prevention and potential intervention in persistent hepatitis b virus infection using a pre-s/s...

Prevention and potential intervention in persistent hepatitis B virus infection using a Pre-S/S vaccine

Daniel ShouvalLiver Unit

Hadassah-Hebrew University Hospital Jerusalem, Israel

Disclosures

Member of ABs or consultant- BMS, J&J,MSD,IntecPharma, SciVac, GSK

Outline

• Immunogenicity of a 3rd generation Pre S1/Pre-S2/S HBV vaccine in healthy subjects

• Immunogenicity of the PreS/S vaccine in special risk groups

• Potential intervention in persistent HBV infection using a combination of anti-viral therapy and active immunization with a PreS/S vaccine

By 2010 179/193 WHO member countries added HBV vaccination in their national immunization program

Weekly epidemiological record No. 40, 2009, 84, 405

Control of hepatitis B

• Prevention – highly successfulVs• Intervention -

– Nucleot(s)ide analogues (HBV-DNA suppression)- successful but eradication of int-ranuclear cccHBV DNA still elusive

– Targeting of the HBV receptor (still experimental)L-protein’s myristoylated N-terminal Pre-S1 domain binds to the sodium-taurocholate co-transporting polypeptide (NTCP)

Hepatitis B vaccination: immunogenicity• Several hundred million doses of HB vaccine have been

administered worldwide with an excellent record of safety and efficacy

• Seroprotection rates to anti-HBs are close to 100% in children and ~95% in healthy young adults

• Most vaccinated individuals have a preserved immune memory and show a strong anamnestic response (boosterability) up to 25 years after primary immunization

• Recently, an increasing number of booster failures has been reported. Anti-HBc seroconversion with or without transient viremia may occur in vaccinees exposed to HBV .Yet clinically significant breakthrough infections after successful vaccination are rare

• People who are elderly, obese, heavy smokers, on hemodialysis or immuno-compromised, have suboptimal responses

Breakthrough* infections followingimmunization

VHPB meeting report in Viral Hepatitsi 2012;20 No 2

Other reports:•Italy –estimated breakthrough 5-6 cases/year notified over 10 years•Gambia- 5-20% anti-HBc seroconversions over 15-20 years•Taiwan- 4.0-5.7% anti-HBc seroconversions

*No standard definition yet: An HBV infection in an appropriately immunized individual that is confirmed at least by anti-HBc seroconversion (with or

without HBsAg and/or HBV-DNA

History of HBV vaccines

• I. 1980-1982 : Plasma derived vaccines

• II. 1986 : Recombinant HBV DNA vaccine expressed in yeasts

• III. 1988 : Recombinant pre-S/S vaccines expressed in mammalian cells

Experimental:• New adjuvants• DNA vaccines• "Therapeutic” HBV vaccines

III

III

Zanetti AR,Van Damme P, Shouval D: The global impact of vaccination against hepatitis B: A historical overviewVaccine 2008;26:6266

Selected HBV vaccines

Country Manufacturer Envelope Protein(s) Source/Expression in

Brand Name

Belgium GSK S Plasma Engerix B

US MSD S Yeast Hepatavax-B

US MSD S Yeast Recombivax

Cuba Centro D.I.G Biotecnologia

S Yeast Heberbiovac

France Pasteur-M *S/Pre-S2 CHO GenHevac B

Israel Scigen S/PreS-1/PreS-2* CHO Sci B Vac

US/G Dynavax S Yeast Heplisav

*Glycosylated

Factors Related to NonFactors Related to Non--response to response to Hepatitis B Vaccines*Hepatitis B Vaccines*

• Genetically determined resistance• Advanced age• Overweight• Age• Gender• Smoking• Immune suppression• Chronic liver disease• Miscellaneous (RF, systemic disease etc.)

Pre

-S

enhancing

attenuating

*Craven DE, et al. Ann Int Med1986;105:356Alper CA et al. N Eng J Med 1989, 321:708-712Milich DR Immunol Today 1988, 9:380-33 Hohler Tet al.Hum Immunol 1998, 59:212-218

Populations of non-responders to conventional vaccination • Cancer patients (children) ~57%

(31% on Rx 88% off Rx)

• Acute lymphocytic leukemia ~ 10%

• Bone marrow transplant patients 15-68%

• Chronic renal failure & dialysis 34-81%

• Patients with chronic liver disease ~50%

• Pre-transplantation candidates 28-36%

• Post-transplantation patients ~10%

• HIV (children & adolescents) ~30%

• Miscellaneous

– Older adults and individuals at risk with overweight

– Patients with IBD

– Candidates for chemo and immunotherapy

HBV vaccine escape mutants- VEMs (I) • Sporadic reports have documented the emergence of escape mutants to the HBV to the small S antigen.

