WO2011076923A1

WO2011076923A1 - Inhibitor for inosine monophosphate (imp) dehydrogenase and/or viral rna polymerase for treatment of hepatitis e

Info

Publication number: WO2011076923A1
Application number: PCT/EP2010/070660
Authority: WO
Inventors: Vincent Mallet; Stanislas Pol
Original assignee: Institut National De La Sante Et De La Recherche Medicale; Centre National De La Recherche Scientifique (C.N.R.S); Assistance Publique - Hopitaux De Paris; Universite Paris Descartes
Priority date: 2009-12-23
Filing date: 2010-12-23
Publication date: 2011-06-30
Also published as: WO2011076923A9

Abstract

The present invention relates to an inhibitor of IMP dehydrogenase, an inhibitor of viral RNA polymerase and a compound of formula (I) for use in the treatment of an infection caused by a virus from the genus Hepevirus in a subject, in particular for use in the treatment of Hepatitis E.

Description

INHIBITOR OF INOSINE MONOPHOSPHATE (IMP) DEHYDROGENASE AND/OR VIRAL RNA

POLYMERASE FOR TREATMENT OF HEPATITIS E

The present invention relates to methods for treating hepatitis E. Viral hepatitis is an important cause of morbidity and mortality. Two viruses have been identified as agents responsible for enterically transmitted hepatitis: hepatitis A virus and hepatitis E virus (HEV).

HEV is a small non-enveloped, polyadenylated, single-stranded, positive-sense RNA virus containing a genome of 7.2 kb. It is icosahedral in shape and its diameter is of 27-34 nm. To date, it is the only member of the Hepevirus genus of the Hepeviridae family. At least four genotypes of HEV have been identified: genotypes 1 and 2 are strictly human, genotype 3 is probably of swine origin but can also infect humans, and genotype 4 is of avian origin and has not been identified as infecting humans yet. All these genotypes belong to only one serotype. HEV is mainly transmitted enterically through contaminated water or food but may also be transmitted parenterally or from mother to child (Dalton et al. (2008) Lancet Infect. Dis. 8:698-709).

The clinical presentations of HEV infection range from a self-limited acute hepatitis with no long-term sequelae to severe forms, including jaundice, dark urine, anorexia, nausea, vomiting and fever lasting for 1 to 6 weeks and sometimes to fulminant hepatic failure. The incubation period lasts usually from 3 to 6 weeks. The reported mortality rates are lower (1 -4%) in endemic countries where infection happens early in life (Mast et at. (1996) Annu. Rev. Med. 47:257-266) than in developed countries (8-1 1 %) were infection happens later (Dalton et al. (2008) Eur. J. Clin. Microbiol. Infect. Dis. 27:579-585). Mortality rates reach high levels in pregnant women (20%), in particular in their third trimester (Mast et al. (1996) Annu. Rev. Med. 47:257-266). HEV superinfection in patients with chronic liver disease has a reported mortality rate of 70% (Dalton et al. (2007) Lancet 369:1260; Peron et al. (2007) J. Viral. Hepat. 14:298-303).

Serologic tests showed that HEV is the most important cause of acute clinical hepatitis among adults throughout Central and Southeast Asia and the second most important cause, behind hepatitis B virus, throughout the Middle East and North Africa. In industrialized countries, hepatitis E is considered an emerging disease due to travel to highly endemic regions. Nevertheless, a growing number of sporadic cases of HEV infection have been identified in developed countries as caused by spreading of autochtonous viral strains (Borgen et al. (2008) BMC Infect. Dis. 8:61 ). Additionally, HEV has been identified as a zoonosis, in particular in swine which may serve as a reservoir for human HEV infection (Feagins et al. (2007) J. Gen. Virol. 88:912-917). Recently, cases of persistent HEV infection with chronic liver disease, sometimes leading very rapidly to cirrhosis, were identified in patients with reduced immune surveillance, such as patients under chemotherapy or immunosuppression, in particular in patients with solid organ transplant recipients (Kamar et al. (2008) N. Engl. J. Med. 358:81 1 -81 7; Haagsma et al. (2008) Liver Transpl. 14:547-553).

Nevertheless, there is currently no specific treatment for HEV infection. In particular, treatments used against other hepatitis have not been identified as efficient against HEV infection.

There is accordingly a need for new treatments against hepatitis E, in particular against chronic hepatitis E, which would be in particular useful in immunodeficient people.

Ribavirin, also known as 1 -[(2/^:?,3/^:?,4S,5/^:?)-3,4-dihydroxy-5- (hydroxymethyl)oxolan-2-yl]-1 H-1 ,2,4-triazole-3-carboxamide or 1 -( -D-ribofuranosyl)-1 H- 1 ,2,4-triazole-3-carboxamide, and commercialized under the brand names Copegus®, Rebetol®, Ribasphere®, Vilona® and Virazole® is an anti-viral drug indicated for severe respiratory syncytial virus (RSV) infection and hepatitis C infection (in conjuction with peginterferon oc-2b or peginterferon oc-2a). Its CAS number is 36791 -04-5 and it is represented by the following formula (V):

Ribavirin is a prodrug, which is a mimetic of purine RNA nucleotides when metabolised, thus interfering with RNA metabolism required for viral replication. Ribavirin and derivatives thereof are in particular described in the US patents 3,798,209 and 4,21 1 ,771 , which also describe the efficiency of ribavirin for treating Herpes simplex virus, influenza, parainfluenza, chickenpox, mumps, measles and herpes zoster infections.

Summary of the invention

The present inventors have unexpectedly shown that ribavirin was a potent drug for treating HEV infection, specifically for treating chronic HEV infection. They have in particular demonstrated that ribavirin induced a fast clearance of HEV accompanied by a complete normalization of liver function tests. Without wishing to be bound to a particular theory, this could be due to a depletion in guanosine via an inhibition of the inosine monophosphate (IMP) dehydrogenase and/or to a direct interaction of ribavirin triphosphate with the viral RNA polymerase.

Detailed description of the invention

The present invention thus relates to an inhibitor of inosine monophosphate (IMP) dehydrogenase inhibitor and/or an inhibitor of viral RNA polymerase for use in the treatment of an infection caused by a virus from the genus Hepevirus in a subject.

The present invention also relates to a compound of the following formula (I):

wherein:

R₄ is H, -OH, -O-alkyl, -O-CO-alkyl, an halogen, -0-CO-(W)_n-R₇, -0-OP(OH)₂-CS- (W)_n-R₇, -0-OP(OH)-(W)_n-R₇, -O-SO₂-OH or a protecting group;

R₅ is H, -OH, -O-alkyl, -O-CO-alkyl, an halogen, -0-CO-(W)_n-R₇, -0-OP(OH)₂-CS- (W)_n-R₇, -0-OP(OH)-(W)_n-R₇, -O-SO₂-OH, -SH, -S-alkyl, -N₃ or a protecting group;

R₆ is H, alkyl, -CO-alkyl, phosphate, diphosphate, triphosphate, phosphonate, -CO- (W)_n-R₇, -OP(OH)₂-CS-(W)_n-R₇, -OP(OH)-(W)_n-R₇, -S0₂-OH or a protecting group;

R₇ is H, alkyl, acyl, cycloalkyl, heterocyclic, aryl, NR_8aR_8b, alkenyl or alkynyl; or R₇ is alkyl, acyl, alkenyl or alkynyl substituted by halo, phenyl, cycloalkyl, NR_8aR_8b, hydroxyl or alkoxy; or R₇ is aryl substituted by phenyl, halo, CN, N0₂, OH, R₉, OR₉, CF₃, SH, SR_8a, SOR_8a, S0₂R_8a, NR_8aR_8b, C0₂H, C0₂ ^" M⁺, O^" M⁺, OR_8a or S^" M⁺; or R₇ is -(CHR_8a)_e-(CH₂)_f- CO-OR_8b, -(CHR_8a)_e-(CH₂)_fOR_8b, or -(CHR_8a)_e-(CH₂)_f-NR_8aR_8b;

M⁺ is an alkali metal cation;

R_8a is H, alkyl, acyl, aryl or is alkyl, acyl or aryl substituted by halo, phenyl, CN, N0₂, OH, C0₂H or alkoxy, and R_8b is H, alkyl or aryl or is alkyl or aryl substituted by halo, CN, N0₂, C0₂H, OH or alkoxy; or R_8a and R_8b taken together with N and one of CHR_8a, NR_8a, O, S, SO or S0₂ form a five-, six- or seven- membered ring;

R₉ is H, aryl, alkyl, or alkyl substituted by OH, halo, NR_8aR_8b, or acyl;

Het is a heteroaryl with 5 to 7 ring atoms and at least one nitrogen atom; and e = 0 to 6, f = 0 to 10 and n = 0 or 1 ; or its pharmaceutically acceptable salts, its N-oxide forms; hydrates or hydrated salts, its polymorphic crystalline structures, its tautomers, racemates, diastereisomers or enantiomers;

for use in the treatment of an infection caused by a virus from the genus Hepevirus in a subject.

