Angiogenesis Imaging Using 68Ga-RGD PET/

CT: Therapeutic Implications

Jae Seon Eo, MD, PhD,* and Jae Min Jeong, PhD†

Angiogenesis imaging is important for diagnostic and therapeutic treatment of various

malignant and nonmalignant diseases. The Arg-Gly-Asp (RGD) sequence has been known
to bind with the αvβ3 integrin that is expressed on the surface of angiogenic blood vessels or
tumor cells. Thus, various radiolabeled derivatives of RGD peptides have been developed for
angiogenesis imaging. Among the various radionuclides, 68Ga was the most widely studied for
RGD peptide imaging because of its excellent nuclear physical properties, easy-to-label
chemical properties, and cost-effectiveness owing to the availability of a 68Ge-68Ga
generator. Thus, various 68Ga-labeled RGD derivatives have been developed and applied for
preclinical and clinical studies. Clinical trials were performed for both malignant and
nonmalignant diseases. Breast cancer, glioma, and lung cancer were malignant, and
myocardial infarction, atherosclerosis, and moyamoya disease were nonmalignant among
the investigated diseases. Further, these 68Ga-labeled RGD derivatives could be applied to
assess the effects of antiangiogenic treatment or theragnosis or both, of cancers. In
conclusion, the angiogenesis imaging technology using 68Ga-labeled RGD derivatives might
be useful for the development of new therapeutic assessments, and for diagnostic and
theragnostic applications.
Semin Nucl Med 46:419-427 C 2016 Elsevier Inc. All rights reserved.

Introduction Selection of Radionuclides for Labeling RGD

A ngiogenesis is an important physiological process and is
related to various diseases such as cancer, myocardial
ischemia, cerebrovascular infarction, arthritis, and psoriasis.1
Therefore, its control and imaging has been a major concern of
many researchers. Although angiogenesis is a very complicated
process, it has a characteristic of expressing the αvβ3 integrin on
the cell surface, which can be specifically bound by a peptide
with the Arg-Gly-Asp (RGD) sequence.2 Thus, most of angio-
genesis imaging with radiolabeled agents has been performed
using radiolabeled RGD derivatives.
*Department of Nuclear Medicine, Korea University Guro Hospital, Seoul,
Korea.
†Department of Nuclear Medicine, Seoul National University College of
Medicine, Seoul, Korea.
This work was supported by a Korea University, South Korea, Grant
(K1422321) and the National Research Foundation of Korea, South Korea,
(NRF) grant funded by the Korea government (MEST) (No. NRF-
2013R1A2A1A05006227).
Address reprint requests to Jae Min Jeong, PhD, Department of Nuclear
Medicine, Seoul National University College of Medicine, 101 Daehak-ro,
Jongno-gu, Seoul 110-744, Korea. E-mail: jmjng@snu.ac.kr
Derivatives
Radioiodine is one of the most easily labeled radionuclides
for peptides. 123I is an excellent radionuclide for SPECT,
whereas 124I is best for PET, and 131I for therapy. However,
the stability of radioiodine-labeled peptides is often a
problem faced in diagnostic or therapeutic applications.
Thus, other radionuclides such as 111In for SPECT, 18F and
68
Ga for PET, and 90Y and 177Lu for therapy are used. Among
these radionuclides, 68Ga is one of the most promising
radionuclides for PET.
68
Ga emits positrons with 89% frequency accompanied by
gamma emission (1.077 MeV) with 3.2% frequency. It has a
low radiation dose because of a short half-life (68 minutes).
Thus, 68Ga is an ideal positron emitter for labeling peptides
such as RGD derivatives. Furthermore, the distribution of 68Ge
or 68Ga generators provides it with great practical applicability.
Bifunctional Chelating Agents
To radiolabel peptides such as RGD derivatives, bifunctional
chelating agents should be used to conjugate 68Ga and the
peptide. The most widely used bifunctional chelating agents

http://dx.doi.org/10.1053/j.semnuclmed.2016.04.001 419
0001-2998/& 2016 Elsevier Inc. All rights reserved.
420 J.S. Eo and J.M. Jeong

Figure 1 Chemical structures of NOTA-RGD monomeric, dimeric, and tetrameric derivatives for 68Ga labeling.

were traditionally 1,4,7,10-tetraazacyclododecane-1,4,7,10-
tetraacetic acid (DOTA) derivatives. However, the ionic radius
of Gaþþþ is too small to fit optimally in the DOTA cage, and
thus, it makes a less stable complex with 68Ga. Therefore, the
biodistribution of the 68Ga-DOTA-RGD peptide was some-
times unfavorable. Relatively high protein binding activity
restrained the wide use of 68Ga-DOTA-RGD,3 but recently
developed remote-control systems provide fast and easy
production of the 68Ga-DOTA-RGD peptide.4
1,4,7-triazacyclononane-1,4,7-triacetic acid (NOTA), which
has a smaller triazacyclononane cage than DOTA, was
preferred for labeling with 68Ga (Fig. 1).5 Jeong et al reported
the feasibility of 68Ga-NOTA-RGD as an excellent imaging
agent for tumor angiogenesis (Fig. 2). 68Ga-NOTA-RGD

