Patent Publication Number: US-9840511-B2

Title: 2-(aryl or heteroaryl-)phenyl (AZA)benzofuran compounds for the treatment of hepatitis C

Description:
CROSS REFERENCE TO RELATED APPLICATION 
     This non-provisional application claims the benefit of U.S. Provisional Application Ser. No. 62/001,243 filed May 21, 2014 which is herein incorporated by reference in its entirety. 
    
    
     FIELD OF THE INVENTION 
     The invention relates to novel compounds, including their salts, which have activity against hepatitis C virus (HCV) and which are useful in treating those infected with HCV. The invention also relates to compositions and methods of making and using these compounds. 
     BACKGROUND OF THE INVENTION 
     Hepatitis C virus (HCV) is a major human pathogen, infecting an estimated 170 million persons worldwide-roughly five times the number infected by human immunodeficiency virus type 1.A substantial fraction of these HCV infected individuals develop serious progressive liver disease, including cirrhosis and hepatocellular carcinoma (Lauer, G. M.; Walker, B. D. N.  Engl. J. Med.  2001, 345, 41-52). 
     HCV is a positive-stranded RNA virus. Based on a comparison of the deduced amino acid sequence and the extensive similarity in the 5′-untranslated region, HCV has been classified as a separate genus in the Flaviviridae family. All members of the Flaviviridae family have enveloped virions that contain a positive stranded RNA genome encoding all known virus-specific proteins via translation of a single, uninterrupted, open reading frame. 
     Considerable heterogeneity is found within the nucleotide and encoded amino acid sequence throughout the HCV genome. At least six major genotypes have been characterized, and more than 50 subtypes have been described. The major genotypes of HCV differ in their distribution worldwide, and the clinical significance of the genetic heterogeneity of HCV remains elusive despite numerous studies of the possible effect of genotypes on pathogenesis and therapy. 
     The single strand HCV RNA genome is approximately 9500 nucleotides in length and has a single open reading frame (ORF) encoding a single large polyprotein of about 3000 amino acids. In infected cells, this polyprotein is cleaved at multiple sites by cellular and viral proteases to produce the structural and non-structural (NS) proteins. In the case of HCV, the generation of mature non-structural proteins (NS2, NS3, NS4A, NS4B, NS5A, and NS5B) is effected by two viral proteases. The first one is believed to be a metalloprotease and cleaves at the NS2-NS3 junction; the second one is a serine protease contained within the N-terminal region of NS3 (also referred to as NS3 protease) and mediates all the subsequent cleavages downstream of NS3, both in cis, at the NS3-NS4A cleavage site, and in trans, for the remaining NS4A-NS4B, NS4B-NS5A, NS5A-NS5B sites. The NS4A protein appears to serve multiple functions, acting as a cofactor for the NS3 protease and possibly assisting in the membrane localization of NS3 and other viral replicase components. The complex formation of the NS3 protein with NS4A seems necessary to the processing events, enhancing the proteolytic efficiency at all of the sites. The NS3 protein also exhibits nucleoside triphosphatase and RNA helicase activities. NS5B (also referred to as HCV polymerase) is a RNA-dependent RNA polymerase that is involved in the replication of HCV. The HCV NS5B protein is described in “Structural Analysis of the Hepatitis C Virus RNA Polymerase in Complex with Ribonucleotides (Bressanelli; S. et al.,  Journal of Virology  2002, 3482-3492; and Defrancesco and Rice,  Clinics in Liver Disease  2003, 7, 211-242. 
     Currently, the most effective HCV therapy employs a combination of alpha-interferon and ribavirin, leading to sustained efficacy in 40% of patients (Poynard, T. et al.  Lancet  1998, 352, 1426-1432). Recent clinical results demonstrate that pegylated alpha-interferon is superior to unmodified alpha-interferon as monotherapy (Zeuzem, S. et al.  N. Engl. J. Med.  2000, 343, 1666-1672). However, even with experimental therapeutic regimens involving combinations of pegylated alpha-interferon and ribavirin, a substantial fraction of patients do not have a sustained reduction in viral load. Thus, there is a clear and important need to develop effective therapeutics for treatment of HCV infection. 
     HCV-796, an HCV NS5B inhibitor, has shown an ability to reduce HCV RNA levels in patients. The viral RNA levels decreased transiently and then rebounded during dosing when treatment was with the compound as a single agent but levels dropped more robustly when combined with the standard of care which is a form of interferon and ribavirin. The development of this compound was suspended due to hepatic toxicity observed during extended dosing of the combination regimens. U.S. Pat. No. 7,265,152 and the corresponding PCT patent application WO2004/041201 describe compounds of the HCV-796 class. Other compounds have been disclosed; see for example, WO2009/101022, as well as WO 2012/058125. 
     What is therefore needed in the art are additional compounds which are novel and are effective against hepatitis C. Additionally, these compounds should provide advantages for pharmaceutical uses, for example, with regard to one or more of their mechanism of action, binding, inhibition efficacy, target selectivity, solubility, safety profiles, and/or bioavailability. Also needed are new formulations and methods of treatment which utilize these compounds. 
     SUMMARY OF THE INVENTION 
     One aspect of the invention is a compound of Formula I including pharmaceutically acceptable salts thereof: 
                         
