hbv therapy & approaches to cure - virology...
TRANSCRIPT
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,
Merck
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
100
80
60
40
20
0 5 10 15 20 25
Su
rviv
al p
rob
ab
ilit
y (
%) 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
mu
lati
ve H
CC
In
cid
ence
(%
)
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
% s
up
pre
ssed
HB
V D
NA
% s
up
pre
sse
d H
BV
DN
A
Suppressive therapy is not a cure
1378 Korean patients on LAM/ETV vs 1014 inactive CHB
Cho Gut 2014
Complete respondersInactive
CHB
p<0.001 p<0.015
Non-cirrhotic Cirrhotic
Cu
mu
lati
ve in
cid
en
ce o
f H
CC
Cu
mu
lati
ve in
cid
en
ce o
f H
CC
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?
O-
5’Cap (A)n 3’
Translocation
dAdAdG
new (-) strand DNA synthesis
pgRNA
DNA Synthesis
Encapsidationof pg RNA
Golgi complex
Release
CCC DNA
DNArepair
HBV RNATranscripts
PregenomicRNA
Attachment andPenetration
S Ag
e Ag
HBV Virion
EnvelopeProteins
S, M, L
e Ag
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
HBV
• Well tolerated, very
effective therapy – high
bar
• 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
O-
5’Cap (A)n 3’
Translocation
dAdAdG
new (-) strand DNA synthesis
pgRNA
DNA Synthesis
Encapsidationof pg RNA
Golgi complex
Release
CCC DNA
DNArepair
HBV RNATranscripts
PregenomicRNA
Attachment andPenetration
S Ag
e Ag
HBV Virion
EnvelopeProteins
S, M, L
e Ag
PolymeraseProteinCore
Protein
uncoating
transport to cellnucleus
Block Entry
Target cccDNA- Destruction- Inactivation
Target HBV RNA
Target packaging
Target DNA synthesis
Target Assembly/Export
Stimulation of innate and/or
adaptive immunity
Potential targets in the lifecycle
O-
5’Cap (A)n 3’
Translocation
dAdAdG
new (-) strand DNA synthesis
pgRNA
DNA Synthesis
Encapsidationof pg RNA
Golgi complex
Release
CCC DNA
DNArepair
HBV RNATranscripts
PregenomicRNA
Attachment andPenetration
S Ag
e Ag
HBV Virion
EnvelopeProteins
S, M, L
e Ag
PolymeraseProteinCore
Protein
uncoating
transport to cellnucleus
Block Entry
Target cccDNA- Destruction- Inactivation
Target HBV RNA
Target packaging
Target DNA synthesis
Target Assembly/Export
Stimulation of innate and/or
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
e Ag
PolymeraseProtein
Core Protein
1. Loss of sAg = loss of sAg transcription silent/absent cccDNA = our goal
Caveats with sAg loss
HBV RNATranscripts
PregenomicRNA
EnvelopeProteins (sAg)
S, M, L
e Ag
PolymeraseProtein
Core Protein
1. Loss of sAg = loss of sAg transcription silent/absent cccDNA = our goal
Elimination
Caveats with sAg loss
CCC DNA
HBV RNATranscripts
PregenomicRNA
EnvelopeProteins (sAg)
S, M, L
e Ag
PolymeraseProtein
Core Protein
x x
1. Loss of sAg = loss of sAg transcription silent/absent cccDNA = our goal Transcriptional
Silencing
Caveats with sAg loss
CCC DNA
HBV RNATranscripts
PregenomicRNA
EnvelopeProteins (sAg)
S, M, L
e Ag
PolymeraseProtein
Core Protein
xx
1. Loss of sAg = loss of sAg transcription silent/absent cccDNA = our goal
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)
Caveats with sAg loss
CCC DNA
EnvelopeProteins (sAg)
S, M, L
HBV RNATranscripts
PregenomicRNA
e Ag
PolymeraseProtein
Core Protein
sAgx x
1. Loss of sAg = loss of sAg transcription silent/absent cccDNA = our goal
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
Potential targets in the lifecycle
O-
5’Cap (A)n 3’
Translocation
dAdAdG
new (-) strand DNA synthesis
pgRNA
DNA Synthesis
Encapsidationof pg RNA
Golgi complex
Release
CCC DNA
DNArepair
HBV RNATranscripts
PregenomicRNA
Attachment andPenetration
S Ag
e Ag
HBV Virion
EnvelopeProteins
S, M, L
e Ag
PolymeraseProteinCore
Protein
uncoating
transport to cellnucleus
Block Entry
Target cccDNA- Destruction- Inactivation
Target HBV RNA
Target packaging
Target DNA synthesis
Target Assembly/Export
Stimulation of innate and/or
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
Potential targets in the lifecycle
O-
5’Cap (A)n 3’
Translocation
dAdAdG
new (-) strand DNA synthesis
pgRNA
DNA Synthesis
Encapsidationof pg RNA
Golgi complex
Release
CCC DNA
DNArepair
HBV RNATranscripts
PregenomicRNA
Attachment andPenetration
S Ag
e Ag
HBV Virion
EnvelopeProteins
S, M, L
e Ag
PolymeraseProteinCore
Protein
uncoating
transport to cellnucleus
Block Entry
Target cccDNA- Destruction- Inactivation
Target HBV RNA
Target packaging
Target DNA synthesis
Target Assembly/Export
Stimulation of innate and/or
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