Figure 2 Small-animal PET of 68Ga-NOTA-RGD injected into mice bearing SNU-C4 xenografts at 1 and 2 hours after
injection. Arrows indicate tumor positions. (This research was originally published in JNM. Reprinted with permission from
Jeong et al.5 © by the Society of Nuclear Medicine and Molecular Imaging, Inc.)
Angiogenesis imaging using 68Ga-RGD PET/CT 421

showed better performance because of less background activity
compared with that of 68Ga-DOTA-RGD. Various 68Ga-
NOTA-RGD derivatives were developed including dimers
and tetramers of RGD to increase affinity (Fig. 1).6 To promote
pharmacokinetics, glycosylated or PEGylated derivatives were
also developed.7 Affinities to various tumor cell lines such as
prostate cancer,8 glioblastoma,6,9,10 fibrosarcoma, and colon
cancer were tested.11 The specific binding of the 68Ga-RGD
peptide to the αvβ3 integrin was studied via the comparison of
uptakes to various tumor cell lines. The αvβ3-positive tumors
have higher 68Ga-RGD uptake than αvβ3-negative tumors.4,12
RGD peptides conjugated with NODAGA, a derivative of
NOTA, showed a better tumor-to-blood ratio than the original
DOTA-RGD peptide.13,14
In addition, new RGD peptides have been tested and
developed with labeling 68Ga via PTCA,15 FSC,16,17 TRAP,18
NH22sar,19 H2dedpa,20 NS3, and OXO-DO3A chelators.21
Some studies reported better tracer kinetics than traditional
NOTA, DOTA, or NODAGA chelators.14,15
RGD Derivatives
First-generation linear form RGD peptides suffered from low
selectivity and stability. The improvement and reforming was
achieved by the development of the cyclic pentapeptide c
(RGDfV).22 68Ga-labeled RGD peptides for PET imaging were
introduced using NOTA-c(RGDyK) and DOTA-c(RGDfK).4,5
Among the variable cyclopeptides, cRGDfK and cRGDyK are
the most extensively studied derivatives for NOTA, DOTA,
and other bifunctional chelating agents (Table 1).
Multimeric cyclic RGD peptides have been synthesized to
enhance the binding affinity to tumor cells (Fig. 1). An in vitro

Table 1 Animal Studies of 68Ga-Labeled RGD Peptides

Chelate RGD Form No. of RGD Addon Agent Subjects Purpose/Target Disease Reference
NOTA c(RGDyK) 1 None Mouse Biodistribution 5
Mouse Feasibility: SiPM PET 25
Rat Myocardial infarct 37
Mouse Atherosclerosis 41
c(RGDyK) 2 None Mouse Glioblastoma 9
c(RGDyK) 2 None Mouse Glioblastoma 10
c(RGDyK) 1, 2, and 4 None Mouse Glioblastoma 6
c(RGDyK) 1 BBN Mouse Biodistribution 52
Mouse Biodistribution 52
c(RGDyK) 1 GE11 Development 53
c(RGDfK) 1 None Mouse Prostate cancer 8
c(RGDfK) 2 None Mouse Glioblastoma, breast cancer 7
c(RGDfK) 2 None Mouse Fibrosarcoma, colon cancer 11
c(RGDfK) TIMP Mouse Biodistribution 55
DB58 1 None Rat Myocardial infarct 39

DOTA c(RGDfK) 1 None Mouse Melanoma 4

c(RGDfK) 2 None Rat Myocardial infarct 38
c(RGDfK) 2 None Mouse Melanoma, breast cancer 12
c(RGDfK) 2 None Mouse Therapeutic response 44
c(RGDfK) 2 None Mouse Feasibility: Ga generator 27
c(RGDfK) 1,2 None Rat Comparison: NOTA 57
c(RGDfK) 1,2,4 None Mouse Renal cell cancer 24
c(RGDfC) 1,2,4,8,16 None Development 23

NODAGA c(RGDyK) 1 None Mouse Feasibility: automated process 26

c(RGDyK) 2 None Mouse Carcinoid, glioblastoma 48
c(RGDyK) 1 None Mouse Pancreatic cancer 56
c(RGDfK) 1 None Mouse Comparison: DOTA 14
Therapeutic response 45
c(RGDfK) 1 None Mouse Comparison: DOTA 13
c(RGDfK) 2 None Mouse Renal cell cancer 58

FSC c(RGDfK) 3 None Development 17

None Mouse Comparison: NODAGA 16

PTCA c(RGDyK) 1 None Mouse Comparison: NOTA 15

(NH2)2sar c(RGDfK) 2 None Mouse Biodistribution 19
NS3 c(RGDfK) 1 None Mouse Biodistribution 21
Oxo-DO3A c(RGDfK) 1 None Mouse Biodistribution 21
TRAP c(RGDfK) 3 None Mouse Biodistribution 18
H2dedpa c(RGDyK) 1,2 None Mouse Biodistribution 20
422 J.S. Eo and J.M. Jeong

