Ragnarsosson, J. of Chem. Soc, Perkin 1, 1996, 537.

All publications referred to in this application are fully incorporated by reference.

Compounds of formula I are prepared using known methods, for example, as illustrated below in Scheme I.

In accordance with Scheme I, an acid of formula i is reacted with a deuterated amino alcohol-amide compound of formula ii in the presence of a condensing reagent, such as, for example, EDCI and HOSu, to produce a hydroxyamide of formula iii . In some embodiments, the percentage of deuterium enrichment (D) shown in ii is greater than 10%. In other embodiments, enrichment is from 10% to 99.95%, from 40% to 99.95%, from 50% to 99.95%, from 60% to 99.95%, from 80% to 99.95% , from 90% to 99.95%, from 93% to 99.95%, from 97% to 99.95% or 99-99.95%, or 99.95% or more. Oxidation iii with a suitable oxidizing reagent provides compounds of formula I. Suitable oxidizing agents include, for example, Dess-Martin or TEMPO periodinan and sodium hypochlorite.

The deuterated amino alcohol amides of formula ii shown in Scheme 1 can be prepared using known methods and, for example, as shown below in Scheme II.

In accordance with Scheme II, the conversion of the glycine imino ester iv to the deuterated sult of formula vii is carried out in accordance with the procedures described previously (Y. Elemes and U. Ragnarsson, J. Chem. Soc, Perkin I, 1996, 6, p. 537). Sequential treatment of compounds of formula vii with acid and base as previously described (L. Lankiewicz, et. Al., J. Chem. Soc, Perkin I, 1994, 17, p. 2503), followed by treatment of an intermediate amino acid (not shown) benzyloxycarbonyl chloride gives the protected deuterated amino acid viii . The interaction of viii with methoxymethylamine in the presence of a CDI condensing reagent gives Weinreb amide of formula ix . Reduction of compound vi , for example, with diisobutylaluminum hydride or lithium aluminum hydride gives aldehyde x . Using a procedure similar to that described previously (see, for example, WO 02/18369), aldehyde x is converted to cyanohydrin xi and then to protected hydroxy amino acid xii . Acid xii is converted to the protected amide xiii , which is deprotected to give aminoamide ii .

Alternatively, the deuterated aminoamide ii shown in Scheme I, where R ₂ is H, can be prepared, for example, as shown in Scheme III.

In accordance with Scheme III, propivyl alcohol xiv is reduced with sodium bis (2-methoxyethoxy) aluminum hydride followed by quenching of the reaction with deuterium oxide to obtain deuterated allyl alcohol xv . Oxidation of compound xv with manganese dioxide gives the aldehyde xvi , which is further oxidized to acid xvii with sodium chlorite (NaClO ₂ ) in the presence of sodium phosphate and 2-methyl-2-butene. The reaction of acid xvii with isobutyl chloroformate (ICBF) in the presence of N-methylmorpholine followed by the reaction of the intermediate mixed anhydride with amine HNR ⁴ R ⁵ gives the amide xviii . The epoxidation of compound xviii to give xix epoxide can be carried out using urea hydrogen peroxide (UHP) in the presence of trifluoroacetic acid and p-toluenesulfonic acid. The reaction of compound xix with sodium azide gives the intermediate azide compound xx , which is then reduced to give the racemic amino alcohol xxi by catalytic hydrogenation in the presence of palladium on carbon. The racemic amino alcohol xxi can be separated using known methods, such as chiral chromatography, the preparation of optically active derivatives, or the formation of salts with the optically active acid HA, followed by crystallization from an organic solvent. Suitable optically active organic acids for the preparation of salts include, for example, L-tartaric acid, L-malic acid, (S)-mandelic acid, (1S) - (+) - 10-camphorsulfonic acid, hydrate (-) 2.2: 4,6-di-O-isopropylidene-2-keto-L-gulonic acid, N-acetyl-L-leucine, deoxycholic acid, (+) - O, O'-dibenzoyl-D-tartaric acid, O, O ' -di- (4-toluoyl) -D-tartaric acid, S - (+) 1,1-binaphthyl-2-2-phosphoric acid, L-lactic acid, D-gluconic acid, lactobionic acid, dipivaloyl-L-tartaric acid, S - (+) - O-acetylmindal acid and S - (-) 2- (phenylcarbamoyloxy) propionic acid. Examples of suitable organic solvents for recrystallization include dimethylacetamide, ethyl acetate and acetone.

The deuterated compounds thus obtained can be identified by conventional analytical methods, for example, NMR and mass spectroscopy. NMR can be used to determine the structure of a compound, while mass spectroscopy can be used to determine the number of deuterium atoms in a compound compared to its non-deuterated form.

