4 th Annual Clinical Care Options for Hepatitis Symposium: HCV Highlights

4th Annual Clinical Care Options for Hepatitis Symposium: HCV Highlights

clinicaloptions.com/hep


Raymond T. Chung, MDDirector of HepatologyMassachusetts General HospitalAssociate Professor of MedicineHarvard Medical SchoolBoston, Massachusetts

Interferon and Ribavirin:Mechanisms of Action, Resistance, and Why It Matters



IRF-3 activation

Innate Immunity: An IFN Amplification Loop

Adapted from Gale M Jr, et al. Nature. 2005;436:939-945.

RIG-I

ST

AT

2

ST

AT

1

IKK-

TBK1

IRF-7

IRF-7P

IRF-7P

IRF-7P

IRF-7P

IRF-7

IRF-3

IRF-3P

IRF-3P

IRF-3P

IRF-3P

IRF-9

IRF-9

Tyk2 Jak1

IFN- IFN-

IFN- IFN-IFN-

IFN-

IFN-S

TA

T 1

P

TLR3

Hepatitis Virus

Nucleus

Cytoplasm

Viral PAMP:dsRNA

ST

AT

2

P

ISGF3

JAK-STATpathway

VRE IFN-PRD ISRE IFN-stimulated genes:

OAS, IRF-7, PKR, ISG56 etc

IFN- production

IFN-/

IFNAR-2IFNAR-1

IFN signalingJak-STAT

ISG expression;IFN amplification

loop

IFN-



The Adaptive (Cellular) Immune Response Finishes the Job

Adapted from Liang TJ, et al. Ann Intern Med. 2000;132;296-305.

HCV

Viral entry

MHC II

TCR

TCR

MHC I

B cell

CD4+ Th cell

CD8+CTL

CD8+CTL

CD4+ Th cell Clonal

expansion(Th1 or Th2)

Th2 cytokines(IL-4, IL-5, IL-6,IL-9, IL-10, IL-13)

Activation,differentiation

Th1 cytokines

Lysis

NeutralizingHCV antibodies

Clonal expansion

Hepatocyte

IFN-(IFN-γTNF-α)



Viral Kinetics After IFN Therapy

Adapted from Feld JJ, et al. Nature. 2005;436:967-972.

Viral kinetics

IFN (efficacy =

HC

V R

NA

(lo

g I

U m

L-1)

Days After Start of Therapy

Two phases ofviral decline

1st phase: antiviral efficacy () (innate)

2nd phase: clearance ofinfected hepatocytes ()

(adaptive)

0

-1

-2

-3

-4-7 0 7 14 21 28



Ribavirin

Initially developed as an antiviral–guanosine analogue

No antiviral activity but improved ALT when given as monotherapy

Combination with IFN improved ETR but greatly enhanced SVR rates by decreasing relapse

– Does not alter 1st phase kinetics appreciably

– Modest of PEG-IFN antiviral effect (0.5-1.0 log)

Mechanistic models must explain clinical observations



Ribavirin: Proposed Mechanisms of Action

Adapted from Feld JJ, et al. Nature. 2005;436:967-972.

Inhibition of HCV RdRp (1st phase)

Immunomodulation (2nd phase)

Defective HCV particles (decreased fitness)

Hepatocyte

RDP RTPRMPRibavirin(-)

IMPIMPDH

GMP

GTP

HCV RNA

HCV RNA

TH1 CTL

RNA Mutagen

RdRp

Replication

IFN-, TNF-

Inhibition of IMPDH (1st phase)

RNA mutagenesis (2nd phase)

RibavirinTH2



HCV NS3-4A Blocks IFN Induction at Multiple Levels


IRF-3P

IRF-3P

IRF-3P

IRF-3P

IRF-3P

CBP/p300

CBP/p300

IRF-3IKK-

TBK1

NS3/4A

NF-l

TRIF

TLR3

IRF-3

NF-

RIG-I MDA5

RIP-1

RIP-1

HCV NS3/4A

IKK

l P

FADD

TRAF6

IRF-3/NF-Target genes

(IFN-)