trial using the hexadecimer showed more than a 100-fold Table 2 Clinical Studies of 68Ga-Labeled RGD Derivatives
increase in avidity relative to the monomer.23 The dimers Chelate RGD No. of Purpose/ Refer-
and tetramers of the 68Ga-DOTA-RGD complex showed Form RGD Target ence
relatively low blood pool activities.24 However, multimeriza- Disease
tion not only has merits but also has disadvantages. The NOTA c(RGDyK) 1 Dosimetry 31
tetrameric 68Ga-RGD peptides conjugated with either NOTA Cerebral infarct 42
or DOTA showed high uptakes in the kidneys in micro-PET Atherosclerosis 41
images.6,24 Therefore, multimers of more than tetramers were Breast cancer 32
not preferred. Moyamoya 43
disease
c(RGDyK) 2 Myocardial 40
Feasibility Test infarct
Variable 68Ga-labeled RGD peptides were used as a feasibility Glioma 34
test for the new technique. Verification of new instruments, Lung cancer 35
such as the small-animal PET using a silicon photomultiplier, DOTA c(RGDfK) 2 Breast cancer 33
was performed using 68Ga-NOTA-RGD peptides.25 Further-
more, the DOTA-RGD peptide dimer was labeled very well
with 68Ga eluted from a nanoceria-polyacrylonitrile–based
generator.26 Recently, radiopharmacy acquired current good patients with lung cancer and lymphoma. The uptake of 68Ga-
manufacturing practice or good radiopharmaceutical practice. NOTA-RGD was the highest in the kidneys (71.6 ⫾ 28.4 μGy/
The current good manufacturing practice or good radio- MBq) and urinary bladders (239.6 ⫾ 56.58 μGy/MBq). The
pharmaceutical practice–compliant automated process for mean effective doses using the International Commission of
radiopharmaceutical production was also verified for the Radiation Protection 60 and 103 weighting factors are 22.4 ⫾
68
Ga-NODAGA-RGD monomer with high yield, high purity, 3.8 to 25.0 ⫾ 4.4 μSv/MBq, which are acceptable effective
and remarkable specific activity.27 radiation doses.31
The first clinical trial for 43 patients with advanced
breast cancer was reported by Yoon et al.32 In this study,
Tumor Angiogenesis Imaging 68
Ga-NOTA-RGD uptake was significantly higher in the
human epidermal growth factor receptor 2 (HER2)–
Angiogenesis is an important target for drug development and
positive patient group and had positive correlation with
diagnosis of malignancy because of its critical role in tumor
the HER2/CEP17 ratio, whereas FDG-PET had higher
growth and metastasis. More than 40 antiangiogenic drugs
uptake in the estrogen receptor–negative and progester-
have entered into clinical trials.28 Therefore, angiogenesis
one receptor–negative groups. In the estrogen receptor–
imaging is necessary for the prediction and noninvasive
negative, progesterone receptor–negative, and HER2-
monitoring of antiangiogenic agents. 18F-labeled RGD pep-
negative groups (triple negative), FDG-PET showed sig-
tides, such as 18F-galacto-RGD, have been used at early time of
nificantly higher uptake than that of others, but 68Ga-
angiogenesis imaging. However, 18F has limitations, such as
NOTA-RGD PET showed significantly lower uptake than that
the requirement of an expensive cyclotron for production and
of other groups. Another small group study for 5 patients with
the difficulties in the radiolabeling procedure. Because of the
breast cancer with in-house generator–produced 68Ga-DOTA-
convenience of labeling and easy accessibility with 68Ge-68Ga
RGD PET showed good radiochemical yields and proper tracer
generators, 68Ga-labeled RGD derivatives are currently more
uptakes (maximum standardized uptake value [SUVmax]:
popular for angiogenesis imaging.
6.34-21) in tumors.33
A study with the 68Ga-NOTA-RGD dimer reported the
Preclinical Studies sensitivity for gliomas.34 68Ga-RGD PET/CT was able to
Recently, various Ga-labeled RGD peptides were newly evaluate the glioma demarcation more specifically than
developed and introduced for preclinical studies (Table 1). FDG-PET/CT because the 68Ga-RGD peptide specifically
68
Ga-labeled RGD peptides are composed of a RGD peptide to accumulated in brain tumors, but not in the brain parenchyma
bind the αvβ3 integrin and a bifunctional chelator for 68Ga other than the choroid plexus. Further, the SUVmax of
68
labeling.3,29,30 Most preclinical studies of novel 68Ga-RGD Ga-RGD were significantly correlated with the grade of
peptides using the new chelator or RGD forms were verified glioma.
with an animal tumor model and biodistribution study.22 A recent clinical study of 91 patients with lung cancer
reported that 68Ga-NOTA-PRGD2 PET/CT showed positive
and negative predictive values of 90.0% and 93.8%, respec-
Clinical Studies tively, in assessing mediastinal lymph node metastases,
Although various radiopharmaceuticals have been tried for whereas those of FDG-PET/CT were 30.2% and 90.5%,
preclinical studies, clinical trials of 68Ga-RGD tumor imaging respectively. These results indicate that 68Ga-NOTA-RGD
were less active (Table 2). Kim et al31 reported the human PET has a significant advantage over FDG-PET/CT in judging
radiation dosimetry of the 68Ga-NOTA-RGD monomer for lymph node metastases (Fig. 3).35
Angiogenesis imaging using 68Ga-RGD PET/CT 423

