Erik DE CLERCQRega Institute for Medical Research, K.U.Leuven

B-3000 Leuven, Belgium

ANTI-HERPESVIRUS THERAPIES:BASIC STRATEGIES AND APPLICATIONS

HerpesviridaeHuman herpesviruses (HHV)

HHV-1: Herpes simplex virus type 1 (HSV-1)HHV-2: Herpes simplex virus type 2 (HSV-2)HHV-3: Varicella-zoster virus (VZV)HHV-4: Epstein-Barr virus (EBV)HHV-5: Human cytomegalovirus (HCMV)HHV-6: Human herpes simplex virus type 6 (HSV-6)HHV-7: Human herpes simplex virus type 7 (HSV-7)HHV-8: Kaposi’s sarcoma herpesvirus (KSHV)

OHO

O

H2N N

N

N

HN

Acyclovir, Aciclovir (ACV), Acycloguanosine9-[(2-Hydroxyethoxy)methyl]guanine

Zovirax®

HN

N

N

N

O

H2N

OHO

HN

N

N

N

O

H2N

OOCOCHH2N

CHH3C CH3

HN

N

N

N

O

H2N

OOCOCH2H2N

HN

N

N

N

O

H2N

OOCOCH

CH3

H2N

Acyclovir Acyclovir valyl esterValaciclovir

Acyclovir glycyl ester Acyclovir alanyl ester

ValaciclovirValine ester of acyclovir

Valtrex®, Zelitrex®

O

O

H2N N

N

N

HN

OC

OHCH2N

CHH3C CH3

Antiviral activity spectrum of valaciclovir(acyclovir)

Herpesviridae! Herpes simplex virus type 1 (HSV-1)! Herpes simplex virus type 2 (HSV-2)! Varicella-zoster virus (VZV)! Epstein-Barr virus (EBV)" Human cytomegalovirus (HCMV)" Human herpesvirus type 6 (HHV-6)" Human herpesvirus type 7 (HHV-7)" Human herpesvirus type 8 (HHV-8)" Thymidine kinase-deficient HSV (TK- HSV)" Thymidine kinase-deficient VZV (TK- VZV)

Major clinical indications of valaciclovir

Systemic (oral) treatment of HSV and VZV infections:

• primary and recurrent genital herpes, mucocutaneousHSV-1 and HSV-2 infections in immuno-compromised and –competent patients

• varicella-zoster in immuno-compromised and –competent patients

OH

N

N

N

HN

H2N

HO

O

N

N

N

N

H2N

H3C O

O

OH3C

O

Famciclovir Penciclovir Famvir®

OH

HO

N

N

N

NH2N

OH

HO

HN

N

N

NH2N

O

Xanthineoxidase

Xanthineoxidase

Xanthineoxidase

Esterase

EsteraseEsterase

Esterase

OH

OH3C

O

HN

N

N

NH2N

O

OH

OH3C

O

N

N

N

NH2N

O

O

H3C

O

H3C

O

N

N

N

NH2N

O

O

H3C

O

H3C

O

HN

N

N

NH2N

O

Intravenous penciclovir versus acyclovir in the treatment of HSV infections in immunocompromised patients

Lazarus et al., Antimicrob. Agents Chemother. 43: 1192-1197 (1999)

Kaplan-Meier plot of time to healing of lesions for penciclovir at 5 mg/kg q12 h(long-dashed line), penciclovir at 5 mg/kg q8h (short-dashed line), and acyclovir at 5 mg/kg q8h (solid line)

O

OH

N

N

N

HN

H2N

HO

O

Ganciclovir (GCV)9-(1,3-dihydroxy-2-propoxymethyl)guanine (DHPG)

Cymevene®, Cytovene®

ValganciclovirValcyte®

O

OH

N

N

N

HN

H2 N

O

O

C

OHCH2N

CHH3C CH3

Nucleoside kinase

3 Na+ P-O

O

O-

CO

O-

FoscarnetPhosphonoformate (trisodium salt)

PFAFoscavir®

N

NH

N

N

O

NH2

OHO O

OH

HO

OH

HO

HO

OH

HO O

OH

HO HO

HO

HO

OHSynguanol A-5021 D/L-Cyclohexenyl guanine

Penciclovir Lobucavir Anhydrohexitol guanine

Acyclovir Ganciclovir H2GHO

OH

Lobucavir

HO

OH

HN

N

N

N

O

H2N

HN

N

N

N

O

H2N

OH

HO

(-) 2HM-HBGH2G

(-)-9-[4-hydroxy-2-(hydroxymethyl)butyl]guanine

N

N

N

N

NH2

HO N

NH

N

N

O

NH2HO

Synadenol Syngenol

Methylene cyclopropane nucleoside analogues

D/L-Cyclohexenyl guanine

HO

OH

N

NH

N

N

O

NH2

A-5021

N

NH

N

N

O

NH2

HO

HO

N

NH

N

N

O

NH2

OH

HO

N

N

N

N

NH2

OH

HO

AV-038 A-5021

Comparative potency of different antiherpetic compoundsagainst different herpesviruses