wherein
     m is 0 or 1;   X is N or C—R 10 ;   R 1  is methyl;   R 2  is Ar 1 ;   Ar 1  is phenyl, 5-membered heteroaryl or 6-membered heteroaryl, and is substituted with 0-3 substituents selected from the group of cyano, halo, alkyl, cycloalkyl, haloalkyl, OH, OR 101 , haloalkoxy, NH 2 , NR 102 R 103 , COOR 102 R 103 , S(O) 2 NR 102 R 103 , and NR 101 CONR 102 R 103 ;   R 101  is hydrogen, alkyl or cycloalkyl with 0-3 substituents selected from halo, hydroxyl, alkoxy, and haloalkoxy;   R 102 , R 103  are each independently hydrogen, alkyl or cycloalkyl with 0-3 substituents selected from halo, hydroxyl, alkoxy, and haloalkoxy; or   R 102  and R 103  can form a ring by joining two atoms, one from each of R 102  and R 103 ;   R 3  is hydrogen, halo, or alkyl;   R 4 , R 5 , R 6 , R 7 , R 8  are each independently selected from the group of hydrogen, halo, alkyl, cycloalkyl, haloalkyl, halocycloalkyl, hydroxyalkyl, hydroxycycloalkyl, alkoxyalkyl, alkoxycycloalkyl, alkoxy, hydroxyalkyloxy, alkoxyalkyloxy, COOR 201  and CON(R 202 )(R 203 );   R 201  is hydrogen, alkyl or cycloalkyl with 0-3 substituents selected from halo, hydroxyl, alkoxy, and haloalkoxy;   R 202 , R 203  are each independently selected from the group of hydrogen, alkyl, cycloalkyl, alkenyl, cycloalkyl alkynyl, cyclic ether, cyclic amine, lactame, fused bicyclic alkyl, bridged bicyclic alkyl, spiro bicyclic alkyl, fused bicyclic ether, bridged bicyclic ether, spiro bicyclic ether, fused bicyclic amine, bridged bicyclic amine and spiro bicyclic amine, with 0-3 substituents selected from the group of halo, OH, OR 104 , NH 2 , NR 105 R 106 , COOR 104 , CONR 105 R 106 , S(O) 2 R 104 , S(O) 2 NR 105 R 106 , NR 104 CONR 105 R 106 ,   OR 104 CONR 105 R 106 , C(═NR 107 )NR 105 R 106 , NR 108 C(═NR 107 )NR 105 R 106 , haloalkoxy, Ar 2 , O—Ar 2 , and NR 105 —Ar 2 ; or   R 202  and R 203  can form a ring by joining two atoms, one from each of R 202  and R 203 ;   R 202  and R 203  can also form bicyclic or tricyclic rings by joining multiple atoms from each of R 202  and R 203 ;   R 104  is hydrogen, alkyl or cycloalkyl with 0-3 substituents selected from halo, hydroxyl, alkoxy, and haloalkoxy;   R 105 , R 106  are each independently hydrogen, alkyl or cycloalkyl with 0-3 substituents selected from halo, hydroxyl, alkoxy, and haloalkoxy; or   R 105 R 106  can form a ring by joining two atoms, one from each of R 105  and R 106 ;   R 107 , R 108  are each independently hydrogen, alkyl or cycloalkyl with 0-3 substituents selected from halo, hydroxyl, alkoxy, and haloalkoxy; or   R 107  and R 108  can form a ring by joining two atoms, one from each of R 107  and R 108 ;   Ar 2  is phenyl, 5-membered heteroaryl or 6-membered heteroaryl, and is substituted with 0-3 substituents selected from the group of cyano, halo, alkyl, cycloalkyl, haloalkyl, OH, OR 204 , haloalkoxy, NH 2 , NR 205 R 206 , COOR 204 , CONR 205 R 206 , S(O) 2 R 204 , S(O) 2 NR 205 R 206 , and NR 204 CONR 205 R 206 ;   R 204  is hydrogen, alkyl or cycloalkyl with 0-3 substituents selected from halo, hydroxyl, alkoxy, and haloalkoxy;   R 205 , R 206  are each independently hydrogen, alkyl or cycloalkyl with 0-3 substituents selected from halo, hydroxyl, alkoxy, and haloalkoxy; or   R 205  and R 206  can form a ring by joining two atoms, one from each of R 205  and R 206 ;   R 9  is selected from the group of hydrogen, halo, alkyl, haloalkyl, cycloalkyl, alkoxy, Ar 3  and NR 301 R 302 ;   R 301  is selected from the group of hydrogen, alkyl, cycloalkyl, (cycloalkyl)alkyl, benzyl, alkylcarbonyl, haloalkyl carbonyl, phenyl carbonyl, (alkoxyphenyl)carbonyl, alkylsulfonyl, phenylsulfonyl, (alkoxyphenyl)sulfonyl and (haloalkoxyphenyl)sulfonyl;   R 302  is hydrogen, alkyl, hydroxyalkyl, or alkoxyalkyl; or   R 301  and R 302  taken together with the nitrogen to which they are attached is oxazolidinonyl or dioxothiazinyl;   Ar 3  is phenyl, 5-membered heteroaryl or 6-membered heteroaryl, and is substituted with 0-3 substituents selected from cyano, halo, alkyl, cycloalkyl, haloalkyl, OH, OR 303 , haloalkoxy, NH 2 , NR 304 R 305 , COOR 303 , CONR 304 R 305 , S(O) 2 R 303 , S(O) 2 NR 304 R 305 , and NR 303 CONR 304 R 305 ;   R 303  is hydrogen, alkyl or cycloalkyl with 0-3 substituents selected from halo, hydroxyl, alkoxy, and haloalkoxy;   R 304 , R 305  are each independently hydrogen, alkyl or cycloalkyl with 0-3 substituents selected from halo, hydroxyl, alkoxy, and haloalkoxy; or   R 304  and R 305  can form a ring by joining two atoms, one from each of R 304  and R 305 ;   R 10  is hydrogen, alkyl, halo, N(R 401 )(R 402 ), or alkylsulfonyl;   R 401  and R 402  are each independently selected from the group of hydrogen, alkyl, hydroxyalkyl, alkoxyalkyl, alkylsulfonyl, and alkylsulfonylalkyl;   or N(R 401 )(R 402 ) taken together is azetidinyl, pyrrolidinyl, piperidinyl, or piperazinyl, and is substituted with 0-2 substituents selected from alkyl, hydroxyalkyl, and hydroxy.   

     The invention also relates to pharmaceutical compositions comprising a compound of Formula I, including a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier, excipient and/or diluent. 
     In addition, the invention provides one or more methods of treating hepatitis C infection comprising administering a therapeutically effective amount of a compound of Formula I to a patient. 
     Also provided as part of the invention are one or more methods for making the compounds of Formula I. 
     The present invention is directed to these, as well as other important ends, hereinafter described. 
    
    
     DETAILED DESCRIPTION OF THE EMBODIMENTS 
     As used herein, the singular forms “a”, “an”, and “the” include plural reference unless the context specifically dictates otherwise. 
     Unless otherwise specifically set forth elsewhere in the application, the following terms may be used herein and shall have the following meanings: “Hydrogen” or “H” refers to hydrogen, including its isotopes, such as deuterium. “Halo” means fluoro, chloro, bromo, or iodo. “Alkyl” means a straight or branched alkyl group composed of 1 to 6 carbons. “Alkenyl” means a straight or branched alkyl group composed of 2 to 6 carbons with at least one double bond. “Cycloalkyl” means a monocyclic ring system composed of 3 to 7 carbons. “Hydroxyalkyl,” “alkoxy” and other terms with a substituted alkyl moiety include straight and branched isomers composed of 1 to 6 carbon atoms for the alkyl moiety. “Halo” includes all halogenated isomers from monohalo substituted to perhalo substituted in substituents defined with halo, for example, “Haloalkyl” and “haloalkoxy”, “halophenyl”, “halophenoxy.” “Aryl” means a monocyclic or bicyclic aromatic hydrocarbon groups having 6 to 12 carbon atoms, or a bicyclic fused ring system wherein one or both of the rings is a phenyl group. Bicyclic fused ring systems consist of a phenyl group fused to a four- to six-membered aromatic or non-aromatic carbocyclic ring. Representative examples of aryl groups include, but are not limited to, indanyl, indenyl, naphthyl, phenyl, and tetrahydronaphthyl. “Heteroaryl” means a 5 to 7 membered monocyclic or 8 to 11 membered bicyclic aromatic ring system with 1-5 heteroatoms independently selected from nitrogen, oxygen, and sulfur. Parenthetic and multiparenthetic terms are intended to clarify bonding relationships to those skilled in the art. For example, a term such as ((R)alkyl) means an alkyl substituent further substituted with the substituent R. Substituents which are illustrated by chemical drawing to bond at variable positions on a multiple ring system (for example a bicyclic ring system) are intended to bond to the ring where they are drawn to append. 
     The invention includes all pharmaceutically acceptable salt forms of the compounds. Pharmaceutically acceptable salts are those in which the counter ions do not contribute significantly to the physiological activity or toxicity of the compounds and as such function as pharmacological equivalents. These salts can be made according to common organic techniques employing commercially available reagents. Some anionic salt forms include acetate, acistrate, besylate, bromide, camsylate, chloride, citrate, fumarate, glucouronate, hydrobromide, hydrochloride, hydroiodide, iodide, lactate, maleate, mesylate, nitrate, pamoate, phosphate, succinate, sulfate, tartrate, tosylate, and xinofoate. Some cationic salt forms include ammonium, aluminum, benzathine, bismuth, calcium, choline, diethylamine, diethanolamine, lithium, magnesium, meglumine, 4-phenylcyclohexylamine, piperazine, potassium, sodium, tromethamine, and zinc. 
     Some of the compounds of the invention possess asymmetric carbon atoms. The invention includes all stereoisomeric forms, including enantiomers and diastereomers as well as mixtures of stereoisomers such as racemates. Some stereoisomers can be made using methods known in the art. Stereoisomeric mixtures of the compounds and related intermediates can be separated into individual isomers according to methods commonly known in the art. The use of wedges or hashes in the depictions of molecular structures in the following schemes and tables is intended only to indicate relative stereochemistry, and should not be interpreted as implying absolute stereochemical assignments. 
     The invention is intended to include all isotopes of atoms occurring in the present compounds. Isotopes include those atoms having the same atomic number but different mass numbers. By way of general example and without limitation, isotopes of hydrogen include deuterium and tritium. Isotopes of carbon include  13 C and  14 C. Isotopically-labeled compounds of the invention can generally be prepared by conventional techniques known to those skilled in the art or by processes analogous to those described herein, using an appropriate isotopically-labeled reagent in place of the non-labeled reagent otherwise employed. Such compounds may have a variety of potential uses, for example as standards and reagents in determining biological activity. In the case of stable isotopes, such compounds may have the potential to favorably modify biological, pharmacological, or pharmacokinetic properties. 
     As set forth above, the invention is directed to one or more compounds of Formula I, including pharmaceutically acceptable salts thereof: 
                         
