Compound for treating arbovirus infections

Abstract

The present invention relates to a compound which is effective in inhibiting the function of the ubiquitin-like modifier activating enzyme 5 (UBA5) for use in a method of treating or preventing a disease or condition mediated by an arbovirus infection, such as a flavivirus, alphavirus or bunyavirus infection. The invention is further directed to a pharmaceutical composition which is effective in inhibiting the function of UBA5 for use in a method for treating or preventing a disease or condition mediated by an arbovirus infection such as a flavivirus infection. The invention further relates to in vitro methods for identifying pharmaceutically active compounds that are useful for treating or preventing a disease or condition mediated by an arbovirus infection such as a flavivirus infection.

Description

Compound for treating arbovirusinfectionsThe present invention relates to a compound which is effectivein inhibiting the function of the ubiquitin-like modifier acti-vating enzyme 5 (UBA5) and the use of such compound in treatingor preventing a disease or condition mediated by an arbovirusinfection, such as a flavivirus, alphavirus or bunyavirus in-fection. The invention is further directed to a pharmaceuticalcomposition which is effective in inhibiting the function ofUBA5 for use in a method for treating or preventing a disease orcondition mediated by an arbovirus infection, such as a fla-vivirus, alphavirus or bunyavirus infection. The invention fur-ther relates to in vitro methods for identifying pharmaceuti-cally active compounds that are useful for treating or preventinga disease or condition mediated by an arbovirus infection, suchas a flavivirus, alphavirus or bunyavirus infection.Background Several of the most prevalent viral infections in the world arecaused by arboviruses, especially flaviviruses, alphaviruses andbunyaviruses, such as e.g. Chikungunya virus, Rift valley fevervirus, Dengue virus, Yellow fever virus, West Nile virus andZika virus. Most of the flaviviruses are primarily transmitted to humans by arthropods which are typically endemic in tropical and subtropical regions of Africa, South America and Asia. There- fore, diseases caused by flaviviruses, such as vascular shocksyndrome, encephalitis, microencephaly and fetal death, have sofar mainly been a threat to the Global South. In recent years,however, as the earth continues to become warmer, the habitatsof tropical and subtropical arthropods have started to spread,increasing also the risk from diseases mediated by flavivirusesand other arboviruses in the Global North.HAMBURG: TEL.: (040) 899 654-0 FAX: (040) 899 654-88 POSTMASTER@UEX.DE WWW.UEX.DEMÜNCHEN: TEL.: (089) 290 917-0 FAX: (089) 290 917-88 MARIA-THERESIA-STRASSE 13 D-81675 MÜNCHENSitz der Partnerschaft mbB: Hamburg – Amtsgericht Hamburg PR 1062Thus, there is a great need for the treatment and prevention ofdiseases mediated by arboviruses, such as flaviviruses, alpha-viruses and bunyaviruses.To date, the most widely used approach to control arbovirusinfections, and in particular flavivirus infections, is vaccina-tion. Currently, there are licensed and commercially availablevaccines for Yellow fever virus, Dengue virus, Japanese enceph- alitis virus, Kyasanur forest disease virus and Tick-borne en- cephalitis virus, and several vaccine candidates have been in- vestigated in preclinical and clinical trials.However, no antiviral compounds for treating diseases caused byflaviviruses have been approved until now. Recently, a drug can-didate for the treatment of Dengue virus infections has enteredclinical trials. The antiviral activity against Dengue virusesof this compound is based on preventing the interaction of the two Dengue virus proteins NS3 and NS4B, which results in theinhibition of Dengue virus replication (Kaptein et al., 2021,Nature, 598, 504-509). Since the active compound was developedspecifically for targeting the NS3 and NS4B proteins of Dengueviruses, it is not effective against other flaviviruses.Until now, it has not been known that the cellular protein“ubiquitin-like modifier activating enzyme 5” (UBA5) is a hostdependency factor for arbovirus infections such as flavivirusinfections. UBA5 is involved in covalently conjugating the ubiq-uitin-like protein “ubiquitin fold modifier 1” (UFM1) to targetproteins in an enzymatic reaction cascade that is referred to as“UFMylation”. In this reaction cascade UBA5 has the role of theE1-type activating enzyme. It has been previously shown thatUFMylation is associated with several cellular activities in- cluding the endoplasmatic reticulum stress response, hematopoi- esis, fatty acid metabolism and the biogenesis of G-protein cou- pled receptors.In two cases, UFMylation has been reported to affect infectionsof viruses which are, however, no arboviruses, and in particularno flaviviruses, alphaviruses or bunyaviruses. Firstly, UFMyla-tion of the ribosomal protein RPL26 was shown to be required forefficient Hepatitis A virus replication (Kulsupatrakul et al.,2021, Cell Rep., 34(11), 108859). Moreover, the E3 ligase of theUFMylation cascade (UFL1) was reported to promote the antiviralimmune response in Sendai virus infections (Snider et al., 2022,Proc. Natl. Acad. Sci. U.S.A., 119(15):e2119531119). In thesestudies, however, no interaction between a viral protein and theUFM1-activating enzyme UBA5 was described. Instead, it was re-ported that the E3 ligase UFL1 of the UFMylation system is re-cruited to intracellular membranes following Sendai virus infec-tion, suggesting a possible interaction between UFL1 and viralcomponents. Sendai viruses belong to the Respirovirus genus ofthe family Paramyxoviridae. They infect rodents and are typi-cally not pathogenic for humans or domestic animals.UBA5-inhibitory compounds have been described in the art. Theuse of such compounds for treating or preventing a disease orcondition mediated by a viral infection has, however, not beenproposed. The small molecule DKM2-93 (2-chloro-N-(3,4-dimethoxybenzyl)ac-etamide) has been reported to impair pancreatic cancer cell sur-vival and in vivo tumor growth, thus making UBA5 a promisingtherapeutic target for treating pancreatic cancer (Roberts et al., 2017, ACS Chem. Biol., 12, 899-904). WO 2015 / 179955 A1 describes UBA5 inhibitors with metal chelatingmoieties and the use of such inhibitors in disrupting cancerprogression and in treating Leishmania parasite infections.In addition, the natural product usenamine A was reported toexhibit UBA5-inhibitory effects and its use for preventing theprogression of breast cancer was described (Fang et al., 2021,Biomolecules, 11, 1348).The present invention is based on the objective of providing acompound for use in a method for the effective treatment orprevention of a wide range of arbovirus infections, and in par-ticular flavivirus, alphavirus and bunyavirus infections. It isdesirable that, in contrast to a vaccine, the compound is ef- fective in the treatment of a flavivirus infection even if thecompound is administered only after viral infection has oc-curred. In addition, it is an object of the present invention toprovide a compound that can prevent infections or the adverseconsequences of infections when the compound is administered already prior to infection.These objects are achieved by a compound for use in a method oftreating or preventing a disease or condition mediated by aninfection caused by an arbovirus, such as a flavivirus, alpha-virus or bunyavirus infection, according to claims 1 to 13. Theinvention also relates to the pharmaceutical compositions ac-cording to claims 14 and 15 and the in vitro methods accordingto claims 16 and 17.Inhibitory compoundsIn a first aspect, the present invention relates to a compoundwhich is effective in inhibiting the function of the ubiquitin-like modifier activating enzyme 5 (UBA5) for use in a method oftreating or preventing a disease or condition mediated by an arbovirus infection in a