Definitions

The term "alkyl" means a saturated or unsaturated aliphatic hydrocarbon group which may be straight or branched having 1 to 12 carbon atoms in the chain. Preferred alkyl groups have 1 to 6 carbon atoms in the chain. "Branched" means that one or lower alkyl groups such as methyl, ethyl or propyl are attached to a linear alkyl chain. The alkyl may be substituted with one or more «alkyl group substituants» which may be the same or different, and include for instance halo, cycloalkyl, hydroxy, alkoxy, amino, acylamino, aroylamino, carboxy.

The terms "arylalkyl" or "aralkyl" refer to an alkyl moiety in which an alkyl hydrogen atom is replaced by an aryl group. Examples of "arylalkyl" or "aralkyl" include benzyl and 9-fluorenyl groups.

The term "halo", "halogen" or "Hal" refers to the atoms of the group 17 of the periodic table (halogens) and includes in particular fluorine, chlorine, bromine, and iodine atom, preferably chlorine, bromine or iodine.

The term "cycloalkyl" as employed herein includes saturated cyclic, bicyclic, tricyclic, or polycyclic hydrocarbon groups having 3 to 12 carbons, wherein any ring atom capable of substitution may be substituted by a substituent. Examples of cycloalkyl moieties include, but are not limited to, cyclohexyl and adamantyl.

The term "aryl" refers to an aromatic monocyclic, bicyclic, or tricyclic hydrocarbon ring system, wherein any ring atom capable of substitution may be substituted by a substituent. Examples of aryl moieties include, but are not limited to, phenyl, naphthyl, and anthracenyl.

The term "heterocyclyl" refers to a nonaromatic 3-10 membered monocyclic, 8-12 membered bicyclic, or 1 1 -14 membered tricyclic ring system having 1 -3 heteroatoms if monocyclic, 1 -6 heteroatoms if bicyclic, or 1 -9 heteroatoms if tricyclic, said heteroatoms selected from O, N, or S (e. g. , carbon atoms and 1 -3, 1 -6, or 1 -9 heteroatoms of N, O, or S if monocyclic, bicyclic, or tricyclic, respectively), wherein any ring atom capable of substitution may be substituted by a substituent.

The term "heteroaryl" refers to an aromatic 5-8 membered monocyclic, 8-12 membered bicyclic, or 1 1 -14 membered tricyclic ring system having 1 -3 heteroatoms if monocyclic, 1 -6 heteroatoms if bicyclic, or 1 -9 heteroatoms if tricyclic, said heteroatoms selected from O, N, or S (e. g. , carbon atoms and 1 -3, 1 -6, or 1 -9 heteroatoms of N, O, or S if monocyclic, bicyclic, or tricyclic, respectively), wherein any ring atom capable of substitution may be substituted by a substituent.

The term "alkoxy" refers to an -O-alkyl radical.

The term "substituents" refers to a group "substituted" on an alkyl, cycloalkyl, heterocyclyl, aryl, aralkyl or heteroaryl group at any atom of that group. Suitable substituents include, without limitation, alkyl, alkenyl, alkynyl, alkoxy, halo, hydroxy, cyano, nitro, amino, S0₃H, sulfate, phosphate, perfluoroalkyl, perfluoroalkoxy, methylenedioxy, ethylenedioxy, carboxyl, oxo, thioxo, imino (alkyl, aryl, aralkyl), S(0)_n alkyl (where n is 0-2), S(0)_n aryl (where n is 0-2), S(0)_n heteroaryl (where n is 0-2), S(0)_n heterocyclyl (where n is 0- 2), amine (mono-, di-, alkyl, cycloalkyl, aralkyl, heteroaralkyl, and combinations thereof), ester (alkyl, aralkyl, heteroaralkyl), amide (mono-, di-, alkyl, aralkyl, heteroaralkyl, and combinations thereof), sulfonamide (mono-, di-, alkyl, aralkyl, heteroaralkyl, and combinations thereof), unsubstituted aryl, unsubstituted heteroaryl, unsubstituted heterocyclyl, and unsubstituted cycloalkyl.

The preferred substituents on aryl or heteroaryl groups are amino, amine, alkoxy, halo, perfluoroalkyl such as CF₃, heterocyclyl, amide, and ester.

The term "acyl" refers to an alkylcarbonyl, cycloalkylcarbonyl, arylcarbonyl, heterocyclylcarbonyl, or heteroarylcarbonyl substituent, any of which may be further substituted by substituents.

The term "oxo" refers to an oxygen atom, which forms a carbonyl when attached to carbon, an N-oxide when attached to nitrogen, and a sulfoxide or sulfone when attached to sulfur.

The term "alkenyl" as employed herein includes partially unsaturated, nonaromatic, hydrocarbon groups having 2 to 12 carbons, preferably 2 to 6 carbons.

The term "alkynyl" as employed herein includes unsaturated, nonaromatic, hydrocarbon groups having 2 to 12 carbons, preferably 2 to 6 carbons, and comprising at least one triple bond.

The term "alkylene" as employed herein refers to a divalent radical comprising from

1 to 12 carbon atoms, and preferably from 1 to 6 carbon atoms. Said radical may be represented by the formula (CH₂)_n wherein n is an integer varying from 1 to 12, and preferably from 1 to 6.

The term "monohalogenated alkynyl" refers to -C≡C-Hal or -C≡alk(Hal) where alk(Hal) denotes a linear or branched C Ci₀ alkyl radical, one hydrogen atom of which being substituted by one halogen atom Hal. The term "dihalogenated alkenyl" is -C(Hal')=CH(Hal), -C(Hal')=C(Hal)alkyl or - C(Hal')=C-alk(Hal), where Hal' is halogen and alk(Hal) denotes a linear or branched d- Cio alkyl radical, one hydrogen atom of which being substituted by one halogen atom Hal. The dihalogenated alkenyl may present a Z or E configuration, the £ configuration being preferred, i.e. the two halogen atoms Hal and Hal' are not on the same side of the planar double bond. Preferred configuration for -C(Hal')=CH(Hal) or -C(Hal')=C(Hal)alkyl is therefore respectively:

In the context of the invention, the term "alkali metal cation" refers in particular to lithium, potassium and sodium cation.

The term "phosphate" refers to the -P(=0)(OR)₃ group, wherein R can be H, alkyl, aralkyl, aryl, or a negative charge.

The term "phosphonate" refers to the -P(=0)(OR)₂ group, wherein R can be H, alkyl, aralkyl, aryl, or a negative charge.

The compounds herein described may have asymmetric centers. Compounds of the present invention containing an asymmetrically substituted atom may be isolated in optically active or racemic forms. It is well-known in the art how to prepare optically active forms, such as by resolution of racemic forms or by synthesis from optically active starting materials. All chiral, diastereomeric, racemic forms and all geometric isomeric forms of a compound are intended, unless the stereochemistry or the isomeric form is specifically indicated.

The term "protecting group" or "protecting group" means a substituent which protects groups, in particular hydroxyl groups, against undesirable reactions during synthetic procedures. Examples of protecting groups include, but are not limited to, substituted methyl ethers, for example, methoxymethyl, benzyloxymethyl, 2- methoxyethoxymethyl, 2-(trimethylsilyl)-ethoxymethyl, benzyl, and triphenylmethyl; tetrahydropyranyl ethers; substituted ethyl ethers, for example, 2,2,2-trichloroethyl and t- butyl; silyl ethers, for example, trimethylsilyl, t-butyldimethylsilyl and t-butyldiphenylsilyl; cyclic acetals and ketals, for example, methylene acetal, acetonide and benzylidene acetal; cyclic ortho esters, for example, methoxymethylene; cyclic carbonates; and cyclic boronates. Commonly used protecting groups are disclosed in T.W. Greene and P.G.M. Wuts, Protective Groups in Organic Synthesis, 3rd edition, John Wiley & Sons, New York (1999). The term "tautomer" as used herein means a proton shift from one atom of a compound to another atom of the same compound wherein two or more structurally distinct compounds are in equilibrium with each other.

"Pharmaceutically acceptable" means it is, within the scope of sound medical judgment, suitable for use in contact with the cells of humans and lower animals without undue toxicity, irritation, allergic response and the like, and are commensurate with a reasonable benefit/risk ratio.