wherein
     m is 0 or 1;   X is N or C—R 10 ;   R 1  is methyl;   R 2  is Ar 1 ;   Ar 1  is phenyl, 5-membered heteroaryl or 6-membered heteroaryl, and is substituted with 0-3 substituents selected from the group of cyano, halo, alkyl, cycloalkyl, haloalkyl, OH, OR 101 , haloalkoxy, NH 2 , NR 102 R 103 , COOR 101 , CONR 102 R 103 , S(O) 2 R 101 , S(O) 2 NR 102 R 103 , and NR 101 CONR 102 R 103 ;   R 101  is hydrogen, alkyl or cycloalkyl with 0-3 substituents selected from halo, hydroxyl, alkoxy, and haloalkoxy;   R 102 , R 103  are each independently hydrogen, alkyl or cycloalkyl with 0-3 substituents selected from halo, hydroxyl, alkoxy, and haloalkoxy; or   R 102  and R 103  can form a ring by joining two atoms, one from each of R 102  and R 103 ;   R 3  is hydrogen, halo, or alkyl;   R 4 , R 5 , R 6 , R 7 , R 8  are each independently selected from the group of hydrogen, halo, alkyl, cycloalkyl, haloalkyl, halocycloalkyl, hydroxyalkyl, hydroxycycloalkyl, alkoxyalkyl, alkoxycycloalkyl, alkoxy, hydroxyalkyloxy, alkoxyalkyloxy, COOR 201  and CON(R 202 )(R 203 );   R 201  is hydrogen, alkyl or cycloalkyl with 0-3 substituents selected from halo, hydroxyl, alkoxy, and haloalkoxy;   R 202 , R 203  are each independently selected from the group of hydrogen, alkyl, cycloalkyl, alkenyl, cycloalkyl alkynyl, cyclic ether, cyclic amine, lactame, fused bicyclic alkyl, bridged bicyclic alkyl, spiro bicyclic alkyl, fused bicyclic ether, bridged bicyclic ether, spiro bicyclic ether, fused bicyclic amine, bridged bicyclic amine and spiro bicyclic amine, with 0-3 substituents selected from the group of halo, OH, OR 104 , NH 2 , NR 105 R 106 , COOR 104 , CONR 105 R 106 , S(O) 2 R 104 , S(O) 2 NR 105 R 106 , NR 104 CONR 105 R 106 , OR 104 CONR 105 R 106 , C(═NR 107 )NR 105 R 106 , NR 108 C(═NR 107 )NR 105 R 106 , haloalkoxy, Ar 5 , O—Ar 2 , and NR 105 —Ar 2 ; or   R 202  and R 203  can form a ring by joining two atoms, one from each of R 202  and R 203 ;   R 202  and R 203  can also form bicyclic or tricyclic rings by joining multiple atoms from each of R 202  and R 203 ;   R 104  is hydrogen, alkyl or cycloalkyl with 0-3 substituents selected from halo, hydroxyl, alkoxy, and haloalkoxy;   R 105 , R 106  are each independently hydrogen, alkyl or cycloalkyl with 0-3 substituents selected from halo, hydroxyl, alkoxy, and haloalkoxy; or   R 105  and R 106  can form a ring by joining two atoms, one from each of R 105  and R 106 ;   R 107 , R 108  are each independently hydrogen, alkyl or cycloalkyl with 0-3 substituents selected from halo, hydroxyl, alkoxy, and haloalkoxy; or   R 107  and R 108  can form a ring by joining two atoms, one from each of R 107  and R 108 ;   Ar 2  is phenyl, 5-membered heteroaryl or 6-membered heteroaryl, and is substituted with 0-3 substituents selected from the group of cyano, halo, alkyl, cycloalkyl, haloalkyl, OH, OR 204 , haloalkoxy, NH 2 , NR 205 R 206 , COOR 204 , CONR 205 R 206 , S(O) 2 R 204 , S(O) 2 NR 205 R 206 , and NR 204 CONR 205 R 206 ;   R 204  is hydrogen, alkyl or cycloalkyl with 0-3 substituents selected from halo, hydroxyl, alkoxy, and haloalkoxy;   R 205 , R 206  are each independently hydrogen, alkyl or cycloalkyl with 0-3 substituents selected from halo, hydroxyl, alkoxy, and haloalkoxy; or   R 205  and R 206  can form a ring by joining two atoms, one from each of R 205  and R 206 ;   R 9  is selected from the group of hydrogen, halo, alkyl, haloalkyl, cycloalkyl, alkoxy, Ar 3  and NR 301 R 302 ;   R 301  is selected from the group of hydrogen, alkyl, cycloalkyl, (cycloalkyl)alkyl, benzyl, alkylcarbonyl, haloalkyl carbonyl, phenyl carbonyl, (alkoxyphenyl)carbonyl, alkylsulfonyl, phenylsulfonyl, (alkoxyphenyl)sulfonyl and (haloalkoxyphenyl)sulfonyl;   R 302  is hydrogen, alkyl, hydroxyalkyl, or alkoxyalkyl; or   R 301  and R 302  taken together with the nitrogen to which they are attached is oxazolidinonyl or dioxothiazinyl;   Ar 3  is phenyl, 5-membered heteroaryl or 6-membered heteroaryl, and is substituted with 0-3 substituents selected from the group of cyano, halo, alkyl, cycloalkyl, haloalkyl, OH, OR 303 , haloalkoxy, NH 2 , NR 304 R 305 , COOR 303 , CONR 304 R 305 , S(O) 2 R 303 , S(O) 2 NR 304 R 305 , and NR 303 CONR 304 R 305 ;   R 303  is hydrogen, alkyl or cycloalkyl with 0-3 substituents selected from halo, hydroxyl, alkoxy, and haloalkoxy;   R 304 , R 305  are each independently hydrogen, alkyl or cycloalkyl with 0-3 substituents selected from halo, hydroxyl, alkoxy, and haloalkoxy; or   R 304  and R 305  can form a ring by joining two atoms, one from each of R 304  and R 305 ;   R 10  is hydrogen, alkyl, halo, N(R 401 )(R 402 ), or alkylsulfonyl;   R 401  and R 402  are independently selected from the group of hydrogen, alkyl, hydroxyalkyl, alkoxyalkyl, alkylsulfonyl, and alkylsulfonylalkyl;   or N(R 401 )(R 402 ) taken together is azetidinyl, pyrrolidinyl, piperidinyl, or piperazinyl, and is substituted with 0-2 substituents selected from alkyl, hydroxyalkyl, and hydroxy.   

