hbv therapy & approaches to cure - virology...

HBV Therapy & Approaches

to Cure

Jordan J. Feld MD MPH

Toronto Centre for Liver Disease

Sandra Rotman Centre for Global Health

University of Toronto

Disclosures

• Research: Abbvie, Gilead, Janssen, Merck, Wako

• Consulting: Abbvie, Contravir, Gilead, Janssen,

Outline

• Goals of therapy

• Why do we need new treatment?

• Approaches to new treatment

– Virological targets

– Immunological targets

What are the goals of therapy?

Learning from natural history

0 5 10 15 20 25

%) Inactive CHB

HBeAg-/HBV DNA+

or HBeAg reversion

HBeAg+ persistence

Time (years)

• Very inactive disease and ideally HBsAg loss associated with excellent long-term and cancer-free survival

• A good goal for therapy

Fattocvich Gut 2008, Yang NEJM 2002

sAg + /eAg +

sAg + /eAg -

sAg - /eAg -Cu

Survival HCC

Jaundice

Fluid RetentionAscites

Esophageal Varices

HepaticEncephalopathy

Liver Cancer

What we’re trying to prevent

Cirrhosis

Goals of Therapy• Cure the infection

– True cure = all traces of HBV gone from the liver (ie. like HCV)

– This is VERY difficult (if not impossible) cccDNA

• Functional cure

– Use the markers of excellent natural history…

1. HBsAg loss (ideally with anti-HBs)

2. Possibly…sustained off treatment inactive disease without

HBsAg loss (HBeAg –ve, DNA undetectable, normal ALT,

normal histology)

Cure not so simple…reasons lie in the virology

Potent HBV DNA suppression

TDF vs TDF/FTC in LAM-R HBV

Long-term therapy with potent nucs leads to suppression in almost all patients (even after resistance)

Chang Hepatology 2010, Fung J Hep 2017

Long-term ETV in eAg +ve HBV

Suppressive therapy is not a cure

1378 Korean patients on LAM/ETV vs 1014 inactive CHB

Cho Gut 2014

Complete respondersInactive

p<0.001 p<0.015

Non-cirrhotic Cirrhotic

Complete responders

Inactive CHB

• Treatment reduces but does not eliminate risk of HCC• Spontaneous (immune) control better than suppressing

HBV DNA with treatment

Why don’t nucs lead to cure?

5’Cap (A)n 3’

Translocation

dAdAdG

new (-) strand DNA synthesis

DNA Synthesis

Encapsidationof pg RNA

Golgi complex

Release

CCC DNA

DNArepair

HBV RNATranscripts

PregenomicRNA

Attachment andPenetration

HBV Virion

EnvelopeProteins

S, M, L

PolymeraseProteinCore

Protein

uncoating

transport to cellnucleus

• Persistence with potent nuc treatment means there is a leak

The leakIntrahepatic HBV DNA during long-term TDF therapy in HIV/HBV co-infection

• Very slow decline and persistence of cccDNA long-term +

detectable intrahepatic non-cccDNA support ongoing

replication despite ‘complete suppression’’ ie the leak!

• cccDNA replenishment - re-circulation + de novo infection

• Implication: Cure requires VERY long-term therapy

cccDNA Total intrahepatic

HBV DNA

Boyd J Hep 2016

Why do we need new treatment?

Nucleoside Analogues

• Effective suppression of HBV DNA

• Minimal or no effect on immune control

• High risk of relapse with stopping before HBsAg loss

• Low rates of HBsAg loss long-term therapy– Safety

– Costs

– Monitoring

– Access in most of the world limited

Interferon

• Higher rate of HBsAg loss

but still low

• Finite therapy but poorly

tolerated

• Patients and providers

don’t want to use it!

Considerations for cureHCV

• Ineffective, poorly

tolerated therapy

• Multiple lifecycle targets

• No long-lasting nuclear

reservoir or integration

• Limited involvement of

immune system

• Well tolerated, very

effective therapy – high

• Single viral enzyme

• cccDNA very persistent

and hard to reach

• Immune control important

…flaresBottom line…it won’t be as easy to cure HBV as it

was to cure HCV!