IPS-1

mitochondria



HCV Blocks IFN Signal Transduction


ST

AT

2

ST

AT

1

SOCS-3

IRF-9

IRF-9

Tyk2 Jak1

ST

AT

1

P

Nucleus

Cytoplasm

ST

AT

2

P

ISGF3

IFN-/

IFNAR-2IFNAR-1

ISRE

PP2APIAS

SOCS-1

ISG expression attenuated

Core

IFN- IFN-IFN- IFN-

IFN-IFN- Core HCV proteins

SOCSInhibition of

Jak-STATsignaling

Block STAT function

IL-8

NS5A

Inhibit P-STAT1 Degrade STAT1



IFN-Stimulated Genes as the Antiviral Workhorses

Adapted from Samuel CE. Clin Microbiol Rev. 2001;14:778-809.

Microarray studies have identified > 100 IFN-stimulated genes

Antiviral Actions of Interferon

IFN IFN

Protein kinase PKRInactive

Oligoadenylate synthetase OAS

dsRNAssRNA

Protein kinase PKRActive

(Ribosome associated)

Oligoadenylate synthetase OASActive

(multiple forms: nuclear and cytoplasmic)

InitiationfactorelF-2

Phosphorylatedinitiation factor

elF-2PPhosphatase

(Soluble)

Pi

mRNA translationinhibition

ATPAMP

2’, 5’-oligoadenylic acid

(2, 5 A)Phosphodiesterase

RNase LInactive

RNA degradation

RNase LActive

InactiveNS5A

E2

HCV

Paucity of recognition sites



Treatment of Hepatitis C inBlacks: SVR

1. Muir AJ, et al. N Engl J Med. 2004;350:2265-227. 2. Jeffers LJ, et al. Hepatology. 2004;39:1702-1708.3. Conjeevaram H, et al. AASLD 2005. Abstract 199.

26%19%

39%

52%

0

20

40

60

80

100

Muir et al[1] Jeffers et al[2]

Black

White

Vir

olo

gic

Res

po

nse

Rat

es

(%)

n = 100 n = 100 n = 78 n = 28

P < .001

PEG-IFN -2b 1.5 g/ kg/wk x 48 weeks +

RBV 1000 800 mg/d100% genotype 1

PEG-IFN -2a 180 g/wk + RBV 1000-

1200 mg/d x 48 wks 98% genotype 1

52%

28%

Virahep-C[3]

n = 196 n = 205

P < .001

PEG-IFN -2a 180 g/ wk + RBV 1000-

1200 mg/d x 48 wks 100% genotype 1



Impaired Host Antiviral Responses as the Basis for Inferior SVR Rates? Differences between A-A and C-A appear to reside

primarily in 1st phase viral decay

These findings suggest intrinsic defects in innate immunity (signal transduction or ISGs)

Search for genetic polymorphisms in innate immunity underway (Virahep-C)



Obesity and Impaired Antiviral Response Rates Obesity also associated with impaired antiviral response

rates (likely related to steatosis)

SOCS-3 as key mediator

– Upregulated in obesity

– Increased by HCV

– Promotes degradation of IRS1 and IRS2 insulin resistance

Kawaguchi T, et al. Am J Path. 2004;165:1499-1508.Walsh MJ, et al. Gut. 2006;55:529-535.



Higher-Dose IFN or Further Refinements in IFN PK Theory: overcome intrinsic blocks to IFN action with higher

doses of exogenous IFN

High dose PEG-IFN + RBV: recent studies showing modest SVR rates in prior PEG-IFN/RBV nonresponders

Albumin-IFN: extend half-life of IFN to permit extended dosing interval

Limitations: tolerability, toxicities, downstream block



Strategies to Improve or Replace RBV

IMPDH inhibitors

– Likely not major mechanism of RBV against HCV

– Antiviral effect may be offset by immunosuppressive effects

Higher RBV doses

– Further decreases in relapse among genotype 1 patients?

