Attached files
| file | filename |
|---|---|
| 8-K - 8-K - ZIOPHARM ONCOLOGY INC | d728674d8k.htm |
| EX-99.1 - EX-99.1 - ZIOPHARM ONCOLOGY INC | d728674dex991.htm |
| INTRATUMORAL REGULATED
EXPRESSION OF IL-12 AS A
GENE THERAPY APPROACH
TO IMMUNOTHERAPY
John
Nemunitis
1
,
John
A.
Barrett
2
,
Francois
Lebel
2
,
Thomas
D.
Reed
3
,
E.
Antonio
Chiocca
4
,
Antonio
M.
Omuro
5
,
Larry
Norton
5
,
Jonathan
Lewis
2
1
Mary
Crowley
Cancer
Research
Centers,
Dallas,
TX,
United
States,
75201
2
ZIOPHARM Oncology Inc., Boston, MA, United States, 02129
3
Intrexon Corporation, Germantown, MD, United States, 20876
4
Brigham and Woman’s Hospital, Harvard Medical School, Boston, MA, United States,
02115 5
Memorial Sloan-Kettering Cancer Center, New York, NY, United States, 10065
Exhibit 99.2 |
| Background & Rationale IL-12 in Glioma
•
Tumors escape the immune system through
the process of immunoediting. Thus,
restoration of the immune system’s ability to
detect the tumor should result in improved
treatment outcomes.
•
Localized IL-12 administration has been
shown to have antitumor activity that is
mediated by direct tumor cell cytotoxicity, and
enhancement of immuno-regulatory activities
including activation of anti-tumor natural killer
(NK) cells, CD4
+
T cells and CD8
+
T cells. |
| Background & Rationale IL-12 in Glioma
•
Roy
&
Kranz
(University
of
Illinois)
IL-12.
Journal
of
Immunology,
2000,
165:
7293–7299.
Model
SV11
Transgenic
mouse
administered
recombinant
mIL-12
i.c.
Findings
Increased
survival
infiltration
of
activated
CD8
and
CD4
T
cells.
•
Vom
Berg
&
Becher
(University
of
Zurich)
J
Exp
Med,
2013,
210:
2803-2811.
Model:
GL261
transduced
to
constitutively
express
IL-12
i.c.
Findings:
Increased
survival
combination of IL-12 + CTLA4 elicits decrease Tregs while increasing Teff.
•
Dimeco
&
Olivi
Johns
Hopkins
School
of
Medicine
&
Istituto
Nazionale
Tumori,
Milan,
Italy)
J
Neurosurg
2000,
92:419–427,
Model:
Rat
9L
gliosarcoma
cells
expressing
IL-12.
Findings
Local
delivery
of
IL-12
in
rat
brain
prolongs
survival
in
animals
challenged
i.c.
with
a
malignant
glioma
cells.
•
Sonabend
&
Lesniak
(University
of
Chicago).
Anti-Cancer
Drugs
2008,
19:133–142.
Model:
GL-261
orthotopic
glioma
model.
Findings:
Synergy
in
survival
with
locally
administered
pmIL-12/PPC
+
biodegradable
carmustine
•
Markert
&
Whitley
(University
of
Alabama)
Journal
of
Virology
2012,86:
5304–5313
Model:
4C8
glioma
cells
orthotopic
B6D2F1
mouse.
Findings:
Hsv
mutant
M002
expressing
IL-12
demonstrated
prolonged
survival
vs.
control
•
Liu
&
Yu
(Cedars
Sinai
Medical
Center)
Cancer
Gene
Therapy
(2002)
9,
9–15
Model:
GL-26
orthotopic
mouse.
Ad5-mIL-12
Findings:
Survival
was
significantly
prolonged
in
Ad-mIL-12–treated
animals
with
increased
CD4+
and
CD8+
T-
cell
infiltration
(
:
:
: |
 1.
The
Switch
Components:
The
RTS®
gene
program
includes
2
receptor
protein
fusions:
VP16-RXR and Gal4-EcR. They form unstable and unproductive heterodimers in the
absence of any ligand.
2.
The
Inducible
Promoter:
A
customizable
promoter
to
which
basal
transcription
proteins
are recruited and the target gene is transcribed.
3.
The
Activator
Ligand
(veledimex):
An
ecdysone
analog,
diacylhydrazine-based
small
molecule functions as an activator. In the presence of the ligand, the protein
heterodimer changes to a stable conformation and binds to the inducible promoter.