Figure 3 18F-FDG-PET/CT and 68Ga-NOTA-PRGD2 PET/CT images and immunohistochemical (IHC) staining of lymph
nodes within lung region. Arrows point to lymph nodes. (A) A 69-year-old woman with moderately differentiated
adenocarcinoma with lymph node metastasis. Both 18F-FDG and 68Ga-NOTA-PRGD2 PET show positive lymph nodes
with positive integrin αvβ3 staining. (B) A 44-year-old woman with highly differentiated adenocarcinoma with lymph
node metastasis. Lymph node is negative on 18F-FDG-PET and positive on 68Ga-NOTA-PRGD2 PET with positive
integrin αvβ3 staining. (C) A 58-year-old woman with adenocarcinoma with no lymph node metastasis. However, 18F-
FDG-PET shows positive lymph nodes, whereas 68Ga-NOTA-PRGD2 PET shows a negative result with negative integrin
αvβ3 staining. (D) A 62-year-old woman with highly differentiated adenocarcinoma with no lymph node metastasis.
Both 18F-FDG and 68Ga-NOTA-PRGD2 PET show negative lymph nodes with negative integrin αvβ3 staining. (This
research was originally published in JNM. Reprinted with permission from Zheng et al.35 © by the Society of Nuclear
Medicine and Molecular Imaging, Inc.)

Angiogenesis Imaging of Cardiovascular Disease

Nonmalignant Diseases
Angiogenesis is also apparent in many nonmalignant diseases
such as myocardial ischemia, cerebrovascular infarction,
arthritis, and psoriasis. Tumor angiogenesis is directly related
to the progression or aggravation of disease. On the contrary,
defects of angiogenesis in ischemia, atherosclerosis, brain
infarction, and ulceration often delay the healing process.1
Therefore, the trial of the improvement of perfusion and
function has been continued with promoting therapeutic
angiogenesis.36
Angiogenesis is a natural repair process that occurs after
myocardial infarction. Eo et al37 reported that the region of
myocardial infarction showed higher 68Ga-NOTA-RGD
uptake than normal and sham-operated myocardia by auto-
radiography and animal PET/CT images. Kiugel et al38
reported the correlation between tracer uptake of myocardial
infarctions and histopathological markers by 68Ga-DOTA-
RGD PET/CT and autoradiography. This study showed
strong correlation between tracer uptake and the amount
of b3 chains, whereas correlation to neovessels could only be
seen at the 4-week time point. Another study with 68Ga-
424 J.S. Eo and J.M. Jeong

Figure 4 68Ga-RGD PET/CT images of representative case. On (A) CT, (B) fusion, and (C) PET images, increased uptake was
observed around the bony flap, most evidently in the craniotomy margin area (arrow). Mild uptake increase was also
observed on the inner surface of the bony flap. (This research was originally published in JNM. Reprinted with permission
from Kim et al.43 © by the Society of Nuclear Medicine and Molecular Imaging, Inc.) This research was originally published
in JNM. Author(s). Title. J Nucl Med. 2014;55:1467-1472. © by the Society of Nuclear Medicine and Molecular Imaging,
Inc.