     In a preferred embodiment of the invention, m is 0. 
     It is further preferred that R 1  is alkyl, and more preferably methyl. 
     It is also preferred that R 4 , R 5 , R 6 , R 7 , R 8  are each selected from CON(R 202 )(R 203 ). More preferably, R 5  is CON(R 202 )(R 203 ). 
     Additionally, it is preferred that R 202  and R 203  are hydrogen or alkyl. 
     In certain embodiments it is also preferred that R 2  is a 5-membered heteroaryl group. More preferably, the heteroaryl group will contain 2 to 3 nitrogens. 
     It is also preferred that R 9  is selected from the group of haloalkyl, Ar 3  and NR 301 R 302 . The haloalkyl is preferably 1-3 fluoroalkyl, and more preferably is 1-3 fluoropropyl. In another embodiment, it is preferred that R 9  is NR 301 , and R 301  is selected from alkyl and alkylsulfonyl. In a further embodiment, it is preferred that Ar 3  is substituted pyrazole. 
     R 10  is preferably hydrogen. 
     Also preferred are compounds of Formula I, including pharmaceutically acceptable salts thereof, which are selected from the group of: 
     
       
         
         
             
             
         
       
       
         
         
             
             
         
       
       
         
         
             
             
         
       
       
         
         
             
             
         
       
     
     Also preferred are compounds of Formula I, including pharmaceutically acceptable salts thereof, which are selected from the group of: 
     
       
         
         
             
             
         
       
     
     Pharmaceutical Compositions and Methods of Treatment 
     The compounds according to the various embodiments herein set forth demonstrate activity against HCV NS5B, and can be useful in treating HCV and HCV infection. Therefore, another aspect of the invention is a composition comprising a compound of Formula I, or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier, excipient and/or diluent. 
     Another aspect of the invention is a composition further comprising an additional compound having anti-HCV activity. 
     Another aspect of the invention is a composition where the compound having anti-HCV activity is an interferon or a ribavirin. Another aspect of the invention is wherein the interferon is selected from interferon alpha 2B, pegylated interferon alpha, consensus interferon, interferon alpha 2A, interferon lambda, and lymphoblastoid interferon tau. 
     Another aspect of the invention is a composition where the compound having anti-HCV activity is a cyclosporin. Another aspect of the invention is where the cyclosporin is cyclosporin A. 
     Another aspect of the invention is a composition where the compound having anti-HCV activity is selected from the group consisting of interleukin 2, interleukin 6, interleukin 12, a compound that enhances the development of a type 1 helper T cell response, interfering RNA, anti-sense RNA, Imiqimod, ribavirin, an inosine 5′-monophospate dehydrogenase inhibitor, amantadine, and rimantadine. 
     Another aspect of the invention is a composition where the compound having anti-HCV activity is effective to inhibit the function of a target selected from HCV metalloprotease, HCV serine protease, HCV polymerase, HCV helicase, HCV NS4B protein, HCV entry, HCV assembly, HCV egress, HCV NS5A protein, IMPDH, and a nucleoside analog for the treatment of an HCV infection. 
     Another aspect of the invention is a composition comprising a compound of Formula I, or a pharmaceutically acceptable salt thereof, a pharmaceutically acceptable carrier, an interferon and ribavirin. 
     Another aspect of the invention is a method of inhibiting the function of the HCV replicon comprising contacting the HCV replicon with a compound of Formula I or a pharmaceutically acceptable salt thereof. 
     Another aspect of the invention is a method of inhibiting the function of the HCV NS5B protein comprising contacting the HCV NS5B protein with a compound of Formula I or a pharmaceutically acceptable salt thereof. 
     Another aspect of the invention is a method of treating an HCV infection in a patient comprising administering to the patient a therapeutically effective amount of a compound of Formula I or a pharmaceutically acceptable salt thereof. In another embodiment the compound is effective to inhibit the function of the HCV replicon. In another embodiment the compound is effective to inhibit the function of the HCV NS5B protein. 
     Another aspect of the invention is a method of treating an HCV infection in a patient comprising administering to the patient a therapeutically effective amount of a compound of Formula I, or a pharmaceutically acceptable salt thereof, in conjunction with (prior to, after, or concurrently) another compound having anti-HCV activity. 
     Another aspect of the invention is the method wherein the other compound having anti-HCV activity is an interferon or a ribavirin. 
     Another aspect of the invention is the method where the interferon is selected from interferon alpha 2B, pegylated interferon alpha, consensus interferon, interferon alpha 2A, interferon lambda, and lymphoblastoid interferon tau. 
     Another aspect of the invention is the method where the other compound having anti-HCV activity is a cyclosporin. 
     Another aspect of the invention is the method where the cyclosporin is cyclosporin A. 
     Another aspect of the invention is the method where the other compound having anti-HCV activity is selected from interleukin 2, interleukin 6, interleukin 12, a compound that enhances the development of a type 1 helper T cell response, interfering RNA, anti-sense RNA, Imiqimod, ribavirin, an inosine 5′-monophospate dehydrogenase inhibitor, amantadine, and rimantadine. 
     Another aspect of the invention is the method wherein the other compound having anti-HCV activity is effective to inhibit the function of a target selected from the group consisting of HCV metalloprotease, HCV serine protease, HCV polymerase, HCV helicase, HCV NS4B protein, HCV entry, HCV assembly, HCV egress, HCV NS5A protein, IMPDH, and a nucleoside analog for the treatment of an HCV infection. 
     Another aspect of the invention is the method wherein the other compound having anti-HCV activity is effective to inhibit the function of target in the HCV life cycle other than the HCV NS5B protein. 
     “Therapeutically effective” means the amount of agent required to provide a meaningful patient benefit as understood by practitioners in the field of hepatitis and HCV infection. 
     “Patient” means a person infected with the HCV virus and suitable for therapy as understood by practitioners in the field of hepatitis and HCV infection. 
     “Treatment,” “therapy,” “regimen,” “HCV infection,” and related terms are used as understood by practitioners in the field of hepatitis and HCV infection. 
     The compounds of this invention are generally given as pharmaceutical compositions comprised of a therapeutically effective amount of a compound or its pharmaceutically acceptable salt and a pharmaceutically acceptable carrier and may contain conventional excipients. Pharmaceutically acceptable carriers are those conventionally known carriers having acceptable safety profiles. Compositions encompass all common solid and liquid forms including for example capsules, tablets, lozenges, and powders as well as liquid suspensions, syrups, elixers, and solutions. Compositions are made using common formulation techniques, and conventional excipients (such as binding and wetting agents) and vehicles (such as water and alcohols) are generally used for compositions. See, for example,  Remington&#39;s Pharmaceutical Sciences , Mack Publishing Company, Easton, Pa., 17th edition, 1985. 
     Solid compositions are normally formulated in dosage units and compositions providing from about 1 to 1000 mg of the active ingredient per dose are preferred. Some examples of dosages are 1 mg, 10 mg, 100 mg, 250 mg, 500 mg, and 1000 mg. Generally, other agents will be present in a unit range similar to agents of that class used clinically. Typically, this is 0.25-1000 mg/unit. 
     Liquid compositions are usually in dosage unit ranges. Generally, the liquid composition will be in a unit dosage range of about 1-100 mg/mL. Some examples of dosages are 1 mg/mL, 10 mg/mL, 25 mg/mL, 50 mg/mL, and 100 mg/mL. Generally, other agents will be present in a unit range similar to agents of that class used clinically. Typically, this is 1-100 mg/mL. 
     The invention encompasses all conventional modes of administration; oral and parenteral methods are preferred. Generally, the dosing regimen will be similar to other agents used clinically. Typically, the daily dose will be about 1-100 mg/kg body weight daily. Generally, more compound is required orally and less parenterally. The specific dosing regimen, however, will be determined by a physician using sound medical judgment. 
     The invention also encompasses methods where the compound is given in combination therapy. That is, the compound can be used in conjunction with, but separately from, other agents useful in treating hepatitis and HCV infection. In these combination methods, the compound will generally be given in a daily dose of 1-100 mg/kg body weight daily in conjunction with other agents. The other agents generally will be given in the amounts used therapeutically. The specific dosing regimen, however, will be determined by a physician using sound medical judgment. 
     Some examples of compounds suitable for compositions and methods as part of the invention herein are listed in Table 1. 
     