The deuterated compounds of the present invention are generally more stable and less susceptible to epimerization than their non-deuterated counterparts. Thus, they can be used for such purposes where a specific spatial configuration in the compounds of the present invention is desired. For example, the deuterated compounds of formula (I) can be used to treat or prevent HCV virus infection or other HCV protease-mediated conditions because they have the ability to inhibit HCV protease. Their inhibitory activity can be measured by conventional enzyme inhibition assays, some of which are described in the publications cited above. See, for example, Perni, R.B. et al., Antimicrobial Agents and Chemotherapy, 2006 (march), 50 (3): 899-909.

In addition, the deuterated compounds of formula (I) can be used as a biological tool for studying the pharmacological properties of their non-deuterated analogues. Accordingly, such applications are also included in the scope of the present invention.

Example 1. Obtaining (1S, 3aR, 6aS) -2 - ((S) -2 - ((S) -2-cyclohexyl-2- (pyrazine-2-carboxamido) acetamido) -3,3-dimethylbutanoyl) -N - ((S) -1- (cyclopropylamino) -1,2-dioxo-3-deutero-hexan-3-yl) octahydrocyclopenta [c] pyrrole-1-carboxamide

Stage a: Getting

Deuterated sultam (i.e., compound vi shown in the diagram below) was prepared by known methods, such as those described by Y. Elemes and U. Ragnarsson, J. of Chem. Soc., Perkin 1, 1996, 6, 537; W. Oppolzer, et al., Helv. Chim. Acta., 1994, 25: 2363, using the corresponding unsubstituted sultam and propyl iodide.

17.32 g of compound vi (45.8 mmol) and 229 ml of THF were then charged into a 500 ml round bottom flask with a magnetic stirrer and inlet for N ₂ . The resulting solution was cooled to −78 ° C. and n-BuLi (31.5 ml of a 1.6 M solution in hexane, 50.3 mmol) was added using a syringe pump over an hour. The resulting yellow solution was kept for 30 minutes, and then an HPMA solution (56 ml) and n-PrI (13.4 ml, 137 mmol) were added thereto over 30 minutes. The mixture was allowed to warm to room temperature over 8 hours and then cooled to -20 ° C before adding D ₂ O (50 ml) to the mixture. The reaction mixture was then extracted with EtOAc (400 ml) and the organic layer was dried over MgSO ₄ and concentrated to obtain 61.3 g of a crude oil. Chromatography on 500 g of silica gel eluting with a 2: 1 heptane / EtOAc mixture followed by concentration of the enriched fraction gave 20.35 g of a white solid. This white solid was then recrystallized from EtOH (210 ml) to obtain 15.39 g of compound vii as a white crystalline solid. Deuterium incorporation was 93%, as determined by ¹ H-NMR.

Stage b: Obtaining (S) -2- (benzyloxycarbonylamino) -2-deuteropentanoic acid, viii

Compound vii (15.39 g, 32.1 mmol) from step a was loaded into THF (100 ml) and 1N. HCl solution (50 ml). The resulting emulsion was stirred overnight at room temperature and then concentrated in vacuo to give a viscous oil. The resulting oil was then dissolved in THF (100 ml) and water (25 ml) and LiOH (3.08 g, 128 mmol) were added to the solution. The resulting solution was again stirred overnight at room temperature and then concentrated to remove THF. A turbid, light yellow emulsion remained. This emulsion was diluted with water (25 ml) and extracted with CH ₂ Cl ₂ (three times, each time 50 ml). The aqueous phase was diluted with THF (200 ml) and cooled to 0 ° C. with rapid stirring and CBZ-Cl (7.6 ml, 54 mmol) was added dropwise over 15 minutes. After stirring for one hour at 0 ° C, the THF solvent was removed in vacuo and the resulting residue was acidified by adding 1N HCl solution (50 ml). The solution was extracted with EtOAc (3 times, 100 ml each time) and the organic phase was dried over Na ₂ SO ₄ and concentrated to give an oil. The resulting residue was dissolved in EtOAc (25 ml) and heptane (150 ml), a seed crystal was added and stirred overnight at room temperature. The solids were collected on a frit, washed with heptane (30 ml) and dried in air to obtain 5.65 g (70%) of compound viii shown in the diagram above. Deuterium incorporation was 93%, as determined by ¹ H-NMR.

Step c: Preparation of (S) -benzyl 1- (methoxy (methyl) amino) -1-oxo-2-deuteropentan-2-ylcarbamate

To a flask containing 1.0 g of (S) -2- (benzyloxycarbonylamino) -2-deuteropentanoic acid (3.97 mmol) in 20 ml of dichloromethane, maintained at 0 ° C, was added 3.0 equiv. N-methylmorpholine (700 μl), 1.5 eq. N, O-dimethylhydroxylamine hydrochloride (581 mg) and 1.5 eq. EDCI (1.14 g). The reaction mixture was stirred overnight at a temperature of from 0 ° C to room temperature. The reaction mixture was then diluted in dichloromethane and washed with HCl (1 N) and brine. The organic layer was dried over MgSO ₄ . The crude mixture was purified by flash chromatography (ethyl acetate 15-75% in hexane) to give 814 mg of pure amide (the title compound). ES + = 296.1, ES- = 295.2. The structure was confirmed by ¹ H-NMR.

