key: cord-0863994-rmxin1da
authors: De Clercq, Erik
title: New Nucleoside Analogues for the Treatment of Hemorrhagic Fever Virus Infections
date: 2019-08-07
journal: Chem Asian J
DOI: 10.1002/asia.201900841
sha: 44a7dd668cca7fb96ba716a660ea19551406b57f
doc_id: 863994
cord_uid: rmxin1da

Eight different compounds, all nucleoside analogues, could presently be considered as potential drug candidates for the treatment of Ebola virus (EBOV) and/or other hemorrhagic fever virus (HFV) infections. They can be considered as either (i) adenine analogues (3‐deazaneplanocin A, galidesivir, GS‐6620 and remdesivir) or (ii) guanine analogues containing the carboxamide entity (ribavirin, EICAR, pyrazofurin and favipiravir). All eight owe their mechanism of action to hydrogen bonded base pairing with either (i) uracil or (ii) cytosine. Four out of the eight compounds (galidesivir, GS‐6620, remdesivir and pyrazofurin) are C‐nucleosides, and two of them (GS‐6620, remdesivir) also contain a phosphoramidate part. The C‐nucleoside and phosphoramidate (and for the adenine analogues the 1′‐cyano group as well) may be considered as essential attributes for their antiviral activity.

Abstract: Eight differentc ompounds, all nucleoside analogues, could presently be considered as potential drug candidates for the treatment of Ebola virus (EBOV) and/oro ther hemorrhagic fever virus (HFV) infections.T hey can be considered as either (i)adenine analogues (3-deazaneplanocin A, galidesivir,G S-6620 andr emdesivir) or (ii)guanine analogues containing the carboxamide entity (ribavirin, EICAR, pyrazofurin and favipiravir). All eight owe their mechanism of action to hydrogen bondedb ase pairing with either (i)uracil or (ii)cytosine. Four out of the eight compounds (galidesivir, GS-6620, remdesivir and pyrazofurin) are C-nucleosides,a nd two of them (GS-6620, remdesivir) also contain ap hosphoramidate part. The C-nucleoside and phosphoramidate (and for the adenine analogues the 1'-cyano group as well) may be considered as essential attributes for their antiviral activity.

The hemorrhagic fever virus (HFV) infections, including Lassa and Ebola, that we highlighted as potential targets for antiviral agents in 1993 [1] have essentially remained unchanged,a nd now,2 7years later,w eare still awaiting the first antiviral drug(s)t ob ef ormally approved for the treatment of HFV infections.T he first antiviral compound ever to be reported to be effective in the treatment of any HFV infection, that is, Lassa, that could be used at any point in the illness as well as for postexposure prophylaxis, was ribavirin. [2] Ribavirin (Virazole, 1-b-d-ribofuranosyl-1,2,4-triazole-3-carboxamide) ( Figure 1 ) was the first synthetic nucleoside analogue ever reported (in 1972) to be active against ab road spectrumo fb oth RNA and DNA viruses. [3] In their final remarks, Sidwell et al. (1972) stated, rightfully that "the antiviral spectrum of Virazole was the broadest ever reported for any syntheticm aterial that does not induce interferon", although the eventual clinicalu se of Virazole would eventually be limited to RNA virus infections. [4] One of the structural analogues mentioned in the latter article wasE ICAR (5- 

EICAR had originally been reported as an antileukemic agent in mice. [5] In its antiviral activity,i ts howed as imilar spectrum as ribavirin, being active against pox-, toga-, arena-, reo-, orthomyxo-and paramyxovirus infections, with ap otency that was 10-to 100-fold greater than that of ribavirin. [6] As originally ascertained for ribavirin, [7] EICAR also provedt ob eapotent inhibitor of IMP dehydrogenase, thus blocking the biosynthesis of GMP. [8] The depletion of the intracellular GTP and dGTP pools, resulting from the decreased biosynthesis of GMP,m ay also explain variouso ther effects of EICAR, such as its potentiating effect on the anti-HIV activity of ddI (2',3'-dideoxyinosine) [9] and its inhibitory effects on the replication of av ariety of viruses such as the double-stranded RNA virus, IPNV (infectious pancreatic necrosis virus). [10, 11] In the latter publication, yet another antiviralc ompound, pyrazofurin, was mentioned to be as pecific inhibitor or IPNV. [11] Pyrazofurin [3-(b-d-ribofuranosyl)-4-hydroxypyrazole-5-carboxamide] (Figure 1 ) is an inhibitor of OMP decarboxylase, a key enzyme in the de novo pyrimidine mononucleotide biosynthesis:i th as provent ob ea ctivea gainst both (+ +)RNA viruses [picorna (polio, CoxsackieB 4), toga (sindbis) and flavi (yellow fever)] and (À)RNA viruses [paramyxo( measles, RSV), orthomyxo (influenza), rhabdo( VSV) and arena (Junin, Tacaribe)]. [4, 12] Pyrazofurinh as been found to be an exquisitely potent inhibitor of VSV (vesicular stomatitis virus), which belongs to af amily (rhabdoviridae), closely relatedt ot he family of the filoviridae to which Ebola virus (EBOV) and Marburg virus belong. VSV can be handled in conventionals afety conditions, whereas EBOV and Marburgv irus require biosafety level 4; VSV could therefore be recommended as ap aradigm for predicting antiviral activity against EBOV. [13] Pyrazofurinh as so far not been evaluated for its potentiala ctivity against EBOV;i t should be done so, as has already been done with the neplanocin Aa nalogues. These analogues are knownt ob et argeted at the S-adenosylhomocysteine hydrolase. [14] The prototype of this class of compounds is 3-deazaneplanocinA( Figure 1 ). Two decades ago, 3-deazaneplanocin Aw as shownt ob ee ffective against al ethal EBOV infection in mice. [15, 16] The compound induced massively increased interferon-a production in EBOV-infected mice, [16] as tartling observation that was never followed up.