subject. In a preferred embodiment, the compound is effective in inhib- iting the function of the ubiquitin-like modifier activating enzyme 5 (UBA5) for use in a method of treating or preventing adisease or condition mediated by a flavivirus, alphavirus orbunyavirus infection in a subject.In accordance with its usual meaning, the term “arbovirus” is used herein to refer to viruses transmitted by arthropod vectors, such as mosquitoes, ticks or sandflies. In the context of the present invention, the term “flavivirus”refers to the genus of the family Flaviviridae which is alsoknown as “orthoflavivirus”. Moreover, “alphaviruses” refers tothe viruses of the alphavirus genus in the Togaviridae family.The term “bunyaviruses” refers to all viruses of the order bun-yavirales. In a more preferred embodiment, the present invention relates to a compound which is effective in inhibiting the function of the ubiquitin-like modifier activating enzyme 5 (UBA5) for use in a method of treating or preventing a disease or condition mediated by a flavivirus. Hence, in any aspect of the present invention described herein, the arbovirus is preferably a flavivirus.The present invention is based on the surprising finding thatproteins of arboviruses, such as flaviviral NS4B proteins, in-teract with UBA5. It was, for example, found that UBA5 interactswith proteins from Zika virus, Dengue virus, Yellow fever virus,Powassan virus and Usutu virus. Importantly, it was found thatthe inhibition of UBA5 results in an inhibition of arbovirusreplication, such as flavivirus, alphavirus or bunyavirus rep-lication. Hence, UBA5 was found to be a host dependency factorfor arbovirus infections, in particular flavivirus, alphavirusand bunyavirus infections. In the light of the above findings,the present invention specifically provides the use of UBA5 in-hibitors in a method of treating or preventing a disease orcondition mediated by an arbovirus infection, such as a fla-vivirus, alphavirus or bunyavirus infection, in a subject.Most approved antiviral compounds, which are so far only avail- able for treating viral infections of other viruses than fla-viviruses, target viral proteins which are required for viralreplication. But targeting host proteins that are involved inthe virus replication cycle is a promising strategy for avoidingvirus resistance and potentially leads to broad-spectrum anti-virals, since viruses from the same virus family often exploitthe same cellular pathways. The compounds according to the first aspect of the invention affect the function of a UBA5 protein, preferably the functionof the human UBA5 protein. The terms “ubiquitin-like modifieractivating enzyme 5” and “UBA5” refer to the E1-type activatingenzyme that is encoded by the UBA5 gene. Activation of UFM1 byUBA5 occurs in a two-step mechanism (Gerakis et al., Trends CellBiol., 2019, Vol. 29, No. 12, 974-986). In a first reaction, UBA5 catalyzes the adenylation of the C-terminus of UFM1. Thisis followed by a second reaction in which UFM1 is attached tothe cystein residue in the active site of UBA5. After activation,UFM1 is transferred from UBA5 to the E2 UFM1-conjugase UFC1 ina transthiolation reaction.Two alternatively spliced transcript variants of UBA5 have beendescribed. The nucleotide sequence of UBA5 is depicted in SEQ IDNO:1. The corresponding amino acid sequences of the transcriptvariants are depicted in SEQ ID NOs:2-3. The compound according to the first aspect of the present in-vention is effective in inhibiting the function of UBA5. Thismeans that the compound causes fewer products to be formed inone or more of the reactions, in which UBA5 is involved, than in the absence of the compound. For example, fewer products can beformed in one or more of (i) the adenylation of UFM1, (ii) theattachment of UFM1 to the active site of UBA5 and (iii) thetransfer of UFM1 to UFC1. Alternatively or in addition, the compound may cause the amount of UBA5 to be reduced. In a preferred embodiment, the compound of the first aspect of the invention decreases the UBA5 activity by at least 10%, at least 20%, at least 30%, at least 40%, at least 50%, at least60%, at least 70%, at least 80%, at least 90% or at least 95% ascompared to the activity in the absence of the compound. In aparticularly preferred embodiment, the compound completelyblocks the UBA5 activity such that there is no detectable trans-fer of UFM1 to UFC1. Preferably, the compound according to the first aspect of thepresent invention is effective in inhibiting the function of aubiquitin-like modifier enzyme (UBA5) comprising, or preferablyconsisting of, an amino acid sequence of any one of SEQ ID NOs:2-3, or an amino acid sequence having at least 80%, preferably atleast 90%, sequence identity to at least one of the sequences ofSEQ ID NOs:2-3.UBA5 inhibition may be determined by any suitable assay. Forexample, UBA5 inhibition can be measured in an immunoblot assay(Zhou et al., STAR Protocols 3, 101074, March 18, 2022) orthrough an in vitro transthiolation assay (Sara et al., Bioor-ganic & Medicinal Chemistry Letters, Volume 26, Issue 18, 2016).Preferably, the compound is specific for UBA5, i.e. the compoundinhibits the function of UBA5, while it does essentially notinhibit the functions of other proteins or enzymes, such as otherE1 activating enzymes. As a result of this UBA5 specificity, thecompounds according to this embodiment elicit no or only toler-able side effects when administered to a subject. The pharmaceutical composition according to the present inven-tion is suitable for the treatment and prevention of infectionscaused by arboviruses, such as flavivirus, alphavirus and bun-yavirus infections. The viral infections may be symptomatic orasymptomatic.Treating an infection caused by an arbovirus, such as a fla-vivirus infection, may involve impairing the viral replicationcycle, reducing the viral load in the infected subject and / orreducing the symptoms caused by the viral infection or by the infected person’s immune response.Preventing an arbovirus infection such as a flavivirus infectionmay involve reducing a subject’s infectiousness, i.e. the like-lihood for an infected subject to infect another individual,and / or a subject’s susceptibility for infection, i.e. the like-lihood for a subject to be infected by another infected indi-vidual.According to the present invention, the compound may be intendedfor administration to a subject with an increased risk of beinginfected with an arbovirus, in particular flavivirus, alphavirusor bunyavirus, e.g. because the subject resides at an area witha high abundance of arbovirus vectors, such as flavivirus, al-phavirus or bunyavirus vectors. The compound may also be intendedfor administration to a subject that is at a particular riskfrom the effects of the viral infection, e.g. due to pre-existinghealth conditions, high age or, particularly in the case of Zikavirus infections, pregnancy. The compound of the first aspect of the present invention mayalso be intended for treating a condition mediated by an infec-tion caused by an arbovirus, such as a flavivirus, alphavirus orbunyavirus infection. Herein, term “condition” refers to a statewhich is not necessarily considered to be a disease, but which deviates from the healthy, uninfected state. For example, thecondition mediated by an arbovirus infection such as an fla-vivirus infection may be an asymptomatic infection or a presymp- tomatic infection, i.e. an infection before the onset of symp- toms.The subject to which the compound of the invention is adminis-tered is preferably a mammal, and in particular a human. In apreferred embodiment, the subject is a pregnant woman. Particu-larly preferably, the subject is a pregnant woman and the com-pound is to be used in a method of treating or preventing adisease or condition mediated by a Zika virus infection.The compound can be selected from all types of inorganic andorganic molecules, including peptides, polypeptides, oligo- orpolysaccharides, nucleic acids, fatty