Therapeutic immunization strategies in chronic hepatitis B
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1 Therapeutic immunization strategies in chronic hepatitis B Mengji Lu Institut für Virologie Universitätsklinkum Essen Falk Symposium Berlin, 04. October 2005
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3 The aim of therapeutic immunization to achieve a long term control of HBV infection by stimulating specific immune responses in chronic HBV carriers What is the rational?
4 Immunological control of HBV during acute infection B cell HBsAg, HBeAg Plasma cells anti-hbs anti-hbe, anti-hbc APC Th cells to HBcAg and HBsAg APC CTLs to HBsAg, HBcAg, and polymerase
5 Impaired immune responses in chronic HBV infection HBsAg, HBeAg B cell anti-hbc APC Specific T-cells anti-hbs? APC
6 Therapeutic vaccination to restore specific immune responses B cell Antigens Plasma cells anti-hbs anti-hbe, anti-hbc APC Th cells to HBcAg and HBsAg Therapeutic Vaccines APC CTLs to HBsAg, HBcAg, and polymerase
7 Clinical experiences: published clinical trials Conventional HBsAg vaccines Pol S. et al.,lancet, 1994, J. Hepatol.,42001 Couline et al., J. Infect. Dis., 2001 Jung C. et al., Vaccine, 2002 Ren F., et al., J. Med. Virol., 2003 Yalcin et al., J. Clin. Gastroenterol., 2003 Dikici et al., J Gastroenterol. Hepatol., 2003 HBsAg vaccine+lamivudine Horiike N., J Clin. Virol., 2005 HBsAg vaccine, oral application Safadi R. et al., American J. Gastroenterol., 2003 CTL vaccine Heathcote et al., Hepatology, 1999 DNA vaccine Mancini-Bourgine et al., Hepatology, 2004 HBsAg-anti-HBs immune complexes Wen Y. et al., Lancet, 1995, Xu D. Z. et al., Vaccine, 2005
8 Clinical trials using HBsAg HBsAg vaccines Reduction of viremia Pol S. et al., 1998, 2001 yes/no Couline et al., 2001 yes Jung C. et al., 2002 Ren F., et al., 2003 yes Yalcin et al., 2003 no Dikici et al., 2003 no 12x vaccinations + lamivudine for 12 m Horiike N., 2005 yes
9 Clinical trials using HBsAg HBsAg vaccines Reduction T-cell response of viremia CTL CD4+ Pol S. et al., 1998, 2001 yes/no Couline et al., 2001 yes yes Jung C. et al., 2002 yes Ren F., et al., 2003 yes no yes Yalcin et al., 2003 no Dikici et al., 2003 no 12x vaccinations + lamivudine for 12 m Horiike N., 2005 yes
10 Clinical trials using HBsAg HBsAg vaccines Reduction T-cell response anti-hbs of viremia CTL CD4+ antibody Pol S. et al., 1998, 2001 yes/no Couline et al., 2001 yes yes (no) Jung C. et al., 2002 yes Ren F., et al., 2003 yes no yes Yalcin et al., 2003 no no Dikici et al., 2003 no no 12x vaccinations + lamivudine for 12 m Horiike N., 2005 yes
11 Conventional vaccines promote mainly Th2 type responses Alum Alum-absorbed vaccines may be effective to stimulate specific Th cells. But appear not to be favorable for therapeutic vaccinations.
12 T-cell responses are crucial to control HBV replication CD8+ CTLs are required for viral clearance T-cell responses are also important to limit HBV replication and liver injury in chronic hepatitis B Manni et al., JEM, 1999 Reciprocal relationship of viral replication/liver damage and specific CTL activity
13 Pathogen associated molecular patterns (PAMPs) as adjuvants
14 Ligands of Toll-like receptors as adjuvants DNA vaccines, CpG ODN PALM2 MPL PALM2: 2x palmitic acid residues MPL: monophosphoryl lipid A
15 CTL vaccine CY-1899 Vitiello A., et al., J. clin. Invest. (1995), 95: 341 2x Palmitic acid residues T Helper Peptide TetTox CTL epitope HBV core KSS-QYIKANSKFIGITE-AAA-FLPSDFFPSV Livingstong B. D., et al., JI (1997), 159: immunizations with 50, 500, and 5000 μg CY-1899 induced efficiently CTLs in healthy persons Dose-response diagram
16 CTL vaccine CY-1899 Vitiello A., et al., J. clin. Invest. (1995), 95: 341 Livingstone B. D., et al., JI (1997), 159: x Palmitic acid residues T Helper Peptide TetTox CTL epitope HBV core KSS-QYIKANSKFIGITE-AAA-FLPSDFFPSV J. Heathcote, Hepatology 30 (1999); Patients n = 90, received 4x immunization with 50, 500, 5000, 10000, and μg Results: mean CTL response max. 10 lytic units no change of viremia no change of viral replication in the liver
17 DNA vaccine expressing HBsAg DNA vaccines, CpG ODN
18 DNA vaccine expressing HBsAg Induction of anti-hbs antibodies and CTL responses (Michel M.L. et a., PNAS, 1995, Schirmbeck et al., 1996) Mancini-Bourgin, Hepatology 40 (2004); Patients n = 10, received 3x or 4x immunization with 1 mg plasmid DNA pcmv S2.S Results: Induction of lymphoproliferative responses in 2 patients Induction of T-cells responding to HBsAg peptides Decrease of HBV DNA titers in 5 patients with one complete clearance
19 Immunotherapy of chronic hepatitis B virus infection Wish....Reality Induction of multiple, effective T- and B- cell responses to HBV proteins Partial Induction of HBVspecific immune responses Sustained responses Transient responses Virus elimination Reduction of virus replication
20 Why are the recent therapeutic approaches not successful? The vaccines used in these studies did not effectively induce all branches of immune responses that are required for control of HBV replication. The high HBV replication may prevent the induction and maintainance of effective immune responses. The specific HBV proteins may interfere with the induction of immune responses.