The term "pharmaceutically acceptable salt" refers to salts which retain the biological effectiveness and properties of the compounds of the invention and which are not biologically or otherwise undesirable. In many cases, the compounds of the invention are capable of forming acid and/or base salts by virtue of the presence of amino and/or carboxyl groups or groups similar thereto. Pharmaceutically acceptable acid addition salts may be prepared from inorganic and organic acids, while pharmaceutically acceptable base addition salts can be prepared from inorganic and organic bases. For a review of pharmaceutically acceptable salts see Berge, et al. ((1977) J. Pharm. Sd, vol. 66, 1 ). The expression "non-toxic pharmaceutically acceptable salts" refers to non-toxic salts formed with nontoxic, pharmaceutically acceptable inorganic or organic acids or inorganic or organic bases. For example, the salts include those derived from inorganic acids such as hydrochloric, hydrobromic, sulfuric, sulfamic, phosphoric, nitric, and the like, as well as salts prepared from organic acids such as acetic, propionic, succinic, glycolic, stearic, lactic, malic, tartaric, citric, ascorbic, pamoic, maleic, hydroxymaleic, phenylacetic, glutamic, benzoic, salicyclic, sulfanilic, fumaric, methanesulfonic, and toluenesulfonic acid and the like. IMP dehydrogenase and viral RNA polymerase inhibitors

In the context of the invention, the term "inosine monophosphate dehydrogenase" or "IMP dehydrogenase" refers to an enzyme which converts inosine monophosphate to xanthosine monophosphate. As known from the skilled person, IMP dehydrogenase catalyses the first unique step in the de novo biosynthesis of guanine nucleotides. Accordingly, an IMP dehydrogenase inhibitor induce a decrease of the level of the GTP pool in the cells.

Examples of IMP deshydrogenase inhibitors are well-known from the one skilled in the art and are notably described in Sintchak and Nimmesgern (2000) (Sintchak and Nimmesgern (2000) Immunopharmacology 47:163-184). They include in particular ribavirin, taribavirin, mizoribine, tiazofurin, selenazofurin, isorawsonol, mycophenolic acid, mycophenolate mofetil, thiazole-4-carboxamide adenine dinucleotide, benzamide riboside and selenophenfurin.

It is within the general knowledge of the one skilled in the art to determine if a test compound is an IMP dehydrogenase inhibitor. Typically, the skilled person may compare the production of xanthosine monophosphote by IMP dehydrogenase from inosine monophosphate in the presence and in the absence of the test compound; or it may compare the intracellular GTP levels in the presence and in the absence of the test compound.

In the context of the invention, the term "viral RNA polymerase" refers to a viral enzyme that produces RNA. Preferably, in the context of the invention, the viral RNA polymerase is an RNA polymerase from a positive-sense single stranded RNA virus. More preferably, the viral RNA polymerase is an RNA polymerase from a virus from the genus Hepevirus. Most preferably, the viral RNA polymerase is an RNA polymerase from HEV.

Inhibitors of viral RNA polymerase can in particular interfere with a RNA polymerase in three ways: (1 ) it could inhibit the enzyme by competing with the natural substrate without being a substrate for the enzyme; (2) it could be an alternative substrate for the polymerase and cause chain termination; or (3) it could be an alternative substrate for the polymerase without causing chain termination.

Examples of inhibitors of viral RNA polymerase are well-known from the one skilled in the art and are notably described in Oberg (2006) (Oberg (2006) Antiviral research 71 :90-95). They include in particular acyclovir, penciclovir, ganciclovir, brivudine, cidofovir, foscarnet, zidovudine, didanosine, zalcitabine, lamivudine, stavudine, abacavir, tenofovir, emtricitabine, nevirapine, delavirdine, efavirenz, adefovir, ribavirin, taribavirin, favipiravir and valopicitabin.

The inhibitor of inosine monophosphate (IMP) dehydrogenase and/or the inhibitor of viral RNA polymerase according to the invention are preferably a compound of formula (I) as defined above.

Compound of formula (I)

Preferably, in the formula (I) defined above, R₄ and R₅ are -OH. Still preferably, R₆ is H. Still preferably, Het is a heteroaryl with 5 or 6 ring atoms and at least one nitrogen atom. More preferably, Het is a heteroaryl with 5 ring atoms and at least one nitrogen atom. More preferably, Het is a heteroaryl with 5 ring atoms and at least two nitrogen atoms. In a preferred embodiment, the compound of formula (I) according to the invention is of the following formula (II):

wherein:

R₄, R₅ and R₆ are as defined above;

and Z together with the nitrogen atom carrying it forms a 5 or 6 membered heteroaryl.

Preferably, Z together with the nitrogen atom carrying it forms a 5 membered heteroaryl. More preferably, Z together with the nitrogen atom carrying it forms a 5 membered heteroaryl containing at least two nitrogen atoms.

In another preferred embodiment, the compound according to the invention is of the following formula (III)

wherein:

R₄, R₅ and R₆ are as defined above;

D is selected from

wherein:

m = 0 or 1 ;

when m = 0, X is CN;

when m = 1 , X is -CO-, -CS- or -C=NH- and Y is NR^, NHOR₁ or OR₁ Ri and R₂ are each independently H, alkyl, acyl, aryl or is alkyl, acyl or substituted by halo, phenyl, CN, N0₂, OH, C0₂H or alkoxy; and R₁₀ is monohalogenated alkynyl or dihalogenated alkenyl. Preferably, when m = 1 , X is -CO- or -C=NH- and Y is NRi R₂, NHOR₁ or OR₁. Still preferably, when m = 1 , X is -CO- or -C=NH- and Y is NRi R₂.

Preferably, Ri and R₂ are each independently H or alkyl. Still preferably, Ri and R₂ are H.

Preferably, Ri₀ represents chloroethynyl, bromoethynyl, iodoethynyl, or

-C(Hal')=CH(Hal), where Hal' is chosen among chlorine, bromine and iodine, and Hal is bromine or iodine.

The compounds according to the invention may be in particular the compounds disclosed in the American patent US 3,798,209, the compounds disclosed in the international application WO 01/68034 and the compounds disclosed in the international application WO 2006/067606.

In a particularly preferred embodiment; the compound according to the invention is of the following formula (IV):

wherein R₄, R₅ and R₆ are as defined above.

Compounds of formulae (I), (II), (III) or (IV) as defined above wherein R₆ represents phosphate, diphosphate, triphosphate and phosphonate are deemed to represents pro-drugs of the corresponding compounds wherein R₆ represents H. Other pro-drugs leading to compounds wherein R₆ is H may also be used in the present invention. Such other pro-drugs include for instance esters, such as aminoacid esters, e.g. alanine esters and glycine esters, quaternary salts of phosphate, and the like. In a most preferred embodiment, the compound according to the invention is of the following formula (V):

This compound of formula (V) is also called ribavirin or 1 -( -D-Ribofuranosyl)-1 H- 1 ,2,4-triazole-3-carboxamide. It is also known by its brand names Copegus, Rebetol, Ribasphere, Vilona and Virazole. Its CAS number is 36791 -04-5.

In another particularly preferred embodiment; the compound according to the invention is of the following formula (VI):

This compound of formula (VI) is also called taribavirin, viramidine, ribamidine, or 1 -( -D-Ribofuranosyl)-1 ,2,4-triazole-3-carboximide.

Preferably, the compound of formula (I), (II) (III), (IV), (V) or (VI) as defined above is an inhibitor of IMP dehydrogenase and/or an inhibitor of viral RNA polymerase.

Treatment of Hepatitis E

In the context of the invention, the expression "genus Hepevirus" refers to a genus of Group IV, single-stranded positive-strand viruses belonging to the Hepeviridae family, the characteristic member of which is the hepatitis E virus (HEV). Preferably, the virus from the genus Hepevirus according to the invention is HEV.

As used herein, the term "hepatitis E virus" or "HEV" refers to an icosahedral, non- enveloped virus approximately 27-34 nm in diameter belonging to the genus Hepevirus. The HEV genome is a single-stranded, positive-sense RNA molecule approximately 7.5 kilobases in length. It comprises three open reading frames (ORF): ORF1 codes for a polyprotein with replicative functions, ORF2 codes for a capsid protein and ORF3 codes for a protein which function is thought to be associated with regulation of cellular protein kinase activity. Two major species of the virus are recognised: mammalian HEV and avian HEV. Preferably, in the context of the invention, HEV refers to mammalian HEV. Four major genotypes of mammalian HEV have been identified. Genotype 1 HEV is particularly present in developing regions of Asia, Africa and South America; genotype 2 HEV has been identified in Mexico, Chad and Nigeria and genotype 4 HEV has been found in industrialised regions of Japan, China and Taiwan. Genotype 3 HEV is observed in many developed regions and has a high prevalence in pig populations worldwide.