Stage d: Obtaining (S) -benzyl 1-oxo-2-deuteropentan-2-ylcarbamate

Using the procedure described in WO 02/18369, the Cbz-protected amino acid from step c was converted to the title compound. Specifically, in a flask containing 1.0 eq. (S) -benzyl 1- (methoxy (methyl) amino) -1-oxo-2-deuteropentan-2-ylcarbamate (810 mg, 2.75 mmol) in 10 ml of anhydrous THF, maintained at 0 ° C (on ice bath), 1.7 equiv. lithium borohydride solution (1.0M) (4.67 ml). After about 10 minutes, the ice bath was removed and the reaction was continued for one hour. The reaction was quenched at 0 ° C by adding 5 ml of KHSO ₄ solution (10%). The solution was then diluted by adding 10 ml of HCl (1 N.). The mixture was stirred for 30 minutes, then extracted 3 times with dichloromethane. The organic phases were combined and washed with HCl solution (1 N), water and brine. The organic phase was then dried over MgSO ₄ and the volatiles were evaporated. Aldehyde was used as such in the next step. ES + = 237.1, ES- = 235.2.

Step e: Preparation of Benzyl (3S) -1- (cyclopropylamino) -2-hydroxy-1-oxo-3-deuterohexan-3-ylcarbamate

Cyclopropylisonitrile was prepared according to the scheme below.

Cyclopropylisonitrile was then coupled with the aldehyde product of step d to afford the title compound as described by JE Semple et al., Org. Lett., 2000, 2 (18), p. 2769; Lumma W., J. Org. Chem., 1981, 46, 3668. "ES + = 322.1.

Step f: Preparation of (3S) -3-amino-N-cyclopropyl-3-deutero-2-hydroxyhexanamide

The hydrogenolysis of the Cbz compound from step e was carried out using a palladium-carbon catalyst in the presence of hydrogen to obtain the title compound. The diagrams below show stages c, d, e, and f .

Step g: Preparation of (1S, 3aR, 6aS) -2 - ((S) -2 - ((S) -2-cyclohexyl-2- (pyrazine-2-carboxamido) acetamido) -3,3-dimethylbutanoyl) -N - ((3S) -1- (cyclopropylamino) -3-deutero-2-hydroxy-1-oxohexan-3-yl) octahydrocyclopenta [c] pyrrole-1-carboxamide

The title compound was obtained from the hydroxyaminoamide product of step f by condensation with a suitable acid in the presence of a coupling reagent, such as, for example, EDCI and HOSu. Specifically, in a flask containing 1.2 eq. (1S, 3aR, 6aS) -2 - ((S) -2 - ((S) -2-cyclohexyl-2- (pyrazine-2-carboxamido) acetamido) -3,3-dimethylbutanoyl) octahydrocyclopenta [c] pyrrole- 1-carboxylic acid (1.59 g) in 20 ml of DMF, 2.5 equiv. diisopropylamine (980 μl), 1.2 eq. N-hydroxybenzotriazole hydrate (411 mg) and 1.3 eq. EDCI (558 mg). After 15 minutes stirring at room temperature, 1.0 eq. (3S) -3-amino-N-cyclopropyl-3-deutero-2-hydroxyhexanamide hydrochloride (500 mg). After another 24 hours, the reaction mixture was diluted in 400 ml of ethyl acetate. The organic phase of the mixture was washed with HCl (1 N), water, saturated sodium bicarbonate, brine, and then dried over MgSO ₄ . The crude product was purified by chromatography on silica (70-100% ethyl acetate in hexane) to give 1.31 g of the title compound as a white solid. ES + = 683.6, ES- = 682.2. The structure was confirmed by ¹ H-NMR.

Step h: Preparation of (1S, 3aR, 6aS) -2 - ((S) -2 - ((S) -2-cyclohexyl-2- (pyrazine-2-carboxamido) acetamido) -3,3-dimethylbutanoyl) -N - ((S) -1- (cyclopropylamino) -1,2-dioxo-3-deutero-hexan-3-yl) octahydrocyclopenta [c] pyrrole-1-carboxamide

The title compound was prepared by oxidizing the product of step g with a suitable oxidizing reagent such as Dess Martin periodinan or TEMPO and sodium hypochlorite. Specifically, in a flask containing 1.31 g of (1S, 3aR, 6aS) -2 - ((S) -2 - ((S) -2-cyclohexyl-2- (pyrazine-2-carboxamido) acetamido) -3, 3-dimethylbutanoyl) -N - ((3S) -1- (cyclopropylamino) -3-deutero-2-hydroxy-1-oxohexan-3-yl) octahydrocyclopenta [c] pyrrole-1-carboxamide in 40 ml of dichloromethane, was added at room temperature 1.06 g of periodinan Dess Martin. After 2 hours of stirring, 50 ml of sodium bisulfite (1 N) was added and the mixture was stirred for 30 minutes. The 2 phases were separated, the organic phase was washed twice with water, brine and dried over Na ₂ SO ₄ . The crude product was purified by chromatography on silica (ethyl acetate 20-100% in hexane) to give 1.07 g of the title compound as a white solid. ES + = 681.5, ES- = 680.0. The structure was confirmed by ¹ H-NMR.