NOTA-DB58, a newly designed cyclic RGD peptide, also
confirmed high tracer uptake in the myocardial infarction
area.39
In clinical trials, Luo et al40 reported interesting 68Ga-
NOTA-RGD dimer PET/CT images for two patients with
myocardial infarction. Uptakes of the 68Ga-NOTA-RGD dimer
were seen in the infarction region in the initial PET/CT scans.
However, in follow-up scans after treatment, the uptake in the
infarcted region was noticed only in the patient who still had
symptoms and was no longer visualized in the asymptomatic
patient.
The feasibility test of atherosclerosis imaging using 68Ga-
NOTA-RGD PET/CT also has been attempted. In the mouse
model, 68Ga-NOTA-RGD PET/CT showed high uptake in the
atherosclerotic aorta as well as in FDG-PET/CT. In 4 patients
with coronary artery disease, the aorta-to-jugular ratios showed
a tendency to correlate with the serum high-sensitivity
C-reactive protein level (r ¼ 0.899, P ¼ 0.102). However,
the potential of atherosclerosis imaging was limited for clinical
applications because of lower imaging contrast and sensitivity
compared with FDG-PET/CT.41
Cerebrovascular Imaging
Angiogenesis is one of the key processes for spontaneous
recovery or therapeutic interventions in ischemic cerebral
disorder. However, only two clinical studies with 68Ga-
NOTA-RGD PET/CT exist for patients with moyamoya
disease, which is a progressive steno-occlusive change in the
internal carotid artery, and which causes compensatory
abnormal vasculature in the brain. The most common treat-
ment of moyamoya disease is the indirect revascularization of
the ischemic area. The efficacy of the treatment can be
evaluated by imaging studies that show collateral vessel
formation or cerebral perfusion, such as angiography, MRI,
and perfusion SPECT.
68
Ga-NOTA-RGD PET/CT has been applied for moyamoya
disease in patients with cerebral infarction, and a significant
correlation was found between the postinfarction time interval
and the lesion-to-control ratio in the recent (within 30 days)
infarction group. It revealed that angiogenic activity is highest
in the early stages and it decreases over time.42 After
reperfusion treatment, angiogenic activity measured by 68Ga-
NOTA-RGD PET gradually decreased with time and it
significantly correlated with the postoperative time interval.
The activity was normalized approximately 6 months after
surgery. Further, compared with perfusion SPECT, angiogenic
activity measured at approximately 3.7 months after surgery
was inversely correlated with perfusion improvement, which
indicates that prolonged angiogenic activity might be caused
by insufficient revascularization (Fig. 4).43
Therapeutic Implications
Response Evaluation
Although many kinds of antiangiogenic drugs have been
approved to inhibit tumor growth, monitoring of angiogenesis
in tumors is still unavailable. Development of monitoring
methods might greatly promote clinical applications. 68Ga-
labeled RGD peptide PET/CT is one of the most useful
techniques for noninvasive monitoring of tumor angiogenesis
and of the efficacy of antiangiogenic drugs. A recent preclinical
study reported that 68Ga-RGD PET could reflect the tumor
response to antiangiogenic therapy much earlier than FDG-
PET (day 3 vs day 14 posttreatment). With this, 68Ga-RGD
PET can contribute to the development of new antiangiogenic
drugs and to the establishment of therapeutic plans and
dosages.44 Another preclinical study with the Ga-NODAGA-
RGD PET peptide reported on antiangiogenic therapy with
bevacizumab for αvβ3-negative cell line tumors. Antiangiogenic
therapy against A-431 tumors inhibited tumor cell growth,
decreased microvessel density, delayed tumor necrosis, and
normalized tumor vasculature. Intriguingly, RGD uptakes
were significantly increased at day 7 of therapy and did not
decrease until after 3 weeks of treatment. In FaDu tumors, the
effect of antiangiogenic therapy was minimal and RGD uptake
was not significantly changed. These discrepancies presumed
that RGD uptake could be affected by normalizing vascular
structure or tumor necrosis during antiangiogenic therapy.
Angiogenesis imaging using 68Ga-RGD PET/CT
Therefore, the interpretation of 68Ga-RGD PET after antiangio-
genic therapy of αvβ3-negative tumors should be handled
carefully.45
Multimodal Approach
The advantage of the 68Ga-labeled peptide is that a new tumor-
targeting radiopharmaceutical can be developed only by
substituting radioisotopes.46 The positron-emitting radionu-
clide 66Ga has been proposed as a PET imaging alternative of
68
Ga. 66Ga has some advantages, such as a long half-life (9.49
hours) and abundant high-energy positron emission (Emax ¼
4.15 MeV, medium range ¼ 7.6 mm), which might be useful
in therapy. A recent preclinical trial with the 66Ga-labeled RGD