Preferably, the inhibitor or the compound of formula (I) according to the invention is for use in the treatment of hepatitis E.

In the context of the invention, the term "hepatitis E" refers to an infection caused by HEV. Hepatitis E is characterized by an incubation period following exposure to HEV usually ranging from 15-60 days. Typical clinical signs and symptoms in patients with symptomatic HEV infection include malaise, anorexia, nausea/vomiting, abdominal pain, fever and hepatomegaly. Other less common signs and symptoms include diarrhea, arthralgia, pruritus and urticarial rash. Additionally, hepatitis E may be characterized by laboratory features comprising a level of serum bilirubin preferably above 17 μηιοΙ/L and a serum level of alanine aminotransferase, and/or of aspartate aminotransferase, and/or of alkaline phosphatase, and/or of gamma-glutamyl transferase preferably above the upper limit of normal range and the detection of serum HEV RNA.

In the context of the invention, the expression "normal range of the serum level of alanine aminotransferase", respectively of aspartate aminotransferase, alkaline phosphatase and gamma-glutamyl transferase, refers to the range of serum level of alanine aminotransferase corresponding to the serum levels of alanine aminotransferase which are the most frequently observed in the healthy population. The normal range of the serum level of alanine aminotransferase is in particular the range wherein the serum level of 95% or 99% of the healthy population is observed.

As used herein, a "healthy population" denotes a population which does not suffer from any hepatitis or liver disease.

As known from the skilled person, the value of the normal range of the serum level of alanine aminotransferase, respectively of aspartate aminotransferase, alkaline phosphatase and gamma-glutamyl transferase depends on the reactive used to evaluate this level. By way of example, the normal range of the serum level of alanine aminotransferase is of 50 IU/L; the normal range of the serum level of aspartate aminotransferase is of 50 IU/L; the normal range of the serum level of alkaline phosphatase is of 120 IU/L; the normal range of the serum level of gamma-glutamyl transferase is of 55 IU/L, in the case described below by the inventors. As used herein, the term "hepatitis E" encompasses both acute and chronic hepatitis E.

As used herein, the term "acute hepatitis E" refers to hepatitis E characterized by clinical signs and laboratory symptoms usually resolving within 6 months after the first infection. Acute hepatitis E may also be characterized by the presence of anti-HEV IgM immunoglobulins switching to anti-HEV IgG immunoglobulins. Preferably, in acute hepatitis E, the level of serum bilirubin decreases below 17 μηιοΙ/Ι_ within 6 months after the first infection. Still preferably, the serum level of alanine aminotransferase decreases below the upper limit of normal range within 6 months after the first infection. Still preferably, the serum level of aspartate aminotransferase decreases below the upper limit of normal range within 6 months after the first infection. Still preferably, the serum level of alkaline phosphatase decreases below the upper limit of normal range within 6 months after the first infection. Still preferably, the serum level of gamma-glutamyl transferase decreases below the upper limit of normal range within 6 months after the first infection. More preferably, serum HEV RNA is not detected anymore within 6 months after the initial infection.

As used herein, the term "chronic hepatitis E" refers to hepatitis E characterized by carriage of HEV by the subject usually for more than 6 months and clinical signs and laboratory symptoms persisting usually 6 months after the initial infection. Chronic hepatitis E may also be characterized by the persistence of the presence of anti-HEV IgM immunoglobulins. Preferably, in chronic hepatitis E, the level of serum bilirubin is still above 17 μηιοΙ/Ι_ 6 months after the first infection. Still preferably, the serum level of alanine aminotransferase is still above the upper limit of normal range 6 months after the first infection. Still preferably, the serum level of aspartate aminotransferase is still above the upper limit of normal range 6 months after the first infection. Still preferably, the serum level of alkaline phosphatase is still above the upper limit of normal range 6 months after the first infection. Still preferably, the serum level of gamma-glutamyl transferase is still above the upper limit of normal range 6 months after the first infection. More preferably, serum HEV RNA is still detected 6 months after the first infection. Chronic hepatitis E may also be characterized by liver histological lesions, such as dense lymphocytic portal infiltrate with piecemeal necrosis. In particular, chronic hepatitis E may be responsible for cirrhosis. More particularly, chronic hepatitis E may be responsible for rapidly progressing liver fibrosis leading to cirrhosis.

As used herein, the term "liver fibrosis" refers to the excessive accumulation of extracellular matrix proteins including collagen that occurs in most types of chronic liver diseases (Battaler et al. (2005) J. Clin. Invest. 115:209-218). As used herein, the term "cirrhosis" refers to a liver disease characterized by the replacement of liver tissue by fibrosis, scar tissue and regenerative nodules. Cirrhosis is most commonly caused by alcoholism, chronic viral hepatitis, and fatty liver disease. Generally, in particular when it is caused by alcoholism, hepatitis B or C and fatty liver disease, cirrhosis develops in 20 to 30 years. Nevertheless, when caused by hepatitis E, extensive liver fibrosis and cirrhosis may rapidly develop in particular in organ-transplant patients. For example, the development of extensive liver fibrosis and cirrhosis within 3 years after the initial HEV infection has been reported (Kamar et at. (2008) Am. J. Transplant. 8:1744-1748). Such a cirrhosis is called in the context of the invention a "rapidly progressing cirrhosis".

Preferably, the inhibitor or the compound according to the invention is for use in the treatment of chronic hepatitis E in a subject. Still preferably, the inhibitor or the compound according to the invention is for use in the treatment for hepatitis E in a subject with extensive liver fibrosis or cirrhosis.

In the context of the invention, the term "treating" or "treatment" means reversing, alleviating, inhibiting the progress of, or preventing the disorder or condition to which such term applies, or one or more symptoms of such disorder or condition. In particular, in the context of the invention, a subject has been efficiently treated when the detection of serum HEV RNA is negative for 24 weeks after the end of the treatment, i.e after the end of the administration of the compound or inhibitor of the invention and during follow-up. As used herein, the term "treating" or "treatment" also encompasses preventing the reactivation of the disorder or condition to which such term applies, after the end of the treatment, i.e. after the end of the administration of the compound or inhibitor of the invention and during follow-up.

A method of treatment of an infection caused by a virus from the genus Hepevirus comprising the administration of a therapeutically effective amount of an inhibitor or a compound as defined above to a subject in need thereof is also included in the present invention.

As used herein, a "therapeutically effective amount" refers to an amount of a compound that 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., the subject gives an indication of or feels an effect).

It will be understood that the total daily usage of the compounds and inhibitors 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; 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 coincidental with the specific compound or inhibitor employed; and like factors well known in the medical arts. For example, it is well known within the skill of the art to start doses of the compound or inhibitor at levels lower than those required to achieve the desired therapeutic effect and to gradually increase the dosage until the desired effect is achieved. However, the daily dosage of the products may be varied over a wide range from 0.01 to 5,000 mg per adult per day. In particular, compositions used for the treatment of the disease of the invention contain 0.01 , 0.05, 0.1 , 0.5, 1 .0, 2.5, 5.0, 10.0, 15.0, 25.0, 50.0, 100, 250, 400, 500, 750, 800 and 1000 mg of the active ingredient for the symptomatic adjustment of the dosage to the patient to be treated. A medicament typically contains from about 0.01 mg to about 1000 mg of the active ingredient, preferably from 1 mg to about 1000 mg of the active ingredient, in particular 400 mg or 800 mg of the active ingredient. An effective amount of the drug is ordinarily supplied at a dosage level from 0.0002 mg/kg to about 25 mg/kg of body weight per day, especially from about 0.001 mg/kg to 21 mg/kg of body weight per day.

Any suitable method of administration known from one skilled in the art may be used. In particular, the inhibitors or compounds according to the invention may be administered for example by the oral route, by inhalation, or by the parenteral route (in particular by intravenous injection).

The compound and inhibitor of the invention may be combined with pharmaceutically acceptable excipients, and optionally sustained-release matrices, such as biodegradable polymers, to form therapeutic compositions.

In the pharmaceutical compositions of the present invention for oral, sublingual, subcutaneous, intramuscular, intravenous, transdermal, local or rectal administration, the active principle, alone or in combination with another active principle, can be administered in a unit administration form, as a mixture with conventional pharmaceutical supports, to animals and human beings. Suitable unit administration forms comprise oral-route forms such as tablets, gel capsules, powders, granules and oral suspensions or solutions, sublingual and buccal administration forms, aerosols, implants, subcutaneous, transdermal, topical, intraperitoneal, intramuscular, intravenous, subdermal, transdermal, intrathecal and intranasal administration forms and rectal administration forms. When the parenteral route is selected, the inhibitors or compounds may be in the form of injectable solutions and suspensions, conditioned in ampoules or flasks. The forms for parenteral delivery are conventionally obtained by mixing the inhibitors or compounds according to the invention with buffers, stabilizers, preservatives, solubilizing agents, isotonic agents and slurrying agents. According to known techniques, these mixtures can then be sterilized and conditioned in the form of intravenous injections.

