# Patent

## Title
Hepatitis C virus inhibitors

## Abstract

The present invention discloses compounds of Formula (I), or pharmaceutically acceptable salts, esters, or prodrugs thereof:




which inhibit RNA-containing virus, particularly the hepatitis C virus (HCV). Consequently, the compounds of the present invention interfere with the life cycle of the hepatitis C virus and are also useful as antiviral agents. The present invention further relates to pharmaceutical compositions comprising the aforementioned compounds for administration to a subject suffering from HCV infection. The invention also relates to methods of treating an HCV infection in a subject by administering a pharmaceutical composition comprising the compounds of the present invention.

## Background
RELATED APPLICATION
This application claims the benefit of U.S. Provisional Application No. 61/286,178 filed Dec. 14, 2009. The entire teachings of the above application are incorporated herein by reference.


TECHNICAL FIELD
The present invention relates to novel antiviral agents. More specifically, the present invention relates to compounds which can inhibit the function of the NS5A protein encoded by Hepatitis C virus (HCV), compositions comprising such compounds, methods for inhibiting HCV viral replication, methods for treating or preventing HCV infection, and processes for making the compounds.
BACKGROUND OF THE INVENTION
Infection with HCV is a major cause of human liver disease throughout the world. In the US, an estimated 4.5 million Americans are chronically infected with HCV. Although only 30% of acute infections are symptomatic, greater than 85% of infected individuals develop chronic, persistent infection. Treatment costs for HCV infection have been estimated at $5.46 billion for the US in 1997. Worldwide over 200 million people are estimated to be infected chronically. HCV infection is responsible for 40-60% of all chronic liver disease and 30% of all liver transplants. Chronic HCV infection accounts for 30% of all cirrhosis, end-stage liver disease, and liver cancer in the U.S. The CDC estimates that the number of deaths due to HCV will minimally increase to 38,000/year by the year 2010.
Due to the high degree of variability in the viral surface antigens, existence of multiple viral genotypes, and demonstrated specificity of immunity, the development of a successful vaccine in the near future is unlikely. Alpha-interferon (alone or in combination with ribavirin) has been widely used since its approval for treatment of chronic HCV infection. However, adverse side effects are commonly associated with this treatment: flu-like symptoms, leukopenia, thrombocytopenia, depression from interferon, as well as anemia induced by ribavirin (Lindsay, K. L. (1997) Hepatology 26 (suppl 1): 71S-77S). This therapy remains less effective against infections caused by HCV genotype 1 (which constitutes  ̃75% of all HCV infections in the developed markets) compared to infections caused by the other 5 major HCV genotypes. Unfortunately, only  ̃50-80% of the patients respond to this treatment (measured by a reduction in serum HCV RNA levels and normalization of liver enzymes) and, of responders, 50-70% relapse within 6 months of cessation of treatment. Recently, with the introduction of pegylated interferon (Peg-IFN), both initial and sustained response rates have improved substantially, and combination treatment of Peg-IFN with ribavirin constitutes the gold standard for therapy. However, the side effects associated with combination therapy and the impaired response in patients with genotype 1 present opportunities for improvement in the management of this disease.
First identified by molecular cloning in 1989 (Choo, Q-L et al (1989) Science 244:359-362), HCV is now widely accepted as the most common causative agent of post-transfusion non-A, non-B hepatitis (NANBH) (Kuo, G et al (1989) Science 244:362-364). Due to its genome structure and sequence homology, this virus was assigned as a new genus in the Flaviviridae family. Like the other members of the Flaviviridae, such as flaviviruses (e.g. yellow fever virus and Dengue virus types 1-4) and pestiviruses (e.g. bovine viral diarrhea virus, border disease virus, and classic swine fever virus) (Choo, Q-L et al (1989) Science 244:359-362; Miller, R. H. and R. H. Purcell (1990) Proc. Natl. Acad. Sci. USA 87:2057-2061), HCV is an enveloped virus containing a single strand RNA molecule of positive polarity. The HCV genome is approximately 9.6 kilobases (kb) with a long, highly conserved, noncapped 5′ nontranslated region (NTR) of approximately 340 bases which functions as an internal ribosome entry site (IRES) (Wang C Y et al ‘An RNA pseudoknot is an essential structural element of the internal ribosome entry site located within the hepatitis C virus 5′ noncoding region’ RNA—A Publication of the RNA Society. 1(5): 526-537, 1995 July). This element is followed by a region which encodes a single long open reading frame (ORF) encoding a polypeptide of  ̃3000 amino acids comprising both the structural and nonstructural viral proteins.
Upon entry into the cytoplasm of the cell, this RNA is directly translated into a polypeptide of  ̃3000 amino acids comprising both the structural and nonstructural viral proteins. This large polypeptide is subsequently processed into the individual structural and nonstructural proteins by a combination of host and virally-encoded proteinases (Rice, C. M. (1996) in B. N. Fields, D. M. Knipe and P. M. Howley (eds) Virology 2nd Edition, p 931-960; Raven Press, N.Y.). There are three structural proteins, C, E1 and E2. The P7 protein is of unknown function and is comprised of a highly variable sequence. There are several non-structural proteins. NS2 is a zinc-dependent metalloproteinase that functions in conjunction with a portion of the NS3 protein. NS3 incorporates two catalytic functions (separate from its association with NS2): a serine protease at the N-terminal end, which requires NS4A as a cofactor, and an ATP-ase-dependent helicase function at the carboxyl terminus. NS4A is a tightly associated but non-covalent cofactor of the serine protease. NS5A is a membrane-anchored phosphoprotein that is observed in basally phosphorylated (56 kDa) and hyperphosphorylated (58 kDa) forms. While its function has not fully been elucidated, NS5A is believed to be important in viral replication. The NS5B protein (591 amino acids, 65 kDa) of HCV (Behrens, S. E. et al (1996) EMBO J. 151 2-22), encodes an RNA-dependent RNA polymerase (RdRp) activity and contains canonical motifs present in other RNA viral polymerases. The NS5B protein is fairly well conserved both intra-typically ( ̃95-98% amino acid (aa) identity across 1b isolates) and inter-typically ( ̃85% aa identity between genotype 1a and 1b isolates). The essentiality of the HCV NS5B RdRp activity for the generation of infectious progeny virions has been formally proven in chimpanzees (A. A. Kolykhalov et al. (2000) Journal of Virology, 74(4): 2046-2051). Thus, inhibition of NS5B RdRp activity (inhibition of RNA replication) is predicted to be useful to treat HCV infection.
Following the termination codon at the end of the long ORF, there is a 3′ NTR which roughly consists of three regions: an  ̃40 base region which is poorly conserved among various genotypes, a variable length poly(U)/polypyrimidine tract, and a highly conserved 98 base element also called the “3′ X-tail” (Kolykhalov, A. et al (1996) J. Virology 70:3363-3371; Tanaka, T. et al (1995) Biochem Biophys. Res. Commun. 215744-749; Tanaka, T. et al (1996) J. Virology 70:3307-3312; Yamada, N. et al (1996) Virology 223:255-261). The 3′ NTR is predicted to form a stable secondary structure which is essential for HCV growth in chimps and is believed to function in the initiation and regulation of viral RNA replication.
Compounds useful for treating HCV-infected patients are desired which selectively inhibit HCV viral replication. In particular, compounds which are effective to inhibit the function of the NS5A protein are desired. The HCV NS5A protein is described, for example, in Tan, S.-L., Katzel, M. G. Virology 2001, 284, 1; and in Rice, C. M. Nature 2005, 435, 374.
Based on the foregoing, there exists a significant need to identify compounds with the ability to inhibit HCV.
SUMMARY OF THE INVENTION
The present invention relates to novel antiviral compounds represented herein below, pharmaceutical compositions comprising such compounds, and methods for the treatment or prophylaxis of viral (particularly HCV) infection in a subject in need of such therapy with said compounds. Compounds of the present invention interfere with the life cycle of the hepatitis C virus and are also useful as antiviral agents.
In its principal aspect, the present invention provides a compound of Formula (I):




or a pharmaceutically acceptable salt thereof, wherein:

Ring A and Ring B are each independently absent or a monocyclic or polycyclic group independently selected from aryl, heteroaryl, heterocyclic, C3-C8 cycloalkyl, and C3-C8 cycloalkenyl, each optionally substituted; preferably, optionally substituted aryl or optionally substituted heteroaryl;
L is absent or selected from the group consisting of optionally substituted C1-C4 alkyl, optionally substituted C2-C4 alkenyl, and optionally substituted C2-C4 alkynyl;
wherein at least one of Ring A, Ring B and L is present;
G and J are each independently an optionally substituted 5-membered heteroaryl or optionally substituted 5/6-membered fused heteroaryl, wherein the 5-membered heteroaryl contains one or more nitrogen atoms, and wherein the 6-membered ring of said 5/6-membered fused heteroaryl is attached to one of Ring A, L and Ring B and is aryl or heteroaryl; preferably, optionally substituted imidazolyl, optionally substituted benzimidazolyl or optionally substituted imidazopyridyl;
X is independently N—OR1, N—N(R1)2, or C(R2)2; preferably, CH2, CF2 or N-OMe;
R1 at each occurrence is independently hydrogen or optionally substituted C1-C4 alkyl;
R2 at each occurrence is independently hydrogen, halogen, optionally substituted C1-C4 alkyl, optionally substituted aryl, or optionally substituted heteroaryl; alternatively, two R2 groups can be taken together with the carbon atom to which they are attached to form an optionally substituted C3-C8 cycloalkyl or optionally substituted heterocyclic ring;
R6 at each occurrence is independently selected from the group consisting of optionally substituted O(C1-C8 alkyl), optionally substituted amino, C1-C8 alkyl, C2-C8 alkenyl, C2-C8 alkynyl, C3-C8 cycloalkyl, C3-C8 cycloalkenyl, heterocyclic, aryl, and heteroaryl, each optionally substituted; preferably, optionally substituted C1-C8 alkyl; more preferably, C1-C8 alkyl optionally substituted with amino, hydroxy, protected amino or O(C1-C4 alkyl);
Q is selected from:




R3 and R4 are each independently selected from the group consisting of hydrogen, optionally substituted C1-C8 alkyl, optionally substituted C2-C8 alkenyl, and optionally substituted C3-C8 cycloalkyl; preferably, hydrogen or optionally substituted C1-C4 alkyl; alternatively, R3 and R4 can be taken together with the carbon atom to which they are attached to form optionally substituted C3-C8 cycloalkyl or optionally substituted heterocyclic;
R5 is independently hydrogen, optionally substituted C1-C8 alkyl, or optionally substituted C3-C8 cycloalkyl; preferably, hydrogen or optionally substituted C1-C4 alkyl;
U is absent or independently selected from O, S, S(O), SO2, NC(O)—(C1-C4 alkyl), C(O), protected carbonyl, OCH2, OCH2CH2, SCH2, SCH2CH2, C(R7)2, C(R7)2C(R7)2, or C═C(R2)2; in an additional embodiment, U is selected from O, S, S(O), SO2, NC(O)—(C1-C4 alkyl), C(O), protected carbonyl, OCH2, OCH2CH2, SCH2, SCH2CH2, C(R7)2, C(R7)2C(R7)2, C═C(R2)2, C═N—O(R1) and C═N—N(R1)2; in yet an additional embodiment, U is selected from O, S, S(O), SO2, NC(O)—(C1-C4 alkyl), C(O), OCH2, OCH2CH2, SCH2, SCH2CH2, C(R7)2, C(R7)2C(R7)2, C═C(R2)2, C═N—O(R1) and C═N—N(R1)2; preferably, U is CH2, C═N—OR1, or C═C(R2)2;
In one embodiment, R7 at each occurrence is independently selected from the group consisting of hydrogen, halogen, cyano, hydroxy, O(C1-C4 alkyl), S(C1-C4 alkyl), amino optionally substituted with one or two C1-C4 alkyl, optionally substituted aryl, optionally substituted heteroaryl, and optionally substituted C1-C4 alkyl; preferably hydrogen, halogen, hydroxyl; in another embodiment, R7 at each occurrence is independently selected from the group consisting of hydrogen, halogen, cyano, hydroxy, O(C1-C4 alkyl), S(C1-C4 alkyl), amino optionally substituted with one or two C1-C4 alkyl, optionally substituted aryl, optionally substituted heteroaryl, optionally substituted C3-C8 cycloalkyl and optionally substituted C1-C4 alkyl; preferably hydrogen, halogen, hydroxyl. or optionally substituted cyclopropyl;
Alternatively two geminal R7 groups can be taken together with the carbon atom to which they are attached to form a spiro, optionally substituted C3-C7 cycloalkyl, spiro, optionally substituted C3-C7 cycloalkenyl or spiro, optionally substituted, 3- to 7-membered heterocyclic ring; preferably, spiro cyclopropyl;
R7a and R7b at each occurrence are each independently selected from the group consisting of hydrogen, optionally substituted aryl, and optionally substituted C1-C4 alkyl; preferably, hydrogen or methyl;
Alternatively, CHR7a—U or CHR7b—U can be taken together to form a group selected from CH═CH, fused and optionally substituted C3-C8 cycloalkyl, fused and optionally substituted aryl, or fused and optionally substituted heterocyclic; preferably fused, optionally substituted cyclopropyl; and
Yet alternatively, U, R7a, and R7b can be taken together with the carbon atoms to which they are attached to form a bridged, optionally substituted 4- to 7-membered ring selected from the group consisting of C4-C7 cycloalkyl, C4-C7 cycloalkenyl and 4- to 7-membered heterocyclic; preferably, bridged cyclopentyl.
Each preferred group stated above can be taken in combination with one, any or all other preferred groups.
In another aspect, the present invention provides a pharmaceutical composition comprising a therapeutically effective amount of a compound or combination of compounds of the present invention, or a pharmaceutically acceptable salt thereof, in combination with a pharmaceutically acceptable carrier or excipient.
In yet another aspect, the present invention provides a method of inhibiting the replication of a RNA-containing virus comprising contacting said virus with said pharmaceutical composition. Particularly, this invention is directed to methods of inhibiting the replication of HCV.
In still another aspect, the present invention provides a method of treating or preventing infection caused by an RNA-containing virus comprising administering to a patient in need of such treatment with said pharmaceutical composition. Particularly, this invention is directed to methods of treating or preventing infection caused by HCV.
Yet another aspect of the present invention provides the use of a compound or combination of compounds of the present invention, or a therapeutically acceptable salt thereof, as defined hereinafter, in the preparation of a medicament for the treatment or prevention of infection caused by RNA-containing virus, specifically HCV.


DETAILED DESCRIPTION OF THE INVENTION
The present invention relates to compounds of Formula (I) as illustrated above, or pharmaceutically acceptable salts thereof.
The compounds of the invention have utility in inhibiting the replication of RNA-containing virus, including, for example, HCV. Other compounds useful for inhibiting the replication of RNA-containing viruses and/or for the treatment or prophylaxis of HCV infection have been described in copending U.S. application Ser. No. 12/702,673 filed Feb. 9, 2010 entitled “Linked Dibenzimidiazole Derivatives”; U.S. application Ser. No. 12/702,692 filed Feb. 9, 2010 entitled “Linked Dibenzimidiazole Derivatives”; U.S. application Ser. No. 12/702,802 filed Feb. 9, 2010 entitled “Linked Dibenzimidiazole Derivatives”; U.S. application Ser. No. 12/707,190 filed Feb. 17, 2010 entitled “Linked Diimidazole Antivirals”; U.S. application Ser. No. 12/707,200 filed Feb. 17, 2010 entitled “Linked Diimidazole Derivatives”; U.S. application Ser. No. 12/707,210 filed Feb. 17, 2010 entitled “Hepatitis C Virus Inhibitors”; U.S. application Ser. No. 12/714,583 filed Mar. 1, 2010 entitled “Novel Benzimidazole Derivatives”; and U.S. application Ser. No. 12/714,576 filed Mar. 1, 2010 entitled “Hepatitis C Virus Inhibitors”; U.S. application Ser. No. 12/816,148 filed Jun. 15, 2010 entitled “Hepatitis C Virus Inhibitors”; U.S. application Ser. No. 12/816,171 filed Jun. 15, 2010 entitled “Hepatitis C Virus Inhibitors”; U.S. application Ser. No. 12/879,025 filed Sep. 10, 2010 entitled “Hepatitis C Virus Inhibitors”; U.S. application Ser. No. 12/879,026 filed Sep. 10, 2010 entitled “Hepatitis C Virus Inhibitors”; U.S. application Ser. No. 12/879,027 filed Sep. 10, 2010 entitled “Hepatitis C Virus Inhibitors”; U.S. application Ser. No. 12/879,028 filed Sep. 10, 2010 entitled “Hepatitis C Virus Inhibitors”; U.S. application Ser. No. 12/879,029 filed Sep. 10, 2010 entitled “Hepatitis C Virus Inhibitors”; U.S. application Ser. No. 12/879,031 filed Sep. 10, 2010 entitled “Hepatitis C Virus Inhibitors”; U.S. Provisional Application Ser. No. 61/297,918 filed Jan. 25, 2010 entitled “Hepatitis C Virus Inhibitors”; U.S. Provisional Application Ser. No. 61/314,304 filed Mar. 16, 2010 entitled “Hepatitis C Virus Inhibitors”; U.S. Provisional Application Ser. No. 61/322,438 filed Apr. 9, 2010 entitled “Hepatitis C Virus Inhibitors”; U.S. Provisional Application Ser. No. 61/351,327 filed Jun. 4, 2010 entitled “Hepatitis C Virus Inhibitors”; U.S. Provisional Application Ser. No. 61/372,999 filed Aug. 12, 2010 entitled “Hepatitis C Virus Inhibitors”; and the contents of each of which are expressly incorporated by reference herein.
As discussed above, a general strategy for the development of antiviral agents is to inactivate virally encoded proteins, including NS5A, that are essential for the replication of the virus. The relevant patent disclosures describing the synthesis of HCV NS5A inhibitors are: US 2009/0202478; US 2009/0202483; US 2010/0233120; US 2010/0260708; WO 2004/014852; WO 2006/079833; WO 2006/133326; WO 2007/031791; WO 2007/070556; WO 2007/070600; WO 2007/082554; WO 2008/021927; WO 2008/021928; WO 2008/021936; WO 2008/048589; WO 2008/064218; WO 2008/070447; WO 2008/144380; WO 2008/154601; WO 2009/020825; WO 2009/020828; WO 2009/034390; WO 2009/102318; WO 2009/102325; WO 2009/102694; WO 2010/017401; WO 2010/039793; WO 2010/065668; WO 2010/065674; WO 2010/065681; WO 2010/091413; WO 2010/096777; WO 2010/096462; WO 2010/096302; WO 2010/111483; WO 2010/111534; WO 2010/117635; WO 2010/111673; WO 2010/117704; WO 2010/132538; WO 2010/132601; WO 2010/138488; WO 2010/138368; WO 2010/138790; and WO 2010/138791, the contents of each of which are expressly incorporated by reference herein.
In another embodiment, the present invention relates to compounds of Formula (I), or pharmaceutically acceptable salts thereof; wherein X is C(R2)2 or N—OR1.
In yet another embodiment, the present invention relates to compounds of Formula (I), or pharmaceutically acceptable salts thereof; wherein X is CH2, CF2, or N-OMe.
In yet another embodiment, the present invention relates to compounds of Formula (I), or pharmaceutically acceptable salts thereof; wherein G and J are each independently optionally substituted five-membered heteroaryl containing one or more nitrogen atoms, and are each C-attached.
In yet another embodiment, the present invention relates to compounds of Formula (I), or pharmaceutically acceptable salts thereof; wherein G and J are each independently optionally substituted 5/6-membered ring fused heteroaryl, wherein the 5-membered ring of said 5/6-membered fused heteroaryl is a heteroaryl containing one or more nitrogen atoms and wherein the 5-membered ring is C-attached, and wherein the 6-membered ring of said 5/6-membered fused heteroaryl is aryl or heteroaryl and is C-attached to one of Ring A, L and Ring B.
In yet another embodiment, the present invention relates to compounds of Formula (I), or pharmaceutically acceptable salts thereof; wherein one of G and J is an optionally substituted five-membered heteroaryl containing one or more nitrogen atoms, and is C-attached; the other one of G and J is an optionally substituted 5/6-membered fused heteroaryl; wherein the 5-membered ring of said 5/6-membered fused heteroaryl is a heteroaryl containing one or more nitrogen atoms and wherein the 5-membered ring is C-attached, and wherein the 6-membered ring of said 5/6-membered fused heteroaryl is aryl or heteroaryl and is C-attached to one of Ring A, L and Ring B.
In yet another embodiment, the present invention relates to compounds of Formula (I), or pharmaceutically acceptable salts thereof; wherein L is absent, Ring A and Ring B are each independently optionally substituted phenyl, monocyclic heteroaryl, naphthyl, or bicyclic heteroaryl.
In yet another embodiment, the present invention relates to compounds of Formula (I), or pharmaceutically acceptable salts thereof; wherein L is optionally substituted C2-C4 alkenyl or optionally substituted C2-C4 alkynyl; and wherein one of Ring A and Ring B is absent, and the other of Ring A and Ring B is independently optionally substituted phenyl, monocyclic heteroaryl, naphthyl, or bicyclic heteroaryl.
In yet another embodiment, the present invention relates to compounds of Formula (I), or pharmaceutically acceptable salts thereof; wherein G and J are each independently illustrated by one of the following heteroaryl groups:




wherein each of the above shown heteroaryl groups is optionally substituted.

In yet another embodiment, the present invention relates to compounds of Formula (I), or pharmaceutically acceptable salts thereof; wherein G and J are each independently optionally substituted imidazolyl, benzimidazolyl or imidazopyridyl.
In yet another embodiment, the present invention relates to compounds of Formulae (Ia-1 ̃Ia-2), or pharmaceutically acceptable salts thereof:




wherein Ring A, Ring B, G, J, L, U, X, R1, R3, R4, R5, R6, R7a and R7b are as previously defined.

In yet another embodiment, the present invention relates to compounds of Formulae (Ia-3 ̃Ia-4), or pharmaceutically acceptable salts thereof:




wherein Ring A, Ring B, G, J, L, U, X, R6, and R7b are as previously defined.

In yet another embodiment, the present invention relates to compounds of Formulae (Ia-5 ̃Ia-6), or pharmaceutically acceptable salts thereof:




wherein Ring A, Ring B, G, J, L, U, X, R6, and R7b are as previously defined.

