The Future Trends in
Molecular Imaging
g g
Dr Juri Gelovani
Dr.
Gelovani, MD,
MD PhD.
PhD
Professor of
Radiology and Neurology
Chairman, Dept. Experimental
Diagnostic Imaging
Director, Center for Advanced
Biomedical Imaging Research
(CABIR)
Molecular And Genetic Imaging of Cancer
E l &A
Early
Accurate
t Di
Diagnosis
i
• To follow Biomarker Screening (blood genomics and proteomics)
• Develop Imaging of Biomarkers (tumor localization, sensitivity)
Tumor Phenotyping
• Tumor Profiling (receptors & signal transduction, invasiveness,
metabolism, proliferation, apoptosis, etc.)
• Stroma Profiling (angiogenesis, tissue rere-organization, etc.)
Imaging for Selection & Monitoring of Therapy
• Drug Target Expression & Activity
• Dose Optimization
• Early Response Evaluation (Rx adjustment)
• Short
Sh
Shortt-term
t
& Long
L
T
Term Prognosis
P
i
• Monitoring Contained/Chronic Disease status
• Early Detection of Relapse
Image-Guided Biopsy, Surgery and Radiotherapy
Image-Guided Biopsy
Image-Guided
Image
Guided
Radiotherapy
Sentinel lymph node mapping in breast carcinoma patients
Sentinel lymph node mapping in breast carcinoma patients
Imaging of angiogenesis in breast tumor
SPECT imaging after injection of 99mTc-RGD peptide
Potential for earlier assessment of response to chemotherapy
• Avoid unnecessary cost and toxicity
• Switch to alternative treatment sooner
Approaches to Optical Imaging of Integrins in Neo-Angiogenesis
SO 3 H
C
Cyclo(RGDfK)-Cy5.5
( G f )C
H O 3S
SO 3 H
SO 3 H
N
N
Gly
A rg
L ys
Our pipeline:
• MMP9
• EGFR
• PDGFR
• IL11Rα
A sp
p he
O
RGD C 5 5
RGD-Cy5.5
6 nmol/mouse
l/
RGD + RGD-Cy5.5
1 hr interval
600 nmoll + 6 nmoll
Cy5 5
Cy5.5
30 nmoll
Structure of 111In-DTPA-K(IRDye800)-c(KRGDf).
Gly
Arg
Asp
O
IRdye800
NH
NH
HN
N
H
O
O
HN
O
H
N
O
HOOC
COOH
N
N
N
HOOC
COOH
COOH
Gamma and near-infrared imaging of 111In-DTPA-K(IRDye800)-c(KRGDf)
in nude mice bearing s.c. human melanoma.
M21 tumor is p
positive in integrin
g
αVβ3
β expression,
p
, while M21-L
tumor is negative in αVβ3 expression.
Selective accumulation of 111In-DTPA-K(IRDye800)-c(KRGDf) in M21
melanoma tumors that express integrin αVβ3.
3
P=0.0082
2
Non-blocked
P=0 54
P=0.54
Blocked
1
0
M21
M21-L
Accumulation of the radiotracer in M21 tumor can be blocked by the parent RGD peptide. On the other
hand, preinjection of the parent RGD peptide do not cause significant reduction in the uptake of 111InDTPA-K(IRDye800)-c(KRGDf)
DTPA
K(IRDye800) c(KRGDf) in αvβ3
αvβ3-negative
negative M21
M21-L
L tumors.
Development of NIR Imaging Probe for Clinical
Applications: RGD Conjugates with TS
TS-ICG
ICG Dyes
SO3-
SO3
SO3-
SO3-
TS-ICG -c(KRGDf)
+
N
N
Et
(CH2)5
CONH-c(KRGDf)
-
SO3-
SO3
SO3-
SO3-
+
N
(CH2)5
CONH-c(KRGDf)
TS-ICG-c(KRGDf)2
N
(CH2)5
CONH-c(KRGDf)
Structures of conjugates of TS-ICG containing one and two
molecules of c(KRGDf) peptide. Ex/Em: 765/796 nm
NIR Imaging using Monomeric and Dimeric
RGD Conjugates
Conj gates
TS-ICG-(KRGDf)2
Chemical structure of L-PG-NIR813 or D-PG-NIR813. L- and DPG-NIR813 differ in their stereoisomeric carbon center (*).