acids, steroids, smallmolecules and the like. For example, inhibitory compounds can beantisense DNA or RNA polynucleotides which bind to the UBA5 geneand block transcription, RNA processing and / or translation of said gene. Also, the UBA5-inhibitory compound of the invention can be an RNA molecule which exerts its effect by RNA interfer- ence. Alternatively, the inhibitory compound of the inventioncan be a ribozyme that cleaves the UBA5-encoding mRNA. Otherclasses of molecules which may give rise to suitable UBA5-in-hibitory compounds include peptides, antibodies and antibodyfragments. Peptides that bind and interfere with the UBA5 proteinmay be conveniently screened in random peptide libraries.It is preferred that the compound of the present invention is asmall molecule. In particular, the compound has a molecularweight of 2,500 Da or less. Further preferred is a compoundhaving a molecular weight of 2,000 Da or less, in particular1,500 Da or less, more preferably 1,000 Da or less and particu-larly preferably 550 Da or less.The UBA5-inhibitory compound is intended to be administered inan amount that is effective to inhibit the function of UBA5 inorder to treat or prevent a disease or condition mediated by aninfection caused by an arbovirus, in particular a flavivirus,alphavirus or bunyavirus infection in a subject, e.g. by inhib-iting arbovirus replication. This amount, which is also referredto as the “therapeutically effective amount” of the UBA5-inhib-itory compound to be administered will depend on several param- eters, such as the mode of administration, the particular typeof arbovirus infection, the particular type of UBA5-inhibitorycompound, the severity of the infection or disease or conditionwhich is mediated by the arbovirus infection, and the age,height, weight, health, and physical condition of the subject tobe treated. A therapeutically effective amount of the UBA5-in-hibitory compound can be determined by one of ordinary skill inthe art without undue experimentation given the disclosure set forth herein. In a preferred embodiment, a therapeutically effective amount ofa compound of the first aspect of the invention decreases arbo-virus replication, such as flavivirus, alphavirus or bunyavirusreplication by at least 20%, in particular at least 50% andparticularly preferably by at least 80%. For example, a thera-peutically effective amount of a compound of the first aspect ofthe invention may decrease arbovirus replication, preferablyflavivirus, alphavirus or bunyavirus replication, by at least5%, at least 10%, at least 15%, at least 20%, at least 25%, atleast 30%, at least 35%, at least 40%, at least 45%, at least50%, at least 55%, at least 60%, at least 65%, at least 70%, atleast 75%, at least 80%, at least 85%, or at least 90%. Thepercentages of decrease of arbovirus replication relate in eachcase to the replication in absence of the compound.For treating or preventing an arbovirus infection, in particulara flavivirus, alphavirus or bunyavirus infection, the UBA5-in-hibitory compound will preferably be administered to the subjectin an amount that ranges from 0.1 µg / kg body weight to 50,000µg / kg body weight, e.g. from 0.5 µg / kg body weight to 30,000µg / kg body weight, from 1.0 µg / kg body weight to 15,000 µg / kgbody weight, from 5.0 µg / kg body weight to 10,000 µg / kg bodyweight, from 7.5 µg / kg body weight to 5,000 µg / kg body weight,from 10 µg / kg body weight to 2,500 µg / kg body weight, from 25µg / kg body weight to 1,500 µg / kg body weight, from 50 µg / kg bodyweight to 1,000 µg / kg body weight, and more preferably from 100µg / kg body weight to 800 µg / kg body weight.In a preferred embodiment, the compound of the invention is used in a method of treating or preventing a disease or condition mediated by an arbovirus infection, preferably by a flavivirus, alphavirus or bunyavirus infection, in particular by an infec-tion caused by a flavivirus, alphavirus or virus of the Phenui-viridae family, most preferably by a flavivirus, alphavirus orphlebovirus infection.In another preferred embodiment, the compound of the inventionis used in a method of treating or preventing a disease or condition mediated by a flavivirus, alphavirus or bunyavirus infection, wherein the flavivirus is selected from the groupconsisting of Zika virus, Japanese encephalitis virus, Powassanvirus, West Nile virus, Yellow fever virus, Dengue virus, Usutuvirus, and Tick-borne encephalitis virus, and wherein the al-phavirus is selected from Chikungunya virus and Mayaro virus andwherein the bunyavirus is Rift valley fever virus and more pref-erably the flavivirus is selected from the group consisting ofZika virus, Powassan virus, Yellow fever virus, Dengue virus,and Usutu virus, the alphavirus is selected from Chikungunyavirus and Mayaro virus and the bunyavirus is Rift valley fever virus.In a particularly preferred embodiment, the arbovirus infectionis a flavivirus or alphavirus infection, preferably a flavivirusinfection, which is in particular a Zika virus infection.The compound according to the first aspect of the invention iseffective in inhibiting the function of UBA5. In a preferredembodiment, the compound is DKM2-93 (CAS No. 65836-72-8, 2-chloro-N-(3,4-dimethoxybenzyl)acetamide). In another preferred embodiment, the compound is usenamine A ((2E,9bR)-6-acetyl-2-(1-aminoethylidene)-7,9-dihydroxy-8,9b-di- methyldibenzofuran-1,3-dione).In even another preferred embodiment, the compound has the struc-ture of Formula (I)is a polyazamacrocycle chelating group,M is a chelatable metal ion,Y is -C(=O)- or -CH2-,L is (i) (C1-C20)-alkylene, wherein(i.a) at least one of the carbon atoms is option-ally replaced with a heteroatom selectedfrom O, NR' and S, wherein R' is H, (C1- C6)-alkyl or -C(=O)-(C1-C6)-alkyl,(i.b) two or three adjacent carbon atoms arejoined together to form a (C3-C10)-cycloal- kyl group or -(C6-C10)-aryl group, and / or(i.c) the (C1-C20)-alkylene group is optionallysubstituted with at least one halo, (ii) (C2-C20)-alkenylene, wherein(ii.a) at least one of the carbon atoms is option-ally replaced with a heteroatom selected from O, NR' and S, wherein R' is H, (C1- C6)-alkyl or -C(=O)-(C1-C6)-alkyl,(ii.b) two or three adjacent carbon atoms are op-tionally joined together to form a (C3-C10)- cycloalkyl group or -(C6-C10)-aryl group,and / or (ii.c) the (C2-C20)-alkenylene group is optionallysubstituted with at least one halo, or (iii) a polyethylene glycol (PEG) moiety,W is (i) -NH-C(=O)-,(ii) -NR'-, wherein R' is H, (C1-C6)-alkylor -C(=O)-(C1-C6)-alkyl,X is (i) -O-,(ii) NR', wherein R' is H, (C1-C6)-alkylor -C(=O)-(C1-C6)-alkyl,(iii) -S-; or -S(=O)2-, or(iv) -C(R")2, wherein each R" is independentlyor simultaneously H, halo or (C1-C6)-alkyl, Raand Ra'are each independently or simultaneously (i) H,(ii) OH,(iii) halo, or(iv) (C1-C3)-alkyl, andG is (i) O,(ii) S,(iii) NR2,R1and R2are each independently or simultaneously (i) H,(ii) (C1-C6)-alkyl,(iii) (C3-C10)-cycloalkyl,(iv) (C3-C10)-heterocycloalkyl,(v) -(CH2)n-(C6-C10)-aryl, wherein n is 1 to 6,preferably 1 to 3, (vi) -(CH2)n-(C5-C10)-heteroaryl, wherein n is 1to 6, preferably 1 to 3, or R1and R2are joined together to form a (vii) guanine or a guanine derivative,(viii) cytosine or a cytosine derivative,(ix) thymine or a thymine derivative,(x) adenine or an adenine derivative,and Q is any suitable counteranion, or a solvate, prodrug and / or stereoisomer thereof.As used herein, the term “polyazamacrocycle chelating group”refers to a macrocycle containing at least 8 atoms, at least two of which (preferably 3, 4, 5 or 6 atoms) are nitrogen atoms, andwhich is able to chelate a metal ion. The polyazamacrocyclechelating group may also have carboxylic acid substituents, suchas ethanoic substituents, bound to the nitrogen atoms. Pol-yazamacrocycle chelating groups as well as suitable chelatable metal ions, which are preferred according to the present inven- tion, are described in WO 2015 / 179955 A1, particularly in para- graphs