In yet another embodiment, the present invention relates to compounds of Formulae (Ia-5 ̃Ia-6), or pharmaceutically acceptable salts thereof; wherein R6 at each occurrence is selected from the group consisting of C1-C8 alkyl, C2-C8 alkenyl, C2-C8 alkynyl, C3-C8 cycloalkyl, C3-C8 cycloalkenyl, heterocyclic, aryl, and heteroaryl, each optionally substituted.
In still another embodiment, the present invention relates to compounds of Formula (Ia-5 ̃Ia-6), or pharmaceutically acceptable salts thereof; wherein R6 at each occurrence is independently C1-C8 alkyl optionally substituted with amino, hydroxy, protected amino, or O(C1-C4 alkyl).
In still another embodiment, the present invention relates to compounds of Formulae (Ib-1 ̃Ib-5), or pharmaceutically acceptable salts thereof:




wherein G, J, Q, X, R1, R6, R7a, and R7b are as previously defined; in Formula (Ib-4), Ring A, Ring B and L are each present and as previously defined; in Formula (Ib-1), Ring A and Ring B are each present and as previously defined; in Formula (Ib-2), Ring B and L are each present and as previously defined; in Formula (Ib-3), Ring A and L are each present and as previously defined; and in Formula (Ib-5), Ring B is present and as previously defined.

In still another embodiment, the present invention relates to compounds of Formulae (Ib-1 ̃Ib-5), or pharmaceutically acceptable salts thereof; wherein R6 is C1-C8 alkyl optionally substituted with amino, hydroxy, protected amino or O(C1-C4 alkyl).
In still another embodiment, the present invention relates to compounds of Formula (Ib-1), or pharmaceutically acceptable salts thereof; wherein Ring A and Ring B are each independently optionally substituted phenyl or monocyclic heteroaryl; and G and J are each independently optionally substituted imidazolyl.
In still another embodiment, the present invention relates to compounds of Formula (Ib-1), or pharmaceutically acceptable salts thereof; wherein Ring A and Ring B are each independently optionally substituted phenyl or monocyclic heteroaryl; and G and J are each independently optionally substituted benzimidazolyl or optionally substituted imidazopyridyl.
In still another embodiment, the present invention relates to compounds of Formula (Ib-1), or pharmaceutically acceptable salts thereof; wherein one of Ring A and Ring B is optionally substituted phenyl or optionally substituted monocyclic heteroaryl, the other of Ring A and Ring B is optionally substituted bicyclic aryl or bicyclic heteroaryl; and G and J are each independently imidazolyl, benzimidazolyl or imidazopyridyl, each optionally substituted.
In still another embodiment, the present invention relates to compounds of Formula (Ib-1), or pharmaceutically acceptable salts thereof; wherein Ring A and Ring B are each independently optionally substituted bicyclic aryl or bicyclic heteroaryl; and G and J are each independently optionally substituted imidazolyl.
In still another embodiment, the present invention relates to compounds of Formula (Ib-2 ̃Ib-3), or pharmaceutically acceptable salts thereof; wherein Ring B or Ring A is optionally substituted phenyl or monocyclic heteroaryl; L is optionally substituted C2-C4 alkenyl or optionally substituted C2-C4 alkynyl; and G and J are each independently imidazolyl, benzimidazolyl or imidazopyridyl, each optionally substituted.
In still another embodiment, the present invention relates to compounds of Formulae (Ib-2 ̃Ib-3), or pharmaceutically acceptable salts thereof; wherein Ring B or Ring A is optionally substituted bicyclic aryl or bicyclic heteroaryl; L is optionally substituted C2-C4 alkenyl or optionally substituted C2-C4 alkynyl; and G and J are each independently imidazolyl, benzimidazolyl or imidazopyridyl, each optionally substituted.
In still another embodiment, the present invention relates to compounds of Formula (Ib-4), or pharmaceutically acceptable salts thereof; wherein Ring A and Ring B are each independently optionally substituted phenyl or monocyclic heteroaryl; L is optionally substituted C2-C4 alkenyl or optionally substituted C2-C4 alkynyl; and G and J are each independently imidazolyl, benzimidazolyl or imidazopyridyl, each optionally substituted.
In still another embodiment, the present invention relates to compounds of Formula (Ib-5), or pharmaceutically acceptable salts thereof; wherein Ring B is independently optionally substituted polycyclic aryl or heteroaryl; and G and J are each independently imidazolyl, benzimidazolyl or imidazopyridyl, each optionally substituted.
In another embodiment, the present invention relates to compounds of Formulae (Ic-1) and (Ic-2) below,




and pharmaceutically acceptable salts thereof, wherein each R, R′, R′′ and R′′′ is independently H, halogen, optionally substituted C1-C6-alkyl, optionally substituted C1-C6-alkoxy, or optionally substituted C1-C6-alkyl-C(O)— and Q, X, R1, R6, R7a and R7b are as previously defined in Formula (I). In a preferred embodiment, each of R, R′, R′′ and R′′′ is H; in another preferred embodiment, each of R, R′, R′′ and R′′′ is independently selected from H, chloro, fluoro, trifluoromethyl, methoxy and acetyl. In yet another embodiment, the present invention relates to compounds of Formulae (Ic-1i) and (Ic-2i):





and pharmaceutically acceptable salts thereof, wherein each R, R′, R′′, R′′′Q, X, R1, R6, R7a and R7b are as previously defined in Formulae (Ic-1) and (Ic-2).

In another embodiment, the present invention relates to compounds of Formula (Ic-3),




and pharmaceutically acceptable salts thereof, wherein Q, X, R1, R6, R7a and R7b are as previously defined in Formula (I) and Aa is selected from the groups set forth below.








In an additional aspect, the invention relates to compounds of Formula (Ic-3i):





and pharmaceutically acceptable salts thereof, wherein Q, X, R1, R6, R7a, R7b and Aa are as previously defined in Formula (Ic-3).

In another embodiment, the present invention relates to compounds of Formula (Ic-4),




and pharmaceutically acceptable salts thereof, wherein Q, X, R1, R6, R7a and R7b are as previously defined in Formula (I) and Aa is selected from the groups set forth below.








In yet an additional aspect, the present invention relates to compounds of Formula (Ic-4i):





or pharmaceutically acceptable salts thereof, wherein Q, X, R1, R6, R7a, R7b and Aa are as previously defined in Formula (Ic-4).

In still another embodiment, the present invention relates to compounds of Formula (Ic-5):




and pharmaceutically acceptable salts thereof, wherein Q, X, R1, R6, R7a and R7b are as previously defined in Formula (I) and Gg is selected from the groups set forth below.





In an additional aspect, the present invention relates to compounds of Formula (Ic-5i):





and pharmaceutically acceptable salts thereof, wherein Q, X, R1, R6, R7a, R7b and Gg are as previously defined in Formula (Ic-5).

In still another embodiment, the present invention relates to compounds of Formulae (I), (Ia-1) to (Ia-6), (Ib-1) to (Ib-5), (Ic-1) to (Ic-5), and (Ic-1i) to (Ic-5i), and pharmaceutically acceptable salts thereof; wherein




at each occurrence is one of the following groups:








Representative compounds of the present invention are those selected from compounds 1-337 compiled in the following tables:







TABLE 1







Compounds 1-219.








 








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128






 


129






 


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133






 


134






 


135






 


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138






 


139






 


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142






 


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144






 


145






 


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150






 


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153






 


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163






 


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219




















TABLE 2







Compounds 220-229.








 
















Entry
R
R′
R′′
U
Entry
R
R′
R′′
U





220
Me
H
H
CH2
221
H
H
H
CF2


 


222
Me
H
H
S
223
H
H
H






 


224
H
Me
H
CH2
225
H
H
H






 


226
H
Ph
H
CH2
227
H
H
H






 


228
H
H
Ph
CH2
229
H
H
H




















TABLE 3







Compounds 230-239.








 














Entry
R
R′
R′′
Entry
R
R′
R′′





230
Me
H
H
231
H
CO2Me
H


232
H
F
H
233
H
H
CO2Me


234
H
H
F
235
H
OMe
H


236
H
Cl
H
237
H
H
OMe


238
H
H
Cl
239
H
CF3
H
















TABLE 4







Compounds 240-249.








 
















Entry
R
R′
R′′
R′′′
Entry
R
R′
R′′
R′′′





240
F
H
H
H
241
F
F
H
H


242
Me
H
H
H
243
Me
Me
H
H


244
H
H
Me
Me
245
H
H
Et
Et


246
CF3
H
H
H
247
CF3
H
CF
H


248
Cl
H
H
H
249
Cl
H
Cl
H
















TABLE 5







Compounds 250-264.








 












Entry
R
Entry
R
Entry
R











250






 


251






 


252






 


253






 


254






 


255






 


256






 


257






 


258






 


259






 


260






 


261






 


262






 


263






 


264




















TABLE 6







Compounds 265-282.








 












Entry
Aa
Entry
Aa
Entry
Aa





 


265




266




267






 


268




269




270






 


271




272




273






 


274




275




276






 


277




278




279






 


280




281




282




















TABLE 7







Compounds 283-303.








 












Entry
Aa
Entry
Aa
Entry
Aa





283




284




285






 


286




287




288






 


289




290




291






 


292




293




294






 


295




296




297






 


298




299




300






 


301




302




303




















TABLE 8







Compounds 304-315.








 












Entry
Gg
Entry
Gg
Entry
Gg





304




305




306






 


307




308




309






 


310




311




312






 


313




314




315




















TABLE 9





Compounds 316-337
















316






 


317






 


318






 


319






 


320






 


321






 


322






 


323






 


324






 