       
         
           
               
               
               
               
             
               
                 TABLE 1 
               
               
                   
               
               
                   
                   
                 Type of Inhibitor or 
                   
               
               
                 Brand Name 
                 Physiological Class 
                 Target 
                 Source Company 
               
               
                   
               
             
            
               
                 NIM811 
                   
                 Cyclophilin Inhibitor 
                 Novartis 
               
               
                 Zadaxin 
                   
                 Immuno-modulator 
                 Sciclone 
               
               
                 Suvus 
                   
                 Methylene blue 
                 Bioenvision 
               
               
                 Actilon 
                   
                 TLR9 agonist 
                 Coley 
               
               
                 (CPG10101) 
               
               
                 Batabulin (T67) 
                 Anticancer 
                 β-tubulin inhibitor 
                 Tularik Inc., South 
               
               
                   
                   
                   
                 San Francisco, CA 
               
               
                 ISIS 14803 
                 Antiviral 
                 antisense 
                 ISIS 
               
               
                   
                   
                   
                 Pharmaceuticals Inc, 
               
               
                   
                   
                   
                 Carlsbad, CA/Elan 
               
               
                   
                   
                   
                 Phamaceuticals Inc., 
               
               
                   
                   
                   
                 New York, NY 
               
               
                 Summetrel 
                 Antiviral 
                 antiviral 
                 Endo 
               
               
                   
                   
                   
                 Pharmaceuticals 
               
               
                   
                   
                   
                 Holdings Inc., 
               
               
                   
                   
                   
                 Chadds Ford, PA 
               
               
                 GS-9132 (ACH- 
                 Antiviral 
                 HCV Inhibitor 
                 Achillion/Gilead 
               
               
                 806) 
               
               
                 Pyrazolopyrimidine 
                 Antiviral 
                 HCV Inhibitors 
                 Arrow Therapeutics 
               
               
                 compounds and 
                   
                   
                 Ltd. 
               
               
                 salts 
               
               
                 From WO- 
               
               
                 2005047288 
               
               
                 26 May 2005 
               
               
                 Levovirin 
                 Antiviral 
                 IMPDH inhibitor 
                 Ribapharm Inc., 
               
               
                   
                   
                   
                 Costa Mesa, CA 
               
               
                 Merimepodib 
                 Antiviral 
                 IMPDH inhibitor 
                 Vertex 
               
               
                 (VX-497) 
                   
                   
                 Pharmaceuticals 
               
               
                   
                   
                   
                 Inc., Cambridge, 
               
               
                   
                   
                   
                 MA 
               
               
                 XTL-6865 (XTL- 
                 Antiviral 
                 monoclonal antibody 
                 XTL 
               
               
                 002) 
                   
                   
                 Biopharmaceuticals 
               
               
                   
                   
                   
                 Ltd., Rehovot, Israel 
               
               
                 Telaprevir 
                 Antiviral 
                 NS3 serine protease 
                 Vertex 
               
               
                 (VX-950, LY- 
                   
                 inhibitor 
                 Pharmaceuticals 
               
               
                 570310) 
                   
                   
                 Inc., Cambridge, 
               
               
                   
                   
                   
                 MA/Eli Lilly and 
               
               
                   
                   
                   
                 Co. Inc., 
               
               
                   
                   
                   
                 Indianapolis, IN 
               
               
                 HCV-796 
                 Antiviral 
                 NS5B Replicase 
                 Wyeth/Viropharma 
               
               
                   
                   
                 Inhibitor 
               
               
                 NM-283 
                 Antiviral 
                 NS5B Replicase 
                 Idenix/Novartis 
               
               
                   
                   
                 Inhibitor 
               
               
                 GL-59728 
                 Antiviral 
                 NS5B Replicase 
                 Gene Labs/ 
               
               
                   
                   
                 Inhibitor 
                 Novartis 
               
               
                 GL-60667 
                 Antiviral 
                 NS5B Replicase 
                 Gene Labs/ 
               
               
                   
                   
                 Inhibitor 
                 Novartis 
               
               
                 2′C MeA 
                 Antiviral 
                 NS5B Replicase 
                 Gilead 
               
               
                   
                   
                 Inhibitor 
               
               
                 PSI 6130 
                 Antiviral 
                 NS5B Replicase 
                 Roche 
               
               
                   
                   
                 Inhibitor 
               
               
                 R1626 
                 Antiviral 
                 NS5B Replicase 
                 Roche 
               
               
                   
                   
                 Inhibitor 
               
               
                 2′C Methyl 
                 Antiviral 
                 NS5B Replicase 
                 Merck 
               
               
                 adenosine 
                   
                 Inhibitor 
               
               
                 JTK-003 
                 Antiviral 
                 RdRp inhibitor 
                 Japan Tobacco Inc., 
               
               
                   
                   
                   
                 Tokyo, Japan 
               
               
                 Levovirin 
                 Antiviral 
                 ribavirin 
                 ICN 
               
               
                   
                   
                   
                 Pharmaceuticals, 
               
               
                   
                   
                   
                 Costa Mesa, CA 
               
               
                 Ribavirin 
                 Antiviral 
                 ribavirin 
                 Schering-Plough 
               
               
                   
                   
                   
                 Corporation, 
               
               
                   
                   
                   
                 Kenilworth, NJ 
               
               
                 Viramidine 
                 Antiviral 
                 Ribavirin Prodrug 
                 Ribapharm Inc., 
               
               
                   
                   
                   
                 Costa Mesa, CA 
               
               
                 Heptazyme 
                 Antiviral 
                 ribozyme 
                 Ribozyme 
               
               
                   
                   
                   
                 Pharmaceuticals 
               
               
                   
                   
                   
                 Inc., Boulder, CO 
               
               
                 BILN-2061 
                 Antiviral 
                 serine protease 
                 Boehringer 
               
               
                   
                   
                 inhibitor 
                 Ingelheim Pharma 
               
               
                   
                   
                   
                 KG, Ingelheim, 
               
               
                   
                   
                   
                 Germany 
               
               
                 SCH 503034 
                 Antiviral 
                 serine protease 
                 Schering Plough 
               
               
                   
                   
                 inhibitor 
               
               
                 Zadazim 
                 Immune modulator 
                 Immune modulator 
                 SciClone 
               
               
                   
                   
                   
                 Pharmaceuticals 
               
               
                   
                   
                   
                 Inc., San Mateo, CA 
               
               
                 Ceplene 
                 Immunomodulator 
                 immune modulator 
                 Maxim 
               
               
                   
                   
                   
                 Pharmaceuticals 
               
               
                   
                   
                   
                 Inc., San Diego, CA 
               
               
                 CellCept 
                 Immunosuppressant 
                 HCV IgG immuno- 
                 F. Hoffmann-La 
               
               
                   
                   
                 suppressant 
                 Roche LTD, Basel, 
               
               
                   
                   
                   
                 Switzerland 
               
               
                 Civacir 
                 Immunosuppressant 
                 HCV IgG immuno- 
                 Nabi 
               
               
                   
                   
                 suppressant 
                 Biopharmaceuticals 
               
               
                   
                   
                   
                 Inc., Boca Raton, FL 
               
               
                 Albuferon-α 
                 Interferon 
                 albumin IFN-α2b 
                 Human Genome 
               
               
                   
                   
                   
                 Sciences Inc., 
               
               
                   
                   
                   
                 Rockville, MD 
               
               
                 Infergen A 
                 Interferon 
                 IFN 
                 InterMune 
               
               
                   
                   
                 alfacon-1 
                 Pharmaceuticals 
               
               
                   
                   
                   
                 Inc., Brisbane, CA 
               
               
                 Omega IFN 
                 Interferon 
                 IFN-ω 
                 Intarcia Therapeutics 
               
               
                 IFN-β and EMZ701 
                 Interferon 
                 IFN-β and EMZ701 
                 Transition 
               
               
                   
                   
                   
                 Therapeutics Inc., 
               
               
                   
                   
                   
                 Ontario, Canada 
               
               
                 Rebif 
                 Interferon 
                 IFN-β1a 
                 Serono, Geneva, 
               
               
                   
                   
                   
                 Switzerland 
               
               
                 Roferon A 
                 Interferon 
                 IFN-α2a 
                 F. Hoffmann-La 
               
               
                   
                   
                   
                 Roche LTD, Basel, 
               
               
                   
                   
                   
                 Switzerland 
               
               
                 Intron A 
                 Interferon 
                 IFN-α2b 
                 Schering-Plough 
               
               
                   
                   
                   
                 Corporation, 
               
               
                   
                   
                   
                 Kenilworth, NJ 
               
               
                 Intron A and 
                 Interferon 
                 IFN-α2b/α1-thymosin 
                 RegeneRx 
               