Michaelis-Menten constant s ((Km)) of the reaction of CB on L-PG-NIR813
with L-PG molecular weight of 17 KD (A) and 56 KD (B).
Dose-dependent inhibition of CB-mediated degradation of L-PG-NIR813
by specific CB inhibitor (Inhibitor II).
Degradation
D
d ti
off PG-NIR813
PG NIR813
(8.3% loading) by U87 cells
culture for 24 hr (1) and 0 hr
(2). Culture medium without
cells was used as a control (3).
In vivo imaging after intravenous injection of L-PG-NIR813 into
U87(TGL) nude mice.
NIRF Images were acquired at 24 hr after i.v. injection of PG-NIR813
(50 nmol/mouse, 17 KD, 8.3 % loading). Note the mouse injection
with L-PG-NIR813, but not the mouse injected with D-PG-NIR813,
exhibits strong NIRF signal.
Co-localization of PG-GdDTPA-NIR813 with isosulfan blue (A-D).
Microphotographs of representative resected lymph node confirming the uptake of
PG-GdDTPA-NIR813 in the LN (E-H). E: H&E stained tissue section. F: DIC image.
G NIRF image.
G:
i
H Overlay
H:
O l
off the
th DIC and
d NIRF images.
i
NIRF signal
i
l is
i
pseudocolored green, and DIC pseudocolored red. Original magnification: 50x.
Dual MR/optical imaging of the axial and branchial lymph nodes
in normal athymic nude mice.
(A) Pre-contrast overlay of white light and NIRF images. (B) Overlay of white light and NIRF images 1 hr after injection
of PG-GdDTPA-NIR813 (0.002 mM/L Gd/kg). (C) NIRF image of the same mouse without skin. (D) Resected lymph
nodes showed bright fluorescence signal. E-F: Representative T1-weighted axial MR images at different times. PGGdDTPA-NIR813 was injected at a dose of 0.02 mmol Gd/kg (E) and 0.002 mmol Gd/kg (F). The arrows indicate SLN.
Visualization of cervical lymph nodes after interstitial injection
of PG-GdDTPA-NIR813
PG-GdDTPA-NIR813 (0.02 mmol Gd/kg) was injected into the tongue of a normal mouse (A-D) and a mouse with a
human DM14 oral squamous carcinoma tumor grown in the tongue (E-H). H&E stained lymph node sections (I-K)
indicating the presence of micrometastases (J) and presumably in-transit metastases in lymphatic duct (K). Note
the difference in the pattern of lymph node enhancement between normal (A) and metastatic (E) nodes in T1weighted MRI.
Detection of Tumors by
Molecular Imaging off
Metabolic Shifts
Comparative PET Imaging of Prostate Carcinomas with 18F-FDG and 11C-Ac
PET images of prostate, lymph node, and bone metastases obtained using 18FFDG and 11C-Ac from 73-y-old man with poorly differentiated (Gleason sum 7)
adenocarcinoma of prostate.
(A)
18F-FDG
PET shows low uptake in prostate, with SUV of 2.87.
(B and C) 18F-FDG uptake in left iliac lymph node metastatic lesion (B) and right
pubic bone metastatic lesion (C).
(D–F) 11C-Acetate PET shows high uptake in prostate (D), with SUV of 5.45; in left
iliac lymph node metastatic lesion (E); and in right pubic bone metastatic
lesion (F). bn = bone; ly = lymph node; pr = prostate.