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337













It will be appreciated that the description of the present invention herein should be construed in congruity with the laws and principals of chemical bonding. In some instances it may be necessary to remove a hydrogen atom in order to accommodate a substitutent at any given location.
It is intended that the definition of any substituent or variable (e.g., R3, R7, etc.) at a particular location in a molecule be independent of its definitions elsewhere in that molecule. For example, when X is C(R7)2, each of the two R7 groups may be the same or different.
It will be yet appreciated that the compounds of the present invention may contain one or more asymmetric carbon atoms and may exist in racemic, diastereoisomeric, and optically active forms. It will still be appreciated that certain compounds of the present invention may exist in different tautomeric forms. All tautomers are contemplated to be within the scope of the present invention.
It should be understood that in some embodiments, the compounds encompassed by the present invention are those that are suitably stable for use as pharmaceutical agent.
It will be further appreciated that reference herein to therapy and/or treatment includes, but is not limited to, prevention, retardation, prophylaxis, therapy and/or cure of the disease. It will further be appreciated that references herein to treatment or prophylaxis of HCV infection includes treatment or prophylaxis of HCV-associated disease such as liver fibrosis, cirrhosis and hepatocellular carcinoma.
A further embodiment of the present invention includes pharmaceutical compositions comprising any single compound a combination of two or more compounds delineated herein, or a pharmaceutically acceptable salt thereof, with a pharmaceutically acceptable carrier or excipient.
Yet a further embodiment of the present invention is a pharmaceutical composition comprising any single compound or a combination of two or more compounds delineated herein, or a pharmaceutically acceptable salt thereof, in combination with one or more agents known in the art, with a pharmaceutically acceptable carrier or excipient.
It will be further appreciated that compounds of the present invention can be administered as the sole active pharmaceutical agent, or used in combination with one or more agents to treat or prevent hepatitis C infections or the symptoms associated with HCV infection. Other agents to be administered in combination with a compound or combination of compounds of the present invention include therapies for disease caused by HCV infection that suppresses HCV viral replication by direct or indirect mechanisms. These agents include, but are not limited to, host immune modulators (for example, interferon-alpha, pegylated interferon-alpha, consensus interferon, interferon-beta, interferon-gamma, CpG oligonucleotides and the like); antiviral compounds that inhibit host cellular functions such as inosine monophosphate dehydrogenase (for example, ribavirin and the like); cytokines that modulate immune function (for example, interleukin 2, interleukin 6, and interleukin 12); a compound that enhances the development of type 1 helper T cell response; interfering RNA; anti-sense RNA; vaccines comprising HCV antigens or antigen adjuvant combinations directed against HCV; agents that interact with host cellular components to block viral protein synthesis by inhibiting the internal ribosome entry site (IRES) initiated translation step of HCV viral replication or to block viral particle maturation and release with agents targeted toward the viroporin family of membrane proteins such as, for example, HCV P7 and the like; and any agent or combination of agents that inhibit the replication of HCV by targeting other proteins of the viral genome involved in the viral replication and/or interfere with the function of other viral targets, such as inhibitors of NS3/NS4A protease, NS3 helicase, NS5B polymerase, NS4A protein and NS5A protein.
According to yet another embodiment, the pharmaceutical compositions of the present invention may further comprise other inhibitor(s) of targets in the HCV life cycle, including, but not limited to, helicase, polymerase, metalloprotease, NS4A protein, NS5A protein, and internal ribosome entry site (IRES).
Accordingly, one embodiment of the present invention is directed to a method for treating or preventing an infection caused by an RNA-containing virus comprising co-administering to a patient in need of such treatment one or more agents selected from the group consisting of a host immune modulator and a second or more antiviral agents, or a combination thereof, with a therapeutically effective amount of a compound or combination of compounds of the present invention, or a pharmaceutically acceptable salt thereof. Examples of the host immune modulator include, but are not limited to, interferon-alpha, pegylated-interferon-alpha, interferon-beta, interferon-gamma, a cytokine, a vaccine, and a vaccine comprising an antigen and an adjuvant, and said second antiviral agent inhibits replication of HCV either by inhibiting host cellular functions associated with viral replication or by targeting proteins of the viral genome. Example of the RNA-containing virus includes, but not limited to, hepatitis C virus (HCV).
A further embodiment of the present invention is directed to a method of treating or preventing infection caused by an RNA-containing virus comprising co-administering to a patient in need of such treatment an agent or combination of agents that treat or alleviate symptoms of HCV infection including cirrhosis and inflammation of the liver, with a therapeutically effective amount of a compound or combination of compounds of the present invention, or a pharmaceutically acceptable salt thereof. Example of the RNA-containing virus includes, but not limited to, hepatitis C virus (HCV).
Yet another embodiment of the present invention provides a method of treating or preventing infection caused by an RNA-containing virus comprising co-administering to a patient in need of such treatment one or more agents that treat patients for disease caused by hepatitis B (HBV) infection, with a therapeutically effective amount of a compound or a combination of compounds of the present invention, or a pharmaceutically acceptable salt thereof. An agent that treats patients for disease caused by hepatitis B (HBV) infection may be for example, but not limited thereto, L-deoxythymidine, adefovir, lamivudine or tenfovir, or any combination thereof. Example of the RNA-containing virus includes, but not limited to, hepatitis C virus (HCV).
A further embodiment of the present invention provides a method of treating or preventing infection caused by an RNA-containing virus comprising co-administering to a patient in need of such treatment one or more agents that treat patients for disease caused by human immunodeficiency virus (HIV) infection, with a therapeutically effective amount of a compound or a combination of compounds of the present invention, or a pharmaceutically acceptable salt thereof. The agent that treats patients for disease caused by human immunodeficiency virus (HIV) infection includes, but is not limited thereto, ritonavir, lopinavir, indinavir, nelfinavir, saquinavir, amprenavir, atazanavir, tipranavir, TMC-114, fosamprenavir, zidovudine, lamivudine, didanosine, stavudine, tenofovir, zalcitabine, abacavir, efavirenz, nevirapine, delavirdine, TMC-125, L-870812, S-1360, enfuvirtide (T-20) or T-1249, or any combination thereof. Example of the RNA-containing virus includes, but is not limited to, hepatitis C virus (HCV).
It can occur that a patient may be co-infected with hepatitis C virus and one or more other viruses, including but not limited to human immunodeficiency virus (HIV), hepatitis A virus (HAV) and hepatitis B virus (HBV). Thus also contemplated is combination therapy to treat such co-infections by co-administering a compound according to the present invention with at least one of an HIV inhibitor, an HAV inhibitor and an HBV inhibitor.
In addition, the present invention provides the use of a compound or a combination of compounds of the invention, or a therapeutically acceptable salt thereof, and one or more agents selected from the group consisting of a host immune modulator and a second or more antiviral agents, or a combination thereof, to prepare a medicament for the treatment of an infection caused by an RNA-containing virus in a patient, particularly hepatitis C virus. Examples of the host immune modulator are, but not limited to, interferon-alpha, pegylated-interferon-alpha, interferon-beta, interferon-gamma, a cytokine, a vaccine, and a vaccine comprising an antigen and an adjuvant, and said second antiviral agent inhibits replication of HCV either by inhibiting host cellular functions associated with viral replication or by targeting proteins of the viral genome.
When used in the above or other treatments, combination of compounds or compounds of the present invention, together with one or more agents as defined herein above, can be employed in pure form or, where such forms exist, in pharmaceutically acceptable salt thereof. Alternatively, such combination of therapeutic agents can be administered as a pharmaceutical composition containing a therapeutically effective amount of the compound or combination of compounds of interest, or their pharmaceutically acceptable salt thereof, in combination with one or more agents as defined hereinabove, and a pharmaceutically acceptable carrier. Such pharmaceutical compositions can be used for inhibiting the replication of an RNA-containing virus, particularly Hepatitis C virus (HCV), by contacting said virus with said pharmaceutical composition. In addition, such compositions are useful for the treatment or prevention of an infection caused by an RNA-containing virus, particularly Hepatitis C virus (HCV).
Hence, a still further embodiment of the invention is directed to a method of treating or preventing infection caused by an RNA-containing virus, particularly a hepatitis C virus (HCV), comprising administering to a patient in need of such treatment a pharmaceutical composition comprising a compound or combination of compounds of the invention or a pharmaceutically acceptable salt thereof, and one or more agents as defined hereinabove, with a pharmaceutically acceptable carrier.
When administered as a combination, the therapeutic agents can be formulated as separate compositions which are given at the same time or within a predetermined period of time, or the therapeutic agents can be given as a single unit dosage form.
Antiviral agents contemplated for use in such combination therapy include agents (compounds or biologicals) that are effective to inhibit the formation and/or replication of a virus in a mammal, including but not limited to agents that interfere with either host or viral mechanisms necessary for the formation and/or replication of a virus in a mammal. Such agents can be selected from another anti-HCV agent; an HIV inhibitor; an HAV inhibitor; and an HBV inhibitor.
Other agents to be administered in combination with a compound of the present invention include a cytochrome P450 monooxygenase inhibitor (also referred to herein as a CYP inhibitor), which is expected to inhibit metabolism of the compounds of the invention. Therefore, the cytochrome P450 monooxygenase inhibitor would be in an amount effective to inhibit metabolism of the compounds of this invention. Accordingly, the CYP inhibitor is administered in an amount such that the bioavailiablity of the protease inhibitor is increased in comparison to the bioavailability in the absence of the CYP inhibitor.
In one embodiment, the invention provides methods for improving the pharmacokinetics of compounds of the invention. The advantages of improving the pharmacokinetics of drugs are recognized in the art (see, for example, US Patent Pub. Nos. 2004/0091527; US 2004/0152625; and US 2004/0091527). Accordingly, one embodiment of this invention provides a method for administering an inhibitor of CYP3A4 and a compound of the invention. Another embodiment of this invention provides a method for administering a compound of the invention and an inhibitor of isozyme 3A4 (“CYP3A4”), isozyme 2C19 (“CYP2C19”), isozyme 2D6 (“CYP2D6”), isozyme 1A2 (“CYP1A2”), isozyme 2C9 (“CYP2C9”), or isozyme 2E1 (“CYP2E1”). In a preferred embodiment, the CYP inhibitor preferably inhibits CYP3A4. Any CYP inhibitor that improves the pharmacokinetics of the relevant NS3/4A protease may be used in a method of this invention. These CYP inhibitors include, but are not limited to, ritonavir (see, for example, WO 94/14436), ketoconazole, troleandomycin, 4-methylpyrazole, cyclosporin, clomethiazole, cimetidine, itraconazole, fluconazole, miconazole, fluvoxamine, fluoxetine, nefazodone, sertraline, indinavir, nelfinavir, amprenavir, fosamprenavir, saquinavir, lopinavir, delavirdine, erythromycin, VX-944, and VX-497. Preferred CYP inhibitors include ritonavir, ketoconazole, troleandomycin, 4-methylpyrazole, cyclosporin, and clomethiazole.
It will be understood that the administration of the combination of the invention by means of a single patient pack, or patient packs of each formulation, containing within a package insert instructing the patient to the correct use of the invention is a desirable additional feature of this invention.
According to a further aspect, the invention is a pack comprising at least a compound of the invention and a CYP inhibitor of the invention and an information insert containing directions on the use of the combination of the invention. In an alternative embodiment of this invention, the pharmaceutical pack further comprises one or more of additional agent as described herein. The additional agent or agents may be provided in the same pack or in separate packs.
Another aspect of this involves a packaged kit for a patient to use in the treatment of HCV infection or in the prevention of HCV infection, comprising: a single or a plurality of pharmaceutical formulation of each pharmaceutical component; a container housing the pharmaceutical formulation (s) during storage and prior to administration; and instructions for carrying out drug administration in a manner effective to treat or prevent HCV infection.
Accordingly, this invention provides kits for the simultaneous or sequential administration of a compound of the invention and a CYP inhibitor (and optionally an additional agent) or derivatives thereof are prepared in a conventional manner. Typically, such a kit will comprise, e.g. a composition of each inhibitor and optionally the additional agent (s) in a pharmaceutically acceptable carrier (and in one or in a plurality of pharmaceutical formulations) and written instructions for the simultaneous or sequential administration.
In another embodiment, a packaged kit is provided that contains one or more dosage forms for self administration; a container means, preferably sealed, for housing the dosage forms during storage and prior to use; and instructions for a patient to carry out drug administration. The instructions will typically be written instructions on a package insert, a label, and/or on other components of the kit, and the dosage form or forms are as described herein. Each dosage form may be individually housed, as in a sheet of a metal foil-plastic laminate with each dosage form isolated from the others in individual cells or bubbles, or the dosage forms may be housed in a single container, as in a plastic bottle. The present kits will also typically include means for packaging the individual kit components, i.e., the dosage forms, the container means, and the written instructions for use. Such packaging means may take the form of a cardboard or paper box, a plastic or foil pouch, etc.
DEFINITIONS
Listed below are definitions of various terms used to describe this invention. These definitions apply to the terms as they are used throughout this specification and claims, unless otherwise limited in specific instances, either individually or as part of a larger group.
The term “viral infection” refers to the introduction of a virus into cells or tissues, e.g., hepatitis C virus (HCV). In general, the introduction of a virus is also associated with replication. Viral infection may be determined by measuring virus antibody titer in samples of a biological fluid, such as blood, using, e.g., enzyme immunoassay. Other suitable diagnostic methods include molecular based techniques, such as RT-PCR, direct hybrid capture assay, nucleic acid sequence based amplification, and the like. A virus may infect an organ, e.g., liver, and cause disease, e.g., hepatitis, cirrhosis, chronic liver disease and hepatocellular carcinoma.
The term “immune modulator” refers to any substance meant to alter the working of the humoral or cellular immune system of a subject. Such immune modulators include inhibitors of mast cell-mediated inflammation, interferons, interleukins, prostaglandins, steroids, cortico-steroids, colony-stimulating factors, chemotactic factors, etc.
The term “aryl,” as used herein, refers to a mono- or polycyclic carbocyclic ring system including, but not limited to, phenyl, naphthyl, tetrahydronaphthyl, indanyl, idenyl. A polycyclic aryl is a polycyclic ring system that comprises at least one aromatic ring. Polycyclic aryls can comprise fused rings, covalently attached rings or a combination thereof.
The term “heteroaryl,” as used herein, refers to a mono- or polycyclic ring system comprising at least one aromatic ring having one or more ring atom selected from S, O and N; and the remaining ring atoms are carbon, wherein any N or S contained within the ring may be optionally oxidized. Heteroaryl includes, but is not limited to, pyridinyl, pyrazinyl, pyrimidinyl, pyrrolyl, pyrazolyl, imidazolyl, thiazolyl, oxazolyl, isooxazolyl, thiadiazolyl, oxadiazolyl, thiophenyl, furanyl, quinolinyl, isoquinolinyl, benzimidazolyl, benzooxazolyl, quinoxalinyl. A polycyclic heteroaryl can comprise fused rings, covalently attached rings or a combination thereof.
In accordance with the invention, any of the aryls, substituted aryls, heteroaryls and substituted heteroaryls described herein, can be any aromatic group. Aromatic groups can be substituted or unsubstituted.
The term “bicyclic aryl” or “bicyclic heteroaryl” refers to a ring system consisting of two rings wherein at least one ring is aromatic; and they can be fused or covalently attached.
The term “tricyclic aryl” or “tricyclic heteroaryl” refers to a ring system consisting of three rings wherein at least one ring is aromatic.
The terms “C1-C4 alkyl,” “C1-C6 alkyl,” “C1-C8 alkyl,” “C2-C4 alkyl,” or “C3-C6 alkyl,” as used herein, refer to saturated, straight- or branched-chain hydrocarbon radicals containing between one and four, one and six, one and eight carbon atoms, or the like, respectively. Examples of C1-C8 alkyl radicals include, but are not limited to, methyl, ethyl, propyl, isopropyl, n-butyl, tert-butyl, neopentyl, n-hexyl, heptyl and octyl radicals.
The terms “C2-C8 alkenyl,” “C2-C4 alkenyl,” “C3-C4 alkenyl,” or “C3-C6 alkenyl,” as used herein, refer to straight- or branched-chain hydrocarbon radicals containing from two to eight, or two to four carbon atoms, or the like, having at least one carbon-carbon double bond by the removal of a single hydrogen atom. Alkenyl groups include, but are not limited to, for example, ethenyl, propenyl, butenyl, 1-methyl-2-buten-1-yl, heptenyl, octenyl, and the like.
The terms “C2-C8 alkynyl,” “C2-C4 alkynyl,” “C3-C4 alkynyl,” or “C3-C6 alkynyl,” as used herein, refer to straight- or branched-chain hydrocarbon radicals containing from two to eight, or two to four carbon atoms, or the like, having at least one carbon-carbon triple bond by the removal of a single hydrogen atom. Representative alkynyl groups include, but are not limited to, for example, ethynyl, 1-propynyl, 1-butynyl, heptynyl, octynyl, and the like.
The term “C3-C8-cycloalkyl”, or “C5-C7-cycloalkyl,” as used herein, refers to a monocyclic or polycyclic saturated carbocyclic ring compound, and the carbon atoms may be optionally oxo-substituted. Examples of C3-C8-cycloalkyl include, but not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cyclopentyl and cyclooctyl; and examples of C5-C7-cycloalkyl include, but not limited to, cyclopentyl, cyclohexyl, bicyclo [2.2.1]heptyl, and the like.
The term “C3-C8 cycloalkenyl” or “C5-C7 cycloalkenyl” as used herein, refers to monocyclic or polycyclic carbocyclic ring compound having at least one carbon-carbon double bond and the carbon atoms may be optionally oxo-substituted. Examples of C3-C8 cycloalkenyl include, but not limited to, cyclopropenyl, cyclobutenyl, cyclopentenyl, cyclohexenyl, cycloheptenyl, cyclooctenyl, and the like; and examples of C5-C7 cycloalkenyl include, but not limited to, cyclopentenyl, cyclohexenyl, cycloheptenyl, and the like.
It is understood that any alkyl, alkenyl, alkynyl, cycloalkyl and cycloalkenyl moiety described herein can also be an aliphatic group or an alicyclic group. An “aliphatic” group is a non-aromatic moiety that may contain any combination of carbon atoms, hydrogen atoms, halogen atoms, oxygen, nitrogen or other atoms, and optionally contains one or more units of unsaturation, e.g., double and/or triple bonds, and includes and/or optionally contains one or more functional groups, e.g., O, OH, NH, NH2, C(O), S(O)2, C(O)O, C(O)NH, OC(O)O, OC(O)NH, OC(O)NH2, S(O)2NH, S(O)2NH2, NHC(O)NH2, or the like, and the carbon atoms may be optionally oxo-substituted. An aliphatic group may be straight chained, branched or cyclic and preferably contains between about 1 and about 24 carbon atoms, more typically between about 1 and about 12 carbon atoms. In addition to aliphatic hydrocarbon groups, as used herein, aliphatic groups expressly include, for example, alkoxyalkyls, polyalkoxyalkyls, such as polyalkylene glycols, polyamines, and polyimines, for example. Such aliphatic groups may be further substituted. A linear aliphatic group is an aliphatic group that comprised of only non-cyclic aliphatic group.
The term “alicyclic,” as used herein, denotes a monovalent group derived from a monocyclic or bicyclic saturated carbocyclic ring compound by the removal of a single hydrogen atom, and the carbon atoms may be optionally oxo-substituted. Examples include, but not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, bicyclo [2.2.1]heptyl, and bicyclo [2.2.2]octyl. Such alicyclic groups may be further substituted.
The terms “heterocyclic” or “heterocycloalkyl” can be used interchangeably and referred to a non-aromatic ring or a bi- or tri-cyclic group fused system, where (i) each ring system contains at least one heteroatom independently selected from oxygen, sulfur and nitrogen, (ii) each ring system can be saturated or unsaturated (iii) the nitrogen and sulfur heteroatoms may optionally be oxidized, (iv) the nitrogen heteroatom may optionally be quaternized, (v) any of the above rings may be fused to an aromatic ring and (vi) the remaining ring atoms are carbon atoms which may be optionally oxo-substituted. Representative heterocycloalkyl groups include, but are not limited to, 1,3-dioxolane, pyrrolidinyl, pyrazolinyl, pyrazolidinyl, imidazolinyl, imidazolidinyl, piperidinyl, piperazinyl, oxazolidinyl, isoxazolidinyl, morpholinyl, thiazolidinyl, isothiazolidinyl, quinoxalinyl, pyridazinonyl, and tetrahydrofuryl. Such heterocyclic groups may be further substituted. Heteroaryl or heterocyclic groups can be C-attached or N-attached (where possible).
It is understood that any alkyl, alkenyl, alkynyl, alicyclic, cycloalkyl, cycloalkenyl, aryl, heteroaryl, heterocyclic, aliphatic moiety or the like, described herein can also be a divalent group when used as a linkage to connect two groups or substituents, which can be at the same or different atom(s).
The term “substituted” when used with alkyl, alkenyl, alkynyl, alicyclic, cycloalkyl, cycloalkenyl, aryl, heteroaryl, heterocyclic, or aliphatic as described herein refers to substitution by independent replacement of one, two, or three or more of the hydrogen atoms thereon with substituents including, but not limited to, —F, —Cl, —Br, —I, —OH, protected hydroxy, —NO2, —N3, —CN, —NH2, protected amino, oxo, thioxo, —NH—C1-C12-alkyl, —NH—C2-C8-alkenyl, —NH—C2-C8-alkynyl, —NH—C3-C12-cycloalkyl, —NH-aryl, —NH-heteroaryl, —NH-heterocycloalkyl, -dialkylamino, -diarylamino, -diheteroarylamino, —O—C1-C12-alkyl, —O—C2-C8-alkenyl, —O—C2-C8-alkynyl, —O—C3-C12-cycloalkyl, —O-aryl, —O-heteroaryl, —O-heterocycloalkyl, —C(O)—C1-C12-alkyl, —C(O)—C2-C8-alkenyl, —C(O)—C2-C8-alkynyl, —C(O)—C3-C12-cycloalkyl, —C(O)-aryl, —C(O)-heteroaryl, —C(O)-heterocycloalkyl, —CONH2, —CONH—C1-C12-alkyl, —CONH—C2-C8-alkenyl, —CONH—C2-C8-alkynyl, —CONH—C3-C12-cycloalkyl, —CONH-aryl, —CONH-heteroaryl, —CONH-heterocycloalkyl, —OCO2—C1-C12-alkyl, —OCO2—C2-C8-alkenyl, —OCO2—C2-C8-alkynyl, —OCO2—C3-C12-cycloalkyl, —OCO2-aryl, —OCO2-heteroaryl, —OCO2-heterocycloalkyl, —CO2—C1-C12 alkyl, —CO2—C2-C8 alkenyl, —CO2—C2-C8 alkynyl, CO2—C3-C12-cycloalkyl, —CO2— aryl, CO2-heteroaryl, CO2-heterocyloalkyl, —OCONH2, —OCONH—C1-C12-alkyl, —OCONH—C2-C8-alkenyl, —OCONH—C2-C8-alkynyl, —OCONH—C3-C12-cycloalkyl, —OCONH-aryl, —OCONH-heteroaryl, —OCONH— heterocycloalkyl, —NHC(O)H, —NHC(O)—C1-C12-alkyl, —NHC(O)—C2-C8-alkenyl, —NHC(O)—C2-C8-alkynyl, —NHC(O)—C3-C12-cycloalkyl, —NHC(O)-aryl, —NHC(O)-heteroaryl, —NHC(O)-heterocycloalkyl, —NHCO2—C1-C12-alkyl, —NHCO2—C2-C8-alkenyl, —NHCO2—C2-C8-alkynyl, —NHCO2—C3-C12-cycloalkyl, —NHCO2-aryl, —NHCO2-heteroaryl, —NHCO2— heterocycloalkyl, —NHC(O)NH2, —NHC(O)NH—C1-C12-alkyl, —NHC(O)NH—C2-C8-alkenyl, —NHC(O)NH—C2-C8-alkynyl, —NHC(O)NH—C3-C12-cycloalkyl, —NHC(O)NH-aryl, —NHC(O)NH-heteroaryl, —NHC(O)NH-heterocycloalkyl, NHC(S)NH2, —NHC(S)NH—C1-C12-alkyl, —NHC(S)NH—C2-C8-alkenyl, —NHC(S)NH—C2-C8-alkynyl, —NHC(S)NH—C3-C12-cycloalkyl, —NHC(S)NH-aryl, —NHC(S)NH-heteroaryl, —NHC(S)NH-heterocycloalkyl, —NHC(NH)NH2, —NHC(NH)NH—C1-C12-alkyl, —NHC(NH)NH—C2-C8-alkenyl, —NHC(NH)NH—C2-C8-alkynyl, —NHC(NH)NH—C3-C12-cycloalkyl, —NHC(NH)NH-aryl, —NHC(NH)NH-heteroaryl, —NHC(NH)NH-heterocycloalkyl, —NHC(NH)—C1-C12-alkyl, —NHC(NH)—C2-C8-alkenyl, —NHC(NH)—C2-C8-alkynyl, —NHC(NH)—C3-C12-cycloalkyl, —NHC(NH)-aryl, —NHC(NH)-heteroaryl, —NHC(NH)-heterocycloalkyl, —C(NH)NH—C1-C12-alkyl, —C(NH)NH—C2-C8-alkenyl, —C(NH)NH—C2-C8-alkynyl, —C(NH)NH—C3-C12-cycloalkyl, —C(NH)NH-aryl, —C(NH)NH-heteroaryl, —C(NH)NH-heterocycloalkyl, —S(O)—C1-C12-alkyl, —S(O)—C2-C8-alkenyl, —S(O)—C2-C8-alkynyl, —S(O)—C3-C12-cycloalkyl, —S(O)-aryl, —S(O)-heteroaryl, —S(O)-heterocycloalkyl, —SO2NH2, —SO2NH—C1-C12-alkyl, —SO2NH—C2-C8-alkenyl, —SO2NH—C2-C8-alkynyl, —SO2NH—C3-C12-cycloalkyl, —SO2NH-aryl, —SO2NH-heteroaryl, —SO2NH— heterocycloalkyl, —NHSO2—C1-C12-alkyl, —NHSO2—C2-C8-alkenyl, —NHSO2—C2-C8-alkynyl, —NHSO2—C3-C12-cycloalkyl, —NHSO2-aryl, —NHSO2-heteroaryl, —NHSO2-heterocycloalkyl, —CH2NH2, —CH2SO2CH3, -aryl, -arylalkyl, -heteroaryl, -heteroarylalkyl, -heterocycloalkyl, —C3-C12-cycloalkyl, polyalkoxyalkyl, polyalkoxy, -methoxymethoxy, -methoxyethoxy, —SH, —S—C1-C12-alkyl, —S—C2-C8-alkenyl, —S—C2-C8-alkynyl, —S—C3-C12-cycloalkyl, —S-aryl, —S-heteroaryl, —S-heterocycloalkyl, or methylthiomethyl. It is understood that the aryls, heteroaryls, alkyls, and the like can be further substituted.
The term “halogen,” as used herein, refers to an atom selected from fluorine, chlorine, bromine and iodine.
The term “hydrogen” includes hydrogen and deuterium. In addition, the recitation of an atom includes other isotopes of that atom so long as the resulting compound is pharmaceutically acceptable.
The term “hydroxy protecting group,” as used herein, refers to a labile chemical moiety which is known in the art to protect a hydroxyl group against undesired reactions during synthetic procedures. After said synthetic procedure(s) the hydroxy protecting group as described herein may be selectively removed. Hydroxy protecting groups as known in the art are described generally in T. H. Greene and P. G. M. Wuts, Protective Groups in Organic Synthesis, 3rd edition, John Wiley & Sons, New York (1999). Examples of hydroxyl protecting groups include benzyloxycarbonyl, 4-methoxybenzyloxycarbonyl, tert-butoxycarbonyl, isopropoxycarbonyl, diphenylmethoxycarbonyl, 2,2,2-trichloroethoxycarbonyl, allyloxycarbonyl, acetyl, formyl, chloroacetyl, trifluoroacetyl, methoxyacetyl, phenoxyacetyl, benzoyl, methyl, t-butyl, 2,2,2-trichloroethyl, 2-trimethylsilyl ethyl, allyl, benzyl, triphenylmethyl (trityl), methoxymethyl, methylthiomethyl, benzyloxymethyl, 2-(trimethylsilyl)ethoxymethyl, methanesulfonyl, trimethylsilyl, triisopropylsilyl, and the like.
The term “protected hydroxy,” as used herein, refers to a hydroxy group protected with a hydroxy protecting group, as defined above, including benzoyl, acetyl, trimethylsilyl, triethylsilyl, methoxymethyl groups, for example.
The term “carbonyl protecting group,” as used herein, refers to a labile chemical moiety which is known in the art to protect a carbonyl group against undesired reactions during synthetic procedures. After said synthetic procedure(s) the carbonyl protecting group as described herein may be selectively removed. Carbonyl protecting groups as known in the art are described generally in T. H. Greene and P. G. M. Wuts, Protective Groups in Organic Synthesis, 3rd edition, John Wiley & Sons, New York (1999). Examples of carbonyl protecting groups include acetals, ketals, cyclic acetals, cyclic ketals, mono- or dithioacetals, mono- or dithioketals, optionally substituted hydrazones or oximes.
The term “protected carbonyl,” as used herein, refers to a carbonyl group protected with a carbonyl protecting group, as defined above, including dimethyl acetal, 1,3-dioxolane, 1,3-dioxane, S,S′-dimethylketal, 1,3-dithiane, 1,3-dithiolane, 1,3-oxathiolane, N,N-dimethylhydrazone, oxime, for example.
The term “amino protecting group,” as used herein, refers to a labile chemical moiety which is known in the art to protect an amino group against undesired reactions during synthetic procedures. After said synthetic procedure(s) the amino protecting group as described herein may be selectively removed. Amino protecting groups as known in the art are described generally in T. H. Greene and P. G. M. Wuts, Protective Groups in Organic Synthesis, 3rd edition, John Wiley & Sons, New York (1999). Examples of amino protecting groups include, but are not limited to, methoxycarbonyl, t-butoxycarbonyl, 9-fluorenylmethoxycarbonyl, benzyloxycarbonyl, and the like.
The term “protected amino,” as used herein, refers to an amino group protected with an amino protecting group as defined above.