21 WHV/Woodchuck (Mamota monax) a mammalian animal model for immunological approaches Woodchuck hepatitis virus (WHV) and HBV have high similarities in morphology, genome structure, replication cycle, etc. WHV and HBV infections resemble in natural history of infection: chronicity, HCC host immune respones at B- and T-cell levels
22 Therapeutic immunizations in the woodchuck model Vaccines Outcomes Inhibition of replication Induction of Anti-WHsAg WHcAg particles 1 WHsAg with a Th epitope 2 WHsAg and L-FMAU 3 WHsAg with MPL as adjuvant 4 DNA vaccines expressing WHsAg / WHcAg 5 WHsAg/Anti-WHs immune complexes 5 partial no partial no transient transient no no/yes yes yes no yes 1) Roggendorf et al. Intervirology, 1995; 2) Hervas-Stubbs et al., ) Menne et al. J. Virol., 2002; 4) Lu et al. J Hepatol., 2004; 5) Lu et al. 2005
23 New therapeutic appraoches should include Potent CTL inducer: CTL inducers DNA vaccines expressing surface antigen, core antigen and interferon-gamma DNA vaccines Viral vectors Adjuvants like CpG ODN Virus-like particles
24 New therapeutic appraoches should include CTL inducers DNA vaccines expressing surface antigen, core antigen and interferon-gamma Add proteins to enhance specific antibody responses immune complexes of surface antigen-antibody! DNA vaccines are poor induces for antibody responses
25 New therapeutic appraoches should include CTL inducers DNA vaccines expressing surface antigen, core antigen and interferon-gamma Add proteins to enhance specific antibody responses immune complexes of surface antigen-antibody Antiviral treatment with nucleoside analoga lamivudine Boni C. et al., JCI, 1998; Hepatology, 2001 Lamivudine enhances T-cell Proliferation and function
26 New therapeutic appraoches should include CTL inducers DNA vaccines expressing surface antigen, core antigen and interferon-gamma Add proteins to enhance specific antibody responses immune complexes of surface antigen-antibody Antiviral treatment with nucleoside analoga lamivudine week Lam 15 mg/day immunization immunization immunization Lam stop Monitoring Virus loads Surface antigen Antibody to surface antigen Lymphoproliferation
27 Controls lamivudine Control woodchucks treated with lamivudine alone anti-whs OD Week lamivudine Con
28 DNA vaccine anti-whs OD lamivudine DNA vaccine Immunizations of chronic carrier woodchucks with DNA vaccines did not induce anti- WHs antibody Week Immunizations at week 9, 13, and
29 Anti-WHs(OD 490) 2 1 Transient decrease of titers of viral DNA without woodchuck induction of anti-whs titer in WH17502 I I I WHsAg(ug/ml) 1,00E+06 1,00E I I week I 1,00E+04 1,00E+03 1,00E I=Immunization DNA lamivudine
30 Woodchuck was showed lymphoproliferative responses to WHV Core, WHsAg, and five different peptides after immunization with DNA vaccines : DNA vaccine neg sag cag Sp31 Sp29 Sp27 Sp25 Sp23 Sp21 Sp19 Sp17 Sp15 Sp13 Sp11 Sp9 Sp7 Sp5 Sp3 Sp1 Cp19 Cp18 Cp17 Cp16 Cp15 Cp14 Cp13 Cp12 Cp11 Cp10 Cp6 Cp1 T-cell responses are critical for down regulation of hepadnavirus replication
31 anti-whs OD Immune complex/dna lamivudine IC/DNA Immunizations of chronic carrier woodchucks with IC/DNA induced anti-whs antibody Immunizations at week 8, 12, and Week Anti-WHs antibodies were detected in 3 of 4 woodchucks after Immunizaitons
32 Transient decrease of titers of WHsAg and viral DNA in paralell with increase of anti-whs titer in WH I 1000 Anti-WHs(OD 490) 1 0 1,00E+09 I I I week WHsAg(ug/ml) 1,00E+08 1,00E+07 I PCR 1,00E+06 1,00E+05 1,00E+04 I I=Immunization IC/DNA lamivudine 1,00E
33 What we learnt from this experiment? Good news Antiviral treatment Therapeutic immunizations Anti-S antibodies Lymphoproliferative responses Reduction of virus load and antigenemia Bad news No indication for immunopathology No sustained response
34 More potent antiviral drug and repeated immunizations CTL inducers DNA vaccines expressing surface antigen, core antigen and interferon-gamma Add proteins to enhance specific antibody responses surface antigen with adjuvant promoting Th1 responses Antiviral treatment with nucleoside analoga Entacavir week Entacavir immunization immunization immunization immunization immunization immunization Entacavir stop Outcomes?
35 University Hospital Essen
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