• Relatively rare breakthrough events were observed in Taiwan, Singapore, China, Italy, Japan and Alaska

•Mutations at the B and T cell epitopes of the envelope genes lead to substitution of amino acids within the "a" determinant of HBsAg which generate a conformational change affecting the binding of HBsAg to neutralizing antibodies

•The most frequent substitution is the sG145R mutation in the S gene, defined by a glycine to arginine switch

HBV vaccine escape mutants – VEMs (II)The main risk factors include:• Wild-type intra-uterine HBV transmission and high viral load in

HBsAg+/HBeAg+ pregnant women who transmit HBV to their newborns despite passive/active or active immunization

• Immunologic selection pressure through the host or vaccine induced immune responses (including hepatitis B immune globulin)

• Treatment with nucleoside analogues leading to mutations in the HBV polymerase gene which overlaps with the envelope gene coding for the "a" determinant of HBsAg

• Immune suppression

• Non-response to vaccination

• Spontaneous emergence of mutations due to the error prone function of the viral polymerase.

HBV envelope Genes Proteins and Particles

Michel ML, Thiollais P Pathologie Biologie 2010;58:288

A Pres1/PreS2/S mammalian cell (CHO) derived HBV vaccine

• Sci B VacTM

(Bio-Hep BTM , HepimmuneTM )

Shouval D, Ilan Y, Adler R, Deepen R, Panet A, Gorecki M, Even-Chen Z, Gerlich WH. Improved immunogenicity in mice of a mammalian cell derived recombinant hepatitis

B vaccine containing pre S1 and pre S2 antigens as compared to conventional yeast derived vaccines. Vaccine 12:1453–1459, 1994

*Shapira MY, Zeira E, Adler R, Shouval D. Rapid seroprotection against hepatitis B following the first dose of Pre-S1/Pre-S2/S vaccine. J. Hepatology 34(1):123-127, 2

Comparative immunogenicity of a Pre-S1/ Pre-S2/S HBV vaccine to a yeast derived vaccine

Impact of body weight on anti-HBs levels

Quantitation of anti-HBs

A B

Immunogenicity of the PreS/S vaccine in neonates

Anti-

HBs

mIU

/ml N=205

A B

Dose Range Study: Neonates Vietnam

Cellular and humoral immune response to a third generation hepatitis B vaccine

Schumann A et al. Journal of Viral Hepatitis 2007; 14, 592-598

Cellular and humoral immune response to Sci B Vac

A B

Schumann A et al. Journal of Viral Hepatitis 2007; Volume 14, 592-598

Comparative immunogenicity of a PreS/S hepatitis B vaccine in non- and low responders to conventional

vaccine

Pamela Rendi-Wagner, MD, DTM&H, MSc

Institute of Specific Prophylaxis & Tropical MedicineCenter for Physiology and Pathophysiology

Medical University of Vienna, Austria

Tel Aviv - 8 Jan, 2007

Induction of a robust T- and B-cell immune response in non- and low-responders to

conventional vaccination against hepatitis B by using a third generation PreS/S vaccine.

Krawczyk A, Ludwig C, Jochum C, Fiedler M, Heinemann FM, Shouval D, Roggendorf M, Roggendorf H, Lindemann M

.Vaccine 2014, : 5077-82 32

Humoral immune response in 19 non responders immunized with Sci B Vac

A.Krawczyk et al. Vaccine 2014; 32:5077

Cellular immune response in 19 non-responders immunized with Sci B Vac

A.Krawczyk et al. Vaccine 2014; 32:5077.