The term “substituted amino,” as used herein, refers to substitution by replacement of one or two hydrogen atoms of —NH2 with substituents independently selected from the group consisting of optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted aryl, optionally substituted cycloalkyl, optionally substituted heteroaryl, and optionally substituted heterocyclic; alternatively, when disubstituted, the two substitutents can be optionally taken together with the nitrogen atom to which they are attached to form an optionally substituted heterocyclic group.
The term “leaving group” means a functional group or atom which can be displaced by another functional group or atom in a substitution reaction, such as a nucleophilic substitution reaction. By way of example, representative leaving groups include chloro, bromo and iodo groups; sulfonic ester groups, such as mesylate, tosylate, brosylate, nosylate and the like; hydroxy; imidazolyl; and acyloxy groups, such as acetoxy, trifluoroacetoxy and the like.
The term “aprotic solvent,” as used herein, refers to a solvent that is relatively inert to proton activity, i.e., not acting as a proton-donor. Examples include, but are not limited to, hydrocarbons, such as hexane and toluene, for example, halogenated hydrocarbons, such as, for example, methylene chloride, ethylene chloride, chloroform, and the like, heterocyclic compounds, such as, for example, tetrahydrofuran and N-methylpyrrolidinone, and ethers such as diethyl ether, bis-methoxymethyl ether. Such compounds are well known to those skilled in the art, and it will be obvious to those skilled in the art that individual solvents or mixtures thereof may be preferred for specific compounds and reaction conditions, depending upon such factors as the solubility of reagents, reactivity of reagents and preferred temperature ranges, for example. Further discussions of aprotic solvents may be found in organic chemistry textbooks or in specialized monographs, for example: Organic Solvents Physical Properties and Methods of Purification, 4th ed., edited by John A. Riddick et al., Vol. II, in the Techniques of Chemistry Series, John Wiley & Sons, NY, 1986.
The term “protic solvent’ as used herein, refers to a solvent that tends to provide protons, such as an alcohol, for example, methanol, ethanol, propanol, isopropanol, butanol, t-butanol, and the like. Such solvents are well known to those skilled in the art, and it will be obvious to those skilled in the art that individual solvents or mixtures thereof may be preferred for specific compounds and reaction conditions, depending upon such factors as the solubility of reagents, reactivity of reagents and preferred temperature ranges, for example. Further discussions of protogenic solvents may be found in organic chemistry textbooks or in specialized monographs, for example: Organic Solvents Physical Properties and Methods of Purification, 4th ed., edited by John A. Riddick et al., Vol. II, in the Techniques of Chemistry Series, John Wiley & Sons, NY, 1986.
Combinations of substituents and variables envisioned by this invention are only those that result in the formation of stable compounds. The term “stable”, as used herein, refers to compounds which possess stability sufficient to allow manufacture and which maintains the integrity of the compound for a sufficient period of time to be useful for the purposes detailed herein (e.g., therapeutic or prophylactic administration to a subject).
The synthesized compounds can be separated from a reaction mixture and further purified by a method such as column chromatography, high pressure liquid chromatography, or recrystallization. As can be appreciated by the skilled artisan, further methods of synthesizing the compounds of the Formula herein will be evident to those of ordinary skill in the art. Additionally, the various synthetic steps may be performed in an alternate sequence or order to give the desired compounds. Synthetic chemistry transformations and protecting group methodologies (protection and deprotection) useful in synthesizing the compounds described herein are known in the art and include, for example, those such as described in R. Larock, Comprehensive Organic Transformations, 2nd Ed. Wiley-VCH (1999); T. W. Greene and P. G. M. Wuts, Protective Groups in Organic Synthesis, 3rd Ed., John Wiley and Sons (1999); L. Fieser and M. Fieser, Fieser and Fieser's Reagents for Organic Synthesis, John Wiley and Sons (1994); and L. Paquette, ed., Encyclopedia of Reagents for Organic Synthesis, John Wiley and Sons (1995), and subsequent editions thereof.
The term “subject” as used herein refers to an animal. Preferably the animal is a mammal. More preferably the mammal is a human. A subject also refers to, for example, dogs, cats, horses, cows, pigs, guinea pigs, fish, birds and the like.
The compounds of this invention may be modified by appending appropriate functionalities to enhance selective biological properties. Such modifications are known in the art and may include those which increase biological penetration into a given biological system (e.g., blood, lymphatic system, central nervous system), increase oral availability, increase solubility to allow administration by injection, alter metabolism and alter rate of excretion.
The compounds described herein contain one or more asymmetric centers and thus give rise to enantiomers, diastereomers, and other stereoisomeric forms that may be defined, in terms of absolute stereochemistry, as (R)— or (S)—, or as (D)- or (L)- for amino acids. The present invention is meant to include all such possible isomers, as well as their racemic and optically pure forms. Optical isomers may be prepared from their respective optically active precursors by the procedures described above, or by resolving the racemic mixtures. The resolution can be carried out in the presence of a resolving agent, by chromatography or by repeated crystallization or by some combination of these techniques which are known to those skilled in the art. Further details regarding resolutions can be found in Jacques, et al., Enantiomers, Racemates, and Resolutions (John Wiley & Sons, 1981). When the compounds described herein contain olefinic double bonds, other unsaturation, or other centers of geometric asymmetry, and unless specified otherwise, it is intended that the compounds include both E and Z geometric isomers or cis- and trans-isomers. Likewise, all tautomeric forms are also intended to be included. Tautomers may be in cyclic or acyclic. The configuration of any carbon-carbon double bond appearing herein is selected for convenience only and is not intended to designate a particular configuration unless the text so states; thus a carbon-carbon double bond or carbon-heteroatom double bond depicted arbitrarily herein as trans may be cis, trans, or a mixture of the two in any proportion.
Certain compounds of the present invention may also exist in different stable conformational forms which may be separable. Torsional asymmetry due to restricted rotation about an asymmetric single bond, for example because of steric hindrance or ring strain, may permit separation of different conformers. The present invention includes each conformational isomer of these compounds and mixtures thereof.
As used herein, the term “pharmaceutically acceptable salt” refers to those salts which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of humans and lower animals without undue toxicity, irritation, allergic response and the like, and are commensurate with a reasonable benefit/risk ratio. Pharmaceutically acceptable salts are well known in the art. For example, S. M. Berge, et al. describes pharmaceutically acceptable salts in detail in J. Pharmaceutical Sciences, 66: 1-19 (1977). The salts can be prepared in situ during the final isolation and purification of the compounds of the invention, or separately by reacting the free base function with a suitable organic acid. Examples of pharmaceutically acceptable salts include, but are not limited to, nontoxic acid addition salts are salts of an amino group formed with inorganic acids such as hydrochloric acid, hydrobromic acid, phosphoric acid, sulfuric acid and perchloric acid or with organic acids such as acetic acid, maleic acid, tartaric acid, citric acid, succinic acid or malonic acid or by using other methods used in the art such as ion exchange. Other pharmaceutically acceptable salts include, but are not limited to, adipate, alginate, ascorbate, aspartate, benzenesulfonate, benzoate, bisulfate, borate, butyrate, camphorate, camphorsulfonate, citrate, cyclopentanepropionate, digluconate, dodecylsulfate, ethanesulfonate, formate, fumarate, glucoheptonate, glycerophosphate, gluconate, hemisulfate, heptanoate, hexanoate, hydroiodide, 2-hydroxy-ethanesulfonate, lactobionate, lactate, laurate, lauryl sulfate, malate, maleate, malonate, methanesulfonate, 2-naphthalenesulfonate, nicotinate, nitrate, oleate, oxalate, palmitate, pamoate, pectinate, persulfate, 3-phenylpropionate, phosphate, picrate, pivalate, propionate, stearate, succinate, sulfate, tartrate, thiocyanate, p-toluenesulfonate, undecanoate, valerate salts, and the like. Representative alkali or alkaline earth metal salts include sodium, lithium, potassium, calcium, magnesium, and the like. Further pharmaceutically acceptable salts include, when appropriate, nontoxic ammonium, quaternary ammonium, and amine cations formed using counterions such as halide, hydroxide, carboxylate, sulfate, phosphate, nitrate, alkyl having from 1 to 6 carbon atoms, sulfonate and aryl sulfonate.
As used herein, the term “pharmaceutically acceptable ester” refers to esters which hydrolyze in vivo and include those that break down readily in the human body to leave the parent compound or a salt thereof. Suitable ester groups include, for example, those derived from pharmaceutically acceptable aliphatic carboxylic acids, particularly alkanoic, alkenoic, cycloalkanoic and alkanedioic acids, in which each alkyl or alkenyl moiety advantageously has not more than 6 carbon atoms. Examples of particular esters include, but are not limited to, formates, acetates, propionates, butyrates, acrylates and ethylsuccinates.
The term “pharmaceutically acceptable prodrugs” as used herein refers to those prodrugs of the compounds of the present invention which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of humans and lower animals with undue toxicity, irritation, allergic response, and the like, commensurate with a reasonable benefit/risk ratio, and effective for their intended use, as well as the zwitterionic forms, where possible, of the compounds of the present invention. “Prodrug”, as used herein means a compound which is convertible in vivo by metabolic means (e.g. by hydrolysis) to a compound of the invention. Various forms of prodrugs are known in the art, for example, as discussed in Bundgaard, (ed.), Design of Prodrugs, Elsevier (1985); Widder, et al. (ed.), Methods in Enzymology, vol. 4, Academic Press (1985); Krogsgaard-Larsen, et al., (ed). “Design and Application of Prodrugs, Textbook of Drug Design and Development, Chapter 5, 113-191 (1991); Bundgaard, et al., Journal of Drug Deliver Reviews, 8:1-38 (1992); Bundgaard, J. of Pharmaceutical Sciences, 77:285 et seq. (1988); Higuchi and Stella (eds.) Prodrugs as Novel Drug Delivery Systems, American Chemical Society (1975); and Bernard Testa & Joachim Mayer, “Hydrolysis In Drug And Prodrug Metabolism: Chemistry, Biochemistry And Enzymology,” John Wiley and Sons, Ltd. (2002).
The present invention also relates to solvates of the compounds of Formula (I), for example hydrates.
This invention also encompasses pharmaceutical compositions containing, and methods of treating viral infections through administering, pharmaceutically acceptable prodrugs of compounds of the invention. For example, compounds of the invention having free amino, amido, hydroxy or carboxylic groups can be converted into prodrugs. Prodrugs include compounds wherein an amino acid residue, or a polypeptide chain of two or more (e.g., two, three or four) amino acid residues is covalently joined through an amide or ester bond to a free amino, hydroxy or carboxylic acid group of compounds of the invention. The amino acid residues include but are not limited to the 20 naturally occurring amino acids commonly designated by three letter symbols and also includes 4-hydroxyproline, hydroxylysine, demosine, isodemosine, 3-methylhistidine, norvalin, beta-alanine, gamma-aminobutyric acid, citrulline, homocysteine, homoserine, ornithine and methionine sulfone. Additional types of prodrugs are also encompassed. For instance, free carboxyl groups can be derivatized as amides or alkyl esters. Free hydroxy groups may be derivatized using groups including but not limited to hemisuccinates, phosphate esters, dimethylaminoacetates, and phosphoryloxymethyloxycarbonyls, as outlined in Advanced Drug Delivery Reviews, 1996, 19, 115. Carbamate prodrugs of hydroxy and amino groups are also included, as are carbonate prodrugs, sulfonate esters and sulfate esters of hydroxy groups. Derivatization of hydroxy groups as (acyloxy)methyl and (acyloxy)ethyl ethers wherein the acyl group may be an alkyl ester, optionally substituted with groups including but not limited to ether, amine and carboxylic acid functionalities, or where the acyl group is an amino acid ester as described above, are also encompassed. Prodrugs of this type are described in J. Med. Chem. 1996, 39, 10. Free amines can also be derivatized as amides, sulfonamides or phosphonamides. All of these prodrug moieties may incorporate groups including but not limited to ether, amine and carboxylic acid functionalities.
Pharmaceutical Compositions
The pharmaceutical compositions of the present invention comprise a therapeutically effective amount of a compound of the present invention formulated together with one or more pharmaceutically acceptable carriers or excipients.
As used herein, the term “pharmaceutically acceptable carrier or excipient” means a non-toxic, inert solid, semi-solid or liquid filler, diluent, encapsulating material or formulation auxiliary of any type. Some examples of materials which can serve as pharmaceutically acceptable carriers are sugars such as lactose, glucose and sucrose; starches such as corn starch and potato starch; cellulose and its derivatives such as sodium carboxymethyl cellulose, ethyl cellulose and cellulose acetate; powdered tragacanth; malt; gelatin; talc; excipients such as cocoa butter and suppository waxes; oils such as peanut oil, cottonseed oil, safflower oil, sesame oil, olive oil, corn oil and soybean oil; glycols such as propylene glycol; esters such as ethyl oleate and ethyl laurate; agar; buffering agents such as magnesium hydroxide and aluminum hydroxide; alginic acid; pyrogen-free water; isotonic saline; Ringer's solution; ethyl alcohol, and phosphate buffer solutions, as well as other non-toxic compatible lubricants such as sodium lauryl sulfate and magnesium stearate, as well as coloring agents, releasing agents, coating agents, sweetening, flavoring and perfuming agents, preservatives and antioxidants can also be present in the composition, according to the judgment of the formulator.
The pharmaceutical compositions of this invention may be administered orally, parenterally, by inhalation spray, topically, rectally, nasally, buccally, vaginally or via an implanted reservoir, preferably by oral administration or administration by injection. The pharmaceutical compositions of this invention may contain any conventional non-toxic pharmaceutically-acceptable carriers, adjuvants or vehicles. In some cases, the pH of the formulation may be adjusted with pharmaceutically acceptable acids, bases or buffers to enhance the stability of the formulated compound or its delivery form. The term parenteral as used herein includes subcutaneous, intracutaneous, intravenous, intramuscular, intraarticular, intraarterial, intrasynovial, intrasternal, intrathecal, intralesional and intracranial injection or infusion techniques.
Liquid dosage forms for oral administration include pharmaceutically acceptable emulsions, microemulsions, solutions, suspensions, syrups and elixirs. In addition to the active compounds, the liquid dosage forms may contain inert diluents commonly used in the art such as, for example, water or other solvents, solubilizing agents and emulsifiers such as ethyl alcohol, isopropyl alcohol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1,3-butylene glycol, dimethylformamide, oils (in particular, cottonseed, groundnut, corn, germ, olive, castor, and sesame oils), glycerol, tetrahydrofurfuryl alcohol, polyethylene glycols and fatty acid esters of sorbitan, and mixtures thereof. Besides inert diluents, the oral compositions can also include adjuvants such as wetting agents, emulsifying and suspending agents, sweetening, flavoring, and perfuming agents.
Injectable preparations, for example, sterile injectable aqueous or oleaginous suspensions, may be formulated according to the known art using suitable dispersing or wetting agents and suspending agents. The sterile injectable preparation may also be a sterile injectable solution, suspension or emulsion in a nontoxic parenterally acceptable diluent or solvent, for example, as a solution in 1,3-butanediol. Among the acceptable vehicles and solvents that may be employed are water, Ringer's solution, U.S.P. and isotonic sodium chloride solution. In addition, sterile, fixed oils are conventionally employed as a solvent or suspending medium. For this purpose any bland fixed oil can be employed including synthetic mono- or diglycerides. In addition, fatty acids such as oleic acid are used in the preparation of injectables.
The injectable formulations can be sterilized, for example, by filtration through a bacterial-retaining filter, or by incorporating sterilizing agents in the form of sterile solid compositions which can be dissolved or dispersed in sterile water or other sterile injectable medium prior to use.
In order to prolong the effect of a drug, it is often desirable to slow the absorption of the drug from subcutaneous or intramuscular injection. This may be accomplished by the use of a liquid suspension of crystalline or amorphous material with poor water solubility. The rate of absorption of the drug then depends upon its rate of dissolution, which, in turn, may depend upon crystal size and crystalline form. Alternatively, delayed absorption of a parenterally administered drug form is accomplished by dissolving or suspending the drug in an oil vehicle. Injectable depot forms are made by forming microencapsule matrices of the drug in biodegradable polymers such as polylactide-polyglycolide. Depending upon the ratio of drug to polymer and the nature of the particular polymer employed, the rate of drug release can be controlled. Examples of other biodegradable polymers include poly(orthoesters) and poly(anhydrides). Depot injectable formulations are also prepared by entrapping the drug in liposomes or microemulsions that are compatible with body tissues.
Compositions for rectal or vaginal administration are preferably suppositories which can be prepared by mixing the compounds of this invention with suitable non-irritating excipients or carriers such as cocoa butter, polyethylene glycol or a suppository wax which are solid at ambient temperature but liquid at body temperature and therefore melt in the rectum or vaginal cavity and release the active compound.
Solid dosage forms for oral administration include capsules, tablets, pills, powders, and granules. In such solid dosage forms, the active compound is mixed with at least one inert, pharmaceutically acceptable excipient or carrier such as sodium citrate or dicalcium phosphate and/or: a) fillers or extenders such as starches, lactose, sucrose, glucose, mannitol, and silicic acid, b) binders such as, for example, carboxymethylcellulose, alginates, gelatin, polyvinylpyrrolidinone, sucrose, and acacia, c) humectants such as glycerol, d) disintegrating agents such as agar-agar, calcium carbonate, potato or tapioca starch, alginic acid, certain silicates, and sodium carbonate, e) solution retarding agents such as paraffin, f) absorption accelerators such as quaternary ammonium compounds, g) wetting agents such as, for example, cetyl alcohol and glycerol monostearate, h) absorbents such as kaolin and bentonite clay, and i) lubricants such as talc, calcium stearate, magnesium stearate, solid polyethylene glycols, sodium lauryl sulfate, and mixtures thereof. In the case of capsules, tablets and pills, the dosage form may also comprise buffering agents.
Solid compositions of a similar type may also be employed as fillers in soft and hard-filled gelatin capsules using such excipients as lactose or milk sugar as well as high molecular weight polyethylene glycols and the like.
The active compounds can also be in micro-encapsulated form with one or more excipients as noted above. The solid dosage forms of tablets, dragees, capsules, pills, and granules can be prepared with coatings and shells such as enteric coatings and other coatings well known in the pharmaceutical formulating art. They may optionally contain opacifying agents and can also be of a composition that they release the active ingredient(s) only, or preferentially, in a certain part of the intestinal tract, optionally, in a delayed manner. Examples of embedding compositions that can be used include polymeric substances and waxes.
Dosage forms for topical or transdermal administration of a compound of this invention include ointments, pastes, creams, lotions, gels, powders, solutions, sprays, inhalants or patches. The active component is admixed under sterile conditions with a pharmaceutically acceptable carrier and any needed preservatives or buffers as may be required. Ophthalmic formulation, ear drops, eye ointments, powders and solutions are also contemplated as being within the scope of this invention.
The ointments, pastes, creams and gels may contain, in addition to an active compound of this invention, excipients such as animal and vegetable fats, oils, waxes, paraffins, starch, tragacanth, cellulose derivatives, polyethylene glycols, silicones, bentonites, silicic acid, talc and zinc oxide, or mixtures thereof.
Powders and sprays can contain, in addition to the compounds of this invention, excipients such as lactose, talc, silicic acid, aluminum hydroxide, calcium silicates and polyamide powder, or mixtures of these substances. Sprays can additionally contain customary propellants such as chlorofluorohydrocarbons.
Transdermal patches have the added advantage of providing controlled delivery of a compound to the body. Such dosage forms can be made by dissolving or dispensing the compound in the proper medium. Absorption enhancers can also be used to increase the flux of the compound across the skin. The rate can be controlled by either providing a rate controlling membrane or by dispersing the compound in a polymer matrix or gel.
For pulmonary delivery, a therapeutic composition of the invention is formulated and administered to the patient in solid or liquid particulate form by direct administration e.g., inhalation into the respiratory system. Solid or liquid particulate forms of the active compound prepared for practicing the present invention include particles of respirable size: that is, particles of a size sufficiently small to pass through the mouth and larynx upon inhalation and into the bronchi and alveoli of the lungs. Delivery of aerosolized therapeutics, particularly aerosolized antibiotics, is known in the art (see, for example U.S. Pat. No. 5,767,068 to VanDevanter et al., U.S. Pat. No. 5,508,269 to Smith et al. and WO 98/43650 by Montgomery, all of which are incorporated herein by reference). A discussion of pulmonary delivery of antibiotics is also found in U.S. Pat. No. 6,014,969 incorporated herein by reference.
Antiviral Activity
An inhibitory amount or dose of the compounds of the present invention may range from about 0.01 mg/Kg to about 500 mg/Kg, alternatively from about 0.1 to about 50 mg/Kg. Inhibitory amounts or doses will also vary depending on route of administration, as well as the possibility of co-usage with other agents.
According to the methods of treatment of the present invention, viral infections are treated or prevented in a patient such as a human or another animal by administering to the subject a therapeutically effective amount of a compound of the invention, in such amounts and for such time as is necessary to achieve the desired result. An additional method of the present invention is the treatment of biological samples with an inhibitory amount of a compound of composition of the present invention in such amounts and for such time as is necessary to achieve the desired result.
The term “therapeutically effective amount” of a compound of the invention, as used herein, means an amount of the compound which confers a therapeutic effect on the treated subject, at a reasonable benefit/risk ratio applicable to any medical treatment. The therapeutic effect may be objective (i.e., measurable by some test or marker) or subjective (i.e., subject gives an indication of or feels an effect).
The term “inhibitory amount” of a compound of the present invention means a sufficient amount to decrease the viral load in a biological sample or a subject (e.g., resulting in at least 10%, preferably at least 50%, more preferably at least 80%, and most preferably at least 90% or 95%, reduction in viral load). It is understood that when said inhibitory amount of a compound of the present invention is administered to a subject it will be at a reasonable benefit/risk ratio applicable to any medical treatment as determined by a physician. The term “biological sample(s),” as used herein, means a substance of biological origin intended for administration to a subject. Examples of biological samples include, but are not limited to, blood and components thereof such as plasma, platelets, subpopulations of blood cells and the like; organs such as kidney, liver, heart, lung, and the like; sperm and ova; bone marrow and components thereof; or stem cells. Thus, another embodiment of the present invention is a method of treating a biological sample by contacting said biological sample with an inhibitory amount of a compound or pharmaceutical composition of the present invention.
Effective doses will also vary depending on route of administration, as well as the possibility of co-usage with other agents. It will be understood, however, that the total daily usage of the compounds and compositions of the present invention will be decided by the attending physician within the scope of sound medical judgment. The specific therapeutically effective dose level for any particular patient will depend upon a variety of factors including the disorder being treated and the severity of the disorder; the activity of the specific compound employed; the specific composition employed; the age, body weight, general health, sex and diet of the patient; the time of administration, route of administration, and rate of excretion of the specific compound employed; the duration of the treatment; drugs used in combination or contemporaneously with the specific compound employed; and like factors well known in the medical arts.
The total daily dose of the compounds of this invention administered to a human or other animal in single or in divided doses can be in amounts, for example, from 0.01 to 50 mg/kg body weight or more usually from 0.1 to 25 mg/kg body weight. Single dose compositions may contain such amounts or submultiples thereof to make up the daily dose. In general, treatment regimens according to the present invention comprise administration to a patient in need of such treatment from about 10 mg to about 1000 mg of the compound(s) of this invention per day in single or multiple doses.
Upon improvement of a patient's condition, a maintenance dose of a compound, composition or combination of this invention may be administered, if necessary. Subsequently, the dosage or frequency of administration, or both, may be reduced, as a function of the symptoms, to a level at which the improved condition is retained when the symptoms have been alleviated to the desired level. Patients may, however, require intermittent treatment on a long-term basis upon any recurrence of disease symptoms.
When the compositions of this invention comprise a combination of a compound of the Formula (I) described herein and one or more additional therapeutic or prophylactic agents, both the compound and the additional agent should be present at dosage levels of between about 1 to 100%, and more preferably between about 5 to 95% of the dosage normally administered in a monotherapy regimen. The additional agents may be administered separately, as part of a multiple dose regimen, from the compounds of this invention. Alternatively, those agents may be part of a single dosage form, mixed together with the compounds of this invention in a single composition.
The said “additional therapeutic or prophylactic agents” includes but not limited to, immune therapies (eg. interferon), therapeutic vaccines, antifibrotic agents, anti-inflammatory agents such as corticosteroids or NSAIDs, bronchodilators such as beta-2 adrenergic agonists and xanthines (e.g. theophylline), mucolytic agents, anti-muscarinics, anti-leukotrienes, inhibitors of cell adhesion (e.g. ICAM antagonists), anti-oxidants (eg N-acetylcysteine), cytokine agonists, cytokine antagonists, lung surfactants and/or antimicrobial and anti-viral agents (eg ribavirin and amantadine). The compositions according to the invention may also be used in combination with gene replacement therapy.
Combination and Alternation Therapy for HCV
It has been recognized that drug-resistant variants of HCV can emerge after prolonged treatment with an antiviral agent. Drug resistance most typically occurs by mutation of a gene that encodes for a protein such as an enzyme used in viral replication, and most typically in the case of HCV, RNA polymerase, protease, or helicase.
Recently, it has been demonstrated that the efficacy of a drug against a viral infection, such as HIV, can be prolonged, augmented, or restored by administering the drug in combination or alternation with a second, and perhaps third, antiviral compound that induces a different mutation from that caused by the principal drug. Alternatively, the pharmacokinetics, biodistribution, or other parameter of the drug can be altered by such combination or alternation therapy. In general, combination therapy is typically preferred over alternation therapy because it induces multiple simultaneous stresses on the virus.
A compound of the present invention can also be administered in combination or alternation with antiviral agent. Exemplary antiviral agents include ribavarin, interferon, interleukin or a stabilized prodrug of any of them. More broadly described, the compound can be administered in combination or alternation with any of the anti-HCV drugs listed in Table 10 below.