Oyama, et al. J Nucl Med (2002)
Metabolism of Fluoro-Acetate
Aconitase
Glycolysis
FDG
Fatty Acid
Synthesis
F-Acetate
In vitro 3H-Acetate and 14C-Fluoroacetate Dual
Dual--Label Radiotracer
Accumulation Studies in Breast Carcinoma Cell Lines
M B231
SKBr3
6.0
6.0
5.0
3H-Acetic Acid
4.0
C/M Ratio
3.0
Linear (3H-Acetic
Acid)
20
2.0
Linear (14C-FAcetic Acid)
1.0
3H-Acetic Acid
4.0
14C-F-Acetic Acid
14C-F-Acetic Acid
3.0
Linear (3H-Acetic
Acid)
2.0
Linear (14C-FAcetic Acid)
1.0
0.0
0.0
0
50
100
150
0
200
50
100
Tim e
Tim e
SKOV3
6.0
3H-Acetic Acid
5.0
C/M/Ratio
C ell/Medium
5.0
4.0
14C-F-Acetic Acid
3.0
Linear (3H-Acetic
Acid)
2.0
Linear (14C-FAcetic Acid)
1.0
0.0
0
50
100
Tim e
150
200
150
200
18F-Fluoroacetate
in a Multi-Tumor Model
PC3
SKBr3
#1
U87
#2
15 min
#3
40 min
60 min
120 min
MB435
Kinetics of [18F]-FAc derived radioactivity accumulation
(tumor/muscle ratio) in different tumors
Accumulation of F-18 fluoroacetate
in mice bearing human carcinoma xenografts
10
Log (Ratios)
L
PC3
SKBr3
U87
MB435
1
15
40
60
120
Time (min)
STAGES OF IMAGING AGENT DEVELOPMENT
Dynamic PET/CT Imaging of [18F]-FA
Mouse
Macaque
Kinetics of Biodistribution of 18F-FAc in Monkeys
1.000%
Blood
Liver
Kidney
Lung
0.100%
%ID/g
Bone
Muscle
0.010%
0.001%
Time (min)
0
20
40
60
80
100
120
140
160
180
200
Analysis of Blood Plasma Metabolites
Whole Blood
0.100%
F 18 FA
F-18
F0.010%
0 001%
0.001%
0.060%
0.000%
0
50
100
Time (min)
150
200
%ID/g
%ID
D/g
1.000%
Whole Blood
0.050%
F-18 FA
F-
0.040%
0.030%
0.020%
0.010%
0.000%
0
50
100
Ti (min)
Time
( i )
150
200
Pharmacological Dose Estimates for [18F]Fluoroacetate
Calculated Specific Activity and Mass.
The total mass per dose and specific activity of 18F-fluoroacetate prepared via
no-carrier added synthesis were calculated as follows:
Avogadro's Number
1Ci in Bq
Lambda (decay constant)
Molecular weight of the
compound
Isotope T1/2 (sec)
Dose in mCi
N of atoms per dose
Moles per dose
Total weight per dose (g)
Calculated Sp.Act.
p
(Ci/mol)
(
)
6.02E+23
3 70E+10
3.70E+10
0.000105022
78
6600
10
3.52306E+12
5.85E-12
4.56E-10
1.71E+09
The fluoroacetate used for imaging may have a mass of 0.456 ng which is about
1/1000 of fluoroacetate that may be present in a cup of tea.
Phase I study using PET/CT with 18F18F-FAcetate in
Tumors that don’t
don t avidly accumulate FDG
• Prostate carconoma, lymph node mets
• Breast carcinoma
carcinoma, lobular
lobular, lymph node mets
• Brain tumors, GBM
30 patients (10 patients per tumor type)
Primary objectives:
Pharmacokinetics
Radiolabeled metabolites
Biodistribution
Radiation dosimetry
Dose optimization
Secondary objectives:
Feasibility of tumor detection
as compared to CT and MRI or biopsy
TUMOR DETECTION:
NEW IMAGING AGENTS
FOR
STROMAL BIOMARKERS
Pancreatic Carcinoma: Problems in Diagnosis & Therapy
•
Pancreatic cancer patients seldom exhibit disease
disease--specific
symptoms until late in the course of the disease process
•
Despite significant advances in the treatment of many other human
tumors, the 5-year survival rate in pancreatic cancer remains <5%.
•
At present, no imaging modality is sensitive for visualization of early
stage pancreatic carcinomas or small metastases in the lymph nodes,
peritoneum, and on the surface or within the liver.
liver.
•
Novel biomarker screening and diagnostic imaging methods for
potential earlier detection and localization of pancreatic carcinomas
must be developed in conjunction with new image
image--guided therapies that
would eradicate early neoplastic lesions if survival from this disease is
to be improved.
improved.