Pre S/S Vaccine in Non-respondersSummary –

• Significantly higher immunogenicity after 2 additional injections of 3rdgeneration PreS/S vaccine compared to conventional S vaccine at anti-HBs 10 and 100 IU/l level

• Influence of age, BMI and gender less pronounced in 3rdgeneration PreS/S vaccinees

• Higher reactogenicity after 3rdgeneration compared to conventional vaccine

• Confirmes link between non response and DRB1*03 and *07, DQB1*02 HLA loci

Rendi-Wagner P & Shouval D Vaccine 2006;24:2781 A.Krawczyk et al. Vaccine 2014; 32:5077

Immunogenicity of Sci B Vac in immune suppressed patients

Enhanced immunogenicity of Sci-B-Vacin dialysis patients with kidney failure

Sci-B-Vac – 20 g (n=9)

HB-Vax- II – 10 g (n=12)

Anti-

HBs

, mIU

/ml

0 30 60 90 180 210

days

10000

1000

100

10

1

Efficacy of a Pre-S /S Sci B vac Vaccine in PatientsReceiving Lamivudine Prophylaxis after Liver

Transplantation for Chronic Hepatitis B

Lo et al. Am J Transplantation 2007;7:434

50% response

Is there any clinical evidence to support development of a therapeutic vaccine against HBV?

1. Restored function of HBV-Specific T cells after long-term effective therapy with nucleos(t)ide analogues

2. Persistent HBV infection may be controlled through adoptive transfer of immunity from immune competent donors to bone marrow or stem-cell transplanted patients

3. Report on partial success of therapeutic vaccination in Woodchucks

Boni eta al. GASTROENTEROLOGY 2012;143:963–973Ilan Y et al. Gastroenterology 1993; 104: 1818; Lau GK et al. Hepatology 1997;25 : 1497Schumann A et al. Transplantation 2009 ; 87:103; Roggendorf M et al. J Viral Hepatitis 2007;14:51S; Menne C,Cote PJ. W J Gastroenterol 2007

Restored Function of HBV-Specific T Cells After Long-term Effective Therapy With Nucleos(t)ide Analogues

CAROLINA BONI,* DILETTA LACCABUE,* PIETRO LAMPERTICO,‡ TIZIANA GIUBERTI,* MAURO VIGANÒ,‡ SIMONA SCHIVAZAPPA,* ARIANNA ALFIERI,* MARCO PESCI,* GIOVANNI B. GAETA,§ GIUSEPPINA BRANCACCIO,§ MASSIMO COLOMBO,‡ GABRIELE MISSALE,* and CARLO FERRARI

In vitro reactivity can be restored to T cells from patients with suppressed HBV infection following long-term treatment with nucleos(t)ide analogues, despite prolonged exposure to large loads of HBsAg antigen

Immune therapies that increase the antiviral T-cell

response might increase the likelihood of complete HBV control in patients undergoing long-term nucleos(t)ideanalogue treatment

GASTROENTEROLOGY 2012;143:963–973

Is there any clinical evidence to support development of a therapeutic vaccine against HBV?

1. Restored function of HBV-Specific T cells after long-term effective therapy with nucleos(t)ide analogues

2. Persistent HBV infection may be controlled through adoptive transfer of immunity from immune competent donors to bone marrow or stem-cell transplanted patients

3. Report on partial success of therapeutic vaccination in Woodchucks

Boni eta al. GASTROENTEROLOGY 2012;143:963–973Ilan Y et al. Gastroenterology 1993; 104: 1818; Lau GK et al. Hepatology 1997;25 : 1497Schumann A et al. Transplantation 2009 ; 87:103; Roggendorf M et al. J Viral Hepatitis 2007;14:51S; Menne C,Cote PJ. W J Gastroenterol 2007

*Ilan & Shouval., Gastroenterology 1993; 104:1818

2 )Clearance of serum HBV markers in an HBsAg carrier following bone marrow transplantation from an HBV immune donor

Anti-HBs

HBsAg/sAb +/- +/- +/- +/- +/- +/- +/- +/- -/- -/+ -/+ -/+ -/+HBeAg +/eAb +/- +/- +/- +/- +/- +/- +/- -/+ -/- -/+ -/+ -/+ -/+

0

100

200

300

400

500

-1 0 4 8 12 16 20 24 28 32 36 40 44 48 52

Weeks after BMT

Seru

m A

LT le

vels

(IU

/L)

BMT

-1 0 4 8 12 16 20 24 28 32 36

Prospective follow-up of a HBsAg+ BMT recipient following adoptive transfer of immunity