Approaches to therapyViral targets - DAA

• Viral entry

• cccDNA

formation/transcription/de

gradation

• RNA intermediates

• Encapsidation

• DNA replication

• Assembly

• Release

Immunomodulators

• Innate immune response

– IFN

– TLR agonists

– RIG-I agonists

• Adaptive immune

response

– Anti-antagonists

(checkpoint inhibitors)

– Vaccination

Potential targets in the lifecycle

5’Cap (A)n 3’

Translocation

dAdAdG

DNA Synthesis

Golgi complex

Release

CCC DNA

DNArepair

HBV RNATranscripts

PregenomicRNA

HBV Virion

EnvelopeProteins

S, M, L

Protein

uncoating

Block Entry

Target cccDNA- Destruction- Inactivation

Target HBV RNA

Target packaging

Target DNA synthesis

Target Assembly/Export

Stimulation of innate and/or

adaptive immunity

5’Cap (A)n 3’

Translocation

dAdAdG

DNA Synthesis

Golgi complex

Release

CCC DNA

DNArepair

HBV RNATranscripts

PregenomicRNA

HBV Virion

EnvelopeProteins

S, M, L

Protein

uncoating

Block Entry

Target HBV RNA

Target packaging

adaptive immunity

HBV Entry: Discovery of HBV

receptor to the first entry blocker

Primary Human Hepatocytes • Pre-S1 of HBsAg binds to sodium taurocholate co-transporting peptide – NTCP

• Predominantly expressed in liver

• Myristylated lipopeptidecontaining aa 2-48 of HBV large surface antigen Myrcludex B

• Blocks HBV and HDV entry

Yan eLife Science 2012 Gripon PNAS 2002, Urban J Virol 2005, Glebe Gastro 2005, Schutze Gastro 2007

Clinical data for Myrcludex B

Bogolomov J Hep 2016, Urban AASLD 2016

• Modest HBV DNA decline (0.8 log), no effect on HBsAg, safety concerns but ALT normalization (55%) and greater effect in HDV

• Not likely enough on its own but maybe an adjunct therapy…

Myrcludex 10 mg OD x 24 weeks in HBV and HDV

HDV HBV

If sAg loss is the goal…we

should be sure we are always

talking about the same thing…

Caveats with sAg loss

CCC DNA

HBV RNATranscripts

PregenomicRNA

EnvelopeProteins (sAg)

S, M, L

PolymeraseProtein

Core Protein

1. Loss of sAg = loss of sAg transcription silent/absent cccDNA = our goal

HBV RNATranscripts

PregenomicRNA

S, M, L

PolymeraseProtein

Core Protein

Elimination

CCC DNA

HBV RNATranscripts

PregenomicRNA

S, M, L

PolymeraseProtein

Core Protein

1. Loss of sAg = loss of sAg transcription silent/absent cccDNA = our goal Transcriptional

Silencing

CCC DNA

HBV RNATranscripts

PregenomicRNA

S, M, L

PolymeraseProtein

Core Protein

2. Loss of sAg = loss of sAg translation siRNA…unclear what this means…may still be very helpful but unknown if the same as 1 (our usual sAg loss)

TranslationalSilencing

(siRNA)

CCC DNA

S, M, L

HBV RNATranscripts

PregenomicRNA

PolymeraseProtein

Core Protein

sAgx x

2. Loss of sAg = loss of sAg translation siRNA…unclear what this means…may still be very helpful but unknown if the same as 1 (our usual sAg loss)

3. sAg may still be made from integrated HBV DNA –makes 1 and 2 hard to confirm!