Current understanding of the molecular changes in
the multi-step
p p
progression
g
model of pancreatic
p
adenocarcinoma
Molecular Imaging of Stromal Biomarkers:
Volume Amplification Approach
1 mm
5 mm
Intraepithelial Neoplasia
Peritumoral Tissue
Normal Tissue
Gene Expression in parenchyma adjacent to infiltrating
pancreatic ductal adenocarcinoma and chronic pancreatitis
(Fukushima, et al. Mod Pathol 18, 779-787, 2005)
Synthesis of [18F]-Lactose
Radiolabeling of [18F]-Lactose
Synthesis of a precursor
HO
OH
O
HO
HO
1) Ac2O, NaOAc
120 deg, 2h
OH
AcO
AcO
AcO
2) PhSH, BF3-Et2O
rt, 5 h
1) NH3, MeOH
rt, overnight
OAc
O
OH
O
O
PMP
O
HO
2) 4-MeO-PhCH(OMe)2
p-TsOH, 60 deg, 3h
SPh
SPh
nBu2SnO
reflux, 3h
PMP
OH
O
O
O
PMBO
Bu4NI, PMBCl
reflux 2h
3
PMP
NaH BnBr
NaH,
OBn
O
O
O
PMBO
reflux, 2h
BzO
O
BzO
Bu4N F
OAc
OEt
OH
HO
HO
O
HO
OH
O
OEt
18
F
17
Ac2O, 1h
OEt
nBu2SnO
reflux, 3h
OBn
O
HO
HO
PMBO
OEt
6
Bu4NI, PMBCl
reflux 2h
Co-injection of purified 16 with cold version of 14
OBn
O
PMBO
HO
PMBO
OEt
ADC1 A
A, ADC1 CHANNEL A (LG\LG000009
(LG\LG000009.D)
D)
mAU
1600
1400
8
7
1200
1000
800
600
O
AcO
9
AcO
200
OAc
SEt
PMBO
O
OAcPMBO
O
OAc
AcO
NIS, TfOH
OAc
AcO
OBn
O
BzO
O
O
B O
OAc BzO
1) DDQ, rt, 1h
OEt
2) BzCl, Py, 5h
0
AcO
10
2
DAD1 A, Sig=254,4 Ref=360,100 (LG\LG000009.D)
OBn
O
4
6
8
10
4
6
8
10
12
14
16
min
12
14
16
min
10.988
OAc
6.072
400
AcO
mAU
-8
OEt
-8.5
-9
11
-9.5
-10
-10.5
OAc
2
BzO
O
OAc BzO
H2, 10% Pd/C
O
AcO
rt, 12 h
AcO
OH
O
OEt
OAc
BzO
O
OAc BzO
Tf2O, Py
rt, 3h
O
AcO
OTf
O
OEt
QC Chromatogram of compound 17
ADC1 A, ADC1 CHANNEL A (LG\LG000009.D)
mAU
1600
12
13
11.206
AcO
1400
1200
1000
800
OAc
80¡ãC, 15m
AcO
OAc
14
HO
O
200
O
OEt
F
400
NaOMe
Reflux, 20m
HO
OH
HO
O
HO
15
0
O
2
DAD1 A, Sig=254,4 Ref=360,100 (LG\LG000009.D)
F
4
6
8
10
4
6
8
10
12
14
16
min
12
14
16
min
mAU
OEt
1600
1400
1200
1000
800
600
400
200
10.988
O
600
BzO
O
BzO
OH
6.072
Bu4NF
AcO
F
16
O
80¡ãC, 5m
O
18
80¡ãC, 15m
HO
Tf2O, EtOH
BzO
O
BzO
11.206
OBn
O
O
O
PMBO
AcO
OEt
4
PMP
O
18
NaOMe
5
SPh
OAc
OTf
O
13
BSP, TTBP
3 A MS, -60 deg
SPh
OA
OAc
AcO
OAc
O
AcO
2
1
AcO
0
2
PET imaging of pancreatic carcinoma
Axial
Coronal
Saggittal
Tumor
Blood
Muscle
PAP immunostain provided by
Dr. Craig Logsdon, MDACC
Time-activity curves
PET imaging of mice bearing orthotopic
MPanc96 human pancreatic cacinoma
xenografts
ft demonstrated
d
t t d higher
hi h retention
t ti
of [18F]FDL (100 uCi i.v.) in the tumorinvaded pancreas, which corresponded
with high levels of HIP/PAP protein
expression in the peritumoral pancreatic
tissue as evidenced by comparative
tissue,
immunohistochemical analysis in situ.
The [18F]FDL was eliminated via the renal
clearance.