Lau et al Gastroenterology 2002

6/17 cleared HBV(35%), 4/6 “permanently” All 6 patients received anti-HBc+/anti-HBs+ BM

Enhanced immunogenicity of Sci B Vac as demonstrated through adoptive transfer of immunity from organ donors to recipients

Protocol: Immunization of organ donor with Sci B Vac Follow-up of anti-HBs seroconversion in organ recipient

Adoptive transfer of immunity to HBV in mice: Comparison of yeast derived S to CHO derived S-pre S2–pre S1 vaccines

• Immunecompetent BALB/c mice (N=20) were selected to receive 1 mg of S or S – pre S2 –pre S1 vaccine in alum, given i.p. x 3

• Bone marrow from seroconverted donor mice was transplanted to naive immune-suppressed (XR, 600 rd) recipient mice

• Anti-HBs was measured in BMT recipient mice before and after booster vaccination with homologous vaccine

Adoptive transfer of immunity to HBV in mice: The humoral responseComparison of yeast to CHO derived hepatitis B vaccines

21 37 47 76 88 106101

102

103

104

105

S – yeast derived HBV vaccine

preS1/preS2/S – CHO derived HBV vaccine

day after BMT

booster

Ant

i-HB

s,m

IU/m

l

Day of BMT

Humoral immune response

Anti-HBs titer (quantitative IU/l) Chemiluminescent Microparticle Immunoassay

(CMIA, Architect)

Cellular immune response

IFN-γ production by ELISpot

Proliferation assay

Methods

0 2 4 6 weeks

Detection of HBV-specific immune responses

Transplantation

Schumann et al. 2009 Jtransplantation

Adoptive immune transfer of hepatitis B virus specific immunity from immunized living liver donors to liver recipients

©2009 Lippincott Williams & Wilkins, Inc. Published by Lippincott Williams & Wilkins, Inc. 2

Adoptive Immune Transfer of Hepatitis B Virus Specific Immunity From Immunized Living Liver Donors to Liver Recipients

.

Schumann, A. et al.Transplantation. 87(1):103-111,, 2009.

FIGURE 1. Humoral and cellular immune responses of 46 potential living liver donors (LLDs). The x-axis indicates the number of vaccinations or booster-vaccinations. Thirty-two donors (*) were excluded from transplantation because of medical or psychological reasons. Fourteen donors ([white up pointing small triangle] SYMBOL) donated a part of their liver to the corresponding recipients. Values in donors one to three who transferred their hepatitis B virus (HBV)-specific immunity are depicted as hollow triangles with numbers 1-3. (A) Antibodies to hepatitis B virus surface antigen (anti-HBs)-titers in the LLDs before transplantation or after the last immunization, (B) cellular immune response in the proliferation-assay, and (C) in the IFN-[gamma]-ELISpot. The horizontal lines indicate the cut-off (stimulation index of 2.5 or 10 spots, respectively).

Donor Recipient

pre pre 0.5 1 2 4 5 6 11 12 13 months post transplantation

Case 1

Donor Recipient

pre pre 0.5 1 2 4 5 6 11 12 13 months post transplantation

Cut-off

Immunosuppression

Sti

mu

lati

on in

dex Booster immunization

anti

-HB

s (I

U/L

)

n.t.

1

10

100

1,000

10,000

100,000

012345

1050

HBsAg - - - - - - - - - - -

Anti-HBc - - - - - - - - - - -

Proliferation assay

Rehermann et al., Nat Med 1996

HBV was detectable for 2-3 decades after acute hepatitis B

and serological recovery

Hepatitis B Virus (HBV): A “stealth” virus

B. Rehermann et al., Nat Med 1996

HBV DNA detectable for 2-3 decades after acute hepatitis B infection and serological anti-HBs recovery

An elusive goal

•Question: Can HBV vaccines be utilized for restoration of immune control over viral replication and bypass of tolerance in HBsAg carriers and finite suppression of cccHBV-DNA?

Can vaccines be used as an intervention in persistent HBV infection ?