               
                 Zadaxin 
                   
                   
                 Biopharma. Inc., 
               
               
                   
                   
                   
                 Bethesda, MD/ 
               
               
                   
                   
                   
                 SciClone 
               
               
                   
                   
                   
                 Pharmaceuticals Inc, 
               
               
                   
                   
                   
                 San Mateo, CA 
               
               
                 Rebetron 
                 Interferon 
                 IFN-α2b/ribavirin 
                 Schering-Plough 
               
               
                   
                   
                   
                 Corporation, 
               
               
                   
                   
                   
                 Kenilworth, NJ 
               
               
                 Actimmune 
                 Interferon 
                 INF-γ 
                 InterMune Inc., 
               
               
                   
                   
                   
                 Brisbane, CA 
               
               
                 Interferon-β 
                 Interferon 
                 Interferon-β-1a 
                 Serono 
               
               
                 Multiferon 
                 Interferon 
                 Long lasting IFN 
                 Viragen/ 
               
               
                   
                   
                   
                 Valentis 
               
               
                 Wellferon 
                 Interferon 
                 Lympho-blastoid IFN- 
                 GlaxoSmithKline 
               
               
                   
                   
                 αn1 
                 plc, Uxbridge, UK 
               
               
                 Omniferon 
                 Interferon 
                 natural IFN-α 
                 Viragen Inc., 
               
               
                   
                   
                   
                 Plantation, FL 
               
               
                 Pegasys 
                 Interferon 
                 PEGylated IFN-α2a 
                 F. Hoffmann-La 
               
               
                   
                   
                   
                 Roche LTD, Basel, 
               
               
                   
                   
                   
                 Switzerland 
               
               
                 Pegasys and 
                 Interferon 
                 PEGylated IFN-α2a/ 
                 Maxim 
               
               
                 Ceplene 
                   
                 immune modulator 
                 Pharmaceuticals 
               
               
                   
                   
                   
                 Inc., San Diego, CA 
               
               
                 Pegasys and 
                 Interferon 
                 PEGylated IFN- 
                 F. Hoffmann-La 
               
               
                 Ribavirin 
                   
                 α2a/ribavirin 
                 Roche LTD, Basel, 
               
               
                   
                   
                   
                 Switzerland 
               
               
                 PEG-Intron 
                 Interferon 
                 PEGylated IFN-α2b 
                 Schering-Plough 
               
               
                   
                   
                   
                 Corporation, 
               
               
                   
                   
                   
                 Kenilworth, NJ 
               
               
                 PEG-Intron/ 
                 Interferon 
                 PEGylated IFN- 
                 Schering-Plough 
               
               
                 Ribavirin 
                   
                 α2b/ribavirin 
                 Corporation, 
               
               
                   
                   
                   
                 Kenilworth, NJ 
               
               
                 IP-501 
                 Liver protection 
                 antifibrotic 
                 Indevus 
               
               
                   
                   
                   
                 Pharmaceuticals 
               
               
                   
                   
                   
                 Inc., Lexington, MA 
               
               
                 IDN-6556 
                 Liver protection 
                 caspase inhibitor 
                 Idun 
               
               
                   
                   
                   
                 Pharmaceuticals 
               
               
                   
                   
                   
                 Inc., San Diego, CA 
               
               
                 ITMN-191 (R-7227) 
                 Antiviral 
                 serine protease 
                 InterMune 
               
               
                   
                   
                 inhibitor 
                 Pharmaceuticals 
               
               
                   
                   
                   
                 Inc., Brisbane, CA 
               
               
                 GL-59728 
                 Antiviral 
                 NS5B Replicase 
                 Genelabs 
               
               
                   
                   
                 Inhibitor 
               
               
                 ANA-971 
                 Antiviral 
                 TLR-7 agonist 
                 Anadys 
               
               
                 Boceprevir 
                 Antiviral 
                 serine protease 
                 Schering Plough 
               
               
                   
                   
                 inhibitor 
               
               
                 TMS-435 
                 Antiviral 
                 serine protease 
                 Tibotec BVBA, 
               
               
                   
                   
                 inhibitor 
                 Mechelen, Belgium 
               
               
                 BI-201335 
                 Antiviral 
                 serine protease 
                 Boehringer 
               
               
                   
                   
                 inhibitor 
                 Ingelheim Pharma 
               
               
                   
                   
                   
                 KG, Ingelheim, 
               
               
                   
                   
                   
                 Germany 
               
               
                 MK-7009 
                 Antiviral 
                 serine protease 
                 Merck 
               
               
                   
                   
                 inhibitor 
               
               
                 PF-00868554 
                 Antiviral 
                 replicase inhibitor 
                 Pfizer 
               
               
                 ANA598 
                 Antiviral 
                 Non-Nucleoside 
                 Anadys 
               
               
                   
                   
                 NS5B Polymerase 
                 Pharmaceuticals, 
               
               
                   
                   
                 Inhibitor 
                 Inc., San Diego, CA, 
               
               
                   
                   
                   
                 USA 
               
               
                 IDX375 
                 Antiviral 
                 Non-Nucleoside 
                 Idenix 
               
               
                   
                   
                 Replicase Inhibitor 
                 Pharmaceuticals, 
               
               
                   
                   
                   
                 Cambridge, MA, 
               
               
                   
                   
                   
                 USA 
               
               
                 BILB 1941 
                 Antiviral 
                 NS5B Polymerase 
                 Boehringer 
               
               
                   
                   
                 Inhibitor 
                 Ingelheim Canada 
               
               
                   
                   
                   
                 Ltd R&amp;D, Laval, 
               
               
                   
                   
                   
                 QC, Canada 
               
               
                 PSI-7851 
                 Antiviral 
                 Nucleoside 
                 Pharmasset, 
               
               
                   
                   
                 Polymerase Inhibitor 
                 Princeton, NJ, USA 
               
               
                 PSI-7977 
                 Antiviral 
                 Nucleotide NS5B 
                 Pharmasset, 
               
               
                   
                   
                 Polymerase Inhibitor 
                 Princeton, NJ, USA 
               
               
                 VCH-759 
                 Antiviral 
                 NS5B Polymerase 
                 ViroChem Pharma 
               
               
                   
                   
                 Inhibitor 
               
               
                 VCH-916 
                 Antiviral 
                 NS5B Polymerase 
                 ViroChem Pharma 
               
               
                   
                   
                 Inhibitor 
               
               
                 GS-9190 
                 Antiviral 
                 NS5B Polymerase 
                 Gilead 
               
               
                   
                   
                 Inhibitor 
               
               
                 Peg-interferon 
                 Antiviral 
                 Interferon 
                 ZymoGenetics/Bristol- 
               
               
                 lambda 
                   
                   
                 Myers Squibb 
               
               
                   
               
            
           
         
       
     
     Synthesis Methods 
     The compounds may be made by methods available in the art, as well as those described below. Some reagents and intermediates are available in the art. Other reagents and intermediates can be made by methods available in the art using commercially available materials. The variables (e.g. numbered “R” substituents) used to describe the synthesis of the compounds are intended only to illustrate how to make these compounds, and are not to be confused with variables used in the claims or in other sections of the specification. Abbreviations used within the schemes generally follow conventions used in the art. 
     Abbreviations used in the schemes generally follow conventions used in the art. Chemical abbreviations used in the specification and examples are defined as follows: “NaHMDS” for sodium bis(trimethylsilyl)amide; “DMF” for N,N-dimethylformamide; “MeOH” for methanol; “NBS” for N-bromosuccinimide; “Ar” for aryl; “TFA” for trifluoroacetic acid; “LAH” for lithium aluminum hydride; “DMSO” for dimethylsulfoxide; “h” for hours; “rt” for room temperature or retention time (context will dictate); “min” for minutes; “EtOAc” for ethyl acetate; “THF” for tetrahydrofuran; “EDTA” for ethylenediaminetetraacetic acid; “Et 2 O” for diethyl ether; “DMAP” for 4-dimethylaminopyridine; “DCE” for 1,2-dichloroethane; “ACN” for acetonitrile; “DME” for 1,2-dimethoxyethane; “HOBt” for 1-hydroxybenzotriazole hydrate; “DIEA” for diisopropylethylamine. 
     For the section of compounds in the 0000 series all Liquid Chromatography (LC) data were recorded on a Shimadzu LC-10AS or LC-20AS liquid chromotograph using a SPD-10AV or SPD-20A UV-Vis detector and Mass Spectrometry (MS) data were determined with a Micromass Platform for LC in electrospray mode. 
       HPLC Method  (i.e., compound isolation). Compounds purified by preparative HPLC were diluted in methanol (1.2 mL) and purified using a Shimadzu LC-8A or LC-10A or Dionex APS-3000 or Waters Acquity™ automated preparative HPLC system. 
     EXAMPLES 
     Preparation of Compound 1001: 
                         