The inclusion of deuterium, determined by the MS method, was 93%. The diastereoisomeric ratio was determined by normal phase chiral HPLC, and it amounted to more than 99% di (diastereoisomeric excess).

The diagram below shows the reactions of steps g and h .

Example 2. Obtaining the hydrochloride of (2S, 3S) -3-amino-3-deutero-N-cyclopropyl-2-hydroxyhexanamide

The above diagram illustrates the general synthesis of the title compound. Each stage is described in detail below.

Stage 1 : Obtaining 3-deutero (E) -hex-2-en-1-ol

2-hexin-1-ol (10 g, 0.1 mol) and THF (100 ml, 10 vol.) Were charged into a 250 ml three-necked round bottom flask equipped with a mechanical stirrer and reflux condenser. The resulting mixture was cooled to 0 ± 5 ° C and then Red-Al (65% in toluene, 32 ml, 1.6 eq.) Was slowly added in a nitrogen atmosphere at a temperature in the range from 0 ° C to 20 ° C. The resulting mixture was allowed to warm to 25 ° C and stirred for 5 hours. The reaction mixture was cooled to -5 ± 5 ° C and D ₂ O (8.2 g, 4 equiv.) Was added dropwise at a temperature in the range from 0 ° C to 15 ° C. To the resulting mixture were added IPAC (50 ml, 5 vol.) And a saturated solution of NH ₄ Cl (50 ml, 5 vol.). After stirring the mixture for 10 minutes, the resulting white solid was filtered off. The organic layer was separated from the filtrate and the aqueous layer was extracted with IPAC (30 ml, 3 vol). The organic layers were combined and washed with water (30 ml, 3 vol.), Dried over MgSO ₄ and concentrated to obtain 9.8 g of the product (compound 2 ) as a colorless oil. The crude product 2 was used in the next step without further purification.

¹ H-NMR (500 MHz, CDCl ₃ ) δ 5.66 (t, 1H, J = 5.0 Hz), 4.12 (d, 2H, J = 5.0 Hz), 2.04 (t, 2H, J = 5.0 Hz), 1.38 ~ 1.46 (m, 2H), 0.93 (t, 3H, J = 5.0 Hz)

Stage 2 : Obtaining 3-deutero (E) -hex-2-enal

Activated MnO ₂ (87 g) was charged into a 250 ml three-necked round bottom flask equipped with a mechanical stirrer containing 3-deutero-2-hexenol (10 g, 0.1 mol) in CH ₂ Cl ₂ (150 ml, 15 vol.) , 10 equiv.) At room temperature. After vigorous stirring for 1 hour, another portion of MnO ₂ (16 g, 2 equiv.) Was added and shaking continued for 4 hours. The reaction solution was filtered through a pad of celite. The solvent was removed in vacuo (25 ° C., 100 mmHg) to give 8.8 g of a crude aldehyde product (compound 3 ) as a pale yellowish oil. The crude product was used in the next step without further purification.

¹ H-NMR (500 MHz, CDCl ₃ ) δ 9.54 (d, 1H, J = 10.0 Hz), 6.14 (s, 1H), 2.34 (m, 2H), 1.55 ~ 1.60 (m, 2H), 1.00 (t, 3H, J = 5.0 Hz)

Stage 3 : Obtaining 3-deutero (E) -hex-2-enoic acid

In a 500 ml three-necked round-bottom flask equipped with a mechanical stirrer and reflux condenser, 3-deutero-2-hexene-1-al (10 g, 0.1 mol), tert-BuOH (90 ml, 9 vol.) And 2 were charged methyl-2-butene (30 ml, 3 vol.). To the resulting solution was added a freshly prepared aqueous solution of NaClO ₂ (27.4 g, 3 equiv.) And NaH ₂ PO ₄ (62.9 g, 4 equiv.) In water (200 ml) over 30 minutes. The reaction mixture was stirred at room temperature for 2 hours. The reaction solution was cooled to 0 ° C. and a saturated aqueous solution of Na ₂ SO _{3 was} added thereto until the reaction mixture became colorless. Stirring was stopped, the organic layer was separated, and the aqueous layer was extracted with EtOAc (3 vol. × 3). The organic layers were combined and concentrated in vacuo until a total volume of 3 vol. The resulting solution was extracted with 1 N. NaOH solution (3 vol. × 3) and the remaining organic layer was drained. The combined aqueous solution was acidified with 6 N. HCl solution until the pH reaches 1.0. The solution was extracted with CH ₂ Cl ₂ (3 vol. × 5). The combined organic layers were dried over MgSO ₄ and concentrated to give 8.7 g of the product (compound 4 ) as a white solid.