Compound 50% Effective concentrationEC50 (µM)

HSV-1 HSV-2 VZV HCMVAcyclovir +++ +++ ++ +Penciclovir +++ ++ ++ (+)Ganciclovir +++++ +++++ ++ ++Lobucavir ++++ ++++ +++ ++H2G ++++ ++++ ++++ (+)A-5021 ++++ +++ +++ +Synguanol + - + ++Anhydrohexitol G ++(+) +++(+) ++ ++D-Cyclohexenyl G ++++ +++ ++ ++L-Cyclohexenyl G +++(+) +++ ++ ++EC50 (µM)= 10-100 (+), 1-10 (++), 0.1-1 (+++), 0.01-0.1 (++++), 0.001-0.01 (+++++)

N

N N

N

OHO

OH

H2N

S22422-Amino-7-[(1,3-dihydroxy-2-propoxy)methyl]purine

Mycophenolic acid

OC

O

OH

CH3

OCH3

O CH3

OH

Mycophenylate mofetil (Cellcept®)2-Morpholinoethyl ester of mycophenolic acid

OC

O

OH

CH3

OCH3

O CH3

ON

O

MPA-IMPDH interactions

Sintchak et al., Cell 85: 921-930 (1996)

XMP

GMP

GDP

GTP

RNAsynthesis

Deoxyguanosine

dGMP

dGDP

DNA synthesis

dGTP

Guanosine analogues

dGMP analogues

dGDP analogues

Phosphoribosylamine(PRA)

Mycophenolic acid

dGTP analogues

H2O + NAD+

NADH + H+

IMP dehydrogenase

IMP

Virus Control 2.5 µg/ml MMF

1 µg/ml LBV 1 µg/ml LBV + 2.5 µg/ml MMF

Inhibitory effects of mycophenolate mofetil (MMF) and lobucavir (LBV)on cytopathicity of HCMV in human embryonic lung (HEL) cells

Virus Control MMF

ACV MMF + ACV

Inhibitory effects of topical mycophenolate mofetil (MMF) 5% and topical acyclovir (ACV) 0.1% on HSV-1-induced lesions in hairless mice

HN

N

O

O

I

OHO

OH

Idoxuridine5-iodo-2’-deoxyuridine (IDU)

Stoxil®

OHO

OH

HN

N

O

O

CF3

Trifluridine, Trifluorothymidine (TFT)5-Trifluoromethyl-2’-deoxyuridine

Viroptic®, TFT-Ophthiol®

O

HN

N

Br

O

O

HO

HO

Brivudin, Bromovinyldeoxyuridine(E)-5-(2-bromovinyl)-2’-deoxyuridine (BVDU)

Zostex®, Zonavir®

De Clercq, Recent Advances in Nucleosides: Chemistry and Chemotherapy, pp 433-454 (2002)

Antiviral activity spectrum of BVDU

O

HN

N

Br

O

O

HO

HO

HOO

HN

N

Br

O

O

HO

HO

OH

HN

N

Br

O

O

HO

HO

BVaraU BVriboU C-BVDU

De Clercq, Recent Advances in Nucleosides: Chemistry and Chemotherapy, pp 433-454 (2002)

S

HN

N

Br

O

O

HO

HO O

N

N

Br

NH2

O

HO

HO

O

HOOH

HN

N

Br

O

O

S-BVDU BVDC L-BVDU

De Clercq, Recent Advances in Nucleosides: Chemistry and Chemotherapy, pp 433-454 (2002)

S-BVMU Triazolyl BVDC L-BVODDU derivative

S

HN

N

Br

O

O

HO

HO

O

O

HO

HN

N

Br

O

O

O

N

N

Br

O

HO

HO

N

N

N

De Clercq, Recent Advances in Nucleosides: Chemistry and Chemotherapy, pp 433-454 (2002)

BMS-181165 4’-methyl BVDU AV-100

OHO

OH

HN

N

Br

O

O

O

HN

N

Br

O

HO

HO

O

H3C

HN

N

Br

O

O

OH

HO

De Clercq, Recent Advances in Nucleosides: Chemistry and Chemotherapy, pp 433-454 (2002)