A mixture of 1H-pyrazole (54.2 mg), 5-(3-(tert-butylcarbamoyl)phenyl)-6-(N-ethylmethylsulfonamido)-2-(4-fluorophenyl)-N-methylbenzofuran-3-carboxamide (15 mg), K 2 CO 3  (14.66 mg) and copper (3.37 mg) was heated to 140° C. for 6 hours. The mixture was diluted with DMF and filtered to removed solid. The solution of mixture was subjected to purification by preparative HPLC system.
 
                                1001                     MS (M + H) +  Calcd.   614.2       MS (M + H) +  Observ.   614.2       Retention Time   1.35 min                 LC Condition                     Solvent A   90% Water-10% Methanol-0.1% TFA       Solvent B   10% Water-90% Methanol-0.1% TFA       Start % B    50       Final % B   100       Gradient Time   2 min       Flow Rate   1 mL/min       Wavelength   220       Solvent Pair   Water-Methanol-TFA       Column   PHENOMENEX-LUNA 2.0 × 30 mm 3 um                    
Preparation of Compounds 1002 and 1003:
 
                         
Compounds 1002 and 1003 were prepared via the same procedure towards compound 1001, using 1,2,3-triazole. LC condition for compounds 1002 and 1003 was the same for compound 100.
 
     
       
         
           
               
             
               
                   
               
               
                 1002 
               
               
                   
               
             
            
               
                   
               
            
           
           
               
               
               
            
               
                   
                 MS (M + H) +  Calcd. 
                 615.2 
               
               
                   
                 MS (M + H) +  Observ. 
                 615.2 
               
               
                   
                 Retention Time 
                 1.18 min 
               
               
                   
                   
               
            
           
         
       
     
                             1003                                                MS (M + H) +  Calcd.   615.2           MS (M + H) +  Observ.   615.2           Retention Time   1.20 min                        
Preparation of Compound 2001:
 
                         
NaH (8.33 mg, 60%) was added into a solution of 5-(3-(tert-butylcarbamoyl)phenyl)-6-chloro-2-(4-fluorophenyl)-N-methylfuro[2,3-b]pyridine-3-carboxamide (10 mg) and 3-(trifluoromethyl)-1H-pyrazole (28.4 mg) in DMF. The reaction was heated at 85° C. for 24 hour. The product was isolated by preparative HPLC system.
 
                                2001                     MS (M + H) +  Calcd.   696.2       MS (M + H) +  Observ.   696.2       Retention Time   2.38 min                 LC Condition                     Solvent A   90% Water-10% Methanol-0.1% TFA       Solvent B   10% Water-90% Methanol-0.1% TFA       Start % B    0       Final % B   100       Gradient Time   2 min       Flow Rate   1 mL/min       Wavelength   220       Solvent Pair   Water-Methanol-TFA       Column   PHENOMENEX-LUNA 2.0 × 30 mm 3 um                    
Preparation of Compound 2002:
 
Compound 2002 were prepared via the same procedure towards compound 2001, using 4-(trifluoromethyl)-1H-pyrazole. LC condition for compound 2002 was the same for compound 2001.
 
                                2002                                                       MS (M + H) +  Calcd.   696.2       MS (M + H) +  Observ.   696.2       Retention Time    2.40 min                    
Preparation of Compound 3001:
 
                         
A mixture of 1H-1,2,3-triazole (96 mg), 5-(3-(tert-butylcarbamoyl)phenyl)-2-(4-fluorophenyl)-N-methyl-6-(3,3,3-trifluoropropyl)furo[2,3-b]pyridine-3-carboxamide (15 mg), K 2 CO 3  (11.48 mg) and copper (3.52 mg) was heated to 140° C. for 6 hours. The mixture was diluted with DMF and filtered to removed solid. The solution of mixture was subjected to purification by preparative HPLC system.
 
                                3001                     MS (M + H) +  Calcd.   591.2       MS (M + H) +  Observ.   591.2       Retention Time   1.63 min                 LC Condition                     Solvent A   90% Water-10% Methanol-0.1% TFA       Solvent B   10% Water-90% Methanol-0.1% TFA       Start % B    50       Final % B   100       Gradient Time   2 min       Flow Rate   1 mL/min       Wavelength   220       Solvent Pair   Water-Methanol-TFA       Column   PHENOMENEX-LUNA 2.0 × 30 mm 3 um                    
Compounds 3002-3010 were prepared via the same procedure towards compound 3001, using the corresponding heterocycles. LC condition for compounds 3002-3010 was the same for compound 3001.
 
                                                     MS   MS   Retention               (M + H) +     (M + H) +     Time       Cmpd #   Structure   Calcd.   Observ.   (min)                  3002                         605.2   605.2   1.31               3003                         605.2   605.2   1.58               3004                         605.2   605.2   1.73               3005                         590.2   590.2   1.79               3006                         590.2   590.2   1.08               3007                         591.2   591.2   1.64               3008                         604.3   604.2   1.93               3009                         604.3   604.2   1.17               3010                         604.3   604.2   1.89                    
Preparation of Compound 4001:
 
Compound 4001 was prepared via the same procedure towards compound 3001, using pyridin-3-ol. LC condition for compound 4001 was the same for compound 3001.
 
     
       
         
           
               
             
               
                   
               
             
            
               
                 4001 
               
               
                 
                   
                     
                     
                         
                         
                     
                   
                 
               
               
                   
               
            
           
           
               
               
            
               
                 MS (M + H) +  Calcd. 
                 617.2 
               
               
                 MS (M + H) +  Observ. 
                 617.2 
               
               
                 Retention Time 
                  1.10 min 
               
            
           
           
               
            
               
                 LC Condition 
               
            
           
           
               
               
            
               
                 Solvent A 
                  90% Water −10% Methanol-0.1% TFA 
               
               
                 Solvent B 
                  10% Water −90% Methanol-0.1% TFA 
               
               
                 Start % B 
                  50 
               
               
                 Final % B 
                 100 
               
               
                 Gradient Time 
                  2 min 
               
               
                 Flow Rate 
                  1 mL/min 
               
               
                 Wavelength 
                 220 
               
               
                 Solvent Pair 
                 Water - Methanol-TFA 
               
               
                 Column 
                 PHENOMENEX-LUNA 2.0 × 30 mm 3 um 
               
               
                   
               
            
           
         
       
     