• Persistence of cccHBV-DNA: available anti-viral agents are effective in suppressing the clinical and virologic manifestation of persistent infection but do not lead to complete eradication

• The challenge: Reversal of cellular and humoral immune hypo-responsiveness (tolerance) to HBV through indirect or direct manipulations of the adaptive and innate immune response, leading to elimination of residual infected hepatocytes; HBeAg seroconversion,; clearance of HBV-DNA and HBsAg

• Potential tools: – Intervention in HBV infection through immunization using

vaccines with enhanced immunigenicity or DNA vaccines:, so called therapeutic vaccination or “vaccinotherapy”

– Targeting cccHBV-DNA• Record: Many failures; few encouraging pilot studies

Is there any clinical evidence to support development of a therapeutic vaccine against HBV?

•Restored function of HBV-Specific T cells after long-term effective therapy with nucleos(t)ide analogues

•Persistent HBV infection may be controlled through adoptive transfer of immunity from immune competent donors to bone marrow or stem-cell transplanted patients

•Report on partial success of therapeutic vaccination in WoodchucksBoni eta al. GASTROENTEROLOGY 2012;143:963–973Ilan Y et al. Gastroenterology 1993; 104: 1818; Lau GK et al. Hepatology 1997;25 : 1497Schumann A et al. Transplantation 2009 ; 87:103; Roggendorf M et al. J Viral Hepatitis 2007;14:51S; Menne C,Cote PJ. W J Gastroenterol 2007

Studies on therapeutic vaccination in the woodchuck model (II)

Menne C,Cote PJ. W J Gastroenterol 2007

Michel ML et al. J Hepatol 2011;54:1286

Failures • Pol S. Immunotherapy of chronic hepatitis B by anti HBV vaccine Biomed

Pharmacother. 1995;49:105-9

• Vandepapelière P, Lau GK, Leroux-Roels G, Horsmans Y, Gane E, Tawandee T, Merican MI, Win KM, Trepo C, Cooksley G, Wettendorff M, Ferrari C. Therapeutic vaccination of chronic hepatitis B patients with virus suppression by antiviral therapy: a randomized, controlled study of co-administration of HBsAg/AS02 candidate vaccine and lamivudine. Vaccine 2007;25:8585

• Fontaine H, Kahi S, Chazallon C, Bourgine M, Varaut A, Buffet C, Godon O, Meritet JF, Saïdi Y, Michel ML, Scott-Algara D, Aboulker JP, Pol S; for the ANRS HB02 study group. Anti-HBV DNA vaccination does not prevent relapse after discontinuation of NUCS in the treatment of chronic hepatitis B: a randomised trial--ANRS HB02 VAC-ADN.Gut. 2014 Feb 20

.

Restored Function of HBV-Specific T Cells After Long-term Effective Therapy With Nucleos(t)ide Analogues

CAROLINA BONI,* DILETTA LACCABUE,* PIETRO LAMPERTICO,‡ TIZIANA GIUBERTI,* MAURO VIGANÒ,‡ SIMONA SCHIVAZAPPA,* ARIANNA ALFIERI,* MARCO PESCI,* GIOVANNI B. GAETA,§ GIUSEPPINA BRANCACCIO,§ MASSIMO COLOMBO,‡ GABRIELE MISSALE,* and CARLO FERRARI

In vitro reactivity can be restored to T cells from patients with suppressed HBV infection following long-term treatment with nucleos(t)ide analogues, despite prolonged exposure to large loads of HBsAg antigen

Immune therapies that increase the antiviral T-cell

response might increase the likelihood of complete HBV control in patients undergoing long-term nucleos(t)ideanalogue treatment

GASTROENTEROLOGY 2012;143:963–973

Emerging concept

• A high viral load is associated with T cell exhaustion/dysfunction

• Consequently, development of new therapeutic vaccination protocol(s) against HBV require suppression of viral load with a NUC intended to restore a strong T cell function

Rational for combined anti-viral therapy and vaccination

Modified after Michel ML et al. J Hepatol 2011;54:1286

Randomized Controlled Study Investigating Viral Suppression and Serological Response following Pre-S1/Pre-S2/S Vaccine Therapy Combined with Lamivudine Treatment in HBeAg-Positive Patients with Chronic Hepatitis B

Pham Thi Le Hoa,1,2* Nguyen Tien Huy,3 Le The Thu,4 Cao Ngoc Nga,1,2 Kazuhiko Nakao,5Katsumi Eguchi,5 Nguyen Huu Chi,1,2 Bui Huu Hoang,6 and Kenji Hirayama3,7,8*Antimicrob. Agents Chemother. 2009, 53(12):5134

Immunization of HBsAg+ CHB patients with Sci B VacOutline of study design.