dimer showed favorable tumor uptake and rapid blood
clearance, and biodistribution study was available up to 24
hours postinjection (Fig. 5).47 64Cu is a useful positron-
emitting radionuclide with a long half-life (12.7 hours). The
68
Ga-labeled RGD peptide has been compared with the 64Cu-
labeled kind with the same RGD and chelator structure.48,14
Radioisotopes such as 177Lu and 90Y could replace 68Ga
without changing chelators for therapeutic radiopharmaceut-
icals. Moreover, by using NETA beyond traditional NOTA or
DOTA chelators, 90Y and 177Lu, labeled RGD peptides such as
177
Lu-5p-C-NETA-c(RGDyK) and 177Lu-NOTA-c(RGDyK)
have been developed for therapeutic applications.49-51
In addition to RGD alone, combination with various specific
targeting peptides alsoare available. Radiolabeled bombesin
(BBN) analogues target the gastrin-releasing peptide receptor
that is highly expressed in variable human cancers. The
Figure 5 Coronal micro-PET images of [66Ga] DOTA -E-[c(RGDfK)]2 in nude mice bearing C6 tumor xenograft at 0.5 hour
(A), 1 hour (B), 3 hours (C), 5 hours (D), and 24 hours (E) after injection of 20 ⫾ 0.5 MBq of tracer under isoflourane
anesthesia. Acquisition time was 20 minutes for A, B, and C; 30 minutes for D, and 60 minutes for E. (Reprinted with
permission from Lopez-Rodriguez et al.47)
425
development of the 68Ga-labeled NOTA-RGD-BBN hetero-
dimer was introduced by Liu et al52 in 2009. The micro-PET
imaging of prostate, melanoma, and breast cancer cell lines
were performed using the 68Ga-NOTA-RGD-BBN peptide.52
In addition, heterodimeric peptides for dual-targeting 68Ga-
NOTA-RGD-GE11 synthesis was succeeded not only for the
αvβ3 integrin but also for the epidermal growth factor receptor
by dodecapeptide GE11 (Tyr-His-Trp-Tyr-Gly-Tyr-Thr-Pro-
Gln-Asn-Val-Ile).53 The human serum albumin-tissue inhib-
itor of metalloproteinase 2 (HSA-TIMP2) has been reported to
be a biologically active candidate therapeutic agent for
angiogenesis-related diseases such as cancer.54 A complex
68
Ga-NOTA-RGD-HSA-TIMP2 was developed for PET/CT
imaging that might be useful not only for cancer imaging
probes but also for anticancer therapeutic agents.55
Approaches with multimodality for targeting the αvβ3
integrin could be helpful not only for in vivo imaging but also
for intraoperative delineation. Trajkovic-Arsic et al56 used 2
αvβ3 integrin–targeting agents, 68Ga-NODAGA-RGD peptide
for PET/CT imaging, and IntegriSense 680 for intraoperative
fluorescence imaging. With this, preoperative PET/CT and
fluorescence molecular tomography enabled visualization of
pancreatic cancer in the mouse model. Furthermore, intra-
operative optical imaging with IntegriSense 680 allowed good
delineation of tumor borders.
Nonmalignant Diseases
Evaluation of the therapeutic implication of 68Ga-RGD pep-
tides for nonmalignant diseases is rare. In the rat myocardial
426
infarction model, angiogenesis therapy with the basic fibroblast
growth factor was tried and the efficacy of therapy was
monitored by 68Ga-NOTA-RGD PET/CT. The uptake of
68
Ga-NOTA-RGD in the angiogenesis therapy group was
significantly higher than that of the saline-injected group on
the first day. However, no significant difference was observed
between the therapy group and the control group at subse-
quent time points, corresponding to the final infarction size
change. This study presumed only a single dose of angio-
genesis therapy that is not prolonged for a long time and that
does not affect the final prognosis, such as by infarction size
reduction.37 In addition, the disappearance of RGD uptake
after treatment in the asymptomatic patient also suggests that
68
Ga-NOTA-RGD PET/CT is useful for monitoring of the
therapeutic response evaluation and for finding the causes of
symptoms.40
Conclusion
Various 68Ga-labeled RGD derivatives were developed for
angiogenesis imaging and studied for preclinical and clinical
effects. Angiogenesis imaging of malignant diseases such as
gliomas and lung cancer showed their feasibility. Furthermore,
their applicability in the imaging of cardiovascular diseases
such as myocardial infarction and atherosclerosis and cere-
brovascular diseases such as moyamoya disease has also been
proved by clinical studies. The assessment of the therapeutic
effect and theragnosic application fields were also possible.
Thus, angiogenesis imaging using 68Ga-labeled RGD deriva-
tives might be a very useful tool for diagnosis, therapeutic
assessment, and theragnosis of angiogenic diseases.
References
1. Carmeliet P, Jain RK: Angiogenesis in cancer and other diseases. Nature
2000;407(6801):249-257
2. Haubner R, Wester HJ, Reuning U, et al: Radiolabeled αvβ3 integrin
antagonists: A new class of tracers for tumor targeting. J Nucl Med
1999;40(6):1061-1071