Molecular Imaging of Drug Target
Expression-Activity:
Selection of Therapy
Current Diagnostic – Therapeutic Paradigm
PET/CT
24 - 48 hours
2 months
Therapy “A”
X
FDG, FLT
Therapy “B”
Mechanisms of selectivity of EGFR kinase imaging agent
Inactive Enzyme
Active Enzyme
PO4
Cys773
Cys773
Molecular Imaging Agent for Imaging EGFR kinase expression/activity
Non-invasive molecular imaging of EGFR expression
Nonexpression--activity with
124I-mIPQA PET/CT in orthotopic U87 glioma xenografts in mice
uPET/uCT
uPET
uCT
U87 glioma
Brain
Heart (blood)
Non-invasive molecular imaging of EGFR expression-activity with
124I-mIPQA PET/CT in intracerebral U87 glioma xenografts in mice
Wild-type EGFR
+ EGFRvIII mutant
U87 glioma
Brain
Heart (blood)
k1
Tumor
k3
Blood
U87EGFRvIII glioma
Brain
Heart (blood)
EGFR
k1
Tumor
k3
Blood
k2
k4
k2
k4
EGFR
NSCLC - GL with
NSCLCs
ith Iressa
I
WST
T-1
3
PC-14GL
PC
14GL
H-441GL
H 3255GL
H-1975GL
2
1
0
-2
-1
0
1
2
3
Iressa μM (log scale)
EC50
PC-14GL
45.84
H-441GL
46.19
H 3255GL
1.598
H-1975GL
25.09
HN
I
H
N
F
N
O
N
O
6
HO
Cells//Medium
Peg6 IPQA
H441GL
H3255GL
PC14GL
H1975GL
H441GL with Iressa
H3255GL with Iressa
PC14GL with Iressa
H1975GL with Iressa
450
400
350
300
250
200
150
100
50
0
0
25
50
75
100
125
150
Time (min)
In-Vitro Uptake and Washout Study of I-Peg6-IPQA
in NSCLCs
Uptake Study of IPQA in NSCLCs
Cells/Me
edium
20
H441
H3255
PC14
H1975
15
H441GL
H3255GL
PC14GL
H1975GL
H441GL-washout
H3255-washout
PC14GL-washout
H1975GL-washout
400
300
10
200
5
100
0
0
30
60
90
Time (min)
120
150
0
0
30
60
Time (min)
90
120
H441GL
H3255GL
%ID/g
5.0
%ID/g
5.0
Axial
0
%ID/g
1.0
0
C
Coronal
l
0
Baseline
%ID/g
5.0
%ID/g
5.0
Axial
0
%ID/g
1.0
0
Coronal
With Inhibitor
3h
0
Inhibitor: gefitinib, 100mg/kg,i.p.
1h prior to the radiotracer
H441GL
H3255GL
%ID/g
5.0
%ID/g
1.0
%ID/g
1.0
Axial
Axial
0
%ID/g
1.0
0
C
Coronal
l
Baseline
C
Coronal
l
0
0
%ID/g
5.0
%ID/g
1.0
%ID/g
1.0
Axial
Axial
0
%ID/g
1.0
0
Coronal
With Inhibitor
3h
0
Coronal
24h
0
Inhibitor: gefitinib, 100mg/kg,i.p.
1h prior to the radiotracer
A Paradigm for Therapy Selection and Monitoring by Imaging (Part II)
After 1 cycle
(or @ 2424-72h)
FDG
FLT
EGFR
k-Ras mutant
ChemoChemoRadiation
k-Ras
inhibitor
FDG
FLT
EGFR
ChemoChemoRadiation
k-Ras
inhibitor
PET/CT
Key Imaging Targets:
EGFR, DNA synth.
EGFR mutant
FDG
G
FLT
EGFR
EGFR Inhibitor
FDG
FLT
EGFR
EGFR Inhibitor
Integration of Molecular Imaging, Interventional Radiology, and Tumor Biomarkers
HISTOPATHOLOGY
Immunohistochemistry
PET/CT Imaging
PK/PD Modeling
Parametric Imaging
Parametric ImageImage-Guided
Biopsy (or Intraoperative)
Tumor Lysate
Arrays
microPET/CT Imaging
Drug Target
GENOMICS
Radiolabeling
Structural
Biochemistry
In Silico Modeling
Cyclotron
Radionuclides
Radiotracer
Precursors
New Chemistry
HT/HC Screning