Mechanism of action of BVDU

HSV-1 dThd kinase DNA polymerase

BVDU BVDUMP BVDUDP BVDUTP DNA

BVDU BVDUMP

HSV-2 dThd kinase

O

HN

N

C

O

HO

HO

O

CBr

H

H

HN

NH

C

O

O

CBr

H

H

O

HO

HO O

+ Pi BVU +Thymidine

phosphorylase

BVDU

HN

NH

O

F

O

HN

NH

O

F

ODihydrothymine dehydrogenase

BVU

5-Fluorouracil 5-Fluorodihydrouracil

P

Major clinical indications of BVDU(Brivudin)

Topical: - herpetic keratitis (BVDU eyedrops)- herpes labialis (BVDU cream)

Systemic: - mucocutaneous HSV-1 and VZV infections(oral) - immunocompromised patients - immunocompetent patients

Oral brivudin versus intravenous acyclovir in thetreatment of severe herpes zoster in cancer patients

BRIVUDIN: BVDU: (E)-5-(2-bromovinyl)-2’-deoxyuridineorally 125 mg x 4 per day for 5 days

ACYCLOVIR: ACV: 9-(2-hydroxyethoxymethyl)guanineintravenously 10 mg/kg x 3 per day for 5 days

Multicentered, double-blind, randomizedBVDU group (24 patients) also received placebo i.v.ACV group (23 patients) also received placebo p.o.

Wutzler et al., J. Med. Virol. 46: 252-257 (1995)

Double-blind study BVDU versus ACV

Number of days following start of therapy

% P

atie

nts

with

new

ves

icle

s100

80

60

40

20

01 2 3 4 5 6 7

---- ACV (n = 23) — BVDU (n = 24)

Current antiviral therapy for zoster

7 days1 x daily125 mgBrivudin oral

7 days3 x daily250 mgFamciclovir oral

7-10 days3 x daily5-10 mg/kgAcyclovir intravenous

7 days5 x daily800 mgAcyclovir oral

7 days3 x daily1000 mgValaciclovir oral

Gross et al., J. Clin. Virol., in press (2003)

Brivudin compared to acyclovirIncidence of postherpetic pain

1.61 (1.15 – 2.25)Odds ratio (95% Cl)

0.006p value for difference

130 (43.5)101 (32.7)Patients with postherpetic pain n(%)

Acyclovir(n = 299)

Brivudin(n = 309)

Survey patients (n = 608)

A randomized, double-blind post-study survey on the effect of brivudinin the prevention of postherpetic pain in comparison with acyclovir

Prof. Dr. S.W. Wassilew

Brivudin compared to famciclovir

0.0102p value for non-inferiority

1.23 (0.92 – 1.65)Odds ratio (95% Cl)

90 (9.2)109 (11.1)Patients with postherpetic pain n(%)

Famciclovir(n = 974)

Brivudin(n = 980)

ITT (n = 1954)

Prevalence of postherpetic pain

Brivudin compared to famciclovir in the prevention of postherpeticneuralgia: a prospective randomized, double-blind multicenter trial

Prof. Dr. S.W. Wassilew

Brivudin compared to famciclovir

0.049p value for non-inferiority1.05 (0.76 – 1.45)Risk ratio (95% Cl)

72.9 ± 63.754.0

69.3 ± 62.847.0

Duration of postherpetic pain Mean ± SD Median

Famciclovir(n = 90)

Brivudin(n = 109)

ITT (n = 199)

Duration of postherpetic pain

Brivudin compared to famciclovir in the prevention of postherpeticneuralgia: a prospective randomized, double-blind multicenter trial

Prof. Dr. S.W. Wassilew

Prof. Dr. S.W. Wassilew

Brivudin compared to famciclovirDuration of ZAP (Zoster-Associated Pain), n = 1712

0

0,1

0,2

0,3

0,4

0,5

0,6

0,7

0,8

0,9

1

0 30 60 90 120 150 180 210 240 270

D uration of ZA P [days, after start of treatm ent]

B rivud in

F am ciclo vir

Prob

abilit

y

Nucleoside AnaloguesNon-nucleoside Analogues

4-Hydroxyquinoline-3-carboxamides(4-HQC)

PNU-181465

inhibit replication of HSV-1, VZV and HCMV, through inhibitory effect on HSV-1, VZV and HCMV DNA polymerase

Oien et al., Antimicrob. Agents Chemother. 46: 724-730 (2002)

N

HO

OH

NH

O

Cl

BILS 179 BS

inhibits replication of HSV (and other herpesviruses ?) through inhibitoryeffect on helicase/primase (UL5, UL8 and UL52 gene products)

Crutz et al., Nature Medicine 8: 386-391 (2002)

N

N

O

HN

O

H3C N

SNH2

N-[5-(aminosulfonyl)-4-methyl-1,3-thiazol-2-yl]-N-methyl-2-[4-(2-pyridinyl)-phenyl]acetamide inhibits replication of HSV (and other herpesviruses ?) through inhibitory effect on helicase /primase (UL5, UL8 and UL52 gene products)

BAY 57-1293

N O

N

CH3

N

SS NH2

O

O

CH3

Kleymann et al., Nature Medicine 8: 392-398 (2002)

BAY 57-1293 in the rat lethal challenge model

Lewis rats were inoculated with HSV-1walki intranasally and were treated via oral gavage t.i.d. from day 0 to day 4 post infection. Five animals for eachgroup were used.

