7/14/2015
HIFU, Drug Delivery, and
Immunotherapy
Katherine W. Ferrara
UC Davis
Minimally toxic activatable particle
Treat accessible lesions
Big hairy audacious goal (BHAG)
Immune therapy is effective for
melanoma, emerging as effective for
many cancers
• FDA-approved
– Interleukin-2 (RCC, melanoma)
– CTLA-4 antibody (melanoma)
– PD-1 Ab (melanoma)
Durable response in
significant (15%) of patients
• Investigational
– CpG as adjuvant to radiotherapy (melanoma,
companion animals)
– ..
1
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Complexation of Cu(II) and Dox within
liposomes
Dox relative Fl (%)
Problem: Even liposomal doxorubicin has substantial
cardiac toxicity and dose cannot exceed 500 mg/m2 in
lifetime.
Solution: Create a doxorubicin salt that is very stable in
circulation
***
100
Ammonium sulfate Copper gluconate/
TEA method
method
**
50
7
6
5
4
3
**
0
3
5 7
pH
8
pH
Kheirolomoom et al Molecular Pharmaceutics
Outline
Activatable particles
+ hyperthermia
Activatable particles +
hyperthermia +
adjuvants
Particles + ablation
+ adjuvants
Temperature sensitive particles
paired with mild heating
Alter
microenvironment
Cell
Achieve
Inject
death,
and
hyperthermia
release
M1 macrophagesdrug 0-40 minutes
24 hourslater
and later
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Thermally-activated vehicles
CuDox-LTSLs
40 min US
Above Tc
Below Tc
Heart
USTumor
US+
DSPE-PEG-2000
DPPC
Lyso-palmitoyl PC
Drug

Dox
• Direct and local release can
increase concentration and
decrease systemic toxicity
• Small molecule can penetrate
tissue
Heart
Tissue
Autofl.
US-
US+
CuDox-LTSLs
20 min US
Kheirolomoom et al, JCR 2013
CuDox-lipo
Copper-doxorubicin
temperature-sensitive
particles
100 nm
Conventional temperaturesensitive particles
T u m o r + U S / M u s c le
6
D o x o r u b ic in f lu o r e s c e n c e r a t io
D o x o r u b ic in f lu o r e s c e n c e r a t io
Cu-dox temperaturesensitive particles
4
*
2
0
C u D o x -L T S L s
AS D o x -L T S L s
4
Particles
US
M u s c le + U S / T u m o r
*
3
2
1
Increasing
Dox
0
C u D o x -L T S L s
AS D o x -L T S L s
Tissue
Autofl
CuDox-TSL (8 treatments)
Control
Control+US
CuDox-LTSLs
CuDox-LTSLs+US
4500
3500
2500
***
~
~
500
***
1500
-500 0
US
*
Tumor growth, %
5500
10 20 30 40 50 200
250
Day post treatment 120
*
***
p<0.05 compared to control
P<0.001 compared to control
Survival, %
100
Treat 2x/week, 4 weeks,
6 mg/kg
80
Control
60
CuDox-LTSLs
40
CuDox-LTSLs+US
20
0
0
Kheirolomoom et al, JCR 2013
50
100
150
200
Day post treatment
250
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7/14/2015
1 mm
1 mm
1 mm
Days
2 Weeks
100 µm
4 Weeks
100 µm
100 µm
(F4/80)
(Fe)
100 µm
Kheirolomoom et al, JCR 2013
Why do we favor thermally-sensitive
nanoparticles?