[0048] ,

[0049] and

[0060] -

[0064] and in the claims.As used herein, the term "derivative" refers to a compound, groupor moiety which comprises the same basic carbon skeleton and functionality as the parent, but can also bear one or more sub- stituents or substitutions of the parent compound, group or moi- ety. For example, nucleoside derivatives (guanine, cytosine, thymine or adenine), include amino substitutions, alkyl aminesubstitutions, heterocyclic substitutions, aryl or heteroaryl substitutions. As used herein, the term "prodrug" refers to a substance that is prepared in an inactive form that is converted to an active form (i.e., drug) within the body or cells thereof by the action of, for example, endogenous enzymes or other chemicals and / or con- ditions. Prodrug derivatives of the compounds of Formula (I) or pharmaceutically acceptable salts or solvates thereof, can be prepared by methods known to those of ordinary skill in the art, and include esters of any free hydroxyl or carboxyl moieties of the compounds.Further groups M, Y, L, W, G, X, Ra, Ra’, R1, which are preferredaccording to the present invention, are also described in WO2015 / 179955 A1, particularly in the claims.It is further preferred that the compound has the structure ofFormula (I)is a polyazamacrocycle chelating group with the structure,M is Zn2+, Cu2+, Co2+, Pd2+, Gd2+, Tb2+, Eu+ or Mo2+,preferably Zn2+or Cu2+,Y is -C(=O)- or -CH2-,L is (i) (C1-C6)-alkylene,preferably (C3-C6)-alkylene, wherein(i.a) one to three of the carbon atoms are op-tionally replaced with a heteroatom se-lected from O, NR’ and S, wherein R’ is H,(C1-C6)-alkyl or -C(=O)-(C1-C6)-alkyl,(i.b) two or three adjacent carbon atoms arejoined together to form a cyclohexyl groupor phenyl group, and / or(i.c) the alkylene group is optionally substi-tuted with at least one fluoro,or (ii) a polyethylene glycol (PEG) moiety,W is (i) -NH-C(=O)-,(ii) -NR'-, wherein R' is H, (C1-C6)-alkylor -C(=O)-(C1-C6)-alkyl,has one of the following structuresR1 and R2 are each independently or simultaneously(i) H,(ii) (C5-C7)-cycloalkyl,preferably cyclopentyl or cyclohexyl, (iii) (C5-C7)-heterocycloalkyl,Preferably morpholinyl or piperazinyl, (iv) phenyl or –(CH2)-phenyl,(v) naphthyl or –(CH2)-naphthyl,(vi) -(CH2)n-(C5-C6)-heteroaryl, wherein n is 1to 6, preferably 1 to 3, or R1and R2are joined together to form a(vii) guanine or a guanine derivative,(viii) cytosine or a cytosine derivative,(ix) thymine or a thymine derivative,(x) adenine or an adenine derivative,and wherein R1 and R2 are particularly preferably joined to-gether to form an adenosine derivative having the structurewherein V is H, halo, (C1-C6)-alkyl,(C6-C10)-aryl, (C5-C10)-het- eroaryl, (C3-C10)-cycloalkyl, (C3-C10)-heterocycloalkyl, and Q is any suitable counteranion, or a solvate, prodrug and / or stereoisomer thereof.In a particularly preferred embodiment, the compound has thestructure of formula (II)(II),wherein n is an integer in the range of 1 to 5, and preferablyis 4 or 5, particularly preferably 5. In another preferred embodiment, the compound is a stereoisomerof the structure of formula (II), wherein n is in particular 4 or5, more preferably 5.Hence, the UBA5-inhibitory compound may be selected from thegroup consisting of DKM2-93 (CAS No. 65836-72-8), usenamine A,and the compound with the structure of formula (I),wherein M, Y, L, W, G, X, Ra, Ra’, R1 are as defined above.Pharmaceutical compositionThe compound of the present invention that inhibits UBA5 isnormally provided in the form of a pharmaceutical composition. Therefore, in a second aspect, the invention relates to a phar-maceutical composition comprising the compound of the first as-pect of the invention, or its pharmaceutically acceptable salt,solvate, prodrug or ester, for use in a method for treating orpreventing a disease or condition mediated by an arbovirus in-fection, in particular by a flavivirus, alphavirus or bunyavirusinfection, in a subject. The pharmaceutical composition of thepresent invention comprises an amount of the UBA5-inhibitorycompound that is effective to inhibit the function of UBA5 inorder to inhibit arbovirus replication, in particular fla-vivirus, alphavirus or bunyavirus replication.The pharmaceutical composition of the invention normally com-prises one or more excipients. Preferably the pharmaceuticalcomposition comprises one or more excipients selected from thegroup consisting of diluents, solvents, binders, carriers, lub-ricants, fillers, glidant, coatings, surfactants, stabilizingagents, antitacking agents, preservatives, emulsifiers, disper-sants, wetting agents, and adjuvants. Generally, the pharmaceu-tical composition may be administered in any suitable form thatdoes not interfere with the compound’s inhibitory activity.Suitable pharmaceutical formulations can be determined by theskilled person depending on the route of administration and thedesired dosage. See, e.g., Remington’s Pharmaceutical Sciences, 1435-712 (18th ed., Mack Publishing Co, Easton, Pennsylvania, 1990). Formulations may influence the physical state, stability,rate of in vivo release and rate of in vivo clearance of theadministered agents. Depending on the route of administration, a suitable dose may be calculated according to body weight, body surface areas or organ size. Further refinement of the calcula- tions necessary to determine the appropriate treatment dose is routinely made by those of ordinary skill in the art without undue experimentation, especially in light of the dosage infor- mation and assays disclosed herein as well as the pharmacokinetic data obtainable through animal or human clinical trials. The compound can be present in a pharmaceutical composition as a pharmaceutically acceptable salt. As used herein, “pharmaceu-tically acceptable salts” include, for example, base additionsalts and acid addition salts. Pharmaceutically acceptable base addition salts may be formed with metals or amines, such as alkali and alkaline earth metals or organic amines. Pharmaceutically acceptable salts of com- pounds may also be prepared with a pharmaceutically acceptable cation. Suitable pharmaceutically acceptable cations are wellknown to those skilled in the art and include alkaline, alkalineearth, ammonium and quaternary ammonium cations. Carbonates or hydrogen carbonates are also possible. Examples of metals used as cations are sodium, potassium, magnesium, ammonium, calcium, or ferric, and the like. Examples of suitable amines includeisopropylamine, trimethylamine, histidine, N,N'-dibenzyleth- ylenediamine, chloroprocaine, choline, diethanolamine, dicyclo- hexylamine, ethylenediamine, N-methylglucamine, and procaine. Pharmaceutically acceptable acid addition salts include inor- ganic or organic acid salts. Examples of suitable acid salts include the hydrochlorides, formates, acetates, citrates, sa- licylates, nitrates, phosphates. Other suitable pharmaceutically acceptable salts are well known to those skilled in the art and include, for example, formic, acetic, citric, oxalic, tartaric, or mandelic acids, hydrochloric acid, hydrobromic acid, sulfuric acid or phosphoric acid; with organic carboxylic, sulfonic, sulfo or phospho acids or N-substituted sulfamic acids, for ex- ample acetic acid, trifluoroacetic acid (TFA), propionic acid, glycolic acid, succinic acid, maleic acid, hydroxymaleic acid, methylmaleic acid, fumaric acid, malic acid, tartaric acid, lac- tic acid, oxalic acid, gluconic acid, glucaric acid, glucuronic acid, citric acid, benzoic acid, cinnamic acid, mandelic acid, salicylic acid, 4-aminosalicylic acid, 2-phenoxybenzoic acid, 2- acetoxybenzoic acid, embonic acid, nicotinic acid or isonico- tinic acid; and with amino acids, such as the 20 alpha amino acids involved in the synthesis of proteins in nature, for ex- ample glutamic acid or aspartic acid, and also with phenylaceticacid, methanesulfonic acid, ethanesulfonic acid, 2 -hydroxyethanesulfonic acid, ethane 1,2-disulfonic acid, benzenesulfonic acid, 4-methylbenzenesulfonic acid, naphthalene 2-sulfonicacid,naphthalene 1,5-disulfonic acid, 2- or 3-phosphogly cerate, glu-cose 6-phosphate, N-cyclohexylsulfamic acid (with the formation of cyclamates), or with other acid organic compounds, such as ascorbic acid. Pharmaceutical compositions containing the compounds disclosed herein can be manufactured in a conventional manner, e.g., by conventional mixing, dissolving, granulating, dragee-making,levigating, emulsifying, encapsulating, entrapping, or lyophi- lizing processes. Proper formulation is dependent upon the route of administration chosen. The pharmaceutical composition of the present invention may be formulated for oral administration. For oral administration, suitable compositions can be formulatedreadily by combining the compound of the first aspect of theinvention with pharmaceutically acceptable excipients such ascarriers well