3. Cai H, Conti PS, RGD-based PET: Tracers for imaging receptor integrin
αvβ3 expression. J Labelled Comp Radiopharm 2013;56(5):264-279
4. Decristoforo C, Hernandez Gonzalez I, Carlsen J, et al: 68Ga- and 111In-
labelled DOTA-RGD peptides for imaging of αvβ3 integrin expression. Eur
J Nucl Med Mol Imaging 2008;35(8):1507-1515
5. Jeong JM, Hong MK, Chang YS, et al: Preparation of a promising
angiogenesis PET imaging agent: 68Ga-labeled c(RGDyK)-isothiocyanato-
benzyl-1,4,7-triazacyclononane-1,4,7-triacetic acid and feasibility studies
in mice. J Nucl Med 2008;49(5):830-836
6. Li ZB, Chen K, Chen X: 68Ga-labeled multimeric RGD peptides for
microPET imaging of integrin αvβ3 expression. Eur J Nucl Med Mol
Imaging 2008;35(6):1100-1108
7. Liu Z, Niu G, Shi J, et al: 68Ga-labeled cyclic RGD dimers with Gly3 and
PEG4 linkers: Promising agents for tumor integrin αvβ3 PET imaging. Eur
J Nucl Med Mol Imaging 2009;36(6):947-957
8. Israel I, Richter D, Stritzker J, et al: PET imaging with [68Ga]NOTA-RGD
for prostate cancer: A comparative study with [18F]fluorodeoxyglucose
and [18F]fluoroethylcholine. Curr Cancer Drug Targets 2014;14
(4):371-379
9. Lang L, Li W, Guo N, et al: Comparison study of [18F]FAl-NOTA-PRGD2,
[18F]FPPRGD2, and [68Ga]Ga-NOTA-PRGD2 for PET imaging of
U87MG tumors in mice. Bioconjug Chem 2011;22(12):2415-2422
J.S. Eo and J.M. Jeong
10. Guo N, Lang L, Li W, et al: Quantitative analysis and comparison study of
[18F]AlF-NOTA-PRGD2, [18F]FPPRGD2 and [68Ga]Ga-NOTA-PRGD2
using a reference tissue model. PLos One 2012;7(5):e37506
11. Shao Y, Liang W, Kang F, et al: A direct comparison of tumor angiogenesis
with 68Ga-labeled NGR and RGD peptides in HT-1080 tumor xenografts
using microPET imaging. Amino Acids 2014;46(10):2355-2364
12. Alam IS, Witney TH, Tomasi G, et al: Radiolabeled RGD tracer kinetics
annotates differential αvβ3 integrin expression linked to cell intrinsic and
vessel expression. Mol Imaging Biol 2014;16(4):558-566
13. Knetsch PA, Petrik M, Griessinger CM, et al: [68Ga]NODAGA-RGD for
imaging αvβ3 integrin expression. Eur J Nucl Med Mol Imaging 2011;38
(7):1303-1312
14. Dumont RA, Deininger F, Haubner R, et al: Novel 64Cu- and 68Ga-labeled
RGD conjugates show improved PET imaging of αvβ3 integrin expression
and facile radiosynthesis. J Nucl Med 2011;52(8):1276-1284
15. Ferreira CL, Yapp DT, Mandel D, et al: 68Ga small peptide imaging:
Comparison of NOTA and PCTA. Bioconjug Chem 2012;23
(11):2239-2246
16. Knetsch PA, Zhai C, Rangger C, et al: [68Ga]FSC-(RGD)3 a trimeric RGD
peptide for imaging αvβ3 integrin expression based on a novel siderophore
derived chelating scaffold-synthesis and evaluation. Nucl Med Biol
2015;42(2):115-122
17. Zhai C, Summer D, Rangger C, et al: Fusarinine C, a novel siderophore-
based bifunctional chelator for radiolabeling with Gallium-68. J Label
Comp Radiopharm 2015;58(5):209-214
18. Notni J, Pohle K, Wester HJ: Be spoilt for choice with radiolabelled RGD
peptides: Preclinical evaluation of 68Ga-TRAP(RGD)(3). Nucl Med Biol
2013;40(1):33-41
19. Ma MT, Neels OC, Denoyer D, et al: Gallium-68 complex of a
macrobicyclic cage amine chelator tethered to two integrin-targeting
peptides for diagnostic tumor imaging. Bioconjug Chem 2011;22
(10):2093-2103
20. Boros E, Ferreira CL, Yapp DT, et al: RGD conjugates of the H2dedpa
scaffold: Synthesis, labeling and imaging with 68Ga. Nucl Med Biol
2012;39(6):785-794
21. Knetsch PA, Petrik M, Rangger C, et al: [68Ga]NS(3)-RGD and [68Ga]
Oxo-DO3A-RGD for imaging αvβ3 integrin expression: Synthesis, evalua-
tion, and comparison. Nucl Med Biol 2013;40(1):65-72
22. Schottelius M, Laufer B, Kessler H, et al: Ligands for mapping αvβ3-
integrin expression in vivo. Acc Chem Res 2009;42(7):969-980
23. Wangler C, Maschauer S, Prante O, et al: Multimerization of cRGD
peptides by click chemistry: Synthetic strategies, chemical limitations, and
influence on biological properties. Chembiochem 2010;11(15):
2168-2181
24. Dijkgraaf I, Yim CB, Franssen GM, et al: PET imaging of αvβ3 integrin
expression in tumours with 68Ga-labelled mono-, di- and tetrameric RGD
peptides. Eur J Nucl Med Mol Imaging 2011;38(1):128-137
25. Kwon SI, Lee JS, Yoon HS, et al: Development of small-animal PET
prototype using silicon photomultiplier (SiPM): Initial results of phantom
and animal imaging studies. J Nucl Med 2011;52(4):572-579
26. Pohle K, Notni J, Bussemer J, et al: 68Ga-NODAGA-RGD is a suitable
substitute for 18F-Galacto-RGD and can be produced with high specific
activity in a cGMP/GRP compliant automated process. Nucl Med Biol
2012;39(6):777-784
27. Chakraborty S, Chakravarty R, Sarma HD, et al: The practicality of
nanoceria-PAN-based 68Ge/68Ga generator toward preparation of 68Ga-
labeled cyclic RGD dimer as a potential PET radiotracer for tumor