%ID delivered
depends on volume insonified,
time of insonation
100
Diameter
10
0.5 cm
1 cm
2 cm
4 cm
1
%ID
0.1
0.01
0.3
0.7
1.0
2.0
4.0
Time insonation (hrs)
Assumes 5% blood volume in tumor
10 sec tumor blood refresh
5 L blood volume
Weight change, %
*
20
15
10
5
0
-5 0
-10
Control
CuDox-LTSLs
CuDox-LTSLs+US
5
10 15 20 25
Day post treatment
Blood cell count
25
30
25
30
CuDox-LTSLs
20
Control
15
10
5
***
0
WBC, K/µL
RBC,M/µL
Toxicity is low
1600
10
Organ weight, mg
Protein, g/dL
CuDox-LTSLs
8
Control
6
4
2
1200
CuDox-LTSLs
Control
800
400
0
0
Albumin
Total protein
4
7/14/2015
Outline
Activatable particles +
hyperthermia +
adjuvants
Activatable particles
+ hyperthermia
Particles + ablation
+ adjuvants
Use of TLR agonists to enhance Teffector
cell function
Tumor
Tumor
Tumor
antigen
iDC
Teff
Treg
IFN-g
IL-2
IFN-a
TLR
activation
TNF-a
Teff
IL-12
TAM
MDSC
Mature
DC
Th1
IL-12
Macro
DC: Dendritic cell; iDC: Immature dendritic cell; MDSC: Myeloid-derived suppressor cell;
Macro: Monocyte/macrophage; TAM: Tumor-associated macrophage; Teff: Effector T cell;
Treg: Regulatory T cell, US: ultrasound
adapted from Immunotherapy. 2009 November 1; 1(6): 949–964
20000
C o n tr o l
18000
CpG
C p G C o n t r a la t e r a l
16000
C uD ox+U S +C pG
14000
C u D o x + U S + C p G C o n t r a la t e r a l
12000
10000
***
8000
**
6000
4000
*
N D L T u m o r g r o w t h (% )
Copper-doxorubicin temperature-sensitive
particles + CpG (3 treatments)
2000
0
0
5
10
15
20
25
30
35
40
45
50
D a y p o s t tre a tm e n t
5
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Abscopal response
CuDox+US+CpG
CuDox+US+CpG
Contralateral
CpG
CpG
Contralateral
Control
CuDox
2 mm
200 µm
3
MDSCs
Immune cell
populations
2
1
*
*
*
*
*
% IFN-g+
CD8+ T cells of Leukocytes
0
2
**
1.6
**
1.2
0.8
0.4
0
Outline
Activatable particles
+ hyperthermia
Activatable particles +
hyperthermia +
adjuvants
Particles + ablation
+ adjuvants
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Long circulating
nanoparticles
Ablation + 64Cu-LCL
6 hours
20 hours
48 hours
25 % ID/cc
0% ID/cc
Optical Density
(Arbitrary Units)
1cm
1
20
Circle #1
Circle #2
Circle #3
No Tx
0
0
2
4
Location (mm)
6
8
7
7/14/2015
Ablate fraction of tumor + drug
No Tx
Single
Circle
1.4
Tumor Volume (cc)
1.2
LCL
Single + LCL
Circle + LCL
1
0.8
2 mm
0.6
4 mm
0.4
0.2
0
0
50
100
150
Days after First Treatment
4 Protocols, 100% Curative for
local disease
Survival %
100%
Hyp+TSL+Circle
75%
TSL+Circle
50%
TSL+Single
25%
No Tx
0%
Survival %
0
100
200
Days After First Treatment
100%
NT
75%
LCL
Single
50%
Single + LCL
25%
Circle
0%
0
40 80 120 160
Time (days)
Circle + LCL
• Hyperthermia +
TSL in 8 treatments
• Hyperthermia+TSL
+ CpG in <3
treatments
• Hyperthermia +
Ablation + TSL in 3
treatments
• Ablation + LCL
Wong et al, in review
Curative Protocols
Drug
LCL
N/A
<100% cure
<100% cure w/CpG
100% cure w/o CpG
100% cure w/ CpG
TSL
None
N/A
None
Ablation
Hypertherm
Comb
US
8
7/14/2015
1 Protocol- extending survival in
systemic disease, needs
optimization
120
S u r v iv a l ( % )
100
80
p=0.02
60
• Hyperthermia+TSL
+ CpG in <3
treatments
40
C o n tr o l
CpG
C uD ox+U S +C pG
20
0
0
4
8 12 16 20 24 28 32 36 40 44 48 52 56 60 64
D a y p o s t tr e a t m e n t
Kheirolomoom et al, in review
Conclusion
• Imaging essential in developing new
nanotherapies, identifying cell populations,
evaluating efficacy
• Activatable nanotherapies can cure local