known in the art. Such excipients and carriers enable the present compounds to be formulated as tablets, pills, dragees, capsules, liquids, gels, syrups, slurries, suspensions and the like, for oral ingestion by a patient to be treated. Pharmaceutical preparations for oral use can be obtained by add-ing the compound of the first aspect of the invention with asolid excipient, optionally grinding a resulting mixture, and processing the mixture of granules, after adding suitable aux- iliaries, if desired, to obtain tablets or dragee cores. Suitable excipients include, for example, fillers and cellulose prepara- tions. If desired, disintegrating agents can be added. Pharma- ceutically acceptable ingredients are well known for the various types of formulation and may be for example binders (e.g., nat- ural or synthetic polymers), lubricants, surfactants, sweeteningand flavoring agents, coating materials, preservatives, dyes,thickeners, adjuvants, antimicrobial agents, antioxidants and carriers for the various formulation types.When a therapeutically effective amount of the compound of thefirst aspect is administered orally, the composition typicallyis in the form of a solid (e.g., tablet, capsule, pill, powder, or troche) or a liquid formulation (e.g., aqueous suspension, solution, elixir, or syrup).When administered in tablet form, the composition can addition- ally contain a functional solid and / or solid carrier, such as a gelatin or an adjuvant. The tablet, capsule, and powder cancontain about 1 to about 95% by weight of the compound of thefirst aspect, and preferably from about 15 to about 90% by weightof the compound of the first aspect.When administered in liquid or suspension form, a functionalliquid and / or a liquid carrier such as water, petroleum, or oils of animal or plant origin can be added. The liquid form of the composition can further contain physiological saline solution,sugar alcohol solutions, dextrose or other saccharide solutions,or glycols. When administered in liquid or suspension form, thecomposition can contain about 0.5 to about 90% by weight of thecompound of the first aspect, and preferably about 1 to about50% of the compound of the first aspect. In one embodiment con-templated, the liquid carrier is non-aqueous or substantially non-aqueous. For administration in liquid form, the compositionmay be supplied as a rapidly-dissolving solid formulation fordissolution or suspension immediately prior to administration.In another embodiment, the pharmaceutical composition of the present invention may be formulated for parenteral administra- tion.When a therapeutically effective amount of the compound of thefirst aspect of the invention is administered by intravenous,cutaneous, or subcutaneous injection, the composition is in the form of a pyrogen-free, parenterally acceptable aqueous solu-tion. The preparation of such parenterally acceptable solutions,having due regard to pH, isotonicity, stability, and the like, is within the skill in the art. A preferred composition forintravenous, cutaneous, or subcutaneous injection typically con-tains, in addition to a compound disclosed herein, an isotonicvehicle. Such compositions may be prepared for administration assolutions of free base or pharmacologically acceptable salts inwater suitably mixed with a surfactant, such as hydroxypropyl-cellulose. Dispersions also can be prepared in glycerol, liquidpolyethylene glycols, and mixtures thereof and in oils. Underordinary conditions of storage and use, these preparations canoptionally contain a preservative to prevent the growth of mi- croorganisms. The compounds of first aspect can be formulated for parenteral administration by injection (e.g., by bolus injection or con- tinuous infusion). Formulations for injection can be presented in unit dosage form (e.g., in ampules or in multidose contain- ers), with an added preservative. The compositions can take such forms as suspensions, solutions, or emulsions in oily or aqueous vehicles, and can contain formulatory agents such as suspending, stabilizing, and / or dispersing agents. Sterile injectable solutions are prepared by incorporating the active compound in the required amount in the appropriate solvent with various of the other ingredients enumerated above, as re- quired, followed by filtered sterilization. Generally, disper- sions are prepared by incorporating the various sterilized ac- tive ingredients into a sterile vehicle which contains the basic dispersion medium and the required other ingredients from those enumerated above. In the embodiment of sterile powders for the preparation of sterile injectable solutions, the preferred meth- ods of preparation are vacuum-drying and freeze-drying tech- niques which yield a powder of the active ingredient plus any additional desired ingredient from a previously sterile-filtered solution thereof. Injectable compositions can include sterile aqueous solutions,suspensions, or dispersions and sterile powders for the extem-poraneous preparation of sterile injectable solutions, suspen-sions, or dispersions. In all embodiments the form must be ster-ile and must be fluid to the extent that easy syringabilityexists. It must be stable under the conditions of manufactureand storage and must resist the contaminating action of micro-organisms, such as bacteria and fungi, by optional inclusion ofa preservative. The carrier can be a solvent or dispersion mediumcontaining, for example, water, ethanol, polyol (e.g., glycerol,propylene glycol, and liquid polyethylene glycol, and the like),suitable mixtures thereof, and vegetable oils. In one embodimentcontemplated, the carrier is non-aqueous or substantially non- aqueous. The proper fluidity can be maintained, for example, bythe use of a coating, such as lecithin, by the maintenance ofthe required particle size of the compound in the embodiment ofdispersion and by the use of surfactants. The prevention of theaction of microorganisms can be brought about by various anti-bacterial and antifungal agents, for example, parabens, chloro-butanol, phenol, sorbic acid, thimerosal, and the like. In manyembodiments, it will be preferable to include isotonic agents,for example, sugars or sodium chloride. Prolonged absorption ofthe injectable compositions can be brought about by the use inthe compositions of agents delaying absorption, for example,aluminum monostearate and gelatin. Pharmaceutical formulations for parenteral administration in- clude aqueous solutions of the compounds in water-soluble form. Additionally, suspensions of the compounds can be prepared as appropriate oily injection suspensions. Suitable lipophilic sol- vents or vehicles include fatty oils or synthetic fatty acid esters. Aqueous injection suspensions can contain substances which increase the viscosity of the suspension. Optionally, the suspension also can contain suitable stabilizers or agents that increase the solubility of the compounds and allow for the prep- aration of highly concentrated solutions. Alternatively, a pre- sent composition can be in powder form for constitution with a suitable vehicle (e.g., sterile pyrogen-free water) before use.For administration by inhalation, compounds of the first aspectof the present invention are conveniently delivered in the formof an aerosol spray presentation from pressurized packs or anebulizer, with the use of a suitable propellant. In the embod-iment of a pressurized aerosol, the dosage unit can be determinedby providing a valve to deliver a metered amount. Capsules andcartridges of, e.g., gelatin, for use in an inhaler or insuf-flator can be formulated containing a powder mix of the compoundand a suitable powder base such as lactose or starch.Compounds of the first aspect can also be formulated in rectalcompositions, such as suppositories or retention enemas (e.g.,containing conventional suppository bases). In addition to theformulations described previously, the compounds also can beformulated as a depot preparation. Such long-acting formulationscan be administered by implantation (e.g., subcutaneously orintramuscularly) or by intramuscular injection. Thus, for exam-ple, the compounds can be formulated with suitable polymeric orhydrophobic materials (for example, as an emulsion in an ac-ceptable oil) or ion exchange resins, or as sparingly soluble derivatives, for example, as a sparingly soluble salt. In particular, a compound of the first aspect of the inventioncan be administered orally, buccally, or sublingually in theform of tablets containing excipients, such as starch or lactose,or in capsules or ovules, either alone or in admixture withexcipients, or in the form of elixirs or suspensions containingflavoring or coloring agents. Such liquid preparations can be prepared with pharmaceutically acceptable additives, such assuspending agents. A compound also can be injected parenterally,for