¹ H-NMR (500 MHz, CDCl ₃ ) δ 5.84 (s, 1H), 2.23 (t, 2H, J = 5.0 Hz), 1.51 ~ 1.55 (m, 2H), 0.98 (t, 3H, J = 5.0 Hz)

Stage 4 : Obtaining 3-deutero (E) -N-cyclopropylhex-2-enamide

In a 250 ml three-neck round-bottom flask equipped with a mechanical stirrer and reflux condenser, 2-hexenoic acid-3d (10 g, 0.09 mol), IBCF (13 g, 1.1 eq.) In CH ₂ Cl ₂ (100 ml, 10 vol.). The resulting solution was cooled to 0 ° C and NMM (13.2 g, 1.5 equiv) was slowly added, while controlling the temperature in the range from 0 to 20 ° C. Then the mixture was allowed to warm to room temperature and was stirred for 1 hour. Cyclopropylamine (5.9 g, 1.2 equiv.) Was added to the resulting solution, and the solution was stirred for 2 hours. The reaction mixture was washed with 1 N. NaOH solution (3 vol. × 2), 1 N. HCl solution (3 vol. × 2) and saturated salt solution (3 vol.) and water (3 vol.). The organic layer was dried over MgSO ₄ and concentrated to give the crude product as an oil. The crude product was dissolved with heptane (5 vol.) And cooled to -78 ° C with stirring. The precipitated solid was filtered and dried to give 8.7 g of product (compound 5 ) as a white solid.

¹ H-NMR (500 MHz, DMSO) δ 7.92 (s, 1H), 5.78 (s, 1H), 2.66 ~ 2.68 (m, 1H), 2.08 (t, 2H, J = 5.0 Hz), 1.38 ~ 1.42 (m, 2H), 0.87 (t, 3H, J = 5.0 Hz), 0.63 (t, 2H, J = 3.0 Hz), 0.40 (t, 2H, J = 3.0 Hz)

Step 5 : Preparation of 3-Deutero-N-cyclopropyl-3-propyloxyran-2-carboxamide

Into a 250 ml three-neck round-bottom flask equipped with a mechanical stirrer and containing (E) -N-cyclopropylhex-2-enamide-3d (i.e. the product from step 4 ) (10 g, 0.06 mol), urea-hydrogen peroxide (25 g, 4 equiv.) And p-TsOH (12.3 g, 1 equiv.) In CH ₂ Cl ₂ (100 ml, 10 vol.), At 0 ° C was added trifluoroacetic anhydride (40.9 g, 3 equiv.) in CH ₂ Cl ₂ (50 ml, 5 vol.) for 30 minutes. The reaction mixture was heated to a temperature of 40 ± 5 ° C and stirred for 3 hours. After cooling to 0 ° C, the reaction mixture was quenched by slow addition of 6 N. NaOH solution (100 ml, 10 vol.) and was stirred for 30 minutes. The organic layer was separated and washed with brine (5 vol.) And water (5 vol.). The washed organic layer was dried over MgSO ₄ and the solvent was evaporated to give 9.7 g of an epoxy product (i.e., compound 6) as a pale yellow oil. The crude product was used in the next step without further purification.

¹ H-NMR (500 MHz, DMSO) δ 8.01 (s, 1H), 3.09 (s, 1H), 2.63 ~ 2.65 (m, 1H), 1.39 ~ 1.54 ( m, 4H), 0.91 (t, 3H, J = 5.0 Hz), 0.60 (t, 2H, J = 3.0 Hz), 0.45 (t, 2H, J = 3.0 Hz)

Step 6 : Preparation of 3-azido-3-deutero-N-cyclopropyl-2-hydroxyhexanamide

Into a 250 ml three-neck round-bottom flask equipped with a mechanical stirrer and reflux condenser containing epoxide-3d 6 (10 g, 0.06 mol) and anhydrous magnesium sulfate (14.1 g, 2.0 equiv.) In MeOH (100 ml 10 vol.), Sodium azide (15.3 g, 4.0 eq.) Was added in one portion. The resulting mixture was heated to a temperature of 65 ± 5 ° C and stirred for 5 hours. The reaction mixture was cooled to room temperature, IPAC (100 ml, 10 vol) was added and the mixture was stirred for another 10 minutes. The mixture was filtered through a pad of Celite® to remove insoluble salts and the resulting clear solution was concentrated to 3 vol. IPAC (170 ml, 17 vol) was added to the resulting solution, and the mixture was stirred for another 10 minutes. Again, the solution was filtered through a pad of Celite® to give the product, azide-3d (compound 7 ), as a clear solution in IPAC (about 200 ml), which was used in the next step without further purification.