     Biological Methods 
     The compound demonstrated activity against HCV NS5B as determined in the following HCV RdRp assays. 
     HCV NS5B RdRp cloning, expression, and purification. The cDNA encoding NS5B proteins of HCV genotype 1b (Con1), a genotype 1b variant with amino acid 316 mutated from cysteine to asparagine, and genotype 2a (JFH-1), were cloned into the pET21a expression vector. Each untagged protein was expressed with an 18 amino acid C-terminal truncation to enhance the solubility. The  E. coli  competent cell line BL21(DE3) was used for expression of the protein. Cultures were grown at 37° C. for ˜4 hours until the cultures reached an optical density of 2.0 at 600 nm. The cultures were cooled to 20° C. and induced with 1 mM IPTG. Fresh ampicillin was added to a final concentration of 50 μg/mL and the cells were grown overnight at 20° C. 
     Cell pellets (3 L) were lysed for purification to yield 15-24 mgs of purified NS5B. The lysis buffer consisted of 20 mM Tris-HC1, pH 7.4, 500 mM NaCl, 0.5% triton X-100, 1 mM DTT, 1 mM EDTA, 20% glycerol, 0.5 mg/mL lysozyme, 10 mM MgCl 2 , 15 ug/mL deoxyribonuclease I, and Complete TM protease inhibitor tablets (Roche). After addition of the lysis buffer, frozen cell pellets were resuspended using a tissue homogenizer. To reduce the viscosity of the sample, aliquots of the lysate were sonicated on ice using a microtip attached to a Branson sonicator. The sonicated lysate was centrifuged at 100,000×g for 30 minutes at 4° C. and filtered through a 0.2 μm filter unit (Corning). 
     The protein was purified using two sequential chromatography steps: Heparin sepharose CL-6B and polyU sepharose 4B. The chromatography buffers were identical to the lysis buffer but contained no lysozyme, deoxyribonuclease I, MgCl 2  or protease inhibitor and the NaCl concentration of the buffer was adjusted according to the requirements for charging the protein onto the column. Each column was eluted with a NaCl gradient which varied in length from 5-50 column volumes depending on the column type. After the final chromatography step, the resulting purity of the enzyme is &gt;90% based on SDS-PAGE analysis. The enzyme was aliquoted and stored at −80° C. 
     HCV NS5B RdRp enzyme assay. An on-bead solid phase homogeneous assay was used in a 384-well format to assess NS5B inhibitors (WangY-K, Rigat K, Roberts S, and Gao M (2006) Anal Biochem, 359: 106-111). The biotinylated oligo dT 12  primer was captured on streptavidin-coupled imaging beads (GE, RPNQ0261) by mixing primer and beads in 1× buffer and incubating at room temperature for three hours. Unbound primer was removed after centrifugation. The primer-bound beads were resuspended in 3× reaction mix (20 mM Hepes buffer, pH 7.5, dT primer coupled beads, poly A template,  3 H-UTP, and RNAse inhibitor (Promega N2515)). Compounds were serially diluted 1:3 in DMSO and aliquoted into assay plates. Equal volumes (5 μL) of water, 3× reaction mix, and enzyme in 3× assay buffer (60 mM Hepes buffer, pH 7.5, 7.5 mM MgCl 2 , 7.5 mM KCl, 3 mM DTT, 0.03 mg/mL BSA, 6% glycerol) were added to the diluted compound on the assay plate. Final concentration of components in 384-well assay: 0.36 nM template, 15 nM primer, 0.29 μM 3 H-UTP (0.3 μCi), 1.6 U/μL RNAse inhibitor, 7 nM NS5B enzyme, 0.01 mg/mL BSA, 1 mM DTT, and 0.33 μg/μL beads, 20 mM Hepes buffer, pH 7.5, 2.5 mM MgCl 2 , 2.5 mM KCl, and 0.1% DMSO. 
     Reactions were allowed to proceed for 24 hours at 30° C. and terminated by the addition of 50 mM EDTA (5 μL). After incubating for at least 15 minutes, plates were read on an Amersham LEADseeker multimodality imaging system. 
     IC 50  values for compounds were determined using ten different [I]. IC 50  values were calculated from the inhibition using the four-parameter logistic formula y=A+ ((B−A)/(1+((C/x)^D))), where A and B denote minimal and maximal % inhibition, respectively, C is the IC 50 , D is hill slope and x represents compound concentration. 
     Cell lines. The cell lines used to evaluate compounds consist of a human hepatocyte derived cell line (Huh-7) that constitutively expresses a genotype 1b (Con-1) HCV replicon or a genotype 1b (Con-1) HCV replicon with an asparagine replacing the cysteine at amino acid 316, or a genotype 2a (JFH-1) replicon, containing a Renilla luciferase reporter gene. These cells were maintained in Dulbecco&#39;s modified Eagle medium (DMEM) containing 10% FBS, 100 U/mL penicillin/streptomycin and 1.0 mg/mL G418. 
     HCV replicon luciferase assay. To evaluate compound efficacy, titrated compounds were transferred to sterile 384-well tissue culture treated plates, and the plates were seeded with HCV replicon cells (50 μL at a density of 2.4×10 3  cells/well) in DMEM containing 4% FBS (final DMSO concentration at 0.5%). After 3 days incubation at 37° C., cells were analyzed for Renilla Luciferase activity using the EnduRen substrate (Promega cat #E6485) according to the manufacturer&#39;s directions. Briefly, the EnduRen substrate was diluted in DMEM and then added to the plates to a final concentration of 7.5 μM. The plates were incubated for at least 1 h at 37° C. then read on a Viewlux Imager (PerkinElmer) using a luminescence program. The 50% effective concentration (EC 50 ) was calculated using the four-parameter logistic formula noted above. 
     To assess cytotoxicity of compounds, Cell Titer-Blue (Promega) was added to the EnduRen-containing plates and incubated for at least 4 hrs at 37° C. The fluorescence signal from each well was read using a Viewlux Imager. All CC 50  values were calculated using the four-parameter logistic formula. 
     Compound EC 50  data is expressed as A:&lt;100 nM; B=100-1000 nM; C&gt;1000 nM). Representative data for compounds are reported in Table 2. 
     
       
         
           
               
               
               
             
               
                 TABLE 2 
               
               
                   
               
               
                   
                   
                 EC 50  (uM) 
               
               
                 Cmpd # 
                 Structure 
                 lb 
               
               
                   
               
             
            
               
                 1001 
                 
                   
                     
                     
                         
                         
                     
                   
                 
                 0.017 A 
               
               
                   
               
               
                 1002 
                 
                   
                     
                     
                         
                         
                     
                   
                 
                 0.069 A 
               
               
                   
               
               
                 1003 
                 
                   
                     
                     
                         
                         
                     
                   
                 
                 A 
               
               
                   
               
               
                 2001 
                 
                   
                     
                     
                         
                         
                     
                   
                 
                 B 
               
               
                   
               
               
                 2002 
                 
                   
                     
                     
                         
                         
                     
                   
                 
                 C 
               
               
                   
               
               
                 3001 
                 
                   
                     
                     
                         
                         
                     
                   
                 
                 0.035 A 
               
               
                   
               
               
                 3002 
                 
                   
                     
                     
                         
                         
                     
                   
                 
                 C 
               
               
                   
               
               
                 3003 
                 
                   
                     
                     
                         
                         
                     
                   
                 
                 3.32  C 
               
               
                   
               
               
                 3004 
                 
                   
                     
                     
                         
                         
                     
                   
                 
                 B 
               
               
                   
               
               
                 3006 
                 
                   
                     
                     
                         
                         
                     
                   
                 
                 B 
               
               
                   
               
               
                 3007 
                 
                   
                     
                     
                         
                         
                     
                   
                 
                 B 
               
               
                   
               
               
                 3008 
                 
                   
                     
                     
                         
                         
                     
                   
                 
                 C 
               
               
                   
               
               
                 3009 
                 
                   
                     
                     
                         
                         
                     
                   
                 
                 C 
               
               
                   
               
               
                 3010 
                 
                   
                     
                     
                         
                         
                     
                   
                 
                 0.28  B 
               
               
                   
               
               
                 4001 
                 
                   
                     
                     
                         
                         
                     
                   
                 
                 B 
               
               
                   
               
            
           
         
       
     
     It will be evident to one skilled in the art that the present disclosure is not limited to the foregoing illustrative examples, and that it can be embodied in other specific forms without departing from the essential attributes thereof. It is therefore desired that the examples be considered in all respects as illustrative and not restrictive, reference being made to the appended claims, rather than to the foregoing examples, and all changes which come within the meaning and range of equivalency of the claims are therefore intended to be embraced therein.