example, intravenously, intramuscularly, subcutaneously, orintracoronarily. For parenteral administration, the compound isbest used in the form of a sterile aqueous solution which cancontain other substances, for example, salts, or sugar alcohols,such as mannitol, or glucose, to make the solution isotonic with blood. Slow release or sustained release formulations may also be pre- pared in order to achieve a controlled release of the activecompound in contact with the body fluids in the gastrointestinal(GI) tract, and to provide a substantially constant and effectivelevel of the active compound in the blood plasma. For example, release can be controlled by one or more of dissolution, diffu- sion, and ion-exchange. In addition, the slow release approach may enhance absorption via saturable or limiting pathways within the GI tract. For example, the compound may be embedded for this purpose in a polymer matrix of a biological degradable polymer, a water-soluble polymer or a mixture of both, and optionally suitable surfactants. Embedding can mean in this context the incorporation of micro-particles in a matrix of polymers. Con- trolled release formulations are also obtained through encapsu- lation of dispersed micro-particles or emulsified micro-droplets via known dispersion or emulsion coating technologies. The amount of compound administered can be dependent on thesubject being treated, on the subject's age, health, sex, andweight, the kind of concurrent treatment (if any), severity ofthe affliction, the nature of the effect desired, the manner andfrequency of treatment, and the judgment of the prescribing phy-sician. The frequency of dosing also can be dependent on phar-macodynamic effects on arterial oxygen pressures. However, themost preferred dosage can be tailored to the individual subject,as is understood and determinable by one of skill in the art,without undue experimentation. This typically involves adjust-ment of a standard dose (e.g., reduction of the dose if thepatient has a low body weight). While individual needs vary, determination of optimal ranges ofeffective amounts of the compound is within the skill of theart. For administration to a human in the curative or prophy-lactic treatment of the conditions and disorders identifiedherein, for example, typical dosages of the compounds of thepresent invention can be about 0.01 mg / kg / day to about 50mg / kg / day, for example at least 0.05 mg / kg, at least 0.08 mg / kg,at least 0.1 mg / kg, at least 0.2 mg / kg, at least 0.3 mg / kg, atleast 0.4 mg / kg, or at least 0.5 mg / kg, and preferably 50 mg / kgor less, 40 mg / kg or less, 30 mg / kg or less, 20 mg / kg or less,or 10 mg / kg or less, which can be about 2.5 mg / day (0.5 mg / kg x5kg) to about 5000 mg / day (50mg / kg x 100kg), for example. Forexample, dosages of the compounds can be about 0.1 mg / kg / day toabout 50 mg / kg / day, about 0.05 mg / kg / day to about 10 mg / kg / day,about 0.05 mg / kg / day to about 5 mg / kg / day, about 0.05 mg / kg / dayto about 3 mg / kg / day, about 0.07 mg / kg / day to about 3 mg / kg / day,about 0.09 mg / kg / day to about 3 mg / kg / day, about 0.05 mg / kg / dayto about 0.1 mg / kg / day, about 0.1 mg / kg / day to about 1 mg / kg / day,about 1 mg / kg / day to about 10 mg / kg / day, about 1 mg / kg / day toabout 5 mg / kg / day, about 1 mg / kg / day to about 3 mg / kg / day, about3 mg / day to about 500 mg / day, about 5 mg / day to about 250 mg / day,about 10 mg / day to about 100 mg / day, about 3 mg / day to about 10mg / day, or about 100 mg / day to about 250 mg / day. Such doses maybe administered in a single dose or it may be divided into multiple doses.In vitro screening methodThe finding that UBA5-inhibitory compounds inhibit arbovirusreplication, and in particular flavivirus, alphavirus and bun-yavirus replication, in a subject allows the design of screeningassays which identify pharmaceutically active compounds which are effective in treating and / or preventing a disease or condi-tion mediated by an arbovirus such as flavivirus infection in asubject. Thus, the present invention also provides methods foridentifying a pharmaceutically active compound that could be used for treating or preventing a disease or condition mediatedby an arbovirus infection, preferably by an flavivirus, alpha-virus or bunyavirus infection, in a subject. The in vitro methodsof the invention can be used for drug screening approaches that aim to identify new pharmaceutically active compounds for treat- ing or preventing a disease or condition mediated by an arbovirus infection.In a third aspect, the invention relates to an in vitro methodfor identifying a pharmaceutically active compound for treating or preventing a disease or condition mediated by an arbovirusinfection, preferably by a flavivirus, alphavirus or bunyavirusinfection, in a subject, comprising(a) contacting a candidate compound with a ubiquitin-like mod-ifier activating enzyme 5 (UBA5),(b) detecting whether the candidate compound inhibits thefunction of UBA5, wherein a compound which inhibits the function of UBA5 is suit- able for treating or preventing a disease or condition mediatedby an arbovirus infection, preferably by an flavivirus, alpha-virus or bunyavirus infection, in a subject.Contacting a candidate compound with UBA5 may, for example, take place in a cell culture system.UBA5 inhibition may be measured in an immunoblot assay (Zhou etal., STAR Protocols 3, 101074, March 18, 2022). The UBA5 pref-erably comprises, in particular consists of, an amino acid se- quence of any one of SEQ ID Nos:2-3, or an amino acid sequence having at least 80%, in particular at least 90%, sequence iden- tity to at least one of the sequences of SEQ ID Nos:2-3.In an alternative approach, the screening can target compoundswhich interfere with arbovirus replication, preferably with fla-vivirus, alphavirus or bunyavirus replication. The inventiontherefore relates, in a fourth aspect, to an in vitro method foridentifying a pharmaceutically active compound for treating orpreventing a disease or condition mediated by an arbovirus in-fection, preferably by a flavivirus, alphavirus or bunyavirusinfection, in a subject, comprising(a) contacting a candidate compound with a cell that expressesa functional ubiquitin-like modifier activating enzyme 5 (UBA5),(b) infecting the cell with an arbovirus, preferably with aflavivirus, alphavirus or bunyavirus, before or after thecell was contacted with the candidate compound,(c) detecting whether the candidate compound decreases thereplication of the arbovirus, preferably of the fla-vivirus, alphavirus or bunyavirus, in the cell, comparedto a cell which was infected with the arbovirus, preferablywith the flavivirus, alphavirus or bunyavirus, but notcontacted with the candidate compound, wherein a compound which decreases the replication of an arbo-virus, preferably of an flavivirus, alphavirus or bunyavirus, inthe cell, compared to a cell which was infected with the arbo-virus, preferably with the flavivirus, alphavirus or bunyavirus,but not contacted with the candidate compound, is suitable fortreating or preventing a disease or condition mediated by anarbovirus infection, preferably by an flavivirus, alphavirus orbunyavirus infection, in a subject.Whether the proteins expressed in the cell are functional or notcan be determined in an immunoblot assay (Zhou et al., STARProtocols 3, 101074, March 18, 2022). Proteins are consideredfunctional if they at least one of the known reaction productsof UBA5 can be detected.Typically, contacting a candidate compound with a cell that ex- presses functional UBA5 in step (a) also includes that the can- didate compound is contacted with UBA5.The inhibition of arbovirus replication, in particular fla-vivirus, alphavirus or bunyavirus replication, may be determinedby any suitable assay. For example, flavivirus replication maybe determined using a Renilla luciferase reporter assay. TheUBA5 preferably comprises, in particular consists of, an amino acid sequence of any one of SEQ ID Nos:2-3, or an amino acid sequence having at least 80%, in particular at least 90%, se- quence identity to at least one of the sequences of SEQ ID Nos:2- 3.Thus, the in vitro methods of the third and fourth aspect of theinvention include methods, in which the candidate compound is acompound of the first aspect of the invention that is effectivein inhibiting the function of UBA5. The invention disclosed herein is exemplified by the following examples which should not be construed to limit the scope of the invention. Alternative mechanistic pathways and analogous struc- tures will be apparent to those skilled in the art.a) UBA5 interacts with the NS4B protein of multiple flavivirusesThe interaction of UBA5 with NS4B proteins of flaviviruses wasstudied by label-free quantitative proteomics using the iBAQ(intensity-based absolute quantification) method.For sample