Betz et al., Antimicrob. Agents Chemother. 46: 1766-1772 (2002)

_####_ Placebo_≤≤≤≤_ Valaciclovir 45 mg/kg_X_ Valaciclovir 135 mg/kg_ππππ_ BAY 57-1293 0.5 mg/kg_′′′′_ BAY 57-1293 2 mg/kg

BAY 57-1293 in the murine lethal challenge model

Mice were infected intranasally with HSV-2 MS and were treated via oral gavage once-daily from day 0 to day 4 post infection. Ten animals for each group were used.

Betz et al., Antimicrob. Agents Chemother. 46: 1766-1772 (2002)

12

1,4-Dihydroxynaphthalene 1,4-Naphthoquinone

inhibit HSV-1 and HCMV proteases, could be considered as scaffold for development of non-peptidic anti-herpesvirus agents

Matsumoto et al., Biol. Pharm. Bull. 24: 236-241 (2001)

OH

OH

O

O

CMV423

2-Chloro-3-pyridin-3-yl-5,6,7,8-tetrahydroindolizine-1-carboxamide

inhibits HCMV replication, acts at an early stage (coinciding with IE antigensynthesis) of the HCMV replication cycle

Snoeck et al., Antiviral Res. 55: 413-424 (2002)

N

N

Cl

ONH2

Pyrrolopyrimidines

828 951 1028

inhibit HCMV replication, target a viral protein that is expressed early in the HCMV replication cycle

Jacobson et al., Antimicrob. Agents Chemother. 43: 1888-1894 (1999)

N

N N

SNH2NH2

OH3CO

N

N N

SNH2NH2

H3CO

N

N N

CNNH2

H3C

NH2

Naphthyridine and dihydroisoquinoline derivatives

7,8-Dihydroisoquinoline-6-carboxyl-[2-(1-indol-3-yl)ethyl]amide

inhibits HCMV (and HSV) replication, affects early (post-adsorption) event(s) of HCMV replication cycle

Bedard et al., Antimicrob. Agents Chemother. 44, 929-937 (2000)

NHN

O NH

BAY 38-4766

inhibits HCMV replication, targets HCMV DNA maturation via UL89 and UL56 gene products, inhibits cleavage of high-molecular-weight DNA concatemers and packaging of monomeric genomes into procapsids

Buerger et al., J. Virol. 75: 9077-9086 (2001)

3-Hydroxy-2,2-dimethyl-N-[4({[5-dimethylamino)-1-naphthyl]sulfonyl}amino)-phenyl]propanamide

NH3C CH3

SNH

O

HN

O

O

OH

CH3

CH3

BDCRB

1-(ββββ-D-Ribofuranosyl)-2-bromo-5,6-dichlorobenzimidazole

inhibits HCMV replication, targets HCMV DNA maturation via UL89 and UL59 gene products which are responsible for cleavage of high-molecular-weight DNA concatemers and packaging of monomeric genomes into procapsids

Biron et al., Antimicrob. Agents Chemother. 46: 2365-2372 (2002)

O

HO

HO

OH

N

NBr

Cl

Cl

1263W94Maribavir

1-(ββββ-L-Ribofuranosyl)-2-isopropylamino-5,6-dichlorobenzimidazole

inhibits HCMV replication, through interaction with the UL97 gene product(which is required for egress of viral nucleocapsids from the nucleus)

Biron et al., Antimicrob. Agents Chemother. 46: 2365-2372 (2002)[Krosky et al., J. Virol. 77: 905-914 (2003)]

O

HO OH

OH

N

NNH

Cl

Cl

CH3

CH3

Indolocarbazoles

Gö 6976inhibit HCMV replication, target UL97 gene product (which is required for egress of viral nucleocapsids from the nucleus)

Zimmermann et al., Antiviral Res. 48: 49-60 (2000)[Krosky et al., J. Virol. 77: 905-914 (2003)]

N N

HNO

CN

CH3

CONCLUSIONMarketed antiviral drugs for treatment of HSV, VZV and CMVinfections:

HSV VZV CMV

AcyclovirValaciclovirFamciclovirBrivudin

GanciclovirValganciclovirFoscarnetCidofovirFomivirsen