Preferably, the pharmaceutical compositions contain vehicles which are pharmaceutically acceptable for a formulation capable of being injected. These may be in particular isotonic, sterile, saline solutions (monosodium or disodium phosphate, sodium, potassium, calcium or magnesium chloride and the like or mixtures of such salts), or dry, especially freeze-dried compositions which upon addition, depending on the case, of sterilized water or physiological saline, permit the constitution of injectable solutions.

The pharmaceutical forms suitable for injectable use include sterile aqueous solutions or dispersions; formulations including sesame oil, peanut oil or aqueous propylene glycol; and sterile powders for the extemporaneous preparation of sterile injectable solutions or dispersions. In all cases, the form must be sterile and must be fluid to the extent that easy syringability exists. It must be stable under the conditions of manufacture and storage and must be preserved against the contaminating action of microorganisms, such as bacteria and fungi.

Solutions comprising compounds or inhibitors of the invention as free base or pharmacologically acceptable salts can be prepared in water suitably mixed with a surfactant, such as hydroxypropylcellulose. Dispersions can also be prepared in glycerol, liquid polyethylene glycols, and mixtures thereof and in oils. Under ordinary conditions of storage and use, these preparations contain a preservative to prevent the growth of microorganisms.

The compound or inhibitor of the invention can be formulated into a composition in a neutral or salt form. Pharmaceutically acceptable salts include the acid addition salts which are formed with inorganic acids such as, for example, hydrochloric or phosphoric acids, or such organic acids as acetic, oxalic, tartaric, mandelic, and the like. Salts can also be derived from inorganic bases such as, for example, sodium, potassium, ammonium, calcium, or ferric hydroxides, and such organic bases as isopropylamine, trimethylamine, histidine, procaine and the like.

The carrier can also be a solvent or dispersion medium containing, for example, water, ethanol, polyol (for example, glycerol, propylene glycol, and liquid polyethylene glycol, and the like), suitable mixtures thereof, and vegetables oils. The proper fluidity can be maintained, for example, by the use of a coating, such as lecithin, by the maintenance of the required particle size in the case of dispersion and by the use of surfactants. The prevention of the action of microorganisms can be brought about by various antibacterial and antifungal agents, for example, parabens, chlorobutanol, phenol, sorbic acid, thimerosal, and the like. In many cases, it will be preferable to include isotonic agents, for example, sugars or sodium chloride. Prolonged absorption of the injectable compositions can be brought about by the use in the compositions of agents delaying absorption, for example, aluminium monostearate and gelatin.

Sterile injectable solutions are prepared by incorporating the active ingredients in the required amount in the appropriate solvent with various of the other ingredients enumerated above, as required, followed by filtered sterilization. Generally, dispersions are prepared by incorporating the various sterilized active ingredients into a sterile vehicle which contains the basic dispersion medium and the required other ingredients from those enumerated above. In the case of sterile powders for the preparation of sterile injectable solutions, the preferred methods of preparation are vacuum-drying and freeze-drying techniques which yield a powder of the active ingredient plus any additional desired ingredient from a previously sterile-filtered solution thereof.

Upon formulation, solutions will be administered in a manner compatible with the dosage formulation and in such amount as is therapeutically effective. The formulations are easily administered in a variety of dosage forms, such as the type of injectable solutions described above, but drug release capsules and the like can also be employed.

For parenteral administration in an aqueous solution, for example, the solution should be suitably buffered if necessary and the liquid diluent first rendered isotonic with sufficient saline or glucose. These particular aqueous solutions are especially suitable for intravenous, intramuscular, subcutaneous and intraperitoneal administration. In this connection, sterile aqueous media which can be employed will be known to those of skill in the art in light of the present disclosure. For example, one dosage could be dissolved in 1 ml of isotonic NaCI solution and either added to 1000 ml of hypodermoclysis fluid or injected at the proposed site of infusion. Some variation in dosage will necessarily occur depending on the condition of the subject being treated. The person responsible for administration will, in any event, determine the appropriate dose for the individual subject.

Examples of slurrying agents include methylcellulose, acacia and sodium carboxymethylcellulose. Examples of stabilizers include sodium sulphite and sodium metasulphite, and examples of preservatives include sodium p-hydroxybenzoate, sorbic acid, cresol and chlorocresol.

The compound or inhibitor of the invention may be formulated within a therapeutic mixture to comprise about 0.0001 to 1000 mg, or about 0.001 to 800 mg, or about 0.1 to 400 mg or even about 200 mg, 300 mg, 400 mg, 500 mg, 600 mg, 700 mg, 800 mg, 900 mg or 1000 mg per dose or so. Multiple doses can also be administered. The inhibitor or the compound according to the invention may be in particular administered at a daily dose of about 5 mg/kg to 21 mg/kg, preferably at a daily dose of about 10 mg/kg to 15 mg/kg. Preferably, the inhibitor or compound according to the invention is administered at a daily dose of about 12 mg/kg, more preferably at a daily dose of 12.3 mg/kg. The inhibitor or compound according to the invention may be in particular administered in monodose or multiple doses. Preferably, the inhibitor or compound according to the invention is administered in a single daily dose or in two daily doses. The inhibitor or compound according to the invention may also be administered in a single weekly dose or in several weekly doses.

In addition to the compounds of the invention formulated for parenteral administration, such as intravenous or intramuscular injection, other pharmaceutically acceptable forms include, e.g. tablets or other solids for oral administration; liposomal formulations; time release capsules; and any other form currently used.

Subject

In the context of the present invention, a "subject" denotes a human or non-human mammal such as a rodent (rat, mouse, rabbit), a primate (chimpanzee), a porcine (swine), or a non-mammal animal such as a bird. Preferably, the subject is a human or a swine. More preferably, the subject is a human.

Acute hepatitis E is a particular high cause of mortality in pregnant women. Accordingly, the subject according to the invention is preferably a pregnant woman.

It has been demonstrated that chronic hepatitis E developed in particular in immunodeficient patients. Accordingly, the subject according to the invention is preferably immunodeficient.

As used herein, the term "immunodeficient" or "immunodeficiency" or "immune deficiency" refers to a state in which the immune system's ability to fight infectious disease is compromised or entirely absent. The immunodeficiency according to the invention may be an acquired immunodeficiency or an innate immunodeficiency.

In the context of the invention, the term "acquired immunodeficiency" or "secondary immunodeficiency" refers to an immune deficiency that is the result of particular external processes or diseases. Common causes for acquired immunodeficiency are malnutrition, aging and particular medications, such as chemotherapy, disease-modifying antirheumatic drugs, immunosuppressive drugs after organ transplants, glucocorticoids). Many specific diseases also directly or indirectly impair the immune system, such as many types of cancer, particularly those of the bone marrow and blood cells (leukemia, lymphoma, multiple myeloma), and certain chronic infections, such as HIV infection. Preferably, the subject according to the invention is selected from the group consisting of a transplant recipient, a subject suffering from a viral infection and a subject suffering from cancer. When the subject is a transplant recipient, he may in particular be a kidney-pancreas transplant recipient. When the subject suffers from a viral infection, he may in particular suffer from HIV infection. When the subject suffers from cancer, he may in particular suffer from lymphoma. He may also undergo chemotherapy.

In the context of the invention, the term "innate immunodeficiency" or "primary immunodeficiency" refers to disorders in which part of the body's immune system is missing or does not function properly. Most primary immunodeficiencies are genetic disorders; the majority being diagnosed in children under the age of one, although milder forms may not be recognised until adulthood. Primary immunodeficiencies encompass eight classes of diseases, totalling over 120 conditions, and include in particular combined T- and B-cell immunodeficiencies, idiopathy CD4 lymphopenia, antibody deficiencies, Wiskott-Aldrich syndrome, ataxia telangiectasia, ataxia-like syndrome, Nijmegen breakage syndrome, Bloom syndrome, DiGeorge syndrome (when associated with thymic defects), cartilage-hair hypoplasia, Schimke syndrome, Hermansky-Pudlak syndrome type 2, hyper-lgE syndrome, chronic mucocutaneous candidiasis, immune dysregulation diseases, phagocyte disorders, hypohidrotic ectodermal dysplasia such as NEMO deficiency and IKBA deficiency, IRAK-4 deficiency, WHIM syndrome, epidermodysplasia verruciformis, autoinflammatory disorders, complement deficiencies.