EcR
RXR
VP16
Gal4
Basal
Transcription
Proteins
Activator
Ligand
RXR
VP16
Gal4
EcR
Inducible Gene Program
Inducible Gene Program
RheoSwitch Therapeutic System®
(RTS®) is a 3-component transcriptional regulator
Inducible Gene Regulation: RheoSwitch Therapeutic System
®
|
 IL-12 Production is Modulated by Activator Ligand in
HT 1080 Cells
5
On
Off
On
AL=75nM
5 |
 Dose-Dependent Increase in Expression of Tumor IL-12
mRNA & IL-12 Protein in Response to Veledimex
10
2
10
10
4
10
5
10
6
0
2
4
6
8
10
12
14
RTS gDNA
Tumor Veledimex Level
10
0
10
1
10
2
10
3
0
2
4
6
8
10
12
14
Time (days)
Time (days)
Vehicle/Vehicle
Vehicle/Veledimex
150
mg/m
Ad (1e10) + Veledimex 15 mg/m
Ad (1e10) + Veledimex 30 mg/m
Ad (1e10) + Veledimex 75 mg/m
Ad
(1e10)
+
Veledimex
150
mg/m
2
Tumor IL-12 Protein Level
IL-12 RNA
0.1
1
10
0
2
4
6
8
10
12
14
Time (days)
10
0
10
1
10
2
10
3
10
4
0
2
4
6
8
10
12
14
Time (days)
2
2
2
2
3 |
 Ad-RTS-mIL-12 + Veledimex Increases Tumor
CD8
+
& CD4
+
While Decreasing CD4
+
Fox P3
+
TILs
in the 4T1 Syngeneic Mouse
Vehicle
Ad-RTS-mIL-12
1 x 10
10
vp
+ Veledimex
150 mg/m
7
CD8
+
CD4
+
CD4
+
Fox P3
+
2 |
 Dose-Dependent Anti-Tumor Activity of Ad-RTS-
mIL-12 + Veledimex (AL) in Murine 4T1 Model
Start of treatment
Tumor
volume
reached
100-200
mm
3
0
4
8
12
16
20
24
28
32
0
5
10
15
Vehicle/Vehicle
Vehicle/Ad-RTS-mIL12
Vehicle/Veledimex
15
mg/m
2
Ad-RTS-mIL12
+
AL
15
mg/m
2
Ad-RTS-mIL12
+
AL
30
mg/m
2
Ad-RTS-mIL12
+
AL
75
mg/m
2
Ad-RTS-mIL12
+
AL
150mg/m
2
Time (Days) |
| •
High expression of IL-12 mRNA in tumors, tightly controlled by
veledimex dose
•
Tumor
biopsies
show
increased
tumor
infiltrating
lymphocytes
in
both
injected and systemic non-injected lesions
•
Serious adverse events are mechanism-based and consistent with
immunotherapy (Fever, N&V, leukopenia, increased LFTs,
hyponatremia, cytokine release response)
•Serious adverse events reversed within days after stopping veledimex
dosing
•Subjects
who
have
had
IL-12
expression
turned
“off”
have
been
redosed, and IL-12 turned “on”
again
Clinical Observations to Date
9
We can control gene expression to achieve a systemic immune
response
We have seen systemic and fully reversible toxicity |
 Higher
Veledimex Levels Normal and in GL261 Orthotopic Glioma Mouse Brains
Veledimex levels at 24 hr posttreatment
0
1000
2000
3000
4000
5000
6000
450 mg/m
2
/day
1200 mg/m
2
/day |
 Effects of Ad-RTS-mIL-12 + Veledimex (AL)
in the Orthotopic GL261 Mouse
Normal Mouse
Vehicle
BID x 14
Ad-RTS-mIL-12 1x10
10
vp
+
AL
450
mg/m
/day
BID x14
Treatment For 14 days
Day 74 (end of study)
Control Day 20
2 |
 Ad-RTS-mIL-12 + Veledimex (in Chow)
Results in Increased Survival in the GL261
Orthotopic Glioma Mouse Model
Veledimex administered ad lib in chow from Day 4 to EOS at ~ 675 mg/m
/day
Ad-RTS-mIL12 administered on Day 5
0
20
40
60
80
0
20
40
60
80
100
Vehicle/Vehicle
Vehicle/AD (5e9)
Vehicle/Activator chow (1000)
Activator chow + AD (5e9)
Activator chow + AD (1e9)
Activator chow + AD (1e8)
Time (Days)
2 |
 Ad-RTS-mIL-12 + Veledimex (AL) Results in
Increased Survival When Compared to Control
in the GL261 Orthotopic Glioma Mouse Model
Ad-RTS-mIL12
administered
on
Day
5
;
Veledimex
(mg/m
2
)
administered
BID
for
14
days
from
Day 5;
0
20
40
60
80
0
10
20
30
40
50
60
70
80
90
100
maisine/saline
maisine/vector
Veledimex (AL)/saline
AL 150/day + AD 1e10
AL 300/day + AD 1e10
AL 450/day + AD 1e10
AL 675/day + AD 1e10
AL 1200/day + AD 1e10
dexamethasone 6 bidx14
bevacizumab 30 biwkx3
temozolamide 300Qdx5
Time (Days) |
 Ad-RTS-mIL-12 + veledimex significantly
reduces brain cancer stem cells
in GL-261
Orthotopic Glioma Model
Nestin levels (marker for cancer stem cells)
inverse correlation with survival (Pearson r= 0.92)
0
10
20
30
Vehicle/Vehicle
Vehicle/Ad-RTS-mIL-12
Vehicle/Veledimex
Veledime
x 450 mg/m
/day
Veledime
x 600 mg/m
/day
Veledime
x 1200 mg/m
/day
Nestin
2
2
2 |
| 15
Conclusions
•
Ad-RTS-mIL-12 + veledimex PO exhibits controllable
systemic immune activation in human subjects with
melanoma and breast cancer.
•
Veledimex exhibits dose-related increases in plasma and
brain tissue exposure with no accumulation in brain.
•
Ad-RTS-mIL-12 (1x10
10
vp) + veledimex PO improves
survival over temozolomide, dexamethasone and
bevacizumab.
•
Ad-RTS-mIL-12 + veledimex significantly reduces brain
cancer stem cells
•
These findings support the utility of localized, regulatable IL-
12 production as an approach for the treatment of malignant
glioma in human subjects. |