Hoa P T L et al. Antimicrob. Agents Chemother. 2009;53:5134-5140

Hoa P T L et al. Antimicrob. Agents Chemother. 2009;53:5134-5140

Vaccine induced anti-HBs responses after 18 months(10 months after the last vaccine dose)

'In 27/55 patients, anti-HBs levels ranged between 100-1000 mIU/mlHowever-only 4 patients exhibited an HBsAg loss.

HBV DNA reduction after 3 months of treatment using repeated immunization with Sci B Vac in

HBsAg + patients treated by lamivudine

Hoa P T L et al. Antimicrob. Agents Chemother. 2009;53:5134-5140

Intervention in persistent HBV infection (II)

• Persistence of cccHBV-DNA: available anti-viral agents are effective in suppressing the clinical and virologic manifestation of persistent infection but do not lead to complete eradication

• Potential “game changer”:– Myrcludex-B (anHBV entery inhibitor inhibits

amplification of cccDNA

– Blocking agent of the taurochoaste co-transporting polypeptide (blocking of HBV entery receptor in humanized mice)

Ni Y, et al.Gastroenterology. 2014 Apr;146(4):107. Voltz T et al. J Hepatol 2013;58:861. Oehler N et al Hepatology. 2014 Apr 8.

Some remaining goals in global control of HBV infection

•Improvement in worldwide coverage of universal vaccination •Restoration of immune control over viral replication and bypass of tolerance in HBsAg carriers •Finite suppression of cccHBV-DNA•Development of a vaccine against HDV

Conclusions – therapeutic vaccines against HBV

• Multiple strategies have and are being tested for vaccine mediated therapeutic interventions in chronic hepatitis B but resolution of persistent infection remains an elusive goal

• Conventional , yeast derived ,alum hydroxide based recombinant HBV vaccines are ineffective in clearance of chronic HBV infection

• Treatment with a combination of a NUC and a Pre-S1/Pre-S2 HBV vaccine is effective in suppression of HBV replication and partial restoration of immune hyporesponsiveness against HBV

• From data obtained so far, it appears that immunotherapy for CHB will require simultaneous or sequential administration of immune stimulatory agents and Nucs

The Sci B Vac PreS1/PreS2/S HBV vaccineSummary

• Highly immunogenic• Induces faster anti-HBs seroconversion as

compared to yeast derived vaccines• Is effective in non-responders to conventional

HBV vaccines• Is immunogenic in immune-suppressed patients• Enhances HBV DNA suppression in CHB patients

when co-administered with NUCS

Three generations of HBV

S

Pre-S2

Pre-S1

Plasma derivedvaccines

rDNA Yeastderived

rDNA Mammalian cell derived vaccines

AcknowledgementsRuth AdlerYaffa AshurChezi BarenholzDevorah DiminskyMarian GoreckiYaron Ilan Ronit KorenJudy LauGeorge LauArnon NaglerAmos PanetMichael Shapira Raul Raz Ivy Yap

Michael RoggendorfHedwig RoggendorfPamela Rendi-WagnerA.SchumannA. Krawczyk M.Lindemann

The Liver Unit at Hadassah 2011

Targets for therapeutic vaccination:Repair of the exhausted and functionally impaired immune responses in patients with persistent

HBV infection and restoration of a multi-specific CD4 and CD8 T-cell responses

Michel ML et al. J Hepatol 2011;54:1286

Suggested scheme for therapeutic vaccination in chronic HBV

Liu J, Kosinska A, Lu M, Roggendorf M. Virologica Sinica 2014; 29:10-16

Increased Seroprevalence of HBV DNA With Mutations in the S Gene Among Individuals >18

Years of Age After Complete Vaccination

Conclusion – “Universal vaccination effectively controls HBV infection in children and adolescents. However, after adolescence, there is a significant increase in the seroprevalence of anti-HBs, anti-HBc, and HBV DNA, indicating that new preventative strategies are needed for adults”.

Ming-Wei Lai et al. Gastroenterology 2012 ;143:400-7.