Preferably, the acquired or innate immunodeficiency according to the invention is caused by a CD4 lymphocytes deficit.

In another embodiment of the present invention, the subject is preferably a non- human animal. More preferably, the subject is a porcine.

Monotherapy or combination therapy

The inhibitors and compounds according to the invention may be used in monotherapy or in combination therapy.

In the context of the invention, "monotherapy" refers to a therapy using only one therapeutic agent or only one type of therapeutic agent. Accordingly, when the inhibitors or compounds of the invention are used in monotherapy, they are not used in association with other therapeutic agents. In the context of the invention, "combination therapy" refers to a therapy using at least two therapeutic agents or at least two types of therapeutic agents. Accordingly, when an inhibitor or compound of the invention is used in combination therapy, it may be used in association with at least one other therapeutic agent. Said other therapeutic agent may be used for the treatment of the same disease as the inhibitors or compounds according to the invention. In particular the inhibitor or compound according to the invention may be used in association with at least one other inhibitor or compound according to the invention. For example an inhibitor of IMP dehydrogenase according to the invention may be used in association with another inhibitor of IMP dehydrogenase according to the invention and/or with an inhibitor of viral RNA polymerase according to the invention. A compound of formula (I) according to the invention may also be used in association with an inhibitor of IMP dehydrogenase according to the invention and/or with an inhibitor of viral RNA polymerase according to the invention.

The inhibitors and compounds according to the invention may also be used in association with anti-viral agents.

As used herein, "anti-viral agent", "anti-viral drug" or "anti-viral compound" refers to a compound specifically used for treating viral infections. Examples of anti-viral agents are well-known from one skilled in the art and include in particular virus entry inhibitors, uncoating inhibitors, reverse transcriptase inhibitors, integrase inhibitors, viral polymerase inhibitors and protease inhibitors.

Said other therapeutic agent may also be used for the treatment of another condition, which is not directly treated by the compounds according to the invention. Said other condition, which is not directly treated by the compounds according to the invention may be in particular innate or acquired immunodeficiencies, as defined above. Accordingly, the inhibitors and compounds according to the invention may be used in association with anticancerous agents, immunosuppressive agents, antiviral agents, immunostimulative agents.

As used herein, an "immunosuppressive agent" or "immunosuppressive drug" refers to a compound that inhibits or prevents activity of the immune system. Examples of immunosuppressive agents are well-known from one skilled in the art an include in particular glucocorticoids; cytostatics such as alkylating agents, antimetabolites, and cytotoxic antibiotics; cyclosporin, tacrolimus, sirolimus, interferon-β, interferon-γ, opioids, TNF binding proteins, mycophenolic acid, fingolimod, and myriocin.

As used herein, an "immunostimulative agent" or "immunostimulative drug" refers to a compound that induces or stimulates activity of the immune system. Examples of immunostimulative agents are well-known from the skilled person and include in particular interferon a.

Nevertheless, in some cases, in particular in transplant recipients, the compound or inhibitor according to the invention is preferably not used in association with an immunostimulative agent. More particularly, in some cases, the compound or inhibitor according to the invention is preferably not used in association with interferon a.

The invention will be further illustrated by the following non-limiting figures and example.

Description of the figures

Figure 1 displays a graph representing the serum level of alanine aminotransferase (in number of times the upper limit of the normal range) of the patient described in the example 1 over time. The period during which the patient was treated with ribavirin is indicated by a bar. The lower panel displays the results of the Adaltis HEV IgG ELISA assay, of the Adaltis HEV IgM ELISA assay and of the RT Cooper, PCR Taqman assay performed for determining the presence of HEV in the patient. Available frozen samples have been processed retrospectively. Positive (+) and negative (-) results are indicated; NA=not available.

Figure 2 displays a zoom of Figure 1 on the period from January 15, 2008 to December 15, 2009.

Figure 3 displays a graph representing the serum level of alanine aminotransferase (ALT) (in number of times the upper limit of the normal range, ULN) of the patient described in the example 2 over time. The period during which the patient was treated with tacrolimus, mycophenolate mofetil, prednisone and ribavirin is indicated respectively by a bar. The lower panel displays the results of the HEV IgG ELISA assay, of the HEV IgM ELISA assay, of the RT-PCR assay in serum and of the RT-PCR assay in stool, performed for determining the presence of HEV in the patient. Positive (+) and negative (-) results are indicated. Figure 4 displays a graph representing the serum level of alanine aminotransferase (ALT) (in number of times the upper limit of the normal range, ULN) of the patient described in the example 3 over time. The period during which the patient was treated with ribavirin is indicated by a bar. The lower panel displays the results of the HEV IgG ELISA assay, of the HEV IgM ELISA assay, of the RT-PCR assay in serum and of the RT-PCR assay in stool, performed for determining the presence of HEV in the patient. Positive (+) and negative (-) results are indicated.

Figure 5 displays a graph representing the serum level of alanine aminotransferase (in number of times the upper limit of the normal range) of the patient described in the example 4 over time. The period during which the patient was treated with ribavirin is indicated by a bar. The lower panel displays the results of the Adaltis HEV IgG ELISA assay, of the Adaltis HEV IgM ELISA assay, of the RT-PCR assay in serum and of the RT-PCR assay in stool performed for determining the presence of HEV in the patient. Available frozen samples have been processed retrospectively. Positive (+) and negative (-) results are indicated.

Examples Example 1

This example shows the report of the first case of HEV eradication under ribavirin in a dual kidney-pancreas transplant recipient with a seven-year history of chronic HEV infection. In June 1998, a 33-year-old man received a first deceased donor dual kidney and pancreas transplant graft for diabetes mellitus after a 2-year period of haemodialysis. The patient received an initial treatment associating a 10 days course of biological induction of polyclonal antibodies (ATG, Thymoglobulins®, Merieux, France) together with mycophenolate mofetil (Roche, Neuilly-sur Seine, France) 1000 mg per day, adapted thereafter according to clinical event). He also received corticosteroids at an initial dose of 500 mg at day 0, rapidly tapered to reach 10 mg/day at day 45. Finally, he received tacrolimus from day 0 at an initial daily dose of 0.15 mg/kg/day and adapted thereafter to reach a targeted through concentration (CO) between 10 to 15 ng/ml. He was insulin-free after the transplantation and his serum creatinine level was 90 μηιοΙ/L (10 mg/L). He did not experience any acute rejection after transplantation and transplantation was uneventful. Forty-seven months post-transplantation, acute hepatitis was diagnosed fortuitously at a routine visit. There was no alcohol intake, no toxic exposure, and no recent introduction of a new medication. Physical examination was normal. Alanine aminotransferase (ALT) level was 1 .6 times the upper limit of normal (ULN), aspartate aminotransferase (AST), gamma-glutamyl transpeptidase (GT) and alkaline phosphatase (AP) levels were normal. Total bilirubin level was 19 μηιοΙ/L (1 1 mg/L) with a predominance of unconjugated bilirubin. Serum creatinine level was 144 μηιοΙ/L (16 mg/L). Prothrombin time ratio was 1 18% of normal. The patient was tested positive for anti-HCV antibodies and negative for hepatitis B virus (HBV) core antibodies (IgG). Genome amplifications of hepatitis B virus (HBV), hepatitis C virus (HCV), cytomegalovirus, Herpes Simplex virus and Epstein-Barr virus were negative. Ferritin and transferrin saturation blood levels were normal, anti-liver and anti-nuclear antibodies were tested negative, serum alpha-1 -antitrypsin, copper and ceruloplasmin levels were normal, which ruled out hemochromatosis, auto-immune hepatitis, alpha-1 -antitrypsin deficiency and Wilson's disease, respectively. Liver ultrasonography was normal. The patient refused the liver-biopsy and was discharged.

The patient was thereafter seen every three to six months. Liver-enzymes abnormalities persisted with a fluctuation of ALT, AST and GT levels ranging from 1 .25 to 5.5, 0.4 to 2.5 and 0.5 to 1 .6 times the ULN respectively. AP levels remained normal. HCV-RNA amplification was repeated and remained undetectable over time (with a latest limit of detection of 12 international units per milliliter).