TABLE 10







Table of anti-Hepatitis C Compounds in Current Clinical Development









Drug name
Drug category
Pharmaceutical Company





PEGASYS
Long acting interferon
Roche


pegylated interferon


alfa-2a


INFERGEN
Long acting interferon
InterMune


interferon alfacon-1


OMNIFERON
Long acting interferon
Viragen


natural interferon


ALBUFERON
Long acting interferon
Human Genome Sciences


REBIF interferon beta-la
Interferon
Ares-Serono


Omega Interferon
Interferon
BioMedicine


Oral Interferon alpha
Oral Interferon
Amarillo Biosciences


Interferon gamma-lb
Anti-fibrotic
InterMune


IP-501
Anti-fibrotic
InterMune


Merimebodib VX-497
IMPDH inhibitor
Vertex



(inosine monophosphate



dehydrogenase)


AMANTADINE (Symmetrel)
Broad Antiviral Agent
Endo Labs Solvay


IDN-6556
Apotosis regulation
Idun Pharma.


XTL-002
Monclonal Antibody
XTL


HCV/MF59
Vaccine
Chiron


CIVACIR
Polyclonal Antibody
NABI Innogenetics



Therapeutic vaccine


VIRAMIDINE
Nucleoside Analogue
ICN


ZADAXIN (thymosin alfa-1)
Immunomodulator
Sci Clone


CEPLENE (histamine)
Immunomodulator
Maxim


VX 950/LY 570310
Protease inhibitor
Vertex/Eli Lilly


ISIS 14803
Antisense
Isis Pharmaceutical/Elan


IDN-6556
Caspase inhibitor
Idun Pharmaceuticals


JTK 003
Polymerase Inhibitor
AKROS Pharma


Tarvacin
Anti-Phospholipid Therapy
Peregrine


HCV-796
Polymerase Inhibitor
ViroPharma/Wyeth


CH-6
Protease inhibitor
Schering


ANA971
Isatoribine
ANADYS


ANA245
Isatoribine
ANADYS


CPG 10101 (Actilon)
Immunomodulator
Coley


Rituximab (Rituxam)
Anti-CD2O
Genetech/IDEC



Monoclonal Antibody


NM283 (Valopicitabine)
Polymerase Inhibitor
Idenix Pharmaceuticals


HepX TM-C
Monoclonal Antibody
XTL


IC41
Therapeutic Vaccine
Intercell


Medusa Interferon
Longer acting interferon
Flamel Technology


E-1
Therapeutic Vaccine
Innogenetics


Multiferon
Long Acting Interferon
Viragen


BILN 2061
Protease inhibitor
Boehringer-Ingelheim


TMC435350
Protease inhibitor
Tibotec/Medivir


Telaprevir (VX-950)
Protease inhibitor
Vertex


Boceprevir (SCH 503034)
Protease inhibitor
Schering-Plough


ACH-1625
Protease inhibitor
Achillion


ABT-450
Protease inhibitor
Abbott/Enanta


BI-201335
Protease inhibitor
Boehringer-Ingelheim


PHX-1766
Protease inhibitor
Phenomix


VX-500
Protease inhibitor
Vertex


MK-7009
protease inhibitor
Merck


R7227 (ITMN-191)
protease inhibitor
InterMune


Narlaprevir (SCH 900518)
Protease inhibitor
Schering/Merck


Alinia (nitazoxanide)
To be determined
Romark


ABT-072
Polymerase Inhibitor
Abbott


ABT-333
Polymerase Inhibitor
Abbott


Filibuvir (PF-00868554)
Polymerase Inhibitor
Pfizer


VCH-916
Polymerase Inhibitor
Vertex


R7128 (PSI6130)
Polymerase Inhibitor
Roche/Pharmasset


IDX184
Polymerase Inhibitor
Idenix


IDX-189
Polymerase Inhibitor
Inhibitex


PSI-7977
Polymerase Inhibitor
Pharmasset


PSI-938
Polymerase Inhibitor
Pharmasset


R1626
Polymerase inhibitor
Roche


MK-3281
Polymerase inhibitor
Merck


PSI-7851
Polymerase inhibitor
Pharmasset


ANA598
Polymerase inhibitor
Anadys Pharmaceuticals


BI-207127
Polymerase inhibitor
Boehringer-Ingelheim


GS-9190
Polymerase inhibitor
Gilead


VCH-759
Polymerase Inhibitor
Vertex


Clemizole
NS4B inhibitor
Eiger Biopharmaceuticals


A-832
NS5A inhibitor
ArrowTherapeutics


BMS-790052
NS5A inhibitor
Bristol-Myers-Squibb


BMS-824393
NS5A inhibitor
Bristol-Myers-Squibb


GS-5885
NS5A inhibitor
Gilead


ITX5061
Entry inhibitor
iTherx


GS-9450
Caspase inhibitor
Gilead


ANA773
TLR agonist
Anadys


CYT107
immunomodulator
Cytheris


SPC3649 (LNA-antimiR TM-122)
microRNA
Santaris Pharma


Debio 025
Cyclophilin inhibitor
Debiopharm


SCY-635
Cyclophilin inhibitor
Scynexis









Unless otherwise defined, all technical and scientific terms used herein are accorded the meaning commonly known to one of ordinary skill in the art. All publications, patents, published patent applications, and other references mentioned herein are hereby incorporated by reference in their entirety.
ABBREVIATIONS
Abbreviations which may be used in the descriptions of the scheme and the examples that follow are: Ac for acetyl; AcOH for acetic acid; AIBN for azobisisobutyronitrile; BINAP for 2,2′-bis(diphenylphosphino)-1,1′-binaphthyl; Boc2O for di-tert-butyl-dicarbonate; Boc for t-butoxycarbonyl; Bpoc for 1-methyl-1-(4-biphenylyl)ethyl carbonyl; BtOH for 1-hydroxy-benzotriazole; Bz for benzoyl; Bn for benzyl; BocNHOH for tert-butyl N-hydroxycarbamate; t-BuOK for potassium tert-butoxide; Bu3SnH for tributyltin hydride; BOP for (benzotriazol-1-yloxy)tris(dimethylamino)phos-phonium Hexafluorophosphate; Brine for sodium chloride solution in water; Cbz for carbobenzyloxy; CDI for carbonyldiimidazole; CH2Cl2 for dichloromethane; CH3 for methyl; CH3CN for acetonitrile; Cs2CO3 for cesium carbonate; CuCl for copper (I) chloride; CuI for copper (I) iodide; dba for dibenzylidene acetone; dppb for diphenylphosphino butane; DBU for 1,8-diazabicyclo[5.4.0]undec-7-ene; DCC for N,N′-dicyclohexylcarbodiimide; DEAD for diethylazodicarboxylate; DIAD for diisopropyl azodicarboxylate; DIPEA or (i-Pr)2EtN for N,N-diisopropylethyl amine; Dess-Martin periodinane for 1,1,1-tris(acetyloxy)-1,1-dihydro-1,2-benziodoxol-3-(1H)-one; DMAP for 4-dimethylaminopyridine; DME for 1,2-dimethoxy-ethane; DMF for N,N-dimethylformamide; DMSO for dimethyl sulfoxide; DMT for di(p-methoxyphenyl)phenylmethyl or dimethoxytrityl; DPPA for diphenylphosphoryl azide; EDC for N-(3-dimethylaminopropyl)-N′-ethylcarbodiimide; EDC HCl for N-(3-dimethylamino-propyl)-N′-ethylcarbodiimide hydrochloride; EtOAc for ethyl acetate; EtOH for ethanol; Et2O for diethyl ether; Fmoc for 9-fluorenylmethoxycarbonyl; HATU for O-(7-azabenzotriazol-1-yl)-N,N,N′,N′,-tetramethyluronium Hexafluorophosphate; HCl for hydrogen chloride; HOBT for 1-hydroxybenzotriazole; K2CO3 for potassium carbonate; n-BuLi for n-butyl lithium; i-BuLi for i-butyl lithium; t-BuLi for t-butyl lithium; PhLi for phenyl lithium; LDA for lithium diisopropylamide; LiTMP for lithium 2,2,6,6-tetramethylpiperidinate; MeOH for methanol; Mg for magnesium; MOM for methoxymethyl; Ms for mesyl or —SO2—CH3; Ms2O for methanesulfonic anhydride or mesyl-anhydride; NaBH4 for sodium borohydride; NaBH3CN for sodium cyanoborohydride; NaN(TMS)2 for sodium bis(trimethylsilyl)amide; NaCl for sodium chloride; NaH for sodium hydride; NaHCO3 for sodium bicarbonate or sodium hydrogen carbonate; Na2CO3 sodium carbonate; NaOH for sodium hydroxide; Na2SO4 for sodium sulfate; NaHSO3 for sodium bisulfite or sodium hydrogen sulfite; Na2S2O3 for sodium thiosulfate; NH2NH2 for hydrazine; NH4HCO3 for ammonium bicarbonate; NH4Cl for ammonium chloride; NMMO for N-methylmorpholine N-oxide; NaIO4 for sodium periodate; Ni for nickel; OH for hydroxyl; OsO4 for osmium tetroxide; Pd for palladium; Ph for phenyl; PMB for p-methoxybenzyl; POPd for dihydrogen dichlorobis(di-tert-butylphosphinito-κP)palladate(II); Pd2(dba)3 for tris(dibenzylidene-acetone) dipalladium (0); Pd(PPh3)4 for tetrakis(triphenylphosphine)palladium (0); PdCl2(PPh3)2 for trans-dichlorobis(triphenyl-phosphine)palladium (II); Pt for platinum; Rh for rhodium; rt for room temperature; Ru for ruthenium; SEM for (trimethylsilyl)ethoxymethyl; TBAF for tetrabutylammonium fluoride; TBS for tent-butyl dimethylsilyl; TEA or Et3N for triethylamine; Teoc for 2-trimethylsilyl-ethoxy-carbonyl; TFA for trifluoroacetic acid; THF for tetrahydrofuran; TMEDA for N,N,N′,N′-tetramethylethylenediamine; TPP or PPh3 for triphenyl-phosphine; Troc for 2,2,2-trichloroethyl carbonyl; Ts for tosyl or —SO2—C6H4—CH3; Ts2O for tolylsulfonic anhydride or tosyl-anhydride; TsOH for p-tolylsulfonic acid; TMS for trimethylsilyl; or TMSCl for trimethylsilyl chloride.
Synthetic Methods
The compounds and processes of the present invention will be better understood in connection with the following synthetic schemes that illustrate the methods by which the compounds of the invention may be prepared. Starting materials can be obtained from commercial sources or prepared by well-established literature methods known to those of ordinary skill in the art. It will be readily apparent to one of ordinary skill in the art that the compounds defined above can be synthesized by substitution of the appropriate reactants and agents in the syntheses shown below. It will also be readily apparent to one skilled in the art that the selective protection and deprotection steps, as well as the order of the steps themselves, can be carried out in varying order, depending on the nature of the variables to successfully complete the syntheses below. The variables are as defined above unless otherwise noted below.
The compounds of the present invention may be prepared via several different synthetic routes from a variety of 5/6-membered ring fused heteroaryl, 5-membered ring heteroaryl, and related intermediates. An exemplary method is shown in Schemes 1, 2, 3, and 4. A retro-synthesis of those title compounds include direct formation of a suitably linked core structure (5/6-membered ring fused heteroaryl or 5-membered ring heteroaryl) followed by attachment of a suitable capping group (such as —C(O)R6), plus some functional group manipulations in between and/or after. Various 5/6-membered ring fused heteroaryl or 5-membered ring heteroaryl intermediates are known to those skilled in the art, for example see the encyclopedic volumes edited by A. R. Katrizky, et al, “Comprehensive Heterocyclic Chemistry” 1984; “Comprehensive Heterocyclic Chemistry II” 1996; “Comprehensive Heterocyclic Chemistry III” 2008.
A general synthesis and further elaboration of some 6-membered ring fused with imidazole related intermediates are summarized in Scheme 1, in which Z is N or CH.
The synthesis starts from the construction of an optionally substituted imidazopyridine or benzimidazole 1-2, which may be obtained by condensation of an amino acid or its derivatives 1-1.1 or 1-1.2 with 2,3-diaminopyridine or 1,2-diaminobenzene 1-1 under the conditions to those skilled in the art. The imidazole ring closure may be realized either in one pot by heat, optionally in the presence of an acid and/or with a dehydration reagent such as polyphosphoric acid; or in two steps: 1) amide formation between diamine 1-1 and amino acid 1-1.1 or 1-1.2 in the presence of a condensation reagent such as EDC.HCl, DCC or the like; or through mixed anhydride approach by reacting acid 1-1.1 or 1-1.2 with a chloroformate such as methyl chloroformate, isobutyl chloroformate, or the like, in the presence of a base such as TEA, DIPEA, DMAP, N-methylmorpholine, or the like, followed by treating the mixed anhydride with diamine 1-1; and 2) the heterocyclic ring closure in the presence of an acid such as acetic acid, sulfuric acid or the like or a dehydration reagent such as HATU or the like, optionally with heat.




The imidazopyridine or benzimidazole 1-2 may be subjected to Suzuki, Stille or related coupling conditions known to those skilled in the art (see reviews: A. Suzuki, Pure Applied Chem. 1991, 63, 419; A. Suzuki, Handbook of Organopalladium Chemistry for Organic Synthesis 2002, 1, 249; A. Anastasia, et al, Handbook of Organopalladium Chemistry for Organic Synthesis 2002, 1, 311; F. Bellina, et al, Synthesis 2004, 2419; M. G. Organ, et al, Synthesis 2008, 2776; A. T. Lindhardt, et al, Chem.—A European J. 2008, 14, 8756; E. A. B. Kantchev, et al, Angew. Chem. Int. Ed. 2007, 46, 2768; V. Farina, et al, Advances in Metal-Organic Chem. 1996, 5, 1) with different coupling partners to provide a variety of key intermediates. For example, Sonogashira coupling between bromide 1-2 and trimethylsilylacetylene can generate alkyne 1-3 after removal of TMS by K2CO3 in MeOH. Alternatively, bromide 1-2 may be coupled with tributylvinylstanne through Stille reaction conditions known to those skilled in the art to provide alkene 1-4. Analogously, a key allyl intermediates 1-5 may be prepared by Stille reaction from bromide 1-2 with an allylstanne such as allyltributylstanne.
Alternatively, bromide 1-2 may be converted to key intermediate 1-7 by selectively reacting with metallic reagent 1-2.2 under the Suzuki or Stille conditions which are known to those skilled in the art. Yet alternatively, intermediate 1-7 may be prepared by treating bromide 1-2 with dimetallic agent 1-2.1 to afford organometallic 1-6, followed by coupling with bromoiodoaryl compound 1-6.1, both may be under the previously described Suzuki or Stille reaction conditions. The bromide 1-7 may be further converted to organometallic 1-8 with dimetallic agent 1-2.1 using the conditions described above to prepare 1-6.
It should be noted that optionally the NH group of all the imidazopyridine or benzimidazole related intermediates listed above may be protected with an amino protecting group, such as SEM (i.e. SEM-Cl, NaH), Boc, Cbz, Teoc, Troc, or the like.




A typical synthesis of imidazole related intermediates are analogous to that of the imidazopyridine or benzimidazole intermediates. As shown in Scheme 2, bromo-imidazole 2-2 can be synthesized by condensation of amino acid derived aldehyde 2-1.1 or 2-1.2 and glyoxal in the presence of methanolic ammonia; followed by bromination of the imidazole ring under the conditions which are known to those skilled in the art. The bromination of the imidazole ring may be realized either in one pot by NBS, bromine, 2,4,4,6-tetrabromo-2,5-cyclohexadienone, or the like; or in two steps: 1) dibromide formation in the presence of excess bromination reagent such as NBS, bromine, 2,4,4,6-tetrabromo-2,5-cyclohexadienone, or the like, optionally with heat; and 2) reduction of the dibromide to monobromide in the presence of a reducing reagent such as NaHSO3, Na2S2O3, Na2SO3, or the like. Bromide 2-2 then may be served as a common intermediate further elaborable to many other imidazole derivatives using the chemistry discussed in Scheme 1. For example, bromide 2-2 may be coupled with allytin or vinyltin or TMS-acetylene to provide intermediate 2-6. Also, bromide 2-2 may be converted to key intermediate 2-4 by selectively reacting with metallic reagent 2-2.1 under the Suzuki or Stille conditions to provide key intermediate 2-4. Yet alternatively, intermediate 2-4 may be prepared by treating bromide 2-2 with dimetallic agent 2-2.2 to afford organometallic 2-5, followed by coupling with bromoiodoaryl compound 2-5.1, both may be under the previously described Suzuki or Stille reaction conditions. In turn, the bromide 2-4 may be further converted to organometallic 2-7 with dimetallic agent 2-4.1 using the conditions described above for the preparation of intermediate 2-5.
Yet alternatively, aryl or heteroaryl bromide 2-4 may also be derived from bromoketone 2-9, which can be prepared from the corresponding ketone 2-8 in the presence of a bromination reagent such as NBS, bromine, or the like, optionally in the presence of an acid and/or with heating. Bromoketone 2-9 may be either converted to the corresponding amine 2-11, or coupled with protected amino acid 1-1.1 or 1-1.2 in the presence of a base such as Et3N or DIPEA to afford keto-ester 2-10. Similarly, amine 2-11 may be converted to the corresponding keto-amide 2-12 via condensation with appropriate amino acid under standard amide formation conditions. Both 2-12 and 2-13 may be transformed to key intermediate 2-4 via heating with NH4OAc under thermal or microwave conditions.




As shown in Scheme 3, a compound 3-1 containing a hydroxy group substituted at the C4-position of the pyrrolidine ring may be illustrated by intermediates 1-2, 1-3, 1-4, 1-5, 1-7, 2-2, 2-4, and 2-6 when U is CH(OH) as shown in Schemes 1-2. Oxidation of 3-1 by a variety of oxidation agents such as Dess-Martin periodinane optionally in the presence of an acid such as acetic acid or camphorsulfonic acid to afford the ketone 3-2. More reagents and conditions for the oxidation of an alcohol to a ketone can be found in Comprehensive Organic transformations R. C. Larock Ed., Wiley-RCH, 1999, page 1236-1249. 3-2 may then serve as a universal intermediate for further derivatization to olefin 3-3, oxime 3-4 and hydrazone 3-5. The olefination of 3-2 may be realized by various types of Wittig Reaction or Peterson Reaction, a more detailed reagents and conditions can be found in Comprehensive Organic transformations R. C. Larock Ed., Wiley-RCH, 1999, page 327-350.
With a variety of suitably substituted imidazopyridines, benzimidazoles and imidazoles such as those listed in Schemes 1-3 in hand, the compounds of the present invention may be prepared through various coupling strategy or a combination of strategies to connect two fragments, optionally with a suitable cyclic or acyclic linker or formation of a cyclic or acyclic linker. The said strategy may include, but not limited to, Stille coupling, Suzuki coupling, Sonogashira coupling, Heck coupling, Buchwald amidation, Buchwald amination, amide coupling, ester bond formation, William etherification, Buchwald etherification, alkylation, pericyclic reaction with different variations, or the like.




An example of the strategies that may be used to prepare the compounds of the present invention is shown in Scheme 4. Both iodide 4-1.1 and alkyne 4-1.2 can be prepared using similar procedures described in Schemes 1-3. Iodide 4-1.1 can be coupled with benzimidazolylacetylene 4-1.2 under Sonogashira condition in the presence of Pd and Cu-catalysts to generate a core structure 4-2. Compound 4-2 then may be served as a common intermediate for further derivatizations to the title compounds I-1 in two steps: 1) deprotection of Boc groups under hydrolytic conditions in the presence of an acid such as TFA or hydrogen chloride; and 2) the released amine functionality may be acylated with a carboxylic acid under standard acylation conditions, for example a coupling reagent such as HATU in combination with an organic base such as DIPEA can be used in this regard. Various carboxylic acids including amino acids in racemic or optical form are commercially available, and/or can be synthesized in racemic or optical form, see references cited in reviews by D. Seebach, et al, Synthesis 2009, 1; C. Cativiela and M. D. Diaz-de-Villegas, Tetrahedron: Asymmetry 2007, 18, 569; 2000, 11, 645; and 1998, 9, 3517; and experimental examples compiled in patent application WO 08/021,927A2 by C. Bachand, et al, from BMS, which is incorporated herein by reference.
Alternatively, as shown in Scheme 4a, the compounds of the present invention (for example I-1) may also be derived from key intermediates 4-1.1a and 4-1.2a using the Sonogashira coupling procedures described previously. The intermediates 4-1.1a and 4-1.2a have the desired acyl groups already installed from 4-1.1 and 4-1.2 using similar sequences shown in Scheme 4.




Yet alternatively, as shown in Scheme 4b, the compounds of the present invention (for example I-1) may also be derived from key intermediate 4-2b after Sonogashira coupling of 4-1.1 and 4-1.2b using the procedures described previously. The alcohol in 4-2b may be oxidized and olefinated to exocyclic double bond compound 4-3b using the procedures described in Scheme 3. 4-3b can then be further converted to I-1 using similar sequences shown in Scheme 4.




The compounds of the present invention containing five-membered heteroaryl other than imidazole may be prepared using similar procedures described above in Schemes 1-4 and 4a. For example, some intermediates containing a desired, suitably substituted five-membered heteroaryl have been published in US 2008/0311075A1 by C. Bachand, et al from BMS, which is incorporated by reference. These intermediates are compiled in the following Table 11.







TABLE 11













 








 








 








 








 








 








 








 















The synthesis of the compounds of the present invention involves 5/6-membered fused heteroaryl intermediates other than benzimidazoles, various 5/6-membered fused heteroaryl are known in the literature. The synthesis of other 5/6-membered fused heteroaryl intermediates depends on the chemical features of each structure. For example, a typical synthesis of indole intermediate is illustrated in Scheme 5. The commercially available bromoiodoaniline 5-1 may be coupled to the commercially available acetylene 5-1.1 under the Sonogashira conditions to give phenylacetylene 5-2. The latter may be cyclized to indole 5-3 under heat or microwave condition in the presence of a copper catalyst.




It will be appreciated that, with appropriate manipulation and protection of any chemical functionality, synthesis of compounds of Formula (I) is accomplished by methods analogous to those above and to those described in the Experimental section. Suitable protecting groups can be found, but are not restricted to, those found in T W Greene and P G M Wuts “Protective Groups in Organic Synthesis”, 3rd Ed (1999), J Wiley and Sons.
In certain aspects, the invention is directed to a method of preparing a compound of Formula (I) comprising the steps of:

    
    
        i) preparing a compound of Formula (II):
    
    






via a transition-metal catalyzed cross-coupling reaction;
wherein:
G, J, U, X, R1, R3, R4, R5, R7a and R7b are as previously defined in Formula (I);
Ring A1 is absent, optionally substituted aryl or optionally substituted heteroaryl;
Ring B1 is optionally substituted aryl or optionally substituted heteroaryl;
L1 is absent, optionally substituted C2-C4 alkenyl or C2-C4 alkynyl; and
Za and Zb are each independently an amino protecting group or —C(O)—R6;

    
    
        ii) when Za or Zb is an amino protecting group, fully or selectively deprotecting a compound of Formula (II) to give the corresponding amine of Formula (III):
    
    






wherein Z′ is hydrogen, an amino protecting group or —C(O)—R6;

    
    
        iii) capping the released amino group of a compound of Formula (III) with LG-C(O)—R6, wherein LG is a leaving group; to give the compound of Formula (IV):
    
    






wherein Zd is an amino protecting group —C(O)—R6; and

    
    
        iv) repeated reaction sequence of deprotecting and capping (step ii-iii) to give the compound of Formula (V):
    
    






All references cited herein, whether in print, electronic, computer readable storage media or other form, are expressly incorporated by reference in their entirety, including but not limited to, abstracts, articles, journals, publications, texts, treatises, internet web sites, databases, patents, and patent publications.
EXAMPLES
The compounds and processes of the present invention will be better understood in connection with the following examples, which are intended as an illustration only and not limiting of the scope of the invention. Various changes and modifications to the disclosed embodiments will be apparent to those skilled in the art and such changes and modifications including, without limitation, those relating to the chemical structures, substituents, derivatives, formulations and/or methods of the invention may be made without departing from the spirit of the invention and the scope of the appended claims.
Although the invention has been described with respect to various preferred embodiments, it is not intended to be limited thereto, but rather those skilled in the art will recognize that variations and modifications may be made therein which are within the spirit of the invention and the scope of the appended claims.
Example 1




Step 1a.