When reviewing in 2015 possible therapeutics trategiest o block EBOV infections, Im entioned 3-deazaneplanocin A, besides BCX4430 andT -705 (favipiravir). [17] Later added to the list were GS-5734 (remdesivir) and ZMapp. [18] The response to ZMapp, am ixture of three monoclonal antibodies directed against the surfaceg lycoprotein of EBOV,w as beneficial but did not meet the prespecified statistical threshold for efficacy [death occurred in 13 of 35 patients (37 %) who received the current standardo fc are alone as compared to 8o f3 6p atients (22 %) who received the current standard of care plus ZMapp]. [19] Apparently the outbreak of Ebola in 2014-2016 ended before any incontrovertible evidenceo fa ny intervention could be assessed.

In this review Iw ill focus on the potentialo ff avipiravir (T-705), BCX4430 (Galidesivir) and GS-5734 (Remdesivir) on the treatment of EBOV and other HFV infections.

The imino-C-nucleoside BCX4430 (Galidesivir)w as first reported by Warrene tal. [20] to be activea gainst aw ide range of viruses, including filo-, toga-, bunya-, arena-, paramyxo, corona-, flavi-, orthomyxo-andp icornaviruses. It was found to completely protectc ynomolgus macaques from Marburg virus infection when administered as late as 48 hours after infection. [20] It effectively blocked yellow fever virus infection in ah amster model. [21] It is now under development for the treatmento f EBOV infection. [22] BCX4430 (Galidesivir) (Figure 1 ) has also been found effective against Zika virus in cell culture and in a lethal mousem odel. [23] Antivirala ctivity of BCX4430 has also been reported against West Nile virus, at ypical mosquitotransmitted flavivirus, and against tick-bornef laviviruses, Kyasanur Forest disease virus (KFDV). [24] It inhibits infection of Rift Valley fever virus (RVFV), am osquito-borne pathogen that causes severe disease in humansa nd livestock in sub-Saharan Africa and the Arabian Peninsula, in Syrian golden hamsters. [25] The phosphoramidatep rodrug of the pyrrolo[2,1-f][triazin-4amino]a denine C-nucleoside GS-5734 (Remdesivir) ( Figure 1 ) was first reported to protect 100 %o fE BOV-infected rhesus monkeys against al ethal EBOV infection. [26] It wasa lso found active against other emerging viruses such as respiratory syncytial virus (RSV) and hepatitis Cv irus (HCV),a nd the presence of the 1'-cyano group in remdesivir wasf ound to be critical in providing selectivity toward the viral (RNA) polymerases. [27] GS-5734 was then found to be highly inhibitory to various viruses other than filo, that is, pneumo-, paramyxo-and coronaviruses. [28] That GS-5734 inhibited both epidemic and zoonotic coronaviruses, and might prove effective against emerging coronaviruses in the future was emphasized by Sheahane tal. [29] The viral RNA polymerase and the proofreading exoribonuclease were suggested as being responsible fort he coronavirus susceptibility to remdesivir. [30] The viral RNA polymerase has also been identified as the targete nzyme of paramyxoviruses (i.e. Nipah virus) for the antiviral activity of GS-5734. [31] For both EBOV RNA-dependentR NA polymerase (RdRp) and RSV RdRp, chain termination was delayeda nd predominantly seen at position i + 1. [32] The first newborn baby to have survived congenital EBOV infection, received remdesivir in addition to ZMapp and ab uffy coat transfusion from an Ebola survivor. [33] Favipiravir (T-705) ( Figure 1 ) is ap yrazine derivative, that is, 6-fluoro-3hydroxy-2-pyrazinecarboxamide, sharing ac ommon structural feature, that is ac arboxamide entity,w ith ribavirin, EICAR and pyrazofurin, andw hich is also an essential (hydrogen bonding) part of guanine( -HN 1 -C 6 O-). [34, 35] As already mentioned, ribavirin and EICAR would primarily owe their antiviral activity to interference with the IMP dehydrogenase, ak ey enzyme in the biosynthesis of GMP and GTP.Y et, Eriksson et al. [36] reported inhibition of influenzav irus RNA polymerase by ribavirint riphosphate,a nd when Furuta et al. [37] had revealed the in vitro and in vivo activities of T-705 against influenza virus, they attributed the mode of action of T-705 to as pecific inhibition of the influenzavirus RNA polymerase by the T-705 RTP (ribofuranosyl triphosphate. [38] To this end (Scheme 1), T-705 had first to be converted to its ribofuranosyl monophosphate( RMP) by a phosphoribosylt ransferase, and then further processed by kinase(s) to its diphosphate( RDP) and triphosphate (RTP), before the latter would interact in aG TP-competitive manner with the viral RNA synthesis.