¹ H-NMR (500 MHz, DMSO) δ 7.91 (s, 1H), 6.00 (d, 1H, J = 5.0 Hz), 4.03 (d, 1H, J = 5.0 Hz ), 2.66 ~ 2.67 (m, 1H), 1.30 ~ 1.58 (m, 4H), 0.88 (t, 3H, J = 5.0 Hz), 0.60 (t, 2H, J = 3.0 Hz), 0.48 (t, 2H, J = 3.0 Hz)

Step 7 : Preparation of 3-amino-3-deutero-N-cyclopropyl-2-hydroxyhexanamide

Pd / C (10% Pd, water content 50) was charged into a 500 ml hydrogenation reactor, which was an autoclave equipped with a mechanical stirrer, containing azide-3d 7 (200 ml, 0.05 mol) in the IPAC obtained in the previous step. %, 0.8 g). Nitrogen (1.0 atm) was introduced into the solution and then released three times, then hydrogen (3.0 atm) was introduced and released three times. Hydrogen (3 atm) was introduced into the resulting solution and stirred for 5 hours. After the release of hydrogen gas, the solution was purged with nitrogen for 5 minutes. MeOH (30 ml, 3 vol) was added to the resulting solution, and the reaction mixture was heated to a temperature of 50 ± 5 ° C. The reaction mixture was filtered through a pad of celite to give a clear solution. The product was isolated by concentrating the solution at a temperature of 20 ± 5 ° C until 3 vol. solution. The solid was collected by filtration, washed (IPAC, 3 vol) and dried to give 7.7 g of the title compound (compound 8 ) as a white crystalline solid.

¹ H-NMR (500 MHz, DMSO) δ 7.70 (s, 1H), 5.31 (s, 2H), 3.68 (s, 1H), 2.64 ~ 2.66 (m, 1H) , 1.10 ~ 1.50 (m, 4H), 0.82 (t, 3H, J = 5.0 Hz), 0.59 (t, 3H, J = 3.0 Hz), 0.45 ( t, 3H, J = 3.0 Hz)

Step 8 : Preparation of (2S, 3S) -3-amino-3-deutero-N-cyclopropyl-2-hydroxyhexanamide deoxycholate

Deoxycholic acid (15.7 g, 0.75 equiv.) Was charged into a 250 ml three-neck round-bottom flask equipped with a mechanical stirrer and containing racemic (2S, 3S) -3-amino-3-deutero-N-cyclopropyl-2-hydroxyhexanamide from stage 7 (10 g, 0.05 mol) in THF (100 ml, 10 vol.). The reaction mixture was heated to 65 ± 5 ° C and stirred for 1 hour. The resulting homogeneous mixture was cooled to a temperature of 23 ± 2 ° C for 1 hour and left at a temperature in the indicated limits for 1 hour. Precipitated solids were collected by filtration, washed with THF (50 ml, 5 vol.) And dried to obtain 12.4 g of the salt compound (compound 9 ) as a white solid. The product had an enantiomeric ratio (ER) of 2:98.

Step 9 : Preparation of (2S, 3S) -3-amino-3-deutero-N-cyclopropyl-2-hydroxyhexanamide hydrochloride

A dihydrocholate salt (from step 8 ) and 2-propanol (62 ml, 5 vol) were charged into a 250 ml three-necked round-bottomed flask equipped with a mechanical stirrer. The solution was heated to a temperature of 75 ± 5 ° C and 5-6 N was slowly added. a solution of HCl in IPA (12 ml, 3 equiv.) with vigorous stirring. The resulting solution was stirred at the indicated temperature for 1 hour and then cooled to a temperature of 23 ± 2 ° C. The reaction mixture was maintained at this temperature for 1 hour. Precipitated solids were collected by filtration, washed with 2-propanol (36 ml, 3 vol), dried to give 3.0 g of the title compound (enantiomeric ratio = 0: 100) as a white solid. Deuterium incorporation was more than 99% as determined by MS and ¹ H-NMR.

¹ H-NMR (500 MHz, DMSO) δ 8.07 (s, 1H), 7.97 (s, 3H), 6.25 (d, 1H, J = 5.0 Hz), 4.16 (d , 1H, J = 5.0 Hz), 2.67 ~ 2.70 (m, 1H), 1.33 ~ 1.46 (m, 4H), 0.84 (t, 3H, J = 5.0 Hz), 0.61 (t, 3H, J = 3.0 Hz), 0.53 (t, 3H, J = 3.0 Hz).

Example 3. Analysis of the measurement of the rate of epimerization

The deuterated compounds of the present invention undergo slow epimerization as follows:

The rate of epimerization was measured in accordance with the following analysis. Specifically, 100 μl of medium (buffer, rat plasma, dog plasma or human plasma) was added to a 96-well deep plate. Then, 10 μl of an acetonitrile solution containing the test compound (1S, 3aR, 6aS) -2 - ((S) -2 - ((S) -2-cyclohexyl-2- (pyrazine-2-carboxamido) acetamido) - 3,3-dimethylbutanoyl) -N - ((S) -1- (cyclopropylamino) -1,2-dioxo-3-deutero-hexan-3-yl) octahydrocyclopenta [c] pyrrole-1-carboxamide (at a concentration of 1 μM or 10 μM) and 1200 μl of ethyl acetate in a 96-well deep-well plate (2 ml) using a TomTec Fluid Unit (Hamden, CT, USA). The tablet was then tightly closed and shaken to swirl with stirring for 20 minutes, and then centrifuged at 3000 rpm for 10 minutes. After centrifugation, 900 μl of the supernatant was transferred to a new 96-well plate with V-shaped deep wells using TomTec, and then dried in a nitrogen gas atmosphere (flow rate 60 l / min) at 25 ° C for about 30 minutes. The resulting residue was restructured with 100 μl of ethyl acetate and the solution was again transferred to glass inserts in a 96-well plate. 20 μl of the restructured solution was introduced into the installation for analysis by LC-MS / MS to determine the number of epimers. For the LC-MS / MS spectrometer, a ChiralPak AD column (4.6 × 150 mm, 10 μm), a mixture of isopropanol and n-heptane (10:90, 50:50 or 90:10) as the mobile phase and isopropanol as solvent for washing. Also, a deuterated analog of the test compound containing 11 deuterium atoms in the cyclohexyl group (C ₃₆ H ₄₂ D ₁₁ N ₇ O ₆ , MM 690.47) was used in a MS spectrometer.

The test compound had a mass (M + H, m / z) 681.36, while its non-deuterated analogues (with the same or different chiral configurations at the deuterated carbon center) had a mass (M + H, m / z) 680.36. Their LC-MS / MS spectra showed a fragment of 323.30 (with deuterium) and 322.30 (non-deuterated).

At both concentrations (1 μM and 10 μM) and in the same medium (i.e., buffer, rat plasma, dog plasma and human plasma), the tested deuterated compound of formula (I) showed a slower rate of epimerization than its non-deuterated form in the buffer rat plasma and dog plasma; and a much slower rate of epimerization in human plasma. For example, in human plasma and at 1 μM or 10 μM, the deuterated compound underwent epimerization by approximately 30% for 180 minutes, while the non-deuterated form underwent epimerization by almost 40%. In addition, in human plasma, the deuterated compound underwent epimerization at a linear rate for 180 minutes, while the non-deuterated form showed an exponential rate of epimerization during the first 60 minutes, after which the rate was equalized.

Example 4. Analysis of the definition of IR _fifty  in cells with HCV replicon

This analysis used (1S, 3aR, 6aS) -2 - ((S) -2 - ((S) -2-cyclohexyl-2- (pyrazine-2-carboxamido) acetamido) -3,3-dimethylbutanoyl) -N - ((S) -1- (cyclopropylamino) -1,2-dioxo-3-deuterohexan-3-yl) octahydrocyclopenta [c] pyrrole-1-carboxamide and (5S, 8S) -3- (5-chloro-2 , 4-dimethoxyphenyl) -7 - ((S) -2- (2-cyclohexylacetamido) -3,3-dimethylbutanoyl) -N - ((S) -1- (cyclopropylamino) -1,2-dioxo-3-deuterohexane -3-yl) -1-oxa-2,7-diazaspiro [4.4] non-2-en-8-carboxamide as described in Lin, C. et al., J. Biol. Chem., 2004, 279: 17508-17514; Lin, K. et al., Antimicrob. Agents Chemother., 2004, 48: 4784-4792.

Huh-7 cells, including an autonomously replicating, subgenomic replicon of HCV Con1 strain, were maintained in Dulbecco's Modified Eagle Medium (DMEM) containing 10% thermally inactivated fetal bovine serum (FBS), 2 mM L-glutamine and non-essential amino acids (JRH Biosciences, Lenexa, KS), plus 0.25 mg / ml G418 (Invitrogen, Carlsbad, CA). The HCV subgenomic replicon also encodes neomycin phosphotransferase, which allows selective growth of HCV replicon-containing Huh-7 cells compared to HCV replicon-negative Huh-7 cells in the presence of G418. The concentrations of the test compound, in which the level of HCV RNA in cells with replicon decreases by 50% (IC ₅₀ ) or 90% (IC ₉₀ ) or cell viability decreases by 50% (CC ₅₀ ), were determined in cells with HCV Conl subgenomic replicon ( 19) using a program for constructing a curve with 4 parameters (SoftMax Pro). Replicon-containing cells were incubated with a test compound diluted in DMEM containing 2% FBS and 0.5% DMSO (without G418) at 37 ° C. Total cellular RNA was extracted using the RNeasy-96 kit (QIAGEN, Valencia, CA) and the number of copies of HCV RNA was determined by quantitative, real-time, multiple reverse transcription-PCR analysis (QRT-PCR or Taqman). The cytotoxicity of compounds in cells with HCV replicon was measured under the same experimental conditions using a tetrazolium cell viability assay.

The results show that both test compounds had a Ki of less than 50 nM and an IR ₅₀ (over 5 days) of less than 10.0 μM.