Into a mixture of 2-bromo-1-(4-iodophenyl)ethanone (5.00 g, 15.4 mmol) and N-Boc-L-proline (3.48 g, 16.1 mmol) in acetonitrile (40 mL) was added diisopropylethylamine (2.4 mL, 17 mmol). The resulting mixture was stirred at rt for 3 hours before being partitioned between EtOAc and aqueous NaHCO3. The organic phase was separated, dried (Na2SO4) and concentrated to afford a brown oil. It was purified by flash column chromatography (silica, hexane-EtOAc) to give the desired product as a light yellow oil (6.00 g, 86%). ESIMS m/z=481.94 [M+Na]+.
Step 1b.
The mixture of compound from step 1a (6.00 g, 12.5 mmol) and ammonium acetate (15.1 g, 196 mmol) in toluene (80 mL) was stirred at 80° C. for 3 hours before being partitioned between EtOAc and aqueous NaHCO3. The organic phase was separated, dried (Na2SO4) and concentrated to afford a deep red oil. It was purified by flash column chromatography (silica, hexane-EtOAc) to give the desired product as a light yellow solid (5.34 g, 93%). ESIMS m/z=439.83 [M+H]+.
Step 1c.
A mixture of N-Boc-L-4-hydroxyproline (6.182 g, 26.73 mmol) and 4-bromo-1,2-diaminobenzene (5.000 g, 26.73 mmol) in DMF (60 mL) was treated with EDC.HCl (6.662 g, 34.75 mmol) and DMAP (0.327 g, 2.673 mmol) at rt. The mixture was stirred at rt overnight before being partitioned (EtOAc—H2O). The organic layer was extracted with dichloromethane. The combined organic layers were washed with brine (*3), dried (Na2SO4), filtered and evaporated to give the crude desired compound as a dark liquid (14.05 g), which was used directly for next step.
Step 1d.
A solution of the crude compound from step 1c (14.05 g, 26.73 mmol at most) in glacial acetic acid (50 mL) was heated at 50° C. for 2 hours. The volatiles were evaporated off. Et3N (5 mL) was added and the mixture was evaporated again. The residue was purified by chromatography (silica, hexanes-ethyl acetate, with 10% MeOH and 1% Et3N in ethyl acetate) to give the desired compound as a brown solid (7.29 g, 71% over 2 steps). ESIMS m/z=382.06, 384.06 [M+H]+.
Step 1e.
A mixture of the compound from step 1d (0.500 g, 1.308 mmol), trimethylsilyl-acetylene (1.85 ml, 13.08 mmol), CuI (7.5 mg, 0.039 mmol) and Pd(PPh3)4 (75.6 mg, 0.065 mmol) in Et3N (13 mL) and THF (6 mL) was degased and then heated at 90° C. under N2 overnight before being evaporated. The residue was purified by chromatography (silica, hexanes-ethyl acetate, with 10% MeOH and 1% Et3N in ethyl acetate) to give the desired compound as a yellow solid (0.400 g, 79%). ESIMS m/z=400.21 [M+H]+.
Step 1f.
A suspension of the compound from step 1e (0.400 g, 1.002 mmol) and K2CO3 (0.346 g, 2.504 mmol) in methanol (10 mL) was stirred at rt for 2 hour. The volatiles were evaporated off. The residue was partitioned (EtOAc—H2O). The organic layer was washed with brine, dried (Na2SO4), filtered and concentrated. The residue was purified by chromatography (silica, hexanes-ethyl acetate, with 10% MeOH and 1% Et3N in ethyl acetate) to give the desired compound as a yellow foam (0.237 g, 72%). ESIMS m/z=328.09 [M+H]+.
Step 1g.
To a mixture of the compounds from step 1b (0.122 g, 0.278 mmol) and step 1f (0.100 g, 0.305 mmol) in THF (4.5 mL) and triethylamine (1.5 mL) was added tetrakis(triphenylphosphine)palladium(0) (16.0 mg, 0.014 mmol) and copper(I) iodide (1.6 mg, 8.3 μmol). The resulting mixture was degassed and then stirred at room temperature for 1 hours, at 35° C. for 15 hours and at 55° C. for 3 hours before being evaporated. The residue was purified by flash column chromatography (silica, hexanes-ethyl acetate, with 20% MeOH and 1% Et3N in ethyl acetate) to give the desired compound as a yellow solid (0.102 g, 57%). ESIMS m/z=639.36 [M+H]+.
Step 1h.
To a solution of the compound from step 1g (40.0 mg, 62.6 μmol) in CH2Cl2 (4 mL) were added camphorsulfonic acid (29.1 mg, 0.125 mmol) and Dess-Martin periodinane (0.265 g, 0.626 mmol). The mixture was stirred at rt for 3 hours before being quenched with aqueous Na2S2O3 solution. The volatiles were evaporated and the residue was partitioned (EtOAc—H2O). The organics were washed with brine, dried (Na2SO4), filtered and evaporated. The residue was purified by flash column chromatography (silica, CH2Cl2-MeOH) to give the desired compound as a yellow solid (36.5 mg, 92%). ESIMS m/z=637.62 [M+H]+.
Step 1i.
To a suspension of PPh3MeBr (0.315 g, 0.882 mmol) in THF (8 mL) was added t-BuOK (1M in THF, 0.80 mL, 0.802 mmol) at rt. The mixture was stirred at rt for 1 hour before a solution of the compound from step 1h (0.102 g, 0.160 mmol) in THF (6 mL) was added. The resultant mixture was allowed to react for 6 hours before being evaporated to dryness. The residue was partitioned (EtOAc—H2O) and the organics were washed with brine, dried (Na2SO4), filtered and evaporated. The residue was purified by flash column chromatography (silica, hexanes-ethyl acetate) to give the title compound as a light yellow solid (90.0 mg, 88%). ESIMS m/z=635.42 [M+H]+.
Example 83




Step 83a.

A solution of the compound from example 1 (90.0 mg, 0.142 mmol) in CH2Cl2 (4 mL) was treated with HCl in 1,4-dioxane (4 M, 3 mL) for 10 min. The volatiles were evaporated off to give the crude desired compound as a yellow solid which was directly used in the next step. ESIMS m/z=435.37 [M+H]+.
Step 83b.
A mixture of the crude compound from step 83a (0.142 mmol at most) and (S)-(methoxycarbonyl)amino-3-methyl-butyric acid (prepared according to WO 2008/021927, 52.1 mg, 0.298 mmol) in DMF (3 mL) was treated with HATU (0.108 g, 0.284 mmol) in the presence of DIPEA (0.35 mL, 2.84 mmol) for 1 hour at rt. The volatiles were evaporated off to provide a brown syrup, which was purified by flash column chromatography (silica, CH2Cl2-MeOH) to give the title compound as a light yellow solid (79.2 mg, 2 steps 75%). ESIMS m/z=749.59 [M+H]+.
Example 252




The title compound was prepared from the compound of Example 332 using procedures similar to that described in example 83. ESIMS m/z=751.46 [M+H]+.

Example 300




The title compound was prepared from the compound of example 317 using procedures similar to that described in example 83. ESIMS m/z=749.41 [M+H]+.

The title compounds of examples 2-82, 84-251, 253-299, 301-315 can be prepared using the chemistry described in the Synthetic Methods and/or Examples 1, 83, 252, 300 and 316-337.







TABLE 1







Examples 1-219








 








Entry















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TABLE 2







Example 220-229








 
















Entry
R
R′
R′′
U
Entry
R
R′
R′′
U





220
Me
H
H
CH2
221
H
H
H
CF2


 


222
Me
H
H
S
223
H
H
H






 


224
H
Me
H
CH2
225
H
H
H






 


226
H
Ph
H
CH2
227
H
H
H






 


228
H
H
Ph
CH2
229
H
H
H




















TABLE 3







Examples 230-239




















Entry
R
R′
R′′
Entry
R
R′
R′′





230
Me
H
H
231
H
CO2Me
H


232
H
F
H
233
H
H
CO2Me


234
H
H
F
235
H
OMe
H


236
H
Cl
H
237
H
H
OMe


238
H
H
Cl
239
H
CF3
H
















TABLE 4







Examples 240-249








 
















Entry
R
R′
R′′
R′′′
Entry
R
R′
R′′
R′′′





240
F
H
H
H
241
F
F
H
H


242
Me
H
H
H
243
Me
Me
H
H


244
H
H
Me
Me
245
H
H
Et
Et


246
CF3
H
H
H
247
CF3
H
CF3
H


248
Cl
H
H
H
249
Cl
H
Cl
H
















TABLE 5







Examples 250-264








 










Entry
R
Entry
R













250




251






 


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TABLE 6







Examples 265-282








 










Entry
Aa
Entry
Aa













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TABLE 7







Example 283-303








 










Entry
Aa
Entry
Aa





 


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TABLE 8







Examples 304-315








 












Entry
Gg
Entry
Gg
Entry
Gg















304




305




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307




308




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310




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313




314




315













Example 316




Step 316a.

A solution of the compound from step 1g of example 1 (50.0 mg, 78.3 μmol) in 1,4-dioxane (1 mL) was treated with HCl in 1,4-dioxane (4 M, 4 mL) at rt for 2 hours. The volatiles were evaporated off to give the crude desired compound as a yellow solid which was directly used in the next step.
Step 316b.
A mixture of the crude compound from step 316a (78.3 μmol at most) and (S)-(methoxycarbonyl)amino-3-methyl-butyric acid (prepared according to WO 2008/021927, 27.4 mg, 0.157 mmol) in DMF (3 mL) and DIPEA (0.27 mL, 1.566 mmol) was treated with HATU (56.5 mg, 0.149 mmol) for 1 hour at rt. The volatiles were evaporated off to provide a brown syrup, which was purified by flash column chromatography (silica, silica, hexanes-ethyl acetate, with 25% MeOH and 1% Et3N in ethyl acetate) to give the desired compound as a yellow solid (52.7 mg, 89% over 2 steps). ESIMS m/z=753.46 [M+H]+.
Step 316c.
To a solution of the compound from step 316b (38.6 mg, 51.3 μmol) in CH2Cl2 (3 mL) were added camphorsulfonic acid (23.8 mg, 0.103 mmol) and Dess-Martin periodinane (0.131 g, 0.308 mmol). The mixture was stirred at rt for 2 hours before another addition of Dess-Martin periodinane (0.131 g, 0.308 mmol). The resultant mixture was allowed to react for another 3 hours before being quenched with aqueous Na2S2O3 solution. The volatiles were evaporated and the residue was partitioned (EtOAc—H2O). The organics were washed with brine, dried (Na2SO4), filtered and evaporated. The residue was purified by flash column chromatography (silica, CH2Cl2-MeOH) to give the desired compound as a yellow brown solid (33.2 mg, 86%). ESIMS m/z=751.54 [M+H]
Step 316d.
To a solution of the compound from step 316c (4.0 mg, 5.3 μmol) in 1,4-dioxane (2 mL) was added MeONH2.HCl (4.4 mg, 53.0 μmol). The mixture was stirred for 14 hours before being evaporated to dryness. The residue was purified by flash column chromatography (silica, CH2Cl2-MeOH) to give the title compound as a white solid (2.7 mg, 66%). ESIMS m/z=780.49 [M+H]+.
Example 317




The title compound was prepared using procedures similar to that described in example 1. ESIMS m/z=635.30 [M+H]+.

Example 318




Step 318a.

To a suspension of activated zinc powder (2.40 g, 36.7 mmol) in dry THF (50 mL) was added allyl bromide (3.1 mL, 36.7 mmol) dropwise. The resulting light yellow solution was cooled to −30° C. before the (S)-1-tert-butyl-2-methyl 4-oxopyrrolidine-1,2-dicarboxylate (4.16 g, 17.1 mmol) was added dropwise. The reaction mixture was stirred at <−10° C. for 4 hours before being quenched with HCl (1 N). The mixture was partitioned (EtOAc—H2O). The organics were washed with brine, dried (Na2SO4) and evaporated. The residue was purified by flash column chromatography (silica, EtOAc-hexanes) to afford the desired compound as a light yellow oil (4.85 g, 98%). ESIMS m/z=286.15 [M+H]+.
Step 318b.
To a solution of the compound from step 318a (0.200 g, 0.627 mmol) in CH3CN (4 mL) were added NaHCO3 (0.211 g, 2.51 mmol) and iodine (0.477 g, 1.88 mmol). The resultant mixture were heated up to 50° C. for 4 hours before the second addition of NaHCO3 (0.211 g, 2.51 mmol) and iodine (0.477 g, 1.88 mmol). The reaction was kept at 50° C. for another 3 hours before being cooled down and quenched by aqueous Na2S2O3. The volatiles were evaporated and the residue was partitioned (EtOAc—H2O). The organics were washed with brine, dried (Na2SO4), filtered and evaporated. The residue was purified by flash column chromatography (silica, EtOAc-hexanes) to give the desired compound as a colorless oil (79.8 mg, 29%). ESIMS m/z=468.23 [M+Na]+.
Step 318c.
Into a solution of the compound from step 318b (1.18 g, 2.66 mmol) in toluene (50 mL) were added tris(trimethylsilyl)silane (2.05 mL, 6.66 mmol) and 2,2′-azo-bis-isobutyronitrile (26.2 mg, 0.160 mmol). The resultant mixture were degassed and heated up to 90° C. under N2 for 3 hours before being allowed to cool down and evaporated to dryness. The residue was purified by flash column chromatography (silica, EtOAc-hexanes) to give the desired compound as a colorless oil (0.333 g, 39%). ESIMS m/z=320.16 [M+H]+.
Step 318d.
Into a solution of the compound from step 318c (0.170 g, 0.533 mmol) in MeOH (6 mL) were added palladium hydroxide (20 wt % on carbon, 50.0 mg) and Boc2O (0.174 g, 0.799 mmol). The resulting mixture was hydrogenated under 60 psi hydrogen gas at room temperature for 1 day before being filtered through a plug of celite. The filtrate was concentrated and purified by flash column chromatography (silica, EtOAc-hexanes) to give the desired compound as a colorless oil (0.127 g, 84%). ESIMS m/z=308.14 [M+Na]+.
Step 318f.
Into a solution of the compound from step 318d (0.127 g, 0.447 mmol) in EtOH (4 mL) at 0° C. was added lithium hydroxide monohydrate (22.5 mg, 0.536 mmol) in H2O (2 mL). The resulting mixture was gradually warmed up to room temperature for 1 day before being evaporated to dryness. The residue was partitioned (Et2O—H2O) and the aqueous phase was acidified to pH  ̃2 at 0° C. The mixture was then partitioned (CH2Cl2—H2O) and the organics were washed with brine, dried (Na2SO4), filtered and evaporated to give the crude desired compound as a colorless oil (0.122 g, 100%). ESIMS m/z=319.14 [M+Li+CH3CN]+.
Step 318g.
Into a mixture of the crude compound from step 318f (60.5 mg, 0.223 mmol at most) and 4-bromo-1,2-diaminobenzene (46.0 mg, 0.246 mmol) in CH3CN (4 mL) were added EDC.HCl (55.7 mg, 0.291 mmol) and DMAP (5.5 mg, 44.7 μmol). The mixture was stirred at room temperature for 14 hours before being evaporated to dryness. The residue was purified by flash column chromatography (silica, hexanes-ethyl acetate) to give the desired compound as a yellow, brown oil (82.0 mg, 83%). ESIMS m/z=440.11, 442.11 [M+H]+.
Step 318h.
A solution of the compound from step 318g (82.0 mg, 0.186 mmol) in AcOH (8 mL) was heated at 50° C. for 16 hours before being evaporated to dryness. The residue was was partitioned (EtOAc—H2O) and the organics were dried (Na2SO4), filtered and evaporated. The residue was purified by flash column chromatography (silica, hexanes-ethyl acetate) to give the desired compound as a yellow oil (54.5 mg, 69%). ESIMS m/z=422.11, 424.14 [M+H]+.
Step 318i.
A mixture of the compound from step 318h (1.88 g, 4.47 mmol) and ethynyltrimethylsilane (6.32 mL, 44.7 mmol) in Et3N (45 mL) were added CuI (25.5 mg, 0.134 mmol) and Pd(PPh3)4 (0.258 g, 0.223 mmol). The resultant mixture were degassed and heated to 95° C. under N2 for 20 hour. The volatiles were evaporated and the residue was partitioned (EtOAc—H2O). The organics were washed with brine, dried (Na2SO4), filtered and evaporated. The residue was purified by flash column chromatography (silica, hexanes-ethyl acetate) to give the desired compound as a light yellow brown foam (1.79 g, 91%). ESIMS m/z=440.27 [M+H]+.
Step 318j.
A solution of the compound from step 318i (1.79 g, 4.08 mmol) in MeOH (40 mL) was treated with K2CO3 (1.41 g, 10.2 mmol) for 2 hours before being evaporated to dryness. The residue was partitioned (EtOAc—H2O) and the organics were dried (Na2SO4), filtered and evaporated. The residue was purified by flash column chromatography (silica, hexanes-ethyl acetate) to give the desired compound as a light yellow solid (1.35 g, 90%). ESIMS m/z=368.23 [M+H]+.
Step 318k.
Into a solution of (S)-1-(tert-butoxycarbonyl)-4-methylenepyrrolidine-2-carboxylic acid (2.02 g, 9.06 mmol) in acetonitrile (12 mL) and DIPEA (6 mL) was added the 2-bromo-1-(4′-iodophenyl)ethanone (3.1 g, 9.5 mmol). The resulting solution was stirred at room temperature overnight (16 h) before was concentrated to give a brownish oil. This crude product was purified by flash column chromatography (silica, EtOAc-hexanes) to afford the desired compound as a light yellow oil (4.37 g, 98%).
Step 318l.
Into a solution of the compound from step 318k (4.37 g, 9.27 mmol) in toluene (60 mL) was added ammonium acetate (7.9 g, 102 mmol). The resulting mixture was heated at 107° C. for 12 h before being cooled and partitioned (H2O-EtOAc). The organic phase was separated, dried (Na2SO4) and concentrated to afford a brown oil, which was purified by flash column chromatography (silica, EtOAc-hexanes) to afford the desired compound as a yellow foam (3.3 g, 80%). ESIMS m/z=452.01[M+H]+.
Step 318m.
A mixture of compounds from step 318j (45.0 mg, 0.122 mmol), compounds from step 318l (55.3 mg, 0.122 mmol), tetrakis(triphenylphosphine)palladium(0) (7.1 mg, 6.1 μmol) and copper(I) iodide (0.7 mg, 3.7 μmol) in triethylamine (2 mL) and acetonitrile (2 mL) was degassed and heated at 40° C. for 15 hours under N2. The mixture was evaporated. The residue was purified by flash column chromatography (silica, hexanes-EtOAc, with 1% Et3N in EtOAc) to give the title compound as a yellow solid (82.0 mg, 97%). ESIMS m/z=691.45 [M+H]+.
Example 319




The title compound was prepared from the compound of example 318 using procedures similar to that described in example 83. ESIMS m/z=805.45 [M+H]+.

Example 320




Step 320a.

To a solution of LiHMDS (1.0 M in THF, 5.17 mL, 5.17 mmol) in THF (20 mL) at −78° C. was added a solution of (+)-(3R,7aS)-tetrahydro-3-phenyl-3H,5H-pyrrolo[1,2-c]oxazol-5-one (0.500 g, 2.460 mmol) in THF (10 mL) under N2. The mixture was stirred at −78° C. for 30 min before ClCO2Me (0.19 mL, 2.460 mmol) was added at −78° C. After 30 min at −78° C., the reaction was quenched with saturated NH4Cl solution. The mixture was allowed to warm up to rt and the volatiles were evaporated. The residue was partitioned (EtOAc—H2O). The organics were washed with brine, dried (Na2SO4), filtered and evaporated. The residue was purified by flash column chromatography (silica, EtOAc-hexanes) to give the desired compound as a colorless oil (0.598 g, 93%). ESIMS m/z=262.13 [M+H]+.
Step 320b.
To a solution of the compound from step 320a (0.350 g, 1.340 mmol) in THF (13 mL) at 0° C. was added NaH (60% in mineral oil, 64.3 mg, 1.607 mmol). After addition, the mixture was stirred at rt for 15 min before allyl bromide (0.13 mL, 1.474 mmol) was added. After 1 h at rt, the reaction was quenched with saturated NH4Cl solution. The mixture was partitioned (EtOAc—H2O). The organics were washed with brine, dried (Na2SO4), filtered and evaporated. The residue was purified by flash column chromatography (silica, EtOAc-hexanes) to give the desired compounds as two separated diastereomers: minor diastereomer (less polar, 56.0 mg, 14%), (3R,6R,7aS)-methyl 6-allyl-5-oxo-3-phenylhexahydropyrrolo[1,2-c]oxazole-6-carboxylate, ESIMS m/z=302.19 [M+H]+, 1H NMR (CDCl3) 7.44-7.33 (m, 5H), 6.32 (s, 1H), 5.75-5.66 (m, 1H), 5.19-5.18 (m, 1H), 5.16 (s, 1H), 4.28-4.22 (m, 2H), 3.78 (s, 3H), 3.57-3.52 (m, 1H), 2.90 (dd, J=6.7, 13.4 Hz, 1H), 2.85 (dd, J=7.9, 14.1 Hz, 1H), 2.58 (dd, J=6.7, 14.1 Hz, 1H), 1.89 (dd, J=6.6, 13.2 Hz, 1H); major diastereomer (more polar, 0.222 g, 55%), (3R,6S,7aS)-methyl 6-allyl-5-oxo-3-phenylhexahydropyrrolo[1,2-c]oxazole-6-carboxylate, ESIMS m/z=302.19 [M+H]+, 1H NMR (CDCl3) 7.46-7.33 (m, 5H), 6.33 (s, 1H), 5.82-5.73 (m, 1H), 5.23-5.18 (m, 2H), 4.28 (dd, J=6.2, 6.5 Hz, 1H), 4.08-4.02 (m, 1H), 3.82 (s, 3H), 3.67 (t, J=8.3 Hz, 1H), 2.80 (dd, J=7.5, 14.0 Hz, 1H), 2.71 (dd, J=7.1, 14.0 Hz, 1H), 2.54 (dd, J=4.9, 12.8 Hz, 1H), 2.38 (dd, J=7.9, 13.8 Hz, 1H).
Step 320c.
To a solution of the major diastereomer from step 320b (0.160 g, 0.585 mmol) in THF/H2O (1/1, 6 mL) at rt was added OsO4 (4 wt % in H2O, 7.5 μL, 0.012 mmol), followed by NaIO4 (0.263 g, 1.229 mmol). The resulting mixture was stirred at rt for 2 hours before being quenched with saturated Na2S2O3 solution. The mixture was partitioned (EtOAc—H2O). The organics were washed with brine, dried (Na2SO4), filtered and evaporated to afford the desired compound as a colorless oil (0.133 g), which was used directly for next step.
Step 320d.
To a solution of the compound from step 320c (0.133 g, 0.438 mmol at most) in EtOH (5 mL) at 0° C. was added NaBH4 (33.2 mg, 0.877 mmol). After 20 min at 0° C., the resulting mixture was stirred at rt for 2.5 hours. More NaBH4 (16.6 mg, 0.438 mmol) was added. After 2 h at rt, the reaction was quenched with saturated NH4Cl solution. The volatiles were evaporated. The residue was taken up in EtOAc (with 5% MeOH) and filtered. The filtrate was evaporated to dryness. The residue was purified by flash column chromatography (silica, EtOAc-MeOH) to give the desired compound as a white foam (67.6 mg, 46% over 2 steps). ESIMS m/z=278.17 [M+H]+.
Step 320e.
To a solution of the compound from step 320d (0.793 g, 2.860 mmol) in pyridine (28 mL) at rt was added TsCl (0.600 g, 3.145 mmol). The resulting solution was stirred at rt for 40 hours. More TsCl (0.600 g, 3.145 mmol) was added. After 24 hours at rt, the reaction was quenched with saturated NaHCO3 solution. The mixture was evaporated to dryness. The residue was taken up in CH2Cl2 and filtered. The filtrate was directly purified by flash column chromatography (silica, EtOAc-hexanes) to give the desired compound as a colorless oil (0.511 g, 69%). ESIMS m/z=260.16 [M+H]+.
Step 320f.
To a solution of the compound from step 320e (0.540 g, 2.082 mmol) in THF (20 mL) at rt was added LiAlH4 (1.0 M in Et2O, 4.16 mL, 4.16 mmol). The resulting mixture was heated at 60° C. for 2 hours before being allowed to cool down. The reaction was quenched by carefully adding H2O (0.16 mL), followed by 15% NaOH solution (0.16 mL) and then H2O (0.32 mL). The suspension was filtered through a short pad of celite. The filtrate was evaporated to give the desired compound as a white semi-solid (0.572 g). ESIMS m/z=248.20 [M+H]+.
Step 320g.
To a solution of the compound from step 320f (2.082 mmol at most) in MeOH (15 mL) at rt was added HOAc (0.16 mL, 2.71 mmol), followed by Pd/C (10%, 0.100 g). The resulting mixture was stirred at rt under H2 (60 psi) for 2 hours before being filtered through a short pad of celite. The filtrate was evaporated to give the desired compound as a colorless oil, which was used directly for next step. ESIMS m/z=158.11 [M+H]+.
Step 320h.
To a solution of the compound from step 320g (2.082 mmol at most) in 1,4-dioxane/H2O (1/2, 21 mL) at rt was added NaHCO3 (1.399 g, 16.66 mmol), followed by (Boc)2O (0.545 g, 2.498 mmol). The resulting mixture was stirred at rt for 15 hours. The volatiles were evaporated. The residue was partitioned (EtOAc—H2O). The organics were washed with brine, dried (Na2SO4), filtered and evaporated. The residue was purified by flash column chromatography (silica, EtOAc-hexanes) to give the desired compound as a colorless oil (0.252 g, 45% over 3 steps). ESIMS m/z=258.18 [M+H]+.
Step 320i.
To a biphasic mixture of the compound from step 320h (0.252 g, 0.979 mmol) in CCl4/CH3CN/H2O (3/4/5, 12 mL) at rt was added RuCl3.xH2O (4.1 mg, 0.020 mmol), followed by NaIO4 (0.419 g, 1.959 mmol). The resulting mixture was stirred at rt for 2 hours. The volatiles were evaporated. The residue was taken up in EtOAc and filtered. The filtrate was washed with brine, dried (Na2SO4) and filtered. The solid from the filtration dissolved in brine and some H2O, acidified to pH  ̃2 and extracted with EtOAc. The combined organics were washed with brine, dried (Na2SO4), filtered and evaporated. The residue was purified by flash column chromatography (silica, EtOAc-MeOH) to give the desired compound as a colorless oil (0.260 g, 98%). ESIMS m/z=272.24 [M+H]+.
Step 320j.
A mixture of the compound from step 320i (0.260 g, 0.958 mmol) and 4-bromo-1,2-diaminobenzene (0.197 g, 1.054 mmol) in CH3CN (6 mL) was treated with EDC.HCl (0.239 g, 1.246 mmol) and DMAP (11.7 mg, 0.096 mmol) at rt. The mixture was stirred at rt for 3 hours before being evaporated. The residue was purified by flash column chromatography (silica, EtOAc-hexanes) to give the desired compound as a yellow foam (0.277 g, 64%). ESIMS m/z=440.29, 442.29 [M+H]+.
Step 320k.
A solution of the compound from step 320j (0.277 g, 0.629 mmol) in glacial acetic acid (7 mL) was heated at 50° C. for 15 hours. The volatiles were evaporated off. Et3N (5 mL) was added and the mixture was evaporated again. The residue was purified by chromatography (silica, hexanes-ethyl acetate, with 1% Et3N in ethyl acetate) to give the desired compound as a yellow foam (0.247 g, 93%). ESIMS m/z=422.15, 424.15 [M+H]+.
Step 320l.
A mixture of the compound from step 320k (0.247 g, 0.585 mmol), trimethylsilyl-acetylene (1.24 mL, 8.772 mmol), CuI (3.3 mg, 0.018 mmol) and Pd(PPh3)4 (33.8 mg, 0.029 mmol) in Et3N (8 mL) was degased and then heated at 90° C. under N2 overnight. More trimethylsilyl-acetylene (0.41 mL, 2.924 mmol) and CH3CN (3 mL) were added. The mixture was heated at 90° C. for 1.5 hours before being allowed to cool down and evaporated. The residue was purified by chromatography (silica, hexanes-ethyl acetate) to give the desired compound as a yellow oil (0.270 g). ESIMS m/z=440.22 [M+H]+.
Step 320m.
A suspension of the compound from step 320l (0.270 g, 0.585 mmol at most) and K2CO3 (0.202 g, 1.462 mmol) in methanol (6 ml) was stirred at rt for 2 hour. The volatiles were evaporated off. The residue was taken up in CH2Cl2 and filtered through a short pad of celite. The filtrate was directly purified by chromatography (silica, hexanes-ethyl acetate) to give the desired compound as a yellow foam (0.186 g, 87% over 2 steps). ESIMS m/z=368.21 [M+H]+.
Step 320n.
The title compound was prepared from the compound from step 320m and compound from step 3181 using procedures similar to that described in step 318m. ESIMS m/z=691.45 [M+H]+.
Example 321