That T-705 (Favipiravir), as av iral RNA polymerase inhibitor, offers great potential in the treatment of aw idev ariety of RNA virus infections, has been reviewed repeatedly. [39] [40] [41] [42] Sidwell et al. [43] demonstrated that T-705 inhibited al ethal avian influenza A( H5N1)v irus infectioni nm ice. This was confirmed by Kiso et al.. [44] It was ascertainedt hat in the influenzaA(H5N1) virus-infected cells T-705 was metabolized to T-705 RTP. [45] The latter is incorporated into the nascent RNA strand as ap urine nucleotide analog (reminiscent of GMP) and inhibits strand extension. [46] The RTP of T-705 acts as aG TP-competitive inhibitor of the influenza viral RNA polymerase. [47] Favipiravir inhibits in vitro replication of aw ide range of influenzav iruses, including those resistantt oc urrently available drugs [48] andt hose isolated from patients who had previously received favipiravir. [49] Favipiravir has been approved, as Avigan, in Japan for the treatment of influenza. [50] Lethal mutagenesis hasb een proposed to be ak ey mechanism in the activity of T-705 (Favipiravir) againsti nfluenzaA (H1N1)v iruses in vitro. [51] Lethalm utagenesis has also been suggested for the activity of favipiravir against norovirus repli-cation [52] and hepatitis Cv irus. [53] Such mechanism of antiviral activity was originally proposed for ribavirin. [54, 55] However,t he activity of ribavirin against yellow fever virus could not be explained by an error-prone mechanism. [56] Favipiravir has provene ffective againstv irtually all hemorrhagic fever virus infections:a rena-and bunyaviridae, [57] Pichinde virus (an arenavirus), [58, 59] Lassa fever virus, [60] yellow fever virus, [61] Western Equine encephalitis virus, [62] West Nile virus, [63] Punta To ro, ap hlebovirus, [64] the hantavirus Maporal, [65] Rift Valley Fever virus [Phlebovirus (Bunyaviridae)], [66] Crimean-Congo hemorrhagic fever virus [Nairovirus (Bunyaviridae)], [67] SevereF ever with Thrombocytopenia Syndrome virus (SFTSV), ar ecently identified emerging virus, [68] and norovirus (Caliciviridae). [69, 70] Resistance to T-705 has never been reported, except for Chikungunya virus (Alphaviridae) which acquired resistance due to the K291R mutation in the RNA-dependentR NA polymerase (RdRp). [71] Favipiravir proveds uccessful in the treatment of EBOV infection in mice. [72, 73] In nonhuman primates, treated intravenously with favipiravir,f ive of six animals (83 %) survived al ethal Marburg virus infection. [74] In patients with the EBOV infection in www.chemasianj.org Sierra Leone, favipiravir was found to increase the survival rate from 35.3 %( 30/85) to 56.4 %( 22/39). [75] Another study carried out in Guinea ended with the statement that favipiravir monotherapy merits furthers tudy in patients with mediumt oh igh viremia but not in those with very high viremia. [76] Guedj et al. [77] concluded that favipiravir mayh ave ap otentialr ole at high doses in the treatment of EBOV infections in humans.