Seventy months after the initial hepatitis, the patient was tested positive for hepatitis E Virus (HEV) antibodies (Abott HEV IgG Elisa and Abott HEV IgM Elisa) with 97 percent avidity of anti-HEV IgG antibodies. Genome amplification of HEV (RT-PCR Cooper and PCR Taqman) was positive. The genotype was 3f. A post-hoc analysis of all available frozen serum samples confirmed that the initial hepatitis was indeed due to HEV and that the infection had persisted over the last 70 months. Seven months later, a liver biopsy showed on 30 millimetres and 10 portal tracts chronic hepatitis with a moderate activity and an extensive fibrosis (METAVIR A2F3). No steatosis, steatohepatitis or vascular abnormality was seen on the liver biopsy.

Five months later (82 months after the initial infection), ribavirin 400 milligrams twice daily (total daily dose, 12.3 mg/kg) was introduced with the informed consent of the patient. Two weeks later, ALT, AST, GT were strictly normal. Four weeks later, HEV IgM antibodies and genome amplification of HEV were negative. All of these findings were confirmed at week 6 of treatment. Ribavirin was progressively lowered to 200 mg per day because of mild anemia and stopped after a total 12-weeks treatment period. The inventors thus observed clearance of HEV and normalization of liver function tests in a four-week period following the introduction of ribavirin. This observation strongly points to a potent activity of ribavirin against HEV. Ribavirin would thus be the first drug to offer a potentially effective oral treatment for HEV infection.

Ribavirin is phosphorylated in human cells by adenosine kinase to ribavirin monophosphate, which is then converted to ribavirin triphosphate by the successive action of nucleoside mono- and di-phosphate kinase. The phosphorylated derivatives of ribavirin have various antiviral effects. Ribavirin monophosphate inhibits inosine monophosphate deshydrogenase and consequently reduces cellular guanosine triphosphate levels. Guanosine depletion interferes with viral (and host) nucleic acid synthesis. Nucleotide pool imbalance also causes the viral polymerase to substitute alternative nucleotides for guanosine triphosphate, favouring viral mutagenesis. The third mechanism of action is a direct interaction of ribavirin triphosphate with the viral polymerase inducing either a direct inhibition of the enzyme, a nucleic acid chain termination synthesis or the introduction of fraudulent nucleotides in the viral genome leading to an error catastrophe phenomenon. The patient of this report was under mycophenolate mofetil. Mycophenolic acid, as ribavirin, is a potent inhibitor of inosine monophosphate deshydrogenase. The fast clearance of VHE accompanied by a complete normalization of liver function tests is in favour of a direct inhibition of HEV polymerase by ribavirin rather than an antiviral activity based on guanosine depletion.

To sum up, the observations of the inventors demonstrate an antiviral effect of ribavirin in individuals with chronic HEV infection.

Example 2

This example is an update of the clinical story of the case presented in Example 1 . The patient was a diabetic white man aged 40 years who, after 2 years of hemodialysis, received a first simultaneous kidney and pancreas transplant from a deceased donor in November 1998. The immunosuppressive regimen included an initial 10-day course of antithymoglobulins; corticosteroids, 500 mg/d at day 0 followed by rapid tapering to reach 10 mg/d; tacrolimus dosed to achieve a targeted trough concentration between 10 and 15 ng/mL; and mycophenolate mofetil, 1000 mg/d, adapted to clinical outcome. The patient did not have any signs of acute rejection and was free of insulin when discharged, with a serum creatinine level of 90 μηιοΙ/L (1 mg/dL).

In January 2006, the patient presented to the inventors'unit for unexplained hepatitis lasting longer than the previous 38 months. Physical examination was normal. Serum alanine aminotransferase (ALT) level was 1 .6 times the upper limit of normal range. Aspartate aminotransferase (AST), γ-glutamyltransferase (GGT), and alkaline phosphatise levels were normal. Total bilirubin level was 19 μηιοΙ/L (1 .1 mg/dL), with predominance of unconjugated bilirubin. Serum creatinine level was 144 μηιοΙ/L (1 .6 mg/dL). Prothrombin time was normal. The patient tested positive for anti-hepatitis C virus antibodies and negative for anti-hepatitis B virus IgG core antibodies. Genome amplifications of hepatitis B virus, hepatitis C virus, cytomegalovirus, herpes simplex virus, and Epstein-Barr virus were all negative. Ferritin and transferrin saturation blood levels were normal; antiliver and antinuclear antibodies were negative; and serum a antitrypsin, copper, and ceruloplasmin levels were normal— which ruled out hemochromatosis, autoimmune hepatitis, and oci -antitrypsin deficiency and Wilson disease, respectively. Liver ultrasonography findings were normal. Hepatitis C virus genome remained undetectable over time (limit of detection, 12 lU/mL).

Hepatitis E virus infection was diagnosed in September 2008 on the basis of amplification of HEV RNA (genotype 3f; GenBank accession number GU936617) and detection of anti-HEV IgG and anti-HEV IgM antibodies (EIAgen HEV IgG and EIAgen HEV IgM, Adaltis, Bologna, Italy) in the serum (Figure 3). In the present study, HEV RNA was amplified with primers targeting the HEV open reading frame 2 region. Two reverse transcriptase polymerase chain reaction (RT-PCR) methods were used: real-time polymerase chain reaction that was sensitive for all HEV genotypes and conventional RT- PCR, followed by sequence analysis of the 330-nucleotide open reading frame 2 product. The detection limit of RT-PCR was 100 copies of equivalent genome per milliliter of serum by using serial 10-fold dilutions of plasmid DNA (pGEM-HEV) as a standard curve (Enouf et al. (2006) J. Med Virol. 78:1076-82). Amplification of glyceraldehyde 3-phosphate dehydrogenase was used as an internal control to rule out RT-PCR inhibitors. An IgG avidity index at 97% suggested past contact with the virus. Serum and plasma samples collected at the time of the transplantation in 1998 and stored until 2008 at -80 'C were screened retrospectively, and possible exposure to the virus was dated to 2003 based on the detection of both anti-HEV IgM and IgG antibodies; however, HEV RNA could not be amplified in both techniques. In 2007, HEV RNA was detected, showing a 99.5% identity with the genome sequence amplified in 2008. A 30-mm liver biopsy (spanning 10 portal tracts) was done in April 2009 and showed evidence of chronic hepatitis with moderate activity and extensive fibrosis (METAVIR activity [A] and fibrosis [F] scores of A2 and F3). The liver biopsy showed no steatosis, steatohepatitis, or vascular abnormality. The CD4⁺ T-lymphocyte count was 0.72 x 10⁹ cells/L at that time. The CD8⁺ T-lymphocyte count was 0.58 x 10⁹ cells/L. The patient had never traveled to endemic areas and reported frequent consumption of undercooked pork, suggesting an autochthonous zoonotic transmission method of HEV.

In September 2009 (82 months after the presumed date of initial infection), ribavirin treatment, 400 mg twice daily (total daily dose, 12.3 mg/kg of body weight), was initiated with the informed consent of the patient. The immunosuppressive regimen was not modified. Two weeks later, ALT, AST, and GGT levels returned to within-normal limits. Four weeks after the initiation of ribavirin treatment, HEV RNA was undetectable (under the limit of detection of the assay) in the serum, and HEV IgM antibodies were negative. No fecal specimen was available for HEV RNA detection at that time. All of these findings were confirmed at weeks 6, 8, 10, and 12 of treatment. Ribavirin dose was progressively decreased to 200 mg/d because of mild anemia (nadir hemoglobin level, 92 g/L), and treatment was stopped after 12 weeks. One year after cessation of treatment, liver function test results remained normal and HEV RNA was undetected in serum and stool.

This example highlights the long-term outcome of patient described in Example 1 . The absence of HEV RNA detection in the stools and in the serum more than 6 months after the end of the treatment suggests that this patient achieved sustained virological response under ribavirin treatment, which could correspond, in analogy with the hepatitis C virus (HCV), to full HEV eradication— or cure.

Example 3

This example describes a case of HEV clearance under ribavirin in a patient with a primary immune deficiency and chronic HEV infection.

The patient was a white woman aged 57 years with idiopathic CD4⁺ T lymphocytopenia and a primary deficiency in lgG-1 , -2, and -4 subclasses. Her medical history included recurrent upper respiratory tract infections since childhood, with secondary bronchiectasia and moderate chronic respiratory insufficiency, and chronic cutaneous and genital human papillomavirus infection that evolved to chronic epidermodysplasia verruciformis (since age 7 years), Bowen disease (age 47 years), spinocellular cancer (age 47 years), and intraepithelial neoplasia of the vulva (age 48 years). She also reported primary cancer of the breast (age 37 years), appendicular abscess (age 52 years), and several episodes of urolithiasis.