The title compound was prepared from the compound of example 320 using procedures similar to that described in example 83. ESIMS m/z=805.52 [M+H]+.

Example 322




Step 322a.

A mixture of compound from step 318l (1.100 g, 2.437 mmol), trimethylsilylacetylene (0.55 mL, 3.900 mmol), CuI (13.9 mg, 0.073 mmol) and Pd(PPh3)4 (0.141 g, 0.122 mmol) in Et3N (15 mL) and CH3CN (15 mL) was degased and then heated at 40° C. under N2 for 40 hours before being allowed to cool down and evaporated. The residue was purified by chromatography (silica, hexanes-ethyl acetate) to give the desired compound as a yellow solid (0.922 g, 90%). ESIMS m/z=422.15 [M+H]+.
Step 322b.
A suspension of the compound from step 322a (0.922 g, 2.187 mmol) and K2CO3 (0.756 g, 5.467 mmol) in methanol (20 ml) was stirred at rt for 2 hour. The volatiles were evaporated off. The residue was taken up in CH2Cl2 and filtered through a short pad of celite. The filtrate was directly purified by chromatography (silica, hexanes-ethyl acetate) to give the desired compound as a light yellow foam (0.695 g, 91%). ESIMS m/z=350.17 [M+H]+.
Step 322c.
A mixture of (5S,8S)-7-(tert-butoxycarbonyl)-2-oxa-7-azaspiro[4.4]nonane-8-carboxylic acid (prepared from the minor diastereomer from step 320b following the procedure of step 320c-320i, 0.289 g, 1.115 mmol) and 1,2-diamino-4-iodobenzene (0.287 g, 1.226 mmol) in CH3CN (10 mL) was treated with EDC.HCl (0.278 g, 1.449 mmol) and DMAP (13.6 mg, 0.112 mmol) at rt. The mixture was stirred at rt for 2.5 hours before being evaporated. The residue was purified by flash column chromatography (silica, EtOAc-hexanes) to give the desired compound as a yellow solid (0.377 g, 69%). ESIMS m/z=488.16 [M+H]+.
Step 322d.
A solution of the compound from step 322c (0.377 g, 0.774 mmol) in glacial acetic acid (8 mL) was heated at 50° C. for 15 hours. The volatiles were evaporated off. Et3N (5 mL) was added and the mixture was evaporated again. The residue was purified by chromatography (silica, hexanes-ethyl acetate, with 1% Et3N in ethyl acetate) to give the desired compound as a yellow foam (0.324 g, 89%). ESIMS m/z=470.12 [M+H]+.
Step 322e.
The title compound was prepared from the compound from step 322b and the compound from step 322d using procedures similar to that described in step 318m. ESIMS m/z=691.43 [M+H]+.
Example 323




The title compound was prepared from the compound of example 322 using procedures similar to that described in example 83. ESIMS m/z=805.46 [M+H]+.

Example 324




The title compound was prepared from (S)-5-(tert-butoxycarbonyl)-5-azaspiro[2.4]heptane-6-carboxylic acid (prepared according to WO 2009/102325) using procedures similar to that described in step 320j-320n. ESIMS m/z=661.37 [M+H]+.

Example 325




The title compound was prepared from the compound of example 324 using procedures similar to that described in example 83. ESIMS m/z=775.40 [M+H]+.

Example 326




Step 326a.

Into a solution of the compound from step 318a (596 mmol, 2 mmol,  ̃8: 1 diastereomers on C-4) and allyl tent-butyl carbonate (1.26 g, 8 mmol) in THF (10 mL) were added Pd2(dba)3 (46 mg, 0.05 mmol) and DPPB (43 mg, 0.1 mmol). The resultant mixture was degassed and then heated at 75° C. under N2 for 1.5 hours. After being allowed to cool down, the solution was concentrated. The residue was purified by flash column chromatography (silica, hexanes-ethyl acetate) to give the desired compound as a yellow oil (605 mg, 93%). ESIMS m/z=326.26 [M+H]+.
Step 326b.
Into a solution of the compound from step 326a (677 mg, 2.08 mmol) in toluene (650 mL) was added the 1,3-bis(2,4,6-trimethylphenyl)-4,5-dihydroimidazol-2-ylidene[2-(iso-propoxy)-5-(N,N-dimethylaminosulfonyl)phenyl]methylene ruthenium(II) dichloride (76.4 mg, 0.104 mmol). The resultant mixture was degassed and then heated at 75° C. under N2 for 15 hours. After being allowed to cool down, the solution was concentrated. The residue was purified by flash column chromatography (silica, hexanes-ethyl acetate) to give the desired compound as a light yellow oil (585 mg, 94%). ESIMS m/z=298.19 [M+H]+.
Step 326c.
Into a solution of the compound from step 326b (160 mg, 0.538 mmol) in EtOH (2 mL) and H2O (2 mL) was added LiOH.H2O (27.1 mg, 0.646 mmol). The resulting mixture was stirred at room temperature for 3 hours before the volatiles were evaporated. The residue was dissolved in H2O (10 mL) and acidified to pH 3 by adding HCl (4 N). The resulted cloudy mixture was extracted with EtOAc. The organic phase was separated, dried (Na2SO4) and concentrated to afford the desired compound as a colorless oil (142 mg). This oil was used directly in the next step. ESIMS m/z=284.15 [M+H]+.
Step 326d.
Into a solution of the compound from step 326c (142 mg, 0.501 mmol), 4-bromo-1,2-diamine benzene (93.7 mg 0.501 mmol) and EDC.HCl (115 mg, 0.6 mmol) in acetonitrile (4 mL) was added DMAP (6.1 mg, 0.05 mmol). The resulting solution was stirred at room temperature overnight (16 h). The solution was concentrated and was purified by flash column chromatography (silica, EtOAc-hexanes) to afford the desired compound as a brownish solid (196 mg, 80% for two steps). ESIMS m/z=452.01, 454.09[M+H]+.
Step 326e.
A solution of the compound from step 326d in AcOH (4 mL) was heated at 50° C. for 6 hours. After being allowed to cool down, the volatile was evaporated. The crude oil was partitioned (aq. NaHCO3-EtOAc). The organic phase was separated, dried (Na2SO4) and concentrated to afford a brown oil, which was purified by flash column chromatography (silica, EtOAc-hexanes) to afford the desired compound as a yellow foam (116 mg, 62%) as a single isomer. ESIMS m/z=434.13, 436.13[M+H]+.
Step 326f.
Into a solution of the compound from step 326e (116 mg, 0.267 mmol), trimethylsilyl acetylene (0.75 mL, 5.34 mmol) in acetonitrile (3 mL) and triethylamine (2 mL) were added the Pd(PPh3)4 (31 mg, 0.027 mmol), CuI (2.5 mg, 0.014 mmol). The resultant mixture was degassed and heated at 90° C. under N2 for 15 hours. After being allowed to cool down, the solution was concentrated. The residue was purified by flash column chromatography (silica, hexanes-ethyl acetate) to give the desired compound as an orange oil (104 mg, 78%). ESIMS m/z=452.27 [M+H]+.
Step 326g.
Into a solution of the compound from step 326f (104 mg, 0.230 mmol) in methanol (3 mL) was added potassium carbonate (70 mg 0.5 mmol). The resultant mixture was stirred at room temperature for 3 hours. The volatile was evaporated. The resulted mixture was partitioned (aq. NaHCO3-EtOAc). The organic phase was separated, dried (Na2SO4) and concentrated to afford a brown oil, which was purified by flash column chromatography (silica, EtOAc-hexanes) to afford the desired compound as a yellow foam (76 mg, 94%). ESIMS m/z=380.20 [M+H]+.
Step 326h.
To a mixture of the compound from 326g (34.1 mg, 0.076 mmol), the compound from 3181 (23.5 mg, 0.0698 mmol) in actonitrile (2 mL) and triethylamine (2 mL) were added Pd(PPh3)4 (4 mg, 0.035 mmol) and CuI (1 mg). The resultant mixture was degassed and heated at 38° C. under N2 for 16 hours. The volatiles were evaporated and the residue was partitioned (EtOAc—H2O). The organics were washed with brine, dried (Na2SO4), filtered and evaporated. The residue was purified by flash column chromatography (silica, hexanes-ethyl acetate) to give the title compound as a light yellow solid (37.3 mg, 66%). ESIMS m/z=703.25 [M+H]+.
Example 327




The title compound was prepared from the compound of example 326 using procedures similar to that described in example 83. ESIMS m/z=817.32 [M+H]+.

Example 328




Step 328a.

To a solution of the minor diastereomer from step 320b (1.380 g, 4.580 mmol) in THF (25 mL) at rt was added 9-BBN (0.5 M in THF, 22.90 mL, 11.45 mmol). The resulting mixture was stirred at rt for 5 hours before another batch of 9-BBN (0.5 M in THF, 36.64 mL, 18.32 mmol) was added. The solution was stirred at rt for 15 hours. H2O ( ̃40 mL) was added, followed by NaBO3.4H2O (9.642 g, 59.53 mmol). The mixture was stirred at rt for 2 hours before being partitioned (EtOAc—H2O). The aqueous layer was back-extracted with EtOAc. The combined organics were washed with brine, dried (Na2SO4), filtered and evaporated. The residue was purified by flash column chromatography (silica, hexanes-EtOAc) to give (3R,6S,7aS)-6-(hydroxymethyl)-6-(3-hydroxypropyl)-3-phenyltetrahydropyrrolo[1,2-c]oxazol-5(1H)-one as a colorless oil (1.140 g, 85%). ESIMS m/z=292.15 [M+H]+.
Step 328b.
Into a solution of the compound from step 328a (957 mg. 3.28 mmol), Ag2O (1.14 g, 4.92 mmol) and NaI (110 mg. 0.63 mol) in DCM (10 mL) was added a solution of TsCl (688 mg 3.61 mmol) in DCM (5 mL) dropwise. The resulting mixture was stirred for 16 hours at room temperature before being passed through a pad of celite. The filtrate was concentrated. The residue was purified by flash column chromatography (silica, EtOAc-hexanes) to afford the desired compound as a yellow solid (745 mg, 51%). ESIMS m/z=446.23 [M+H]+.
Step 328c.
Into a solution of the compound from step 328b (745 mg, 1.67 mmol) in THF (35 mL) was added NaH (180 mg, 4.5 mmol). The resulting mixture was heated at 30° C. for 16 hours before being allowed to cool down and quenched with ice/water. The mixture was partitioned (EtOAc—H2O). The organic phase was separated, dried (Na2SO4) and concentrated to afford an oil, which was purified by flash column chromatography (silica, EtOAc-hexanes) to afford the desired compound as a white solid (402 mg, 88%). ESIMS m/z=274.16 [M+H]+.
Step 328d.
Into a solution of the compound from step 328c (405 mg, 1.47 mmol) in THF (5 mL) was added LiAlH4 (1 M in THF, 3 mL, 3 mmol). The resulting mixture was heated at 60° C. for 2 hours before being allowed to cool down. H2O (0.1 mL), NaOH (10%, 0.15 mL) and H2O (0.5 mL) were added in sequence. The resulting suspension was passed through a pad of celite. The filtrate was partitioned (H2O-EtOAc). The organic phase was separated, dried (Na2SO4) and concentrated. The residue was purified by flash column chromatography (silica, EtOAc-hexanes) to afford the desired compound as a colorless oil (337 mg, 88%). ESIMS m/z=262.16[M+H]+.
Step 328e.
Into a solution of the compound from step 328d (337 mg, 1.29 mmol) in MeOH (6 mL) was added Pd/C (10%, 30 mg). The resulting mixture was stirred under H2 (60 psi) for 6 hours before being filtered through a pad of celite. The filtrate was concentrated to give a colorless oil and was used directly for next step. ESIMS m/z=172.13[M+H]+.
Step 328f.
Into a solution of the compound from step 328e (1.29 mmol at most) and NaHCO3 (650 mg, 8 mmol) in H2O (4 mL) and dioxane (2 mL) was added Boc2O (340 mg, 1.55 mmol). The resulting solution was stirred at room temperature overnight before the volatiles were evaporated. The crude product was partitioned (EtOAc—H2O). The organic phase was separated, dried (Na2SO4) and concentrated. The residue was purified by flash column chromatography (silica, EtOAc-hexanes) to afford the desired compound as a white solid (305 mg, 87% over two steps). ESIMS m/z=272.18 [M+H]+.
Step 328g.
Into a solution of the compound from step 328f (305 mg, 1.22 mmol) and RuCl3.xH2O (4.6 mg, 0.0225 mmol) in CCl4 (3 mL)/ACN (4 mL)/H2O (5 mL) was added NaIO4 (477 mg, 2.24 mmol). The resulting mixture was stirred at room temperature for 2 hours before being filtered through a pad of celite. The filtrate was partitioned (EtOAc—H2O). The organic phase was separated, dried (Na2SO4) and concentrated to afford an oil (330 mg), which was used directly in the next step. ESIMS m/z=230.11 [M+H−56]+.
Step 328h.
Into a solution of the compound from step 328g (1.12 mmol at the most), 1,2-diamine-4-iodobenzene (288 mg, 1.23 mmol) and EDC.HCl (258 mg, 1.35 mmol) in acetonitrile (6 mL) was added DMAP (14 mg, 0.112 mmol). The resulting solution was stirred at room temperature overnight (16 h). The solution was concentrated. The residue was purified by flash column chromatography (silica, EtOAc-hexanes) to afford the desired compound as a brownish foam (354 mg, 63% over two steps). ESIMS m/z=502.15 [M+H]+.
Step 328i.
A solution of the compound from step 328h in AcOH (5 mL) was heated at 55° C. for 3 hours. After being allowed to cool down, the volatile was evaporated. The crude oil was partitioned (aq. NaHCO3-EtOAc). The organic phase was separated, dried (Na2SO4) and concentrated. The residue was purified by flash column chromatography (silica, EtOAc-hexanes) to afford the desired compound as a yellow foam (326 mg, 95%). ESIMS m/z=484.15 [M+H]+.
Step 328j.
The title compound was prepared from the compound from step 328i and the compound from step 322b using procedures similar to that described in step 318m. ESIMS m/z=705.34 [M+H]+.
Example 329




The title compound was prepared from the compound of example 328 using procedures similar to that described in example 83. ESIMS m/z=819.55 [M+H]+.

Example 330




The title compound was prepared from the major diastereomer from step 320b using procedures similar to that described in example 328. ESIMS m/z=705.34 [M+H]+.

Example 331




The title compound was prepared from the compound of example 330 using procedures similar to that described in example 83. ESIMS m/z=819.55 [M+H]+.

Example 332




Step 332a.

To a mixture of 2,4′-dibromoacetophenone (5.00 g, 18.0 mmol) and N-Boc-L-proline (3.87 g, 18.0 mmol) in CH3CN (60 mL) was added triethylamine (5.40 mL, 37.8 mmol) slowly. The mixture was stirred at rt until the disappearance of the starting material. The volatiles were evaporated and the residue was partitioned (EtOAc-water). The organics were washed with brine, dried (Na2SO4), filtered and evaporated. The residue was purified by flash column chromatography (silica, hexanes-ethyl acetate) to give the desired compound as a light yellow foam (6.73 g, 91%). 1H NMR (CDCl3) 7.76 (t, J=8.0 Hz, 2H), 7.63 (dd, J=5.0, 8.5 Hz, 2H), 5.51, 5.16 (2d, J=16.0 Hz, 1H), 5.32, 5.28 (2d, J=16.5 Hz, 1H), 4.48, 4.40 (dd, J=5.0, 8.5 Hz, 1H), 3.56 (m, 1H), 3.43 (m, 1H), 2.30 (m, 2H), 2.06 (m, 1H), 1.92 (m, 1H), 1.46, 1.43 (2s, 9H).
Step 332b.
To a solution of the compound from step 332a (6.73 g, 16.3 mmol) in toluene (100 mL) was added ammonium acetate (25.1 g, 0.327 mol) and the resultant mixture was heated at 100° C. for 14 hours. The volatiles were evaporated and the residue was partitioned (EtOAc-aq. NaHCO3). The organics were washed with brine, dried (Na2SO4), filtered and evaporated. The residue was purified by flash column chromatography (silica, hexanes-ethyl acetate) to give the desired compound as a yellow foam (6.10 g, 95%). ESIMS m/z=392.24, 394.24 [M+H]+. 1H NMR (CDCl3) 7.57 (bs, 1H), 7.48 (m, 3H), 7.23 (s, 1H), 4.97 (m, 1H), 3.42 (m, 2H), 2.99 (m, 1H), 2.16 (m, 2H), 1.97 (m, 1H), 1.46 (s, 9H).
Step 332c.
To a mixture of the compound from step 332b (1.00 g, 2.55 mmol), bis(pinacolato)diboron (1.35 g, 5.33 mmol) and potassium acetate (0.640 g, 6.53 mmol) in 1,4-dioxane (20 mL) was added Pd(PPh3)4 (0.147 g, 0.128 mmol). The resultant mixture was degassed and heated at 80° C. under N2 for 14 hours. The volatiles were evaporated and the residue was partitioned (EtOAc-water). The organics were washed with brine, dried (Na2SO4), filtered and evaporated. The residue was purified by flash column chromatography (silica, hexanes-ethyl acetate) to give the desired compound as a light yellow solid (0.978 g, 87%). ESIMS m/z=440.39 [M+H]+. 1H NMR (CDCl3) 11.03, 10.55 (2s, 1H), 7.79 (m, 3H), 7.45 (m, 1H), 7.26 (m, 1H), 4.97 (m, 1H), 3.41 (m, 2H), 3.06, 2.91 (2m, 1H), 2.17 (m, 2H), 1.97 (m, 1H), 1.49 (s, 9H), 1.35 (s, 12H).
Step 332d.
To a mixture of the compound from step 318l (106 mg, 0.235 mmol), the compound from step 332c (155 mg, 0.352 mmol) and NaHCO3 (79 mg, 0.94 mmol) in DME (5.2 mL) and H2O (1.7 mL) was added Pd(PPh3)4 (14 mg, 0.012 mmol). The resultant mixture was degassed and heated at 80° C. under N2 for 3 hours. The volatiles were evaporated and the residue was partitioned (EtOAc—H2O). The organics were washed with brine, dried (Na2SO4), filtered and evaporated. The residue was purified by flash column chromatography (silica, hexanes-ethyl acetate) to give the title compound as a light yellow solid (53 mg, 35%). ESIMS m/z=637.40 [M+H]+.
Example 333




Step 333a.