Other options for implementation

It should be clear that although the present invention has been disclosed in connection with its detailed description, the above description of the invention is intended to illustrate it and does not limit the scope of the present invention, which is determined by the scope of the attached claims. Other aspects, advantages and modifications are covered by the following claims.

Claims (32)

1. Enriched in deuterium α-ketoamide compound of the formula (1)

where D is a deuterium atom;
R ¹ represents

Where

represents C ₄ -C ₈ , monocyclic azaheterocyclyl, or C ₃ -C ₁₀ polycyclic azaheterocyclyl, or C ₃ -C ₁₀ polycyclic azaheterocyclenyl, where unsaturation is present in the ring distal to the ring containing the group R ²¹ , and to which is attached - C (O) -N (R ² ) -CDR ³ -C (O) -C (O) -NR ⁴ R ⁵ , wherein each azaheterocyclyl or azaheterocyclenyl is optionally substituted with phenyl, optionally substituted with one or more substituents selected from alkoxy and halogen ;
R ²¹ represents

Where
each R ⁶ and R ⁸ is independently C ₁ -C ₄ alkyl substituted with C ₁ -C ₄ alkyl or C ₃ -C ₁₀ cycloalkyl;
each R ⁷ , R ⁹ and R ¹¹ is independently hydrogen or C ₁ -C ₄ alkyl;
R ¹⁰ represents pyrazine or C ₁ -C ₄ alkyl optionally substituted with C ₃ -C ₁₀ cycloalkyl;
L represents —C (O) - or —NR ¹¹ C (O) -;
n is 0 or 1; and
each of R ² , R ³ and R ⁴ , independently, is H or C _1-6 alkyl; and
R ⁵ represents H or C ₃ -C ₁₀ cycloalkyl,
where the “hetero” atoms in the heteroaryl or heterocycloaliphatic groups are N or O,
and connection:

2. The compound according to claim 1, where n is 1. 3. The compound according to claim 1, where R ⁶ is methylene substituted with isobutyl. 4. The compound according to claim 1, where R ⁷ represents hydrogen. 5. The compound according to claim 1, where R ⁸ represents methylene substituted by C ₃ -C ₁₀ cycloalkyl. 6. The compound according to claim 5, where R ⁸ represents methylene substituted by cyclohexyl. 7. The compound according to claim 1, where R ⁹ represents hydrogen. 8. The compound according to claim 1, where L is —C (O) -. 9. The compound according to claim 1, where R ¹⁰ represents pyrazine. 10. The compound according to claim 9, where R ¹⁰ represents 2-pyrazine. 11. The compound according to claim 1, where

represents C ₄ -C ₈ monocyclic azaheterocyclyl. 12. The compound according to claim 1, where

represents C ₆ -C ₁₀ polycyclic azaheterocyclyl or C ₆ -C ₁₀ polycyclic azaheterocyclyl, optionally substituted with phenyl, optionally substituted with one or more substituents selected from alkoxy and halogen. 13. The compound according to item 12, where

represents

14. The compound according to item 13, where

represents

15. The compound according to claim 1, where R ² represents hydrogen, R ⁴ represents hydrogen, and R ⁵ independently represents hydrogen or cyclopropyl. 16. The compound according to claim 1, where R ³ represents propyl. 17. The compound according to claim 1, where n is 0. 18. The compound according to claim 1, where L is —NR ¹¹ C (O) - and R ¹¹ is hydrogen. 19. The compound of claim 1, wherein R ¹⁰ is C ₁ -C ₄ alkyl optionally substituted with C ₃ -C ₁₀ cycloalkyl. 20. The compound according to claim 19, where R ¹⁰ represents tert-butyl. 21. The compound according to claim 1, where the compound is a

22. The compound according to item 12, where

represents

Where
A represents —CH ₂ -;
B is —CH ₂ -;
each of Y ¹ and Y ² represents hydrogen;
R ²² represents phenyl optionally substituted with one or more substituents selected from alkoxy and halogen. 23. The compound of claim 22, wherein R ²² is phenyl substituted with one or more substituents selected from alkoxy and halogen. 24. The compound according to item 22, where

represents

25. The compound according to paragraph 24, where the compound has the structure:

26. The compound according to claim 1, where the enrichment of deuterium in the compound is at least 50%. 27. The compound of claim 26, wherein the deuterium enrichment is at least 80%. 28. The compound of claim 27, wherein the deuterium enrichment is at least 90%. 29. The compound of claim 28, wherein the deuterium enrichment is at least 99%. 30. A pharmaceutical composition having anti-HCV activity, comprising a pharmaceutically acceptable carrier and a compound according to any one of claims 1, 21, 25, 28 or 29. 31. A method of increasing the bioavailability of a compound, comprising replacing a hydrogen atom that is bonded to a steric carbon atom in the compound, a deuterium atom, wherein the resulting deuterated compound is a compound according to any one of claims 1, 21, 25, 28 or 29. 32. A method of inhibiting HCV protease, comprising contacting the HCV protease with a compound according to any one of claims 1, 21, 25, 28, or 29.