Integration

5’Cap (A)n 3’

Translocation

dAdAdG

DNA Synthesis

Golgi complex

Release

CCC DNA

DNArepair

HBV RNATranscripts

PregenomicRNA

HBV Virion

EnvelopeProteins

S, M, L

Protein

uncoating

Block Entry

Target HBV RNA

Target packaging

adaptive immunity

cccDNA – Approaches

1. Formation- RC to cccDNA- HBV DNA recycling

3. Transcription- pgRNA - replication- mRNA – Ag production

2. Degradation- Destroy existing pool

Boucle Clin Liv Dis 2016

cccDNA - approaches

1. Formation– DSS – disubstituted sulfonamides – rc to cccDNA

– Early days but interesting…

2. Degradation– CRISPR/Cas9

– Directly cleave cccDNA• Mutation or degradation

• Delivery, off-target effects

3. Transcription– Prevent RNA transcription

• pgRNA no replication

• mRNA no proteins (sAg, xAg)

– Histone acetyltransferase inhibitor• Specificity?

Cai Antimicrob. Agents Chemother 2012Lin Int J Mol Sci 2015, Tropberger PNAS 2015

5’Cap (A)n 3’

Translocation

dAdAdG

DNA Synthesis

Golgi complex

Release

CCC DNA

DNArepair

HBV RNATranscripts

PregenomicRNA

HBV Virion

EnvelopeProteins

S, M, L

Protein

uncoating

Block Entry

Target HBV RNA

Target packaging

adaptive immunity

Target RNA - siRNA

• Overlapping reading frames = conserved regions• siRNA targeting can eliminate all HBV gene products

- sAg, pol, core immune function - pgRNA (replication)

Woodell Mol Ther 2013, Arrowhead

Clinical Data with RNAi

HBeAg -ve

HBeAg +ve

Placebo

HBV Protein reduction with ARC520 sAg decline eAg + vs -

• Effective knockdown of all HBV proteins• Much more effective in HBeAg + than HBeAg -

• e+ - initial 1.6 log to 2.9 log cccDNA?• e- - initial 0.5 log to 1.2 log integrated?

• Well tolerated but safety concerns…new delivery system? FDA HOLD• ALT rise off RNAi…?restored immune response?

Yuen AASLD 2015, Yuen EASL 2017

A different RNAi, different results

–0.50

–1.00

–1.501* 29* 57* 85

HBeAg-negative ARB-1467 0.2 mg/kgHBeAg-negative ARB-1467 0.4 mg/kgHBeAg-postive ARB-1467 0.4 mg/kgPlacebo

Efficacy

*Dosing

Different RNAi combination effective with single & multi-doseNo difference between eAg +ve and –ve (but slightly lower sAg decline)

Streinu-Cercel A, et al. EASL 2017, Amsterdam. #SAT-155

ARB-1467 0.2 or 0.4 mg/kg in NA-suppressed patients monthly IV infusion

Bottom line on siRNA for HBV

• Attractive approach

– Inhibits viral protein reduction immune restoration

(need to prove this)

– Inhibits viral replication directly (pgRNA)

– Pan-genotypic, relatively high barrier to resistance

• Major challenges

– Delivery

– Effect in eAg –ve ?integrated sAg

– Safety

5’Cap (A)n 3’

Translocation

dAdAdG

DNA Synthesis

Golgi complex

Release

CCC DNA

DNArepair

HBV RNATranscripts

PregenomicRNA

HBV Virion

EnvelopeProteins

S, M, L

Protein

uncoating

Block Entry

Target HBV RNA

Target packaging

adaptive immunity

Core protein – Attractive target

• Highly conserved

• Lots of functions– Transport to the nucleus

– Uncoating of HBV DNA

– Packaging

– Capsid assembly

– Modulate reverse transcription

– Interacts with sAg

– May also modulate cccDNA and export viral RNA

• All allosteric regulation…1 molecule that affects any function will affect them all!