preparation, JEG-3 human placental cells stably ex-pressing HA-tagged 2k-NS4B proteins of Zika virus (ZIKV, strainH / PF / 2013), Dengue virus (DENV-2, strain 16681), Powassan virus(POWV, strain LB), Usutu virus (USUV, strain Vienna 2001), Yellowfever virus (YFV, strain Asibi) or control proteins (Gaussialuciferase, Gluc; NS4B of Hepatitis C virus (HCV, Jfh-1 strain)were lysed and clarified supernatants were used for HA-specific affinity purification using anti-HA-conjugated agarose beads. After extensive washing, proteins bound to the resin were dena- tured by incubation in 40 μl Urea / Thiourea buffer (6 M Urea / 2 M thiourea in 10 mM HEPES, pH 8.0), and reduced and alkylated in10 mM DTT and 55 mM iodoacetamide, respectively. NS4B- or con-trol-interacting proteins were proteolytically digested with 1 µg LysC (FUJIFILM Wako Chemicals) and 1 µg trypsin (Promega) in 40 mM ABC buffer (50 mM NH4HCO3 in water, pH 8.0) overnight at 25°C. After digestion, peptides were purified on stage tips with 3 layers of C18 Empore filter discs (3M) as previously described (PubMed identifier (PMID): 30177828). Samples were analysed on a nanoElute (plug-in v.1.1.0.27; Bruker) coupled to a trapped ion mobility spectrometry quadru- pole time of flight (timsTOF Pro) (Bruker) equipped with a Cap- tiveSpray source. Peptides were injected into a Trap cartridge (5 mm × 300 μm, 5 μm C18; Thermo Fisher Scientific) and next sepa-rated on a 25 cm × 75 μm analytical column, 1.6 μm C18 beads with apacked emitter tip (IonOpticks). The column temperature was maintained at 50 °C using an integrated column oven (Sonation GmbH). The column was equilibrated using 4 column volumes before loading samples in 100% buffer A (99.9% Milli-Q water, 0.1%formic acid (FA)). Samples were separated at 400 nl  min−1 using alinear gradient from 2 to 17% buffer B (99.9% ACN, 0.1% FA) over 60 min before ramping up to 25% (30 min), 37% (10 min) and 95% of buffer B (10 min) and sustained for 10 min (total separation method time, 120 min). The timsTOF Pro was operated in parallelaccumulation-serial fragmentation (PASEF) mode using Compass Hystar v.5.0.36.0. Settings were as follows: mass range 100– 1700 m / z, 1 / K0 Start 0.6 V⋅s / cm2End 1.6 V⋅s / cm2; ramp time 110.1 ms; lock duty cycle to 100%; capillary voltage 1,600 V; dry gas 3 l min−1; dry temperature 180 °C. The PASEF settings were: 10 tandem mass spectrometry (MS) scans (total cycle time, 1.27 s); charge range 0–5; active exclusion for 0.4 min; scheduling target intensity 10,000; intensity threshold 2,500; collision-induced dissociation energy 42 eV.Raw MS data were processed with the MaxQuant software v.1.6.17.0using the built-in label-free quantitation algorithm and Androm- eda search engine (PMID: 27809316). The search was done against the Homo sapiens proteome containing forward and reverse se- quences plus the exogenous HA-tagged baits. Additionally, the intensity-based absolute quantification (iBAQ) algorithm and match between runs option were used. In MaxQuant, carbamidometh- ylation was set as fixed and methionine oxidation and N-acety- lation as variable modifications. Initial search peptide toler-ance was set at 20 p.p.m. and the main search was set at 10p.p.m.. Experiment type was set as TIMS-DDA with no modification to the default settings. Search results were filtered with a false discovery rate of 0.01 for peptide and protein identifi- cation. Results were plotted as heat map using Perseus (PMID: 27348712)and are shown in Figure 1, which shows the relative intensity ofhuman UBA5 identified across the viral or control baits. Eachcolumn represents an individual biological replicate. (N.I.=Notidentified; n=3-4)Thus, it was found that UBA5 interacts with NS4B proteins frommultiple flaviviruses (Zika virus, Dengue virus, Powassan virus, Usutu virus, Yellow fever virus).b) Zika virus replication is inhibited upon silencing of UBA5 UBA5 was found to be a Zika virus host-dependency factor. Si- lencing UBA5 was shown to inhibit Zika virus replication. JEG-3 human placental cells stably expressing control non-target-ing (NT) shRNA (short-hairpins RNA) or shRNAs against UBA5 wereanalyzed for efficiency of protein expression knock-down by west- ern-blotting using anti-UBA5 or anti-GADPH-specific antibodies. The results are shown in Figure 2A and confirm strongly reduced production of UBA5, i.e. effective silencing of UBA5.The effect of silencing of UBA5 on Zika virus replication wasstudied (see Figure 2B). Wild-type or stable knock-down JEG-3cells depleted of UBA5 were seeded in 24-well plates (8x104cells / well). The day after cell monolayers were infected with ZIKV(PRVABC59 strain) at an MOI (Multiplicity of Infection) of 0.01and 48h later virus-containing supernatants were analyzed by plaque-forming unit assay (PFU) on VeroE6 cells. (n=4; Mann-Whit-ney test p-value < 0.05). Thus, silencing of UBA5 resulted in astrong reduction of Zika virus replication. Specificity of inhibition of Zika virus replication as a resultof silencing UBA5 was further confirmed in a “rescue” experiment,in which stable shUBA5 JEG-3 cells from were reconstituted with shRNA-resistant exogenous Flag-tagged UBA5 and infected with ZIKV (PRVABC59 strain) as described above. Stable JEG-3 cells silencedwith non-targeting shRNAs (NT) or ATP6V0C- and UBA5-specificshRNAs were used as negative and positive controls, respectively. Reconstitution with Flag-tagged UBA5 was found to result in UBA5 protein expression (see Figure 2C) and high Zika virus replication (see Figure 2D).c) UBA5 inhibitor DKM29-3 effectively inhibits Zika virus rep-licationThe effects of a pharmacological modulation of UFMylation, in particular by UBA5 inhibition, on Zika virus replication were studied.In a first experiment, the cytotoxicity of the UBA5 inhibitorDKM29-3 was determined. Naïve JEG-3 human placental cells wereseeded in 24-well plates (8x104 cells / well) and on the next daytreated with increasing amounts of the UBA5 inhibitor DKM29-3. Forty-height hours later, cell viability was assessed by Resazurin assay using a multi-well plate reader to determine the 50% cyto-toxicity concentration (CC50=80.23 ^M) (see Figure 3A).Then, the effect of UBA5-inhibitor DKM29-3 on Zika virus repli-cation was studied. Wild-type JEG-3 cells were seeded as describedabove and infected with ZIKV (PRVABC59 strain) at an MOI (Multi-plicity of Infection) of 0.01 for 1h at 37°C. After virus absorp- tion the cells monolayers were washed with PBS (Phosphate BufferSaline), and cell culture media containing 16 ^M or 32 ^M ofDKM29-3 were added to the cells. Forty-height hours later virus-containing supernatants were analyzed by plaque-forming unit assay (PFU) on VeroE6 cells (n=4; Mann-Whitney test p-value < 0.05). Zika virus replication in cells treated with DKM29-3 was found to be significantly reduced in presence of DKM29-3.d) Impairment of UFMylation inhibits arbovirus replicationUBA5 inhibitor DKM2-93 was also found to be effective in inhib-iting the replication of other arboviruses from the group con-sisting of flaviviruses, alphaviruses and bunyaviruses.Wildtype JEG-3 cells were plated in 24-well plates at a seeding density of 8x104cells / well. The next day the cells were infectedfor 1 h at 37 °C with alphaviruses (Chikungunya Virus, CHIKV LS3strain, at a Multiplicity of Infection (MOI) of 0.0005; or MayaroVirus, MAYV TC625 strain, at an MOI of 0.005) or flaviviruses(Zika Virus, ZIKV PRVABC59 strain, at an MOI of 0.01; DengueVirus-2, DENV-2 16681 strain, at an MOI of 1; or Japanese Enceph-alitis Virus, JEV SA-14, at an MOI of 0.1) or bunyaviruses (RiftValley Fever Virus, RVFV Ar20364 strain, at an MOI of 0.01). Aftervirus absorption, the wells were washed once with Phosphate BufferSaline (PBS), and cell culture media containing 32 µM of DKM2-93 (or control DMSO) was added to the cells. The virus containing supernatants were collected 48 h later and analyzed by plaque assay on VeroE6 cells. Arbovirus replication in cells treated with UBA5 inhibitor DKM29- 3 was found to be significantly reduced as compared to cells treated with DMSO as a control (see Figure 4A). An impairment of arbovirus replication was also observed in UBA5 knock-out cells, which provides further evidence that UBA5 is ahost-dependency factor that arboviruses have in common.UBA5 Knock-out JEG-3 cells (gUBA5) and control cells (gNT) were plated in 24-well plates at a seeding density of 8x104cells / well.The next day the cells were infected as described hereinabove forthe experiments using UBA5 inhibitor DKM2-93 for impairment of UFMylation. After virus adsorption, the wells were washed once with PBS, and cell culture media was added to the cells. The virus containing supernatants were collected 48 h later and analyzed by plaque assay on VeroE6 cells. PFU / mL = Plaque Forming Units / milli Litre. (n=3; Welch’s t-test p-value < 0.05). The resultsare shown in Figure 4B.

