Novel Approaches to Augmenting the Immunogenicity of DNA Vaccines for HIV1 Dan H. Barouch March 18,

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Novel Approaches to Augmenting the Immunogenicity
of DNA Vaccines for HIV-1Dan H. BarouchMarch
18, 2005
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The Expanding Worldwide HIV EpidemicUNAIDS 2004

• 39.4 million people living with HIV/AIDS
• 4.9 million new HIV infections in 2004
• 3.1 million deaths due to AIDS in 2004
• 57 HIV-infected adults in sub-Saharan Africa are
  women
• 90 of HIV-infected individuals worldwide have no
  access to antiretroviral therapy

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Current HIV-1 Vaccine Strategies

• Traditional Strategies
• live attenuated virus
• whole killed virus
• protein subunit
• Novel Strategies
• live recombinant vector
• plasmid DNA

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In Vivo Depletion of CD8 T Lymphocytes Prolongs
SIV Viremia in Rhesus Monkeys
CD8-depleted (lt21 days)
CD8-depleted (gt28 days)
Controls
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1
SIV Gag p27 ng/ml
0.1
CD8-depl.
CD8-depl.
0.01
Days
median range
Schmitz et al. Science 1998
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Current Challenges in the Development of DNA
Vaccines for HIV-1

• DNA vaccines in humans
• Poorly immunogenic
• High doses required
• Prohibitively expensive
• Manufacturing challenges
• Need for improving DNA vaccines
• Improve immunogenicity
• Lower required dose

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Trial of Cytokine-Augmented DNA Vaccination in
Rhesus Monkeys

• Plasmid DNA vaccines (5 mg each)
• SIVmac239 Gag
• HIV-1 89.6P Env
• Immunostimulatory cytokines
• IL-2/Ig protein
• IL-2/Ig plasmid
• Immunization i.m. at weeks 0, 4, 8, 40
• SHIV-89.6P challenge at week 46
• 100 MID50 i.v.

Barouch et al. Science 2000
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Gag-Specific CD8 T Lymphocyte Responses Elicited
by Plasmid IL-2/Ig-Augmented DNA Vaccines in
Rhesus Monkeys
Week
Barouch et al. Science 2000
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Efficacy of Cytokine-Augmented DNA Vaccine
Against SHIV-89.6P Challenge in Rhesus Monkeys
Sham Vaccine
Adjuvanted DNA Vaccine






death
Barouch et al. Science 2000
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HVTN 044A Phase I Clinical Trial to Evaluate
the Safety and Immunogenicity of the HIV-1 DNA
Vaccine VRC-HIVDNA-009-00-VP (Gag-Pol-Nef-Multicla
de Env) with the Plasmid Cytokine Adjuvant
VRC-ADJDNA-004-IL2-VP (IL-2/Ig)
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HVTN 044 Study Outline

• Study Design randomized, double-blinded,
  dose-escalation
• Vaccine VRC Gag-Pol-Nef-Env DNA Vaccine
  Plasmid IL-2/Ig administered i.m. week 0, 4, 8,
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• Group N (vac/ctrl) DNA Vaccine Plasmid IL-2/Ig
• A 5 / 6 4 mg 0.1 mg
• B 5 / 6 4 mg 0.5 mg
• C 10 / 6 4 mg 1.5 mg
• D 10 / 6 4 mg 4 mg
• E 10 / 6 4 mg 4 mg (day 2)
• Each group contains 6 control subjects 2
  placebo, 2 DNA vaccine alone, 2 plasmid IL-2/Ig
  alone

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Recruitment and Activation of Dendritic Cells
Potentiate the Immunogenicity and Protective
Efficacy of DNA Vaccines in Mice
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Enhancement of Antigen Presentation

• Hypothesis
• Availability of dendritic cells (DCs) may be
  rate-limiting for DNA vaccines since DCs are
  typically absent in muscle
• Aims
• Assess inflammatory infiltrates at the site of
  vaccine inoculation and immune responses to DNA
  vaccines when administered with
• MIP-1a chemokine that recruits DCs
• Flt3L growth factor that mobilizes and activates
  DCs

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Histopathology and Immunohistochemistry Study