Ta king into account that rhabdo-and filoviruses are closely related,i ti sn ot surprising that favipiravir has also been advocated for the postexposure prophylaxis of rabiesvirus, as ap otential alternative to rabies immunoglobulin. [78] The compounds described here can be considered as either adenine derivatives (3-deazaneplanocin A, galidesivir,r emdesivir or guanine derivatives (ribavirin), EICAR, pyrazofurin,f avipiravir). This is immediatelyo bvious for the adenine derivatives, which can be expected to interferew ith viral RNA synthesis at the level of the RdRp (RNA-dependentR NA polymerase), where they can serve as competitive inhibitors with respectt o ATP( based on the hydrogen bonds formed between adenine and uracil) ( Figure 2 ). In addition, the adenine analogues can also serve as S-adenosylhomocysteine (SAH) hydrolase inhibitors, thus interfering with the S-adenosylmethionine (SAM)-dependentmethylationr eactions.

That ribavirin, EICAR, pyrazofurin and favipiravir would be able to act as guanined erivatives thus competing with GTP at the RdRp level is less obvious. Yet, all 4c ompounds share a carboxamide group, reminiscent of the carboxamide (-C (6) O-N (1) H-) part of guanine, which could explain the hydrogen bondingw ith cytosine ( Figure 2 ). In addition to their action at the RdRp level, the carboxamide-containing derivatives could also interfere with the IMP dehydrogenase activity,t hus suppressing the biosynthesis of GTP,a sh as been specifically demonstrated for ribavirin and EICAR.

The exact mechanism of action of favipiravir remains to be assessed;i tm ay well vary from one virus to another.A sf ar as the activity of favipiravir RTP against influenzaAvirus polymerase is concerned, this has been ascribed to "ambiguous base-pairing". [79] In 2016, Ip roposed that "C-nucleosidess hould be revisited". [80] This proposal was prompted by the advent of two new C-nucleosides, BCX4430 (Figure 1 ), which has in the meantime, been further pursued (as galidesivir) for the treatment of EBOV infections (see supra), and GS-6620 (compound 2, containing a 2'-C-methyl group imparting specific activity against hepatitis Cv irus (GCV). [80] GS-6620 (Figure 1 ) also contained a1 '-cyano group, which later on was found to be critical in providing selectivity towardv iral (RNA) polymerases. [27] GS-6620 has only been reported for its potentiala sa na nti-HCV agent. [81] [82] [83] It has not been thoroughly explored for its potential activity against EBOV or any other hemorrhagic fever virus (HFV) infections. According to literatured ata, [27] it would only have weaka ctivity against EBOV.

Remdesivir (GS-5734) is at present aleadingd rug candidate for the treatment of EBOV infections. Its chemical attributes are that it is aC -nucleoside, extendedb yaphosphoramidate and equippedw ith aC Ng roup. Galidesivir (BCX4430) is also aCnucleoside, but what would happeni fi tw ould also contain a CN group and would be converted to ap hosphoramidate prodrug?F or favipiravir,i ts N-nucleoside T-1106 is more effective than the free 2-pyrazinecarboxamide against yellow fever virus infection in hamsters, [61] but less so against Punta To ro virus in mice. [64] What would happen if T-1106 would be convertedt o its C-nucleoside, and, furthermore, extended by ap hosphora- www.chemasianj.org 2019 Wiley-VCH Verlag GmbH &Co. KGaA, Weinheim midate entity?A lso, the forgotten C-nucleoside, pyrazofurin, [80] could be converted to its phosphoramidate, and EICAR, which containsa ne thynyl, reminiscent of the cyano group, could be first transformed to its C-nucleoside before being converted to its phosphoramidate (Scheme 2). Any of these chemical interventions may provide potential clues in the design of new medicines against EBOVand other HFV infections.

The (candidate) antiviral compoundsc urrently availablef or the treatment of EBOV and other HFV infections are 3-deazaneplanocin A, galidesivir,G S-6620, remdesivir,r ibavirin, EICAR, pyrazofurin andf avipiravir.They are boxed in (Scheme2).

In attempts to increase their efficacy and potentially lower their toxicity, they could, where applicable, first be converted to their C-nucleoside, and, subsequently,t ot heir phosphoramidate (Scheme 2). For GS-6620 and Remdesivir,w hich,i na ddition, also contain ac yanogroup, these manipulations have already been achieved.

The authordeclares no conflict of interest.

Keywords: antivirals · ebola · hemorrhagic fever viruses · nucleoside analogues

Proc. Natl. Acad.Sci.U SA 1973

Antiviral Res

CurrentC hemotherapy:P roceedings of the 10th International Congress of Chemotherapy

Proc. Jpn. Acad. Ser.B2017

Proc. Natl. Acad.S ci

Proc. Natl. Acad. Sci

Revised manuscript received