Capsid Assembly Modulators (CpAMs)

Feld Antivira Res 2007Katen Chem Bio 2010

Phenylpropanamides (AT130) inhibit packaging – empty capsids

• Not capsid inhibitor – actually accelerate capsid formation• But empty capsids – no RNA or polymerase inside!• Better agents in the pipeline…

5’Cap (A)n 3’

Translocation

dAdAdG

DNA Synthesis

Golgi complex

Release

CCC DNA

DNArepair

HBV RNATranscripts

PregenomicRNA

HBV Virion

EnvelopeProteins

S, M, L

Protein

uncoating

Block Entry

Target HBV RNA

Target packaging

adaptive immunity

New nucs…TAF Tenofovir alofenamide (TAF)

• Very similar kinetics in the blood

• Unclear what is happening in the liver…

• Faster ALT normalization is interesting…

• Unlikely to be ‘the cure’ but may be combined with other agents

improved safety and possibly more potent

• Adding RNaseH activity may improve potency

• Multiple others (TDF variants) in late-stage development…

HBeAg -ve HBeAg +ve

Buti EASL 2016, Chan EASL 2016

5’Cap (A)n 3’

Translocation

dAdAdG

DNA Synthesis

Golgi complex

Release

CCC DNA

DNArepair

HBV RNATranscripts

PregenomicRNA

HBV Virion

EnvelopeProteins

S, M, L

Protein

uncoating

Block Entry

Target HBV RNA

Target packaging

adaptive immunity

Antigen & Virion SecretionGlucosidase Inhibitors• HBV proteins heavily glycosylated

• Mimic glycosylated residues on viral proteins leading to inhibition of α-glucosidase

• Inhibit secretion of HBsAg and HBV virions in vitro and in woodchucks

• Recognized a long time ago…no progress

Newer Approaches• REP 9’AC – blocks secretion of empty HBsAg (not virus)

– Promising HBsAg loss data…but mechanism obscure

– Too good to be true?

• Triazolopyrimidines – also block HBsAg secretion

Block 1998 Nat Med, Mahtab Hepatol 2012, Yu JMC 2011

5’Cap (A)n 3’

Translocation

dAdAdG

DNA Synthesis

Golgi complex

Release

CCC DNA

DNArepair

HBV RNATranscripts

PregenomicRNA

HBV Virion

EnvelopeProteins

S, M, L

Protein

uncoating

Block Entry

Target HBV RNA

Target packaging

adaptive immunity

Immune response to HBV

Hepatocyte

replication

activation

BAnti-HBe, HBc, HBs

Hepatocyte

replication

neutralization

Innate immunity Adaptive immunity

Rehermann Nat Rev Immun 2005, Woltman Gut 2010

ImmunotherapiesInnate

• Cytokine therapy

– IFN

• TLR agonists

– TLR7

• RIG-I agonists

Adaptive

• Therapeutic vaccine

• Checkpoint inhibitors

– PD-1

– PD-L1

Immune restoration through inhibition of viral antigens

Immune restoration with NAs

HC HBV0

100 **

t=0 t=60

NK cell

IFN+IL-12

Viral suppression with ETV restores NK cell response activation

and cytokine production

ETV Tx ETV Tx

Tjwa et al. J Hepatol 2011

Would this justify adding a nuc

to IFN?ETV-suppressed patients randomized to ETV continuation vs adding PegIFN

2 4 3 6 4 8

W e e k s

2 4 3 6 4 8

W e e k s

ETV +PEG-IFN

P<0.001P<0.001

HBV DNA HBsAg

Improved DNA & HBsAg decline BUT no IFN monotherapy arm and only 18% off-treatment response…intriguing but not the answer