• Objective To assess the extent and nature of
  cellular inflammatory infiltrates following DNA
  vaccination with or without plasmid Flt3L and
  plasmid MIP-1a in mice
• Mice injected intramuscularly with
• Saline
• 50 mg HIV-1 Env gp120 DNA vaccine alone
• 50 mg DNA vaccine 50 mg plasmid Flt3L
• 50 mg DNA vaccine 50 mg plasmid MIP-1a
• 50 mg DNA vaccine 50 mg plasmid Flt3L 50 mg
  plasmid MIP-1a
• On day 7 following immunization, injected muscle
  sites analyzed by histopathology and
  immunohistochemistry

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Plasmid MIP-1a and Flt3L Recruit Large Cellular
Infiltrates to the Site of Inoculation (Muscle
Sections, HE, Day 7)
Sham
DNA alone
Flt3L
MIP-1a
MIP/Flt3L
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Plasmid MIP-1a and Flt3L Recruit Large
Infiltrates of CD11b Antigen-Presenting Cells
(APCs) (Muscle Sections, IHC, Day 7)
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Plasmid MIP-1a and Flt3L Recruit Large
Infiltrates of S100 CD83 Dendritic Cells
(Muscle Sections, IHC, Day 7)
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DCs Recruited by Plasmid MIP-1a and Flt3L Exhibit
an Activated and Mature Phenotype
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Immunogenicity Study in Balb/c Mice

• Objective To assess whether increased DC
  recruitment and activation lead to improved DNA
  vaccine immunogenicity
• Mice (N8/group) injected at week 0 with
• Sham DNA
• HIV-1 Env DNA vaccine alone
• DNA vaccine plasmid Flt3L
• DNA vaccine plasmid MIP-1a
• DNA vaccine plasmid Flt3L plasmid MIP-1a
• Mice boosted at week 4 with 106 vp rAd5-Env
• CD8 T lymphocyte responses assessed by Dd/P18
  tetramer binding assays following primary and
  boost immunization

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Plasmid MIP-1a and Flt3L Synergistically Augment
TetramerCD8 T Lymphocyte Responses
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Plasmid MIP-1a and Flt3L Augment TetramerCD8 T
Lymphocyte Responses Following rAd5 Boost
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Vaccinia-Env Viral Challenge Study

• Objective To assess whether increased
  immunogenicity due to DC recruitment improves the
  protective efficacy of DNA vaccines against a
  pathogenic vaccinia virus challenge
• Mice (N4/group) vaccinated at week 0 with
• Sham DNA
• HIV-1 Env DNA vaccine alone
• DNA vaccine plasmid Flt3L plasmid MIP-1a
• Mice challenged at week 12 with 107 pfu rVac-Env
• On day 7 following viral challenge, mice
  sacrificed to quantitate vaccinia virus titers

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Plasmid MIP-1a and Flt3L Augment Anamnestic
TetramerCD8 T Lymphocyte Responses Following
Vaccinia-Env Challenge
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Plasmid MIP-1a and Flt3L Augment Protective
Efficacy of DNA Vaccine Against Vaccinia Challenge
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Conclusions

• A critical limitation of DNA vaccines may be the
  availability of sufficient DCs at the site of
  antigen production
• This limitation can be overcome in mice using
  plasmid MIP-1a and Flt3L adjuvants
• Recruit and activate large numbers of DCs at the
  site of inoculation and antigen production
• Synergistically augment DNA vaccine
  immunogenicity and improve protective efficacy
  against a vaccinia challenge
• Vaccine studies testing this hypothesis in
  nonhuman primates in progress

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Acknowledgements

• Beth Israel Deaconess, Harvard Medical School
• Shawn Sumida
• Diana Truitt
• Michael Kishko
• Janelle Arthur
• Michael Seaman
• Ayako Miura
• Birgit Korioth-Schmitz
• Shawn Jackson
• Darci Gorgone
• Michelle Lifton
• Norman Letvin
• Raphael Dolin

• St. Georges Hospital, London
• Paul McKay
• Vaccine Research Center, National Institutes of
  Health
• Zhi-yong Yang
• Wing-pui Kong
• John Mascola
• Gary Nabel
• HIV Vaccine Trials Network, National Institutes
  of Health