Into a suspension of NaH (60%, 320 mg, 8 mmol) in DME (10 mL) was added 3-bromopropyltriphenylphosphonium bromide (1.86 g, 4 mmol). The resulting suspension was stirred at 70° C. for 6 hours before (S)-1-(tent-butoxycarbonyl)-4-oxopyrrolidine-2-carboxylic acid (508 mg, 2 mmol) was added. The suspension was heated at 70° C. for another 20 hours before being allowed to cool down and quenched with water. The volatile was evaporated. The residue was partitioned (H2O-EtOAc). The organic phase was extracted with K2CO3 (1 M) twice. The combined aqueous phase was cooled in an ice-water bath. HCl (Conc.) was added to adjust the pH to  ̃3. The resulted mixture was extracted with EtOAc (*2). The combined organic phases were dried (Na2SO4) and concentrated to afford a brown oil (280 mg), which was used directly for the next step. ESIMS m/z=254.13 [M+H]+.
Step 333b.
Into a solution of the compound from step 333a (1.1 mmol at most), 1,2-diamine-4-iodobenzene (257 mg, 1.1 mmol) and EDC.HCl (250 mg, 1.3 mmol) in acetonitrile (6 mL) was added the DMAP (13 mg, 0.10 mmol). The resulting solution was stirred at room temperature for 16 h. The volatile was evaporated. The resultant oil was partitioned (H2O-DCM). The organic phase was separated, dried (Na2SO4) and concentrated to afford a brown oil (1.10 g), which was used directly for the next step.
Step 333c.
The solution of the crude product from 333b (1.1 mmol at most) in AcOH was heated at 55° C. for 6 hours. The volatile was evaporated. The crude oil was partitioned between NaHCO3 (aq) and EtOAc. The organic phase was separated, dried (Na2SO4) and concentrated. The residue was purified by flash column chromatography (silica, EtOAc-hexanes) to afford the desired compound as a yellow foam (68 mg, 8% over three steps). ESIMS m/z=452.06 [M+H]+.
Step 333d.
A mixture of the compound from step 332b (0.559 g, 1.425 mmol), trimethylsilylacetylene (0.60 ml, 4.275 mmol), CuI (28.5 mg, 0.150 mmol) and Pd(PPh3)2Cl2 (80.0 mg, 0.114 mmol) in Et3N (15 mL) was heated at 80° C. under N2 for 6 hours before being evaporated. The residue was purified by chromatography (silica, hexanes-ethyl acetate, with 1% Et3N in ethyl acetate) to give the desired compound as a yellow foam (0.484 g, 83%). ESIMS m/z=410.24 [M+H]+.
Step 333e.
A suspension of the compound from step 333d (0.484 g, 1.182 mmol) and K2CO3 (0.408 g, 2.954 mmol) in methanol (12 ml) was stirred at rt for 3 hour. The volatiles were evaporated off. The residue was purified by chromatography (silica, dichloromethane-ethyl acetate) to give the desired compound as a yellow foam (0.370 g, 93%). ESIMS m/z=338.24 [M+H]+.
Step 333f.
The title compound was prepared from the compound from step 333c and the compound from step 333e using procedures similar to that described in step 318m. ESIMS m/z=661.31 [M+H]+.
Example 334




The title compound was prepared from the compound of example 333 using procedures similar to that described in example 83. ESIMS m/z=775.24 [M+H]+.

Example 335




The title compound was prepared using procedures similar to that described in example 1. ESIMS m/z=647.25 [M+H]+.

Example 336




The title compound was prepared from the compound of example 335 using procedures similar to that described in example 83. ESIMS m/z=761.32 [M+H]+.

Example 337




Step 337a.

Into a solution of (+)-(3R,7aS)-tetrahydro-3-phenyl-3H,5H-pyrrolo[1,2-c]oxazol-5-one (2.10 g, 9.85 mmol) in THF (60 mL) at −78° C. was added LiHMDS (1M in THF, 39.4 mL, 39.4 mmol). The resultant mixture was kept at −78° C. for 30 minutes before slow addition of allyl bromide (5.0 mL, 59.1 mmol). The reaction was gradually warmed up to 0° C. and was quenched by aqueous NH4Cl. The volatiles were evaporated and the residue was was partitioned (EtOAc—H2O). The organics were dried (Na2SO4), filtered and evaporated. The residue was purified by flash column chromatography (silica, hexanes-ethyl acetate) to give the desired compound as a very light yellow oil (2.30 g, 78%). ESIMS m/z=284.16 [M+H]+.
Step 337b.
Ozone was bubbled through a solution of the compound from step 337a (2.30 g, 8.11 mmol) in MeOH (85 mL) at −78° C. until the appearance of blue color. The extra O3 was removed by the O2 flow before the addition of NaBH4 (2.46 g, 64.9 mmol) at −78° C. The resultant mixture was gradually warmed up to room temperature for 16 hours and was quenched by 2M aqueous HCl to pH 5. The volatiles were evaporated and the residue was was partitioned (EtOAc—H2O). The organics were dried (Na2SO4), filtered and evaporated. The residue was purified by flash column chromatography (silica, hexanes-ethyl acetate) to give the desired compound as a colorless oil (1.61 g, 68%). ESIMS m/z=292.15 [M+H]+.
Step 337c.
Into a mixture of the compound from step 337b (1.52 g, 5.21 mmol), Ag2O (1.81 g, 7.80 mmol) and KI (0.173 g, 1.04 mmol) in CH2Cl2 (40 mL) was added TsCl (1.09 g, 5.73 mmol) in CH2Cl2 (20 mL) slowly. The resultant mixture was stirred at room temperature for 24 hours before being filtered through a pad of celite. The filtrates were evaporated and the residue was purified by flash column chromatography (silica, hexanes-ethyl acetate) to give the desired compound as a colorless oil (1.38 g, 60%) with the recovery of the compound from step 26b (0.473 g, 31%). ESIMS m/z=446.07 [M+H]+.
Step 337d.
Into a solution of the compound from step 337c (1.38 g, 3.11 mmol) in THF (62 mL) was added NaH (60% in mineral oil, 0.187 g, 4.67 mmol). The resultant mixture was stirred at room temperature for 24 hours before being quenched by aqueous NH4Cl. The volatiles were evaporated and the residue was partitioned (EtOAc—H2O). The organics were dried (Na2SO4), filtered and evaporated. The residue was purified by flash column chromatography (silica, hexanes-ethyl acetate) to give the desired compound as a colorless oil (0.726 g, 86%). ESIMS m/z=274.10 [M+H]+.
Step 337e.
Into a solution of the compound from step 337d (0.726 g, 2.66 mmol) in THF (50 mL) was added LiAlH4 (1M in THF, 5.3 mL, 5.32 mmol). The resultant mixture was heated to 60° C. for 3 hours before being quenched by sequential addition of H2O (0.20 mL), 15% aqueous NaOH (0.20 mL) and H2O (0.60 mL) at 0° C. The mixture was passed through a pad of celite and the filtrates were evaporated. The residue was partitioned (EtOAc—H2O) and the organics were dried (Na2SO4), filtered and evaporated. The residue was purified by flash column chromatography (silica, hexanes-ethyl acetate) to give the desired compound as a colorless oil (0.718 g). ESIMS m/z=262.21 [M+H]+.
Step 337f.
Into a mixture of compound from step 337e (2.66 mmol at most) and AcOH (0.30 mL, 5.32 mmol) in MeOH (16 mL) was added palladium (10 wt % on carbon, 54.8 mg). The resulting mixture was hydrogenated under 60 psi H2 at room temperature for 4 hours before being filtered through a plug of celite. The filtrate was concentrated to give the crude desired compound as a colorless oil (0.782 g). ESIMS m/z=172.17 [M+H]+.
Step 337g.
Into a mixture of the crude compound from step 337f (2.66 mmol at most) and NaHCO3 (1.79 g, 21.3 mmol) in 1,4-dioxane (10 mL) and H2O (20 mL) was added Boc2O (0.696 g, 3.19 mmol). The resultant mixture was stirred at room temperature for 1 day before being evaporated to dryness. The residue was partitioned (EtOAc—H2O) and the organics were dried (Na2SO4), filtered and evaporated. The residue was purified by flash column chromatography (silica, hexanes-ethyl acetate) to give the desired compound as a colorless oil (0.610 g, 3 step 85%). ESIMS m/z=272.26 [M+H]+.
Step 337h.
Into a solution of the compound from step 337g (0.610 g, 2.25 mmol) in carbon tetrachloride (9 mL), CH3CN (12 mL) and H2O (15 mL) were added RuCl3.XH2O (9.3 mg, 45.0 μmol) and NaIO4 (0.963 g, 4.50 mmol). The resultant mixture was stirred at room temperature for 4 hours before being partitioned (CH2Cl2—H2O). The aqueous phase was acidified to pH 3 and was extracted by CH2Cl2 for 3 times. The combined organics were dried (Na2SO4), filtered and evaporated to give the crude desired compound as a light brown foam (0.640 g). ESIMS m/z=286.24 [M+H]+.
Step 337i.
The crude desired compound was prepared from the crude compound from step 337h and 1,2-diamino-4-iodobenzene using procedures similar to that described in step 1c. ESIMS m/z=502.19 [M+H]+.
Step 337j.
The desired compound was prepared from the crude compound from step 337i using procedures similar to that described in step 1d. ESIMS m/z=484.21 [M+H]+.
Step 337k.
The title compound is prepared from the compounds of step 337j and 3181 using procedures similar to that described in examples 318 and 83.
Biological Activity
1. HCV Replicon Cell Lines
HCV replicon cell lines (kindly provided by R. Bartenschlager) isolated from colonies as described by Lohman et. al. (Lohman et al. (1999) Science 285: 110-113, expressly incorporated by reference in its entirety) and used for all experiments. The HCV replicon has the nucleic acid sequence set forth in EMBL Accession No.: AJ242651, the coding sequence of which is from nucleotides 1801 to 8406.
The coding sequence of the published HCV replicon was synthesized and subsequently assembled in a modified plasmid pBR322 (Promega, Madison, Wis.) using standard molecular biology techniques. One replicon cell line (“SGR 11-7”) stably expresses HCV replicon RNA which consists of (i) the HCV 5′UTR fused to the first 12 amino acids of the capsid protein, (ii) the neomycin phosphotransferase gene (neo), (iii) the IRES from encephalomyocarditis virus (EMCV) and (iv) HCV NS2 to NS5B genes and the HCV 3′UTR. Another replicon cell line (“Huh-luc/neo-ET”) described by Vrolijk et. al. (Vrolijk et. al. (2003) Journal of Virological Methods 110:201-209, expressly incorporated by reference in its entirety) stably expresses HCV replicon RNA which consists of (i) the HCV 5′UTR fused to the first 12 amino acids of the capsid protein, (ii) the firefly luciferase reporter gene, (iii) the ubiquitin gene, (iv) the neomycin phosphotransferase gene (neo), (v) the IRES from encephalomyocarditis virus (EMCV) and (vi) HCV NS3 to NS5B genes that harbor cell culture adaptive mutations (E1202G, T12801, K1846T) and the HCV 3′UTR.
These cell lines are maintained at 37° C., 5% CO2, 100% relative humidity in DMEM (Cat#11965-084, Invitrogen), with 10% fetal calf serum (“FCS”, Invitrogen), 1% non-essential amino acids (Invitrogen), 1% of Glutamax (Invitrogen), 1% of 100× penicillin/streptomycin (Cat#15140-122, Invitrogen) and Geneticin (Cat#10131-027, Invitrogen) at 0.75 mg/ml or 0.5 mg/ml for 11-7 and Huh-luc/neo-ET cells, respectively.
2. HCV Replicon Assay—qRT-PCR
EC50 values of single agent compounds and combinations are determined by HCV RNA detection using quantitative RT-PCR, according to the manufacturer's instructions, with a TAQMAN® One-Step RT-PCR Master Mix Reagents Kit (Cat# AB 4309169, Applied Biosystems) on an ABI Model 7500 thermocycler. The TAQMAN primers to use for detecting and quantifying HCV RNA obtained from Integrated DNA Technologies. HCV RNA is normalized to GAPDH RNA levels in drug-treated cells, which is detected and quantified using the Human GAPDH Endogenous Control Mix (Applied Biosystems, AB 4310884E). Total cellular RNA is purified from 96-well plates using the RNAqueous 96 kit (Ambion, Cat# AM1812). Chemical agent cytotoxicity is evaluated using an MTS assay according to the manufacturer's directions (Promega).
3. HCV Replicon Assay—Luciferase
Since clinical drug resistance often develops in viral infections following single agent therapies, there is a need to assess the additive, antagonistic, or synergistic properties of combination therapies. We used the HCV replicon system to assess the potential use of the compound of the present invention or in combination therapies with Interferon alpha, cyclosporine analogs and inhibitors targeting other HCV proteins. The acute effects of a single or combinations of drugs are studied in the “Huh-luc/neo-ET” replicon with each chemical agent titrated in an X or Y direction in a 6 point two-fold dilution curve centered around the EC50 of each drug. Briefly, replicon cells are seeded at 7,000 cells per well in 90 ul DMEM (without phenol red, Invitrogen Cat.#31053-036) per well with 10% FCS, 1% non-essential amino acids, 1% of Glutamax and 1% of 100× penicillin/streptomycin and incubated overnight at 37° C., 5% CO2, 100% relative humidity. 16-20 h after seeding cells, test compounds previously solubilized and titrated in dimethyl sulfoxide (“DMSO”) from each X plate and Y plate are diluted 1:100 in DMEM (without phenol red, Invitrogen Cat.#31053-036) with 10% FCS, 1% non-essential amino acids, 1% of Glutamax and 1% of 100× penicillin/streptomycin and added directly to the 96-well plate containing cells and growth medium at a 1:10 dilution for a final dilution of compound and DMSO of 1:1000 (0.2% DMSO final concentration). Drug treated cells are incubated at 37° C., 5% CO2, 100% relative humidity for 72 hours before performing a luciferase assay using 100 ul per well BriteLite Plus (Perkin Elmer) according to the manufacturer's instructions. Data analysis utilizes the method published by Prichard and Shipman (Antiviral Research, 1990. 14:181-205). Using this method, the combination data are analyzed for antagonistic, additive, or synergistic combination effects across the entire combination surface created by the diluted compounds in combination.
The compounds of the present invention may inhibit HCV by mechanisms in addition to or other than NS5A inhibition. In one embodiment the compounds of the present invention inhibit HCV replicon and in another embodiment the compounds of the present invention inhibit NS5A.
The compounds of the present invention can be effective against the HCV 1b genotype. It should also be understood that the compounds of the present invention can inhibit multiple genotypes of HCV. In one embodiment compound of the present invention are active against the 1a, 1b, 2a, 2b, 3a, 4a, and 5a genotypes. Table 12 shows the EC50 values of representative compounds of the present invention against the HCV 1b genotype from the above described qRT-PCR or luciferase assay. EC50 ranges against HCV 1b are as follows: A>1 nM; B 10-1000 nM; C<10 μM.







TABLE 12







Genotype-1b replicon EC50















Example
Range
EC50
Example
Range
EC50
Example
Range
EC50


















83
C
5.7 pM
252
B
11.2 pM
300
C
 4.4 pM


316
C

319
B

321
B


323
C
9.2 pM
325
C
 5.9 pM
327
C


329
C

331
B
13.7 pM
334
B
13.0 pM


336
C
6.4 pM









While this invention has been particularly shown and described with references to preferred embodiments thereof, it will be understood by those skilled in the art that various changes in form and details may be made therein without departing from the scope of the invention encompassed by the appended claims.

## Claims

1. A compound represented by Formula (I):



or a pharmaceutically acceptable salt thereof, wherein:
L is absent; Ring A and Ring B are each independently optionally substituted phenyl or optionally substituted monocyclic heteroaryl;
R1 is hydrogen or optionally substituted C1-C4 alkyl;
G and J are each independently selected from:





wherein each of said imidazolyl, benzimidazolyl or imidazopyridyl groups are each optionally substituted; and wherein the 6-membered ring of said benzimidazolyl or imidazopyridyl is attached to one of Ring A or Ring B;
X is C(R2)2;
R2 at each occurrence is independently hydrogen, halogen, optionally substituted C1-C4 alkyl, optionally substituted aryl, or optionally substituted heteroaryl;
R6 is C1-C8 alkyl optionally substituted with amino, hydroxy, protected amino, or O(C1-C4 alkyl);
Q is





wherein U is CH2, CHF, CHMe, CF2, C═CH2, or C═CF2;
R7a is hydrogen; and,
R7b is hydrogen or methyl.

2. A compound represented by Formula (I):



or a pharmaceutically acceptable salt thereof, wherein:
L is absent;
one of Ring A and Ring B is optionally substituted phenyl or optionally substituted monocyclic heteroaryl, and the other of Ring A and Ring B is optionally substituted bicyclic aryl or optionally substituted bicyclic heteroaryl;
R1 is hydrogen or optionally substituted C1-C4 alkyl;
G and J are each independently selected from:





wherein each of said imidazolyl, benzimidazolyl or imidazopyridyl groups are each optionally substituted; and wherein the 6-membered ring of said benzimidazolyl or imidazopyridyl is attached to one of Ring A or Ring B;
X is C(R2)2;
R2 at each occurrence is independently hydrogen, halogen, optionally substituted C1-C4 alkyl, optionally substituted aryl, or optionally substituted heteroaryl;
R6 is C1-C8 alkyl optionally substituted with amino, hydroxy, protected amino, or O(C1-C4 alkyl);
Q is





wherein U is CH2, CHF, CHMe, CF2, C═CH7, or C═CF2;
R7a is hydrogen; and,
R7b is hydrogen or methyl.

3. A compound represented by Formula (I):



or a pharmaceutically acceptable salt thereof, wherein:
Ring A is absent; Ring B is phenyl, monocyclic heteroaryl, bicyclic aryl, or bicyclic heteroaryl, each optionally substituted; and L is optionally substituted C2-C4 alkenyl or optionally substituted C2-C4 alkynyl;
R1 is hydrogen or optionally substituted C1-C4 alkyl;
G and J are each independently selected from:





wherein each of said imidazolyl, benzimidazolyl or imidazopyridyl groups are each optionally substituted; and wherein the 6-membered ring of said benzimidazolyl or imidazopyridyl is attached to one of L or Ring B;
X is C(R2)2;
R2 at each occurrence is independently hydrogen, halogen, optionally substituted C1-C4 alkyl, optionally substituted aryl, or optionally substituted heteroaryl;
R6 is C1-C8 alkyl optionally substituted with amino, hydroxy, protected amino, or O(C1-C4 alkyl);
Q is




U is CH2 or C═CH2;
R7a is hydrogen; and,
R7b is hydrogen or methyl.

4. A compound of Formula (Ic-5):



or a pharmaceutically acceptable salt thereof, wherein Gg is selected from the following:




R1 at each occurrence is independently hydrogen or optionally substituted C1-C4 alkyl;
R6 at each occurrence is independently selected from the group consisting of optionally substituted O(C1-C8 alkyl), optionally substituted amino, C1-C8 alkyl, C2-C8 alkenyl, C2-C8 alkenyl, C3-C8 cycloalkyl, C3-C8 cycloalkenyl, heterocyclic, aryl, and heteroaryl, each optionally substituted;
Q is selected from




R3 and R4 are each independently selected from the group consisting of hydrogen, optionally substituted C1-C8 alkyl, optionally substituted C2-C8 alkenyl, and optionally substituted C3-C8 cycloalkyl; alternatively, R3 and R4 can be taken together with the carbon atom to which they are attached to form optionally substituted C3-C8 cycloalkyl or optionally substituted heterocyclic;
R5 is independently hydrogen, optionally substituted C1-C8 alkyl, or optionally substituted C1-C8 cycloalkyl;
R2 at each occurrence is independently hydrogen, halogen, optionally substituted C1-C4 alkyl, optionally substituted aryl, or optionally substituted heteroaryl; alternatively, two R2 groups can be taken together with the carbon atom to which they are attached to form an optionally substituted C3-C8 cycloalkyl or optionally substituted heterocyclic ring;
U is absent or independently selected from O, S, S(O), SO2, NC(O)—(C1-C4 alkyl), C(O), protected carbonyl, OCH2, OCH2CH2, SCH2, SCH2CH2, C(R7)2, C(R7)2C(R7)2, C═C(R2)2, C═N—O(R1) or C═N—N(R1)2;
R7 at each occurrence is independently selected from the group consisting of hydrogen, halogen, cyano, hydroxy, O(C1-C4 alkyl), S(C1-C4 alkyl), amino optionally substituted with one or two C1-C4 alkyl, optionally substituted aryl, optionally substituted heteroaryl, optionally substituted C3-C8 cylcloalkyl, and optionally substituted C1-C4 alkyl;
Alternatively two geminal R7 groups can be taken together with the carbon atom to which they are attached to form a spiro, optionally substituted C3-C7 cycloalkyl, spiro, optionally substituted C3-C7 cycloalkenyl or spiro, optionally substituted, 3- to 7-membered heterocyclic;
R7a and R7b at each occurrence are each independently selected from the group consisting of hydrogen, optionally substituted aryl, and optionally substituted C1-C4 alkyl;
Alternatively, CHR7a—U or CHR7b-U can be taken together to form a group selected from CH═CH, fused and optionally substituted C3-C8 cycloalkyl, fused and optionally substituted aryl, or fused and optionally substituted heterocyclic; and
Alternatively, U, R7a, and R7b can be taken together with the carbon atoms to which they are attached to form a bridged, optionally substituted C4-C7 cycloalkyl, a bridged, optionally substituted C4-C7 cycloalkenyl or a bridged optionally substituted, 4- to 7-membered heterocyclic ring.

5. A compound selected from the group of compounds 1-337 compiled in the following tables:
Compounds I-219

















 








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Compounds 220-229

















 
















Entry
R
R′
R′′
U
Entry
R
R′
R′′
U





220
Me
H
H
CH2
221
H
H
H
CF2


 


222
Me
H
H
S
223
H
H
H






 


224
H
Me
H
CH2
225
H
H
H






 


226
H
Ph
H
CH2
227
H
H
H






 


228
H
H
Ph
CH2
229
H
H
H












Compounds 230-239

















 














Entry
R
R′
R′′
Entry
R
R′
R′′





230
Me
H
H
231
H
CO2Me
H


232
H
F
H
233
H
H
CO2Me


234
H
H
F
235
H
OMe
H


236
H
Cl
H
237
H
H
OMe


238
H
H
Cl
239
H
CF3
H








Compounds 240-249

















 
















Entry
R
R′
R′′
R′′′
Entry
R
R′
R′′
R′′′





240
F
H
H
H
241
F
F
H
H


242
Me
H
H
H
243
Me
Me
H
H


244
H
H
Me
Me
245
H
H
Et
Et


246
CF3
H
H
H
247
CF3
H
CF3
H


248
Cl
H
H
H
249
Cl
H
Cl
H








Compounds 250-264

















 








Entry
R





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260






 


261






 


262






 


263






 


264












Compounds 265-282

















 








Entry
Aa





265






 


266






 


267






 


268






 


269






 


270






 


271






 


272






 


273






 


274






 


275






 


276






 


277






 


278






 


279






 


280






 


281






 


282












Compounds 283-303

















 








Entry
Aa





283






 


284






 


285






 


286






 


287






 


288






 


289






 


290






 


291






 


292






 


293






 


294






 


295






 


296






 


297






 


298






 


299






 


300






 


301






 


302






 


303












Compounds 304-315

















 










Entry
Gg
Entry
Gg





304




305






 


306




307






 


308




309






 


310




311






 


312




313






 


314




315












Compounds 316-337

6. A pharmaceutical composition comprising a compound or a combination of compounds according to claim 5 or a pharmaceutically acceptable salt thereof, in combination with a pharmaceutically acceptable carrier or excipient.