She was first seen in the inventors' unit in July 2009 for abnormal liver function test results and a 10-kg weight loss over the previous 6 months. Apart from the dermatologic lesions, the physical examination was normal. Levels of ALT, AST, GGT, and alkaline phosphatase were 8, 5, 7.5, and 1 times the upper limit of normal range, respectively. Total bilirubin level was 14 μηιοΙ/L (0.8 mg/dL). The CD4⁺ T-lymphocyte count was 0.22 x 10⁹ cells/L. The CD8⁺ T-lymphocyte count was 0.05 x 10⁹ cells/L. The CD19⁺ B- lymphocyte count was 0.03 x 10⁹ cells/L. The serum creatinine level was 63 μηιοΙ/L (0.7 mg/dL). Prothrombin time index was 97%. The patient tested negative for anti-hepatitis A virus IgM antibodies, anti-hepatitis C virus antibodies, anti-hepatitis B virus IgG core antibodies, and anti-HEV antibodies (HEV IgG enzyme-linked immunosorbent assay, Abbott Laboratories, Park, Illinois). Genome amplifications of hepatitis B virus, hepatitis C virus, cytomegalovirus, herpes simplex virus, and Epstein-Barr virus were negative. Ferritin blood level was normal; antiliver and antinuclear antibodies were negative; and serum oci -antitrypsin, copper, and ceruloplasmin levels were normal. Liver ultrasonography findings were normal.

In December 2009, a 9-mm liver biopsy (spanning 10 portal tracts) showed evidence of chronic hepatitis, with moderate activity and mild fibrosis (METAVIR scores of A2 and F1 ). At that time, HEV RNA was amplified (genotype 3c; GenBank accession number HM066937) from both serum and stool samples (Figure 4), and a low anti-HEV IgM reactivity (EIAgen HEV IgM) was detected in the absence of a detectable anti-HEV IgG antibody (EIAgen HEV IgG). Two previous serum samples collected in April and July 2009 (stored at -80 ^<€) were positive for HEV RNA and anti-HEV IgM antibody, confirming chronic HEV infection. The patient reported having travelled to the south of France in November 2008 where she consumed pig liver sausage, suggesting autochthonous zoonotic transmission. Two weeks later (12 months after the presumed date of infection), ribavirin treatment, 600 mg/d divided into 2 doses (total daily dose, 12 mg/kg), was initiated with the informed consent of the patient. Two weeks later, ALT and AST levels returned to normal limits. After 4 weeks of treatment, HEV RNA was undetectable in the serum and stools. Ribavirin treatment was stopped after 12 weeks. Two months after the end of treatment, liver function test results remained normal and HEV RNA was not amplified from either serum or stool. The inventors thus observed a normalization of liver function test results after 2 weeks of treatment, and clearance of HEV RNA was attained by week 4. To the inventors' knowledge, this is the first description of the efficacy of a nucleoside analogue for the management of primary immunocompromised patients with chronic HEV infection. Given the absence of available treatment, these results suggest that ribavirin is a potent strategy for improving the natural course and management of patients with severe forms of chronic HEV infection.

Example 4

This example describes a case of HEV clearance under ribavirin in a patient with acute HEV infection. The patient was a 63-year-old white man who received a first kidney transplant from a deceased donor 15 years earlier because of unexplained chronic kidney disease. His immunosuppressive regimen included corticosteroids 15 mg/d and sirolimus 2 mg/d. Abnormal liver function test results were detected in January 2010 during a routine visit in his transplant unit. Liver function tests were strictly normal 3 months earlier. The patient was referred to the inventor's unit.

The patient reported severe asthenia over the past 4 months. The physical examination was normal. Levels of ALT, AST, GGT, and alkaline phosphatase were respectively 14, 4, 5, and 1 times the upper limit of normal range. Total bilirubin level was 14 μηιοΙ/L (0.8 mg/dL). Prothrombin time index was 100%. HEV RNA was amplified (genotype 3e) from both serum and stool samples and anti-HEV IgM reactivity (HEV IgM enzyme-linked immunosorbent assay, Abbott Laboratories, Park, Illinois) was detected in the absence of a detectable anti-HEV IgG antibody (HEV IgG enzyme-linked immunosorbent assay, Abbott Laboratories, Park, Illinois).

The patient tested negative for anti-hepatitis A virus IgM antibodies, anti-hepatitis C virus antibodies and anti-hepatitis B virus IgG core antibodies. Genome amplifications of hepatitis B virus, hepatitis C virus, cytomegalovirus, herpes simplex virus, and Epstein- Barr virus were negative. Ferritin blood level was normal; anti-liver and anti-nuclear antibodies were negative; and serum oci -antitrypsin, copper, and ceruloplasmin levels were normal. Liver ultrasonography findings were normal.

Two weeks later (less than 6 months after the presumed date of infection), ribavirin treatment, 1000 mg/d divided into 2 doses (total daily dose, 12 mg/kg), was initiated with the informed consent of the patient. Ten days later, ALT and AST levels had returned to normal limits (Figure 5). After 2 weeks of treatment, HEV RNA was undetectable in the serum and stools. Ribavirin was stopped after 4 weeks of treatment. Six months after the end of treatment, liver function test results remained normal and HEV RNA was not amplified from either serum or stool.

The inventors thus observed a normalization of liver function test results after 10 days of treatment with ribavirin and clearance of HEV RNA was attained by week 2. To the inventors' knowledge, this is the first description of the efficacy of a nucleoside analogue for the management of patients with acute HEV infection. Given the absence of available treatment, these results suggest that ribavirin is a potent strategy for improving the natural course and management of patients with severe forms of acute HEV infection.

Claims

1. An inhibitor of inosine monophosphate (IMP) dehydrogenase and/or an inhibitor of viral RNA polymerase for use in the treatment of an infection caused by a virus from the genus Hepevirus in a subject.

2. A compound of the following formula (I):

wherein:

M⁺ is an alkali metal cation;

R₉ is H, aryl, alkyl, or alkyl substituted by OH, halo, NR_8aR_8b, or acyl;

Het is a heteroaryl with 5 to 7 ring atoms and at least one nitrogen atom; and e = 0 to 6, f = 0 to 10 and n = 0 or 1 ;

or its pharmaceutically acceptable salts, its N-oxide forms; hydrates or hydrated salts, its polymorphic crystalline structures, its tautomers, racemates, diastereisomers or enantiomers; for use in the treatment of an infection caused by a virus from the genus Hepevirus in a subject.

3. The compound for use according to claim 2, wherein said compound is of the following formula (II):

wherein:

R₄, R₅ and R₆ are as defined in claim 2;

4. The compound for use according to claim 2 or 3, wherein said compound is of the following formula (III)

wherein:

R₄, R₅ and R₆ are as defined in claim 2;

D is selected from

wherein:

m = 0 or 1 ;

when m = 0, X is CN;

when m = 1 , X is -CO-, -CS- or -C=NH- and Y is NR₁R₂, NHOR₁ or OR₁ Ri and R₂ are each independently H, alkyl, acyl, aryl or is alkyl, acyl or substituted by halo, phenyl, CN, N0₂, OH, C0₂H or alkoxy; and R₁₀ is monohalogenated alkynyl or dehalogenated alkenyl.

5. The compound for use according to any one of claims 2 to 4, wherein said compound is of the following formula (IV):

wherein R₄, R₅ and R₆ are as defined in claim 2.

6. The compound for use according to any one of claims 2 to 5, wherein said compound is of the following formula (V):

7. The compound for use according to any one of claims 2 to 4, wherein said compound is of the following formula (VI):

8. The inhibitor for use according to claim 1 or the compound for use according to any one of claims 2 to 7, wherein the infection caused by a virus from the genus Hepevirus is hepatitis E.

9. The inhibitor for use according to claim 1 or 8 or the compound for use according to any one of claims 2 to 8, for use in the treatment of acute or chronic hepatitis E in a subject.

10. The inhibitor for use according to any one of claims 1 or 8 to 9 or the compound for use according to any one of claims 2 to 9, wherein the subject is a human.

11. The inhibitor for use according to any one of claims 1 or 8 to 10 or the compound for use according to any one of claims 2 to 10, wherein the subject suffers from innate or acquired immunodeficiency.

12. The inhibitor for use according to any one of claims 1 or 8 to 1 1 or the compound for use according to any one of claims 2 to 1 1 , wherein the subject is selected from the group consisting of a transplant recipient, a subject suffering from a viral infection and a subject suffering from cancer.

13. The inhibitor for use according to any one of claims 1 or 8 to 12 or the compound for use according to any one of claims 2 to 12, wherein the subject presents a cirrhosis.

14. The inhibitor for use according to any one of claims 1 or 8 to 9 or the compound for use according to any one of claims 2 to 9, wherein the subject is a non-human animal.

15. The inhibitor or the compound for use according to claim 14, wherein the subject is a porcine.