Inducing Antibodies with Rationally-designed

HIV Vaccines

Susan Zolla-Pazner, Ph. D.New York University School of Medicine

Department of Pathology

• Sequence and antigenic diversity• Conformational masking of critical

epitopes• Evolution to escape the effects of Abs • Poor induction of Abs with broad

anti-viral functions• Inability as a vaccine reagent to

induce a long-lived Ab response (<6 months)

Problems with Whole Env Immunogens

Antigenic Determinants on the HIV Envelope

Modified from D R Burton, R A Weiss Science 2010;329:770-773Published by AAAS

V3 loop

gp41gp120

Structural Vaccinology Approach• Required steps:

– Generate neutralizing monoclonal Abs– Select mAbs with broad reactivity– Crystallize mAbs– Analyze bioinformatics data– Model the epitope– Design and generate recombinant epitope-scaffold immunogens– Immunize animals– Assess immune response

StudyAb

Study Epitope

In vivoStudies

Structural Vaccinology Approach• Required steps:

– Generate neutralizing monoclonal Abs– Select mAbs with broad reactivity– Crystallize mAbs– Analyze bioinformatics data– Model the epitope– Design and generate recombinant epitope-scaffold immunogens– Immunize animals– Assess immune response

StudyAb

Study Epitope

In vivoStudies

Structural Vaccinology Approach• Required steps:

– Generate neutralizing monoclonal Abs– Select mAbs with broad reactivity– Crystallize mAbs– Analyze bioinformatics data– Model the epitope– Design and generate recombinant epitope-scaffold immunogens– Immunize animals– Assess immune response

StudyAb

Study Epitope

In vivoStudies

Crystallographic Analysis of Anti-V3 mAbs Complexed with V3 Peptides

V3/mAb 447:“Ladle-like” V3 binding

V3/mAb 2219:“Cradle-like” V3 binding

(V. Burke et al, Structure, 2009)

Structural Vaccinology Approach• Required steps:

– Generate neutralizing monoclonal Abs– Select mAbs with broad reactivity– Crystallize mAbs– Analyze bioinformatics data– Model the epitope– Design and generate recombinant

epitope-scaffold immunogens– Immunize animals– Assess immune response

Study Epitope

Two-thirds of the 35 Residues in V3 are Conserved

Structural Vaccinology Approach• Required steps:

– Generate neutralizing monoclonal Abs– Select mAbs with broad reactivity– Crystallize mAbs– Analyze bioinformatics data– Model the epitope– Design and generate recombinant

epitope-scaffold immunogens– Immunize animals– Assess immune response

Study Epitope

A

The Conserved Structure of the V3 Crown

Almond et al., ARHR 2010. Jiang et al., Nature Struct. Mol. Biol., 2010

Hydrophilic faceof circlet

Band Arch

Hydrophobic faceof circlet

Structural Vaccinology Approach• Required steps:

– Generate neutralizing monoclonal Abs– Select mAbs with broad reactivity– Crystallize mAbs– Analyze bioinformatics data– Model the epitope– Design and generate recombinant

epitope-scaffold immunogens– Immunize animals– Assess immune response

Study Epitope

Design of Recombinant V3-scaffold Immunogen

M. Totrov et al., Virology, 2010.

Example: V3-Cholera Toxin B

Structural Vaccinology Approach• Required steps:

– Generate neutralizing monoclonal Abs– Select mAbs with broad reactivity– Crystallize mAbs– Analyze bioinformatics data– Model the epitope– Design and generate recombinant epitope-

scaffold immunogens– Immunize animals– Assess immune response

Immunization Protocol

6 weeks 6 weeks 4 weeks

2 weeksPost-boost

P1 P2 P3 B1 B2

gp120 DNA prime V3-CTB boost

Pre-bleed

Tier 1A 1A 1B 1B 1B 1B 1B 1B 1BClade C B B B B C B AG

C

Virus MW965 SF162 BaL Bx08 BZ167 TV1.21 S1196 T271-11 25710CTBwt O O <10 O ND ND <10 ND ND

B O O O O ND O O O ND

3074 O O O O ND O O O ND

C O O O O O O O O OB+3074 O O O O ND ND O <10 ND

C+3074 O O O O O O O O O

50% Neutralizing Ab Response vs. Tier 1 Viruses

1:10-99 1:100-999 1:1000-9999 >1:10,000

O 1/5Responders: O2/5 O O O3/5 4/5 5/5

Clade B B C C B C C C B

Virus 6535 RHPA259.7

ZM135M ZM233M WITO160.33

Du156.12

CAP210 ZM109 QH0692

CTBwt <10 <10 <10 <10 O <10 O O <10

B <10 <10 <10 <10 <10 <10 <10 <10 O3074 O <10 <10 O <10 <10 O O <10

C O <10 O <10 <10 <10 O O <10

B+3074 <10 <10 O O O O O O O

C+3074 O <10 O <10 <10 <10 O O <10

50% Neutralizing Ab Responses vs. Standard Tier 2 Panel of Clade B and C Viruses

NT50 = 1:10-99O 1/5Responders: O2/5 O O O3/5 4/5 5/5

Neutralizing Abs are Detectable 60 Weeks after the Last Boost

6 12 16 25 58 68 760

20

40

60

80

100

% N

eutr

aliz

atio

n

Weeks

Post 3rd

PrimePost 1st

BoostPost 2nd

Boost

D %

Neu

tral

izat

ion

vs. B

x08

Weeks

Antigenic Determinants on the HIV Envelope

Modified from D R Burton, R A Weiss Science 2010;329:770-773Published by AAAS

gp41gp120

V2 loop

Functions of the V2 Loop–Not essential for infectivity

–Binds to α4β7 integrin on activated T cells

–With V3, protects the chemokine receptor binding site

• Use the same structural vaccinology approach as used for V3.

• Identify “hidden” conserved structure within the 2nd variable loop.

• Engraft this generic structure into a scaffold.

• Use V2-scaffold as a boost to elicit cross-reactive V2 Abs with multiple anti-viral functions.

Planned Design of V2 Immunogens

V2

V3

(S. Zolla-Pazner and T. Cardozo, 2010)

V2 and V3 are Similar in Their Patterns of Amino Acid Conservation

V2

V3

• The structural vaccinology approach has succeeded in inducing cross-clade neutralizing Abs based on the use of a gp120 DNA prime and a V3-scaffold protein boost.

• The development of the boost was the result of revealing a generic conserved structure with the third sequence “variable region”.

• This prime/boost vaccine approach can focus the Ab response on selected epitopes.

• Neutralizing Abs were detectable >1 year after the last boost.

Conclusions: #1

• More than one epitope needs to be targeted for an effective vaccine.

• The same principles that guided the successful development of the V3-scaffold immunogens are being applied to V2, and can ultimately be applied to more complex epitopes (QNE, CD4bs, etc.)

Conclusions: #2

CollaboratorsNYU School of

Medicine Mirek GornySandy Sharpe

CohenConnie Williams Barbara Volsky

Xiang-Peng KongXunqing Jiang

Tim O’NealTim Cardozo

David AlmondyJames Swetnam

Suman LaalPhillipe Nyambi

Valicia BurkeXunqing Jiang

Higuang LiJared SampsonBrett SpurrierApril Killikelly

University of Massachusetts School of Medicine

Shan LuShixia Wang

Molsoft, Inc.Max Totrov

Ruben Abagyan

Harvard Medical SchoolMichael Seaman

NYU Medical Center