sAg
crAg
eAg
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
0.00
–0.50
–1.00
–1.501* 29* 57* 85
Day
HB
sAg
(lo
g 10
IU/m
L)
HBeAg-negative ARB-1467 0.2 mg/kgHBeAg-negative ARB-1467 0.4 mg/kgHBeAg-postive ARB-1467 0.4 mg/kgPlacebo
Efficacy
*Dosing
e+e-
e-
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
Potential targets in the lifecycle
O-
5’Cap (A)n 3’
Translocation
dAdAdG
new (-) strand DNA synthesis
pgRNA
DNA Synthesis
Encapsidationof pg RNA
Golgi complex
Release
CCC DNA
DNArepair
HBV RNATranscripts
PregenomicRNA
Attachment andPenetration
S Ag
e Ag
HBV Virion
EnvelopeProteins
S, M, L
e Ag
PolymeraseProteinCore
Protein
uncoating
transport to cellnucleus
Block Entry
Target cccDNA- Destruction- Inactivation
Target HBV RNA
Target packaging
Target DNA synthesis
Target Assembly/Export
Stimulation of innate and/or
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…
Potential targets in the lifecycle
O-
5’Cap (A)n 3’
Translocation
dAdAdG
new (-) strand DNA synthesis
pgRNA
DNA Synthesis
Encapsidationof pg RNA
Golgi complex
Release
CCC DNA
DNArepair
HBV RNATranscripts
PregenomicRNA
Attachment andPenetration
S Ag
e Ag
HBV Virion
EnvelopeProteins
S, M, L
e Ag
PolymeraseProteinCore
Protein
uncoating
transport to cellnucleus
Block Entry
Target cccDNA- Destruction- Inactivation
Target HBV RNA
Target packaging
Target DNA synthesis
Target Assembly/Export
Stimulation of innate and/or
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
Potential targets in the lifecycle
O-
5’Cap (A)n 3’
Translocation
dAdAdG
new (-) strand DNA synthesis
pgRNA
DNA Synthesis
Encapsidationof pg RNA
Golgi complex
Release
CCC DNA
DNArepair
HBV RNATranscripts
PregenomicRNA
Attachment andPenetration
S Ag
e Ag
HBV Virion
EnvelopeProteins
S, M, L
e Ag
PolymeraseProteinCore
Protein
uncoating
transport to cellnucleus
Block Entry
Target cccDNA- Destruction- Inactivation
Target HBV RNA
Target packaging
Target DNA synthesis
Target Assembly/Export
Stimulation of innate and/or
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
Potential targets in the lifecycle
O-
5’Cap (A)n 3’
Translocation
dAdAdG
new (-) strand DNA synthesis
pgRNA
DNA Synthesis
Encapsidationof pg RNA
Golgi complex
Release
CCC DNA
DNArepair
HBV RNATranscripts
PregenomicRNA
Attachment andPenetration
S Ag
e Ag
HBV Virion
EnvelopeProteins
S, M, L
e Ag
PolymeraseProteinCore
Protein
uncoating
transport to cellnucleus
Block Entry
Target cccDNA- Destruction- Inactivation
Target HBV RNA
Target packaging
Target DNA synthesis
Target Assembly/Export
Stimulation of innate and/or
adaptive immunity
Immune response to HBV
Hepatocyte
NK(T)
DC
IFN
Viral
replication
activation
BAnti-HBe, HBc, HBs
Hepatocyte
lysis
Viral
replication
neutralization
Innate immunity Adaptive immunity
CD4+
CD8+
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
20
40
60
80
100 **
IFN p
ro
du
cin
g c
ell
s (
%)
t=0 t=60
20
40
60
80
100 *
CD
69 (
%)
t=0 t=60
25
50
75 *
IFN p
rod
ucin
g c
ell
s (
%)
NK cell
CD69+
CD69+
IFN+IL-12
IL-18
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
-0 .4
-0 .2
0 .0
2 4 3 6 4 8
W e e k s
HB
sA
g d
ec
lin
e (
log
IU
/mL
)
-2 .0
-1 .5
-1 .0
-0 .5
0 .0
2 4 3 6 4 8
W e e k s
HB
V D
NA
de
cli
ne
(lo
g I
U/m
L)
ETV +PEG-IFN
ETV
ETV +PEG-IFN
ETV
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
ImmunotherapiesInnate
• Cytokine therapy
– IFN
• TLR agonists
– TLR7
• RIG-I agonists
Adaptive
• Therapeutic vaccine
• Checkpoint inhibitors
– PD-1
– PD-L1
Early days…so far limited effects seen
ImmunotherapiesInnate
• Cytokine therapy
– IFN
• TLR agonists
– TLR7
• RIG-I agonists
Adaptive
• Therapeutic vaccine
• Checkpoint inhibitors
– 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
-5
-4
-3
-2
-1
-0 .8
-0 .6
-0 .4
-0 .2
0 .0
0 .2
-5
-4
-3
-2
-1
-0 .8
-0 .6
-0 .4
-0 .2
0 .0
0 .2
Nivolumab0.1 mg/kg
Nivolumab0.3 mg/kg
Nivolumab0.3 mg/kg+ GS-4774
Week 12 Week 24
HB
sAg
Ch
ange
fro
m B
L, L
og 1
0 IU
/mL
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…
Gane E, et al. EASL 2017
Learning from 1 patient
1
1 0
1 0 0
1 0 0 0
1 0 0 0 0
0
1 0 0
2 0 0
3 0 0
1 4 8 12 16 20 242 3B L
HB
sAg,
IU/m
L ALT, U
/L
IFN-γ Sp
ots/ 1
06
Ce
lls
Week
• 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…)
Gane E, et al. EASL 2017
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
A+
NA + TLR7
Immune
Target A
Attractive combinations
Nuc
HBV DNAsuppression
+Viral protein
Depletion(s, x, core)
+Immuno-therapy
RNAi
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…