Claims

Claims1. Compound which is effective in inhibiting the function ofubiquitin-like modifier activating enzyme 5 (UBA5) for usein a method of treating or preventing a disease or condi-tion mediated by an arbovirus infection in a subject.

2. Compound for use according to claim 1, wherein the arbo-virus infection is a flavivirus, alphavirus or bunyavirusinfection.

3. Compound for use according to claim 2, wherein the arbo-virus infection is a flavivirus infection.

4. Compound for use according to any one of claims 1 to 3,wherein the subject is a mammal, preferably a human.

5. Compound for use according to any one of claims 1 to 4,wherein the compound is a small molecule.

6. Compound for use according to any one of claims 1 to 5,wherein the compound has a molecular weight of 2,500 Da orless.

7. Compound for use according to any one of claims 1 to 6,wherein the compound is to be administered in an amount of from 0.1 µg / kg body weight to 50,000 µg / kg body weight.

8. Compound for use according to any one of claims 1 to 7,wherein the arbovirus infection is an infection caused bya flavivirus selected from the group consisting of Zikavirus, Japanese encephalitis virus, Powassan virus, West Nile virus, Yellow fever virus, Dengue virus, Usutu virus and Tick-borne encephalitis virus, or by an alphavirusselected from Chikungunya virus and Mayaro virus or by anbunyavirus that is a Rift valley fever virus.

9. Compound for use according to claim 8, wherein the fla-vivirus infection is a Zika virus infection.

10. Compound for use according to any one of claims 1 to 9,wherein the compound is selected from the group consistingof DKM2-93 (CAS No. 65836-72-8, 2-chloro-N-(3,4-dimethox- ybenzyl)acetamide), usenamine A ((2E,9bR)-6-acetyl-2-(1-aminoethylidene)-7,9-dihydroxy-8,9b-dimethyldibenzofuran- 1,3-dione), and compounds with the structure of Formula (I)is a polyazamacrocycle chelating group, Mis a chelatable metal ion,Y is -C(=O)- or -CH2-,L is (i) (C1-C20)-alkylene, wherein(i.a) at least one of the carbon atoms is op-tionally replaced with a heteroatom selected from O, NR' and S, wherein R' is H, (C1-C6)-alkyl or -C(=O)-(C1-C6)-alkyl,(i.b) two or three adjacent carbon atoms arejoined together to form a (C3-C10)-cyclo- alkyl group or -(C6-C10)-aryl group,and / or (i.c) the (C1-C20)-alkylene group is optionallysubstituted with at least one halo, (ii) (C2-C20)-alkenylene, wherein(ii.a) at least one of the carbon atoms is op-tionally replaced with a heteroatom se- lected from O, NR' and S, wherein R' is H, (C1-C6)-alkyl or -C(=O)-(C1-C6)-alkyl,(ii.b) two or three adjacent carbon atoms areoptionally joined together to form a (C3- C10)-cycloalkyl group or -(C6-C10)-arylgroup, and / or (ii.c) the (C2-C20)-alkenylene group is option-ally substituted with at least one halo, or (iii) a polyethylene glycol (PEG) moiety,W is (i) -NH-C(=O)-,(ii) -NR'-, wherein R' is H, (C1-C6)-alkylor -C(=O)-(C1-C6)-alkyl,X is (i) -O-,(ii) NR', wherein R' is H, (C1-C6)-alkylor -C(=O)-(C1-C6)-alkyl,(iii) -S-; or -S(=O)2-, or(iv) -C(R")2, wherein each R" is inde-pendently or simultaneously H, halo or (C1-C6)-alkyl, Raand Ra'are each independently or simultaneously (i) H,(ii) OH,(iii) halo, or(iv) (C1-C3)-alkyl, andG is (i) O,(ii) S,(iii) NR2,R1and R2are each independently or simultaneously (i) H,(ii) (C1-C6)-alkyl,(iii) (C3-C10)-cycloalkyl,(iv) (C3-C10)-heterocycloalkyl,(v) -(CH2)n-(C6-C10)-aryl, wherein n is 1 to6, (vi) -(CH2)n-(C5-C10)-heteroaryl, wherein n is1 to 6, or R1and R2are joined together to form a (vii) guanine or a guanine derivative,(viii) cytosine or a cytosine derivative,(ix) thymine or a thymine derivative,(x) adenine or an adenine derivative,and Q is any suitable counteranion, or a solvate, prodrug and / or stereoisomer thereof.

11. Compound for use according to claim 10, wherein the compoundhas the structure of formula (II)(II), wherein n is an integer in the range of 1 to 5.

12. Compound for use according to claim 10, wherein the com-pound is DKM2-93 (CAS No. 65836-72-8, 2-chloro-N-(3,4-di- methoxybenzyl)acetamide).

13. Compound for use according to any one of claims 1 to 12,wherein ubiquitin-like modifier activating enzyme 5 (UBA5) comprises an amino acid sequence of any one of SEQ IDNos:2-3, or an amino acid sequence having at least 80%,preferably at 90%, sequence identity to at least one ofthe sequences of SEQ ID NOs:2-3.

14. Pharmaceutical composition comprising the compound accord-ing to any one of claims 1, 5, 6 and 10 to 12, or itspharmaceutically acceptable salt, solvate, prodrug or es- ter, for use in a method of treating or preventing a dis-ease or condition mediated by a flavivirus infection in asubject.

15. Pharmaceutical composition according to claim 14 whichcomprises one or more excipients, carriers and diluents.

16. In vitro method for identifying a pharmaceutically activecompound for treating or preventing a disease or conditionmediated by an arbovirus infection, preferably by a fla-vivirus, alphavirus or bunyavirus infection, in a subject,comprising (a) contacting a candidate compound with a ubiquitin-likemodifier activating enzyme 5 (UBA5),(b) detecting whether the candidate compound inhibits thefunction of UBA5,wherein a compound which inhibits the function of UBA5, issuitable for treating or preventing a disease or condition mediated by an arbovirus infection, preferably by a fla-vivirus, alphavirus or bunyavirus infection, in a subject.

17. In vitro method for identifying a pharmaceutically activecompound for treating or preventing a disease or condition mediated by an arbovirus infection, preferably by a fla-vivirus, alphavirus or bunyavirus infection, in a subject,comprising (a) contacting a candidate compound with a cell that ex-presses a functional ubiquitin-like modifier activat-ing enzyme 5 (UBA5), (b) infecting the cell with an arbovirus, preferably witha flavivirus, alphavirus or bunyavirus, before or af-ter the cell was contacted with the candidate compound, (c) detecting whether the candidate compound decreases thereplication of the arbovirus, preferably of the fla-vivirus, alphavirus or bunyavirus, in the cell, com-pared to a cell which was infected with the arbovirus,preferably with the flavivirus, alphavirus or bunya-virus, but not contacted with the candidate compound,wherein a compound which decreases the replication of anarbovirus, preferably of an flavivirus, alphavirus or bun-yavirus, in the cell, compared to a cell which was infectedwith the arbovirus, preferably with the flavivirus, alpha-virus or bunyavirus, but not contacted with the candidatecompound, is suitable for treating or preventing a diseaseor condition mediated by an arbovirus infection, prefera-bly by an flavivirus, alphavirus or bunyavirus infection,in a subject.