DNA Vaccines to Suppress Myeloma
B cell
CD4+ T cell
CD8+ T cell
CD4+ T cell
Molecular Immunology Group
Tenovus Laboratory
University of Southampton UK
Success of passive immune attack in human cancer
Anti-CD20 in lymphoma
Anti-HER2 in breast cancer
B cell
CD4+ T cell
CD8+ T cell
CD4+ T cell
Graft vs leukemia effect
In allotransplantation
Powerful cytotoxic T cells can specifically eliminate tumor cells in vivo
Antigen +ve (green)
tumor cells are killed
killed
by transferred CTL
Biossonnas A et al JEM 2006
Reduction in metastatic melanoma
following lymphodepletion and adoptive transfer of tumor-infiltrating lymphocytes
CT scans of liver
Gattinoni L, Rosenberg SA, Nat Rev Immunol 2006
Is vaccination appropriate for myeloma?
Patients can achieve remission
and recover immune capacity
Advantages:
iMIDs widely used and may
increase immune respo
onses
AlloSCT donor can be vaccinated
Disadvantages:
Incurable in majority of patients
Patients take time to regain immune
so more support for new approaches
capacity and tumor may recur
Idiotypic Ig provides disease marker
Only 50% of patients have HDT/ASCT
and alloSCT rare
Wide range of tumor antigens
including idiotypic Ig
Ig
Tumor is highly
Tolerance unlikely
immunosuppressive
Tumor is accessible to T cells
T-cell responses against TT vaccine in myeloma patients post ASCT
For vaccination against myeloma to succeed:
Clinicians:
1. Tumor load must be reduced with retention of immune capacity.
Could follow chemo/radiotherapy/transplantation.
Could also follow passive MoAb or adoptive T-cell transfer.
Transplant donor could be vaccinated.
Immunologists:
2. Generation of effective immunity will require activation of
innate immunity and induction of T-cell help.
Triple action DNA fusion gene vaccines can induce specific immune attack
Antigen
Microbial sequence
1
CpG
Activates innate immunity
Triple action DNA fusion gene vaccines can induce specific immune attack
Antigen
Microbial sequence
2
Molecular form to elicit
antibody, CD4+ T cells, CD8+ T cells
1
CpG
Activates innate immunity
Triple action DNA fusion gene vaccines can induce specific immune attack
Antigen
Microbial sequence
2
Molecular form to elicit
3 Activates T-cell help
help from
from a large
antibody, CD4+ T cells, CD8+ T cells
undeleted repertoire
Breaks tolerance
1
CpG
Activates innate immunity
What has been achieved so far?
Ig variable regions
scFv
FC
FrC f
o tetanus toxin
VH
VL
B-cell lymphoma
CpG
Single chain Fv-Fragment C fusion genes induce protection against lymphoma
in two mouse models
scFv
FrC
A31 lymphoma
BCL lymphoma
1
280
260
240
ps
p. FvBcl
sFvBcl-1
- -
1 FrC
-
)
220
-3
01
200
xl
p.sFvA31-FrC
180
/m
(U
160
selv 140
le
120
-Idtin 100
A
80
60
40
20
0
XX
XX
X
XXX
p
FA
.scFv 31
A31 FC
-.FrC
p.scFvA31
King et al Nat Med 1998
Safety/feasibiity DNA fusion vaccine trial in patients with lymphoma: LIFTT
scFv
FrC
No significant side effects
Immune responses of 18 patients analyzed [FrC]
1
16 patients analyzed [Idiotype]
Fragment C
Idiotypic antigen
_________________________________________________________________
Ab
CD4+ T cell
Ab
CD4+ T cell
____________
__________
+
/
+ -
Tt
Total
%
+
/
+ -
Tt
Total
%
_________________________________________________________________
[7]
[]
11
2
13
72
2
2
3
6
38
_________________________________________________________________
Idiotypic antigen: individual recombinant scFv-C
A Mander, C Ottensmeier et al, in preparation
What about multiple myeloma?