Prodrugs

– Permit targeted dosing of RBV (viramidine)



New Therapies for Chronic Hepatitis C: Rational Drug DesignGary Davis, MDDirector, Division of HepatologyBaylor University Medical CenterMedical Director, Liver TransplantationBaylor Regional Transplant InstituteDallas, Texas



Role of New Agents in Treating HCV

Primary aim should remain eradication

Chronic suppression may be achievable

Interferon likely to remain foundation of therapy

Combination therapy will be key

Other agents may allow lower doses or shorter duration of poorly tolerated drugs

New agents will be able to target different processes of the HCV replication cycle



Target: Infection of the Hepatocyte



Therapeutics: Infection of the Hepatocyte Polyclonal preparations[1]

– Neutralize infectious inoculae ex vivo

– Inhibit or prevent infection in chimps

– Studies in man disappointing to date

Monoclonal antibodies[2]

– Anti-E2 human monoclonal XTL

– Mild HCV RNA suppression with daily dosing

Vaccine-derived anti-E1E2[3]

– Neutralizing antibody

– In vitro inhibition of CD81 and VSV pseudovirions

1. Davis, et al. Liver Transpl 2005. Willems, et al. J Hepatol. 2002.2. Schiano, et al. Hepatol. 2005.3. DiBisceglie, et al. Hepatol. 2005.



Therapeutics: Infection of the Hepatocyte (cont’d) N-glycans in envelope glycoprotein (E1E2) are

essential for protein folding, secretion/assembly, antigenicity, receptor binding, and cell entry

Mutations in E2 eliminate infectivity[1]

– E2N2, E2N4 by blocking cell entry

MX-3256 (celgosivir, Migenix)[2]

– In vitro synergy with interferon-ribavirin

– No reduction in HCV RNA

1. Goffard, et al. J Virol. 2005.2. Yoshida, et al. Gastroenterology. 2006.



Target: RNA Transport to the ER

3’

5’



Therapeutics: RNA Transport to the ER

Oligonucleotides

– Ribozymes

– Antisense oligos

– siRNA



Therapeutics: RNA Transport to the ER (cont’d)

mRNA

Antisense oligo Ribozyme

Translationarrest

Endogenous ribonucleases destroy ineffective mRNA



Therapeutics: siRNA

Ancient host process of gene silencing, probably evolved for antiviral defense/genome protection

siRNA

– Exogenous short synthetic ds nucleic acid molecules

– Highly modified to increase stability (nuclease resistance), prolong half-life, and reduce nonspecific (off-target) effects

– Incorporated into RNA-induced splicing complex (RISC) which pairs it with target mRNA

– Destruction of mRNA by RNase



Target: Translation and Protein Processing



Therapeutics: Translation and Protein Processing

HCV polyprotein

C E1 E2 p7 NS2 NS3 NS4A NS4B NS5A NS5B

Serine protease (trans)

Serine protease (cis)



Therapeutics: Translation and Protein Processing (cont’d) Conclusions

– Potent antivirals

– Orally bioavailable

– Well tolerated

– Synergy with IFN; increased IFN sensitivity

– Require maintenance of trough concentration

– May be able to shorten course of therapy

– Other PI in development: ITMN B (InterMune)



Target: Viral RNA Transcription

-RNA

SubcellularMembrane

3’5’

+RNA5’3’

SubcellularMembrane

-RNA

3’5’

+RNA5’3’

+RNA5’3’

+RNA5’3’



Therapeutics: Viral RNA Transcription

HCV polymerase inhibitors in development

– NM-283 (valopicitabine, Idenix)

– R1626 (Roche)

– HCV-796 (Viropharma)



Target: Virus Assembly



Therapeutics: Virus Assembly

N-glycans in envelope glycoprotein (E1E2) are essential for protein folding, secretion/assembly, antigenicity, receptor binding, and cell entry

Imino sugars inhibit α-glucosidases and prevent proper glycosylation of viral envelope proteins; may inhibit secretion and infectivity of viruses

– Zitzmann, et al. PNAS 1999; Mehta, et al. FEBS Ltr 1998

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4 th Annual Clinical Care Options for Hepatitis Symposium: HCV Highlights