disease, enhance systemic therapy
• Addition of immune adjuvants may allow
the accessible lesions to be treated,
resulting in systemic benefits
NCI and NIBIB
Charles Caskey
James Chomas
Paul Dayton
Lisa Even
Brett Fite
Karen Gagnon
Xiaowen Hu
Elizabeth Ingham
Sarah Johnson
Azadeh Kheirolomoom
Dustin Kruse
Jiro Kusunose
Chun-Yen Lai
Yu Liu
Aaron Lum
Karen Morgan
Eric Paoli
Shengping Qin
Jai Woong Seo
Susanne Stieger
Kate Watson
Andrew Wong
Guodong Zhang
Hua Zhang
U Bergen Frits Thorsen, Cecilie Brekke, Rolf Reed
UCD John Boone, Sandy Borowsky, Bob Cardiff,
Nipavan Chiamvimonvat, Roy Curry, Greg Farwell,
Bruce Hammock, C. Lebrilla, Claude Meares, Jinyi Qi,
Tingrui Pan, Scott Simon, C. Tepper, So Yamada
LLNL Paul Hoeprich
G Tech/Emory Hanjoong Jo
Northwestern Tom Meade
Burnham/UCSB Juliana Hamzah, Erkki Ruoslahti
UCB He Dong, Ting Xu
Cedars Sinai Gadi Palled, Dan Gazit, Galina Shapiro
CMGI (Dave Kukis)
Pfizer Oncology
Targeson (Jack Rychak)
Image-Guided Therapy (Erik Dumont)
Siemens Medical Solutions
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7/14/2015
Free dox
Before
0
1
5
2
15 min of heat
5 min
10 min after
heat ends
Dox-TSL
Manzoor AA et al, Cancer Res. 2012 Nov 1;72(21):5566-75.
Dewhirst laboratory, Duke University
Intensity map: (0, 0, 55) mm in x-y plane
Intensity map: (0, 5, 55) mm in x-y plane
Thermal effects are very significant
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7/14/2015
Thermal and mechanical
oversight are feasible
MRgFUS insert from
Image Guided Therapy
7T, 3 MHz ultrasound in
Bruker magnet
Fite BZ et al PLoS ONE 7(4): e35509
°C
0.5
4
1
3.5
1.5
With probe motion
3
2
2.5
Heating is ON
from 0 to 20 s
2.5
3
2
3.5
1.5
4
4.5
0.5
1
1.5
2
2.5
3
3.5
4
4.5
1
Foiret et al, submitted
US+ temperature sensitive
liposomes (TSL) or long circulating
liposomes (LCL)
• US hyperthermia+TSL= local cure (100% in 8
treatments)
• US hyperthermia+CpG+TSL= local cure (100%
in 3 treatments) + slow systemic disease
– Needs optimization, many alternatives
• US ablation + LCL or TSL, slow disease or cure
11
7/14/2015
After ablation
4mm
200 µm
200 µm
200 µm
Canine melanoma - Combined radiotherapy
and immunotherapy using CPG
oligodeoxynucleotides and indolamine 2,3
dioxygenase (IDO) blockade
Monjazeb, et al J Immunother Cancer. 2013; 1(Suppl 1): P256.
Why nanoparticles? Why ultrasound?
 Target accumulation
20
18
16
14
12
%ID/g 10
8
6
4
2
0
Paoli et al, JCR 2010
12
7/14/2015
Ultrasound used to guide hyperthermia
-e.g. Multi-frequency arrays
64 elements at 1.5 MHz
0.8 mm pitch, 38.4 mm ; 5mm element.
Therapy
Imaging
128 elements at 5 - 7 MHz
0.27 mm pitch, 38.4mm ; 4mm element
24 mm
43 mm
Therapy
Stephens
et al
PMB
2008,
IEEE
UFFC
2011
MRfUS
Image-Guided Therapy (IGT) system
•
•
•
•
•
•
•
•
•
16 element annular array
Diameter: 45mm
Radius of curvature: 35mm
Aperture: 85 degrees
Operating frequency: 3MHz
Acoustic efficiency > 65%
Power rating: 50Wac
Bandwidth: 300KHz.
Focal spot size (approx.) : 1.5mm x 1.5mm x 2mm
Copper-doxorubicin
temperature-sensitive
particles + CpG
CuDox-lipo
100 nm
Temperature-sensitive particles
+ ultrasound= local cure
Temperature-sensitive particles
+ ultrasound + adjuvant = systemic cure
Particles
US
Adjuvant
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7/14/2015
Day 7
Foxp3+ (Treg) cells
Contralateral tumors- in rim
Directly treatedNone in rim or tumor
Tumor
Lymph node
14