Sonneveld J Hep 2015

TLR7 Agonist

Led to IFN production, few systemic side effects

TLR7- Pathogen recognition receptor- Triggers IFN production – antiviral

response- GS-9620 – oral, nanomolar potency- Active in vivo – chimps, humans

Lanford Gastro 2013

TLR7 in chimps

• Mean max HBV DNA decline 2.2 logs, HBeAg decline

• ALT flares occurred; but limited IFN systemic effects

• Ongoing phase 2 trial in patients on TDF

Lanford Gastro 2013

– IFN

• TLR agonists

– TLR7

• RIG-I agonists

Adaptive

– PD-1

– PD-L1

Early days…so far limited effects seen

– IFN

• TLR agonists

– TLR7

• RIG-I agonists

Adaptive

– PD-1

– PD-L1

Early days…so far limited effects seen

Therapeutic vaccines

Tarmogen- Modified yeast expressing HBV protein- Scalable- Ongoing studies…

Other approaches:- Adeno or AAV vector with HBV proteins- Improved adjuvant- Add TLR agonist to vaccine…- Lots of ideas – limited data

- Results generally disappointing to date

Checkpoint inhibitors

Bertoletti HBV Endpoints 2016

Excessive co-stimulatory signals T cell exhaustion/deletion

• Checkpoint inhibitors – taking off the brake…• leads to immune activation ?control?

Promising target in oncology

PD1 relevant in HBV

• PD1 most relevant checkpoint inhibitor for HBV• Expressed on HBV-specific T cells• PDL1 (ligand for PD1) on liver cells during hepatitis

Bengsh J Hep 2014

PD1 Blockade

Baseline Week 4 Week 16Week 12

Nivolumab 0.3 mg/kgSentinel B, n=2

Sentinel A, n=2 Nivolumab 0.1 mg/kg

Cohort A, n=10 Nivolumab 0.3 mg/kg

Cohort B, n=10 +GS-4774 40 YU

Nivolumab 0.3 mg/kg

NA-suppressed e-neg patients treated with nivolumab +/- GS-4774 (Tarmogen vaccine)

Gane E, et al. EASL 2017

Efficacy

Nivolumab0.1 mg/kg

Nivolumab0.3 mg/kg

Nivolumab0.3 mg/kg+ GS-4774

Week 12 Week 24

Nivolumab0.1 mg/kg

Nivolumab0.3 mg/kg

Nivolumab0.3 mg/kg+ GS-4774

• High receptor occupancy despite relatively low dose• Very modest decline in HBsAg overall…but 1 patient…

Learning from 1 patient

1 0 0 0

1 0 0 0 0

1 4 8 12 16 20 242 3B L

L ALT, U

IFN-γ Sp

ots/ 1

• Cleared HBsAg, stopped TDF 4 weeks after dosing • 8 months later – remains HBsAg-negative and anti-HBs positive• Interesting proof of concept – very low dose given (but pretty big flare…)

It’s not just choosing the right

target/compound…• Other MAJOR issues

1. Correct population

• Highest need?

• Easiest to show an effect?

• Immuntolerant/e+/e-/inactive/NA suppressed…

2. Correct endpoint

• Is sAg loss the same with an NA as with an siRNA?

• Do we need to look in the liver? Do we need new biomarkers –HBV RNA, HBV crAg others?

3. Correct combinations

• Lots of possibilities – a huge matrix!!

4. Safety!

• A major concern…especially with immunotherapies

Combination Approaches

Viral target

A+ Viral target

A NA + NA

Viral target

A+ Viral target

B NA + RNAi

Immune

Target A +TLR7 + Vaccine

Immune

Target B

Viral target

NA + TLR7

Immune

Target A

Attractive combinations

HBV DNAsuppression

+Viral protein

Depletion(s, x, core)

+Immuno-therapy

Nucleic Acid Polymers

cccDNAi

+/- cccDNAi

+/- entry inhibitor

+/- RNAi

+/- CpAM

TLR/RIG-I agonist

αPD1/PDL1

Therapeutic

vaccine

May not need all 3 ‘classes’…may need more…mix and match

Summary for HBV cure• Many virological targets (but all with challenges…)

– Entry

– cccDNA formation/degradation/transcription

– Capsid

– Secretion

• Fewer immunological targets

– Innate – TLR, RIG-I

– Adaptive – Therapeutic vaccine, checkpoint inhibitors

• Much more challenging than HCV…both scientifically

and clinically (much higher bar, difficult to study)

• Interesting times ahead…

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