Secretes Ig
DNA scFv-FrC vaccine induces specific protective immunity
py in 5T33 model
100
p.scFv5T33-FrC
80
al 60
ivrvus 40
%
p.scFv5T33
CFA
20
alone
non-vaccinated
5T33 Ig
IgG/CFA
F
0
20
30
40
50
60
70
Day
King et al Nat Med 1998
A single tested patient with myeloma post autograft showed responses
Now 15 patients recruited, 9 vaccinated
450
400
Vaccinations
350
PBMNC)
300
6 0
250
0
200
150
100
(spots/1T
50
O
0
O
Pre
0
10
20
30
40
150
ELISP-
100
N
50
IF
0
Pre
010
20
30
40
800
700
Abs
Patient Idiotype
600
500
Control Idiotype
400
300
Fragment C
200
100
Anti-FrC
0
Pre
0
10
20
30
Weeks from vaccination
Week 0 = 39 weeks post transplant
Modified strategies for delivering DNA vaccines
Probl
blems of translating principles of DNA vaccit
cina i
tion establi
bli h
s ed in
i
m ce to human subj
bj
t
ec s
DNA vac
ccine
Physical modifications
Recombinant organisms
Microparticles
Vaccinia/MVA
BCG
Fowl pox
Liposomes
Salmonella
Adenovirus
Listeria
Electroporation
Alphavirus replicon vectors
"Prime/boost" strategies
Advantages of physical modifications:
Safe and simple
No problem with pre-existing immunity
Electroporation device for DNA vaccination
Tjelle T et al Vaccine 2005
Electroporation increases antigen expression
in sheep muscle
No EP
EP
Each "needle path" has received 25 g of DNA -gal encoding plasmid
Perfect match between electric field and injection of DNA
Iacob Mathiesen Inovio
Electroporation is effective in "prime/boost" for antibody induction
using DNA scFv-FrC
Buchan et al JI 2005
Electroporation is effective in "prime/boost" for CD8+ T-cell induction
Amplifies responses in rhesus macaques (Otten et al Vaccine 2004)
Succeeds in wide range of large animals and human subjects (DNA Vaccines 2007, Malaga)
Clinical trial in prostate cancer using DNA vaccine against PSMA
So far: well tolerated and immune responses generated
Appear increased following EP [C Ottensmeier]
ARM A - DNA
0
1
2
6
12
18
months
Crossover allowed
depending on immune/
clinical responses
0
1
2
6
12
18
months
ARM B - DNA + EP
Vaccine designs to activate Th-dependent specific pathways
Outcome
Sf
Surface
or secreted
Dom1
Dom2
Antibody, CD4+ T cells
antigen
Contains competitor MHC Class I-binding epitopes
Surfac
ce or
Antibody, CD4+ and CD8+ T cells
Dom
intracellular
antigen
Dom
Higher levels of epitope-specific
CD8+ T cells
Target MHC I-bound peptide
Rice et al JI 2001
Rice et al JI 2002
Stevenson et al PNAS 2004
Stevenson et al Imm Rev 2004
Stevenson and Rice EJI 2006
Ta
T rget
a
antigens in myeloma
m for CD8+ T--cell
cell attack
Viral antigens?
Mutated proto-oncogene peptides
Cancer-testis antigens:
MAGE-A2/3/NY-ESO/PAS-D1
CD8+
Tran
a slocation peptides
Tl
T ce l
ll
Autoantigens: hTERT
CYPIBI
Survivin
Idi t
o
i
yp c
t
pep id
tides
PRAME
Products of genomic instability
Minor histocompatibility antigens
Rice et al Cancer Res 2006
Analysis of cancer-testis antigen expression by RT-PCR, Q-PCR, and
antibody staining in bulk populations and in single MM cells
Comprehensive analysis of expression
patterns [Condomines M et al JI 2007]
MM cells of 98% of patients
%p
express
p
at
least 1 CT gene. 70% at least 3 CT genes
Expression of 6 CT genes is associated
with shorter survival
Sahota, S. S. et al. Blood 2006
DNA fusion gene vaccination against NY-ESO-1 epitope
DOM
NY-ESO-1 epitope
HLA-A2
ELISPOT of HLA-A2 mice immunized with p.DOM-epitope vaccine
600
500
llion
mi 400
per
300
otsp
s
200
IFN
100
0
prime p
pp.DOM
prime p
pp.DOM-epit
p
pprime/boost p.DOM prime/boost p.DOM-epit
Vaccination
"Mix and match" DNA vaccine cassettes to induce immune attack on myeloma
MAGE-A3
Cell surface or secreted protein
Idiotypic Ig
NY-ESO
PAS-D1
Cytotoxic CD8+ T cells
Antibody
HA-
HA 1
CD4+ T
ll
ce s
CMV
Take home messages:
1.
DNA fusion vaccines activate selec
cted immune pathways
able to attack myeloma cells.
2.
Patients with myeloma post
yp
autologous transplantation
gp
are
immune competent and donors of allogeneic transplants can be vaccinated.
3.
DNA idiotypic fusion vaccines induce persistent IFN-producing
CD4+ T-cell responses.
No evidence for deletional tolerance.
4.
More antigens are being tested: CTA/mucins/minor histocompatibility
ti
an gens/CMV.
5. Electroporation is increasing performance of DNA vaccines in patients.
Jason Rice
Jo Roddick
Surinder Sahota
Jane Watkins
Gianfranco di Genova
Juan Campos-Perez
Delin Zhu
Niklas Zojer-Vienna
FF
Francesco Forconi-Siena
Kim Orchard
Orchard
Christian Ottensmeier
Ottensmeier
Feargal McNicholl
Te
T novus
NHS R&D Hope WCT
CANCER RESEARCH UK