imaging. Cancer Biother Radiopharm 2013;28(1):77-83
28. Samant RS, Shevde LA: Recent advances in anti-angiogenic therapy of
cancer. Oncotarget 2011;2(3):122-134
29. Fani M, Maecke HR: Radiopharmaceutical development of radiolabelled
peptides. Eur J Nucl Med Mol Imaging 2012;39:S11-S30 (suppl 1)
30. Cai H, Conti PS: RGD-based PET tracers for imaging receptor integrin αv
β3 expression. J Labelled Comp Radiopharm 2013;56(5):264-279
31. Kim JH, Lee JS, Kang KW, et al: Whole-body distribution and radiation
dosimetry of 68Ga-NOTA-RGD: A positron emission tomography agent
for angiogenesis imaging. Cancer Biother Radiopharm 2012;27(1):65-71
32. Yoon HJ, Kang KW, Chun IK, et al: Correlation of breast cancer subtypes,
based on estrogen receptor, progesterone receptor, and HER2, with
Angiogenesis imaging using 68Ga-RGD PET/CT
functional imaging parameters from 68Ga-RGD PET/CT and 18F-FDG
PET/CT. Eur J Nucl Med Mol Imaging 2014;41(8):1534-1543
33. Vatsa R, Bhusari P, Kumar S, et al: Integrin αvβ3 as a promising target to
image neoangiogenesis using in-house generator-produced positron
emitter 68Ga-Labeled DOTA-arginine-glycine-aspartic acid (RGD) ligand.
Cancer Biother Radiopharm 2015;30(5):217-224
34. Li D, Zhao X, Zhang L, et al: 68Ga-PRGD2 PET/CT in the evaluation of
glioma: A prospective study. Mol Pharm 2014;11(11):3923-3929
35. Zheng K, Liang N, Zhang J, et al: 68Ga-NOTA-PRGD2 PET/CT for integrin
imaging in patients with lung cancer. J Nucl Med 2015;56(12):1823-1827
36. Ahn A, Frishman WH, Gutwein A, et al: Therapeutic angiogenesis: A new
treatment approach for ischemic heart disease—Part I. Cardiol Rev
2008;16(4):163-171
37. Eo JS, Paeng JC, Lee S, et al: Angiogenesis imaging in myocardial infarction
using 68Ga-NOTA-RGD PET: Characterization and application to ther-
apeutic efficacy monitoring in rats. Coron Artery Dis 2013;24(4):303-311
38. Kiugel M, Dijkgraaf I, Kyto V, et al: Dimeric [68Ga]DOTA-RGD peptide
targeting αvβ3 integrin reveals extracellular matrix alterations after
myocardial infarction. Mol Imaging Biol 2014;16(6):793-801
39. Menichetti L, Kusmic C, Panetta D, et al: MicroPET/CT imaging of αvβ3
integrin via a novel 68Ga-NOTA-RGD peptidomimetic conjugate in rat
myocardial infarction. Eur J Nucl Med Mol Imaging 2013;40
(8):1265-1274
40. Luo Y, Sun Y, Zhu Z, et al: Is the change of integrin αvβ3 expression in the
infarcted myocardium related to the clinical outcome? Clin Nucl Med
2014;39(7):655-657
41. Paeng JC, Lee YS, Lee JS, et al: Feasibility and kinetic characteristics of
68
Ga-NOTA-RGD PET for in vivo atherosclerosis imaging. Ann Nucl Med
2013;27(9):847-854
42. Choi H, Phi JH, Paeng JC, et al: Imaging of integrin αvβ3 expression using
68
Ga-RGD positron emission tomography in pediatric cerebral infarct.
Mol Imaging 2013;12(4):213-217
43. Kim YI, Phi JH, Paeng JC, et al: In vivo evaluation of angiogenic activity
and its correlation with efficacy of indirect revascularization surgery in
pediatric moyamoya disease. J Nucl Med 2014;55(9):1467-1472
44. Shi J, Jin Z, Liu X, et al: PET imaging of neovascularization with 68Ga-
3PRGD2 for assessing tumor early response to endostar antiangiogenic
therapy. Mol Pharm 2014;11(11):3915-3922
45. Rylova SN, Barnucz E, Fani M, et al: Does imaging αvβ3 integrin
expression with PET detect changes in angiogenesis during bevacizumab
therapy? J Nucl Med 2014;55(11):1878-1884
427
46. Otte A, Jermann E, Behe M, et al: DOTATOC: A powerful new tool for
receptor-mediated radionuclide therapy. Eur J Nucl Med 1997;24
(7):792-795
47. Lopez-Rodriguez V, Gaspar-Carcamo RE, Pedraza-Lopez M, et al: Prep-
aration and preclinical evaluation of 66Ga-DOTA-E(c(RGDfK))2 as a
potential theranostic radiopharmaceutical. Nucl Med Biol 2015;42
(2):109-114
48. Oxboel J, Brandt-Larsen M, Schjoeth-Eskesen C, et al: Comparison of two
new angiogenesis PET tracers 68Ga-NODAGA-E[c(RGDyK)]2 and 64Cu-
NODAGA-E[c(RGDyK)]2; in vivo imaging studies in human xenograft
tumors. Nucl Med Biol 2014;41(3):259-267
49. Shi J, Zhou Y, Chakraborty S, et al: Evaluation of in-labeled cyclic RGD
peptides: Effects of peptide and linker multiplicity on their tumor uptake,
excretion kinetics and metabolic stability. Theranostics 2011;1:322-340
50. Ju CH, Jeong JM, Lee YS, et al: Development of a 177Lu-labeled RGD
derivative for targeting angiogenesis. Cancer Biother Radiopharm 2010;
25(6):687-691
51. Kang CS, Chen Y, Lee H, et al: Synthesis and evaluation of a new
bifunctional NETA chelate for molecular targeted radiotherapy using 90Y
or 177Lu. Nucl Med Biol 2015;42(3):242-249
52. Liu Z, Niu G, Wang F, et al: 68Ga-labeled NOTA-RGD-BBN peptide for
dual integrin and GRPR-targeted tumor imaging. Eur J Nucl Med Mol
Imaging 2009;36(9):1483-1494
53. Yu HM, Chen JH, Lin KL, et al: Synthesis of 68Ga-labeled NOTA-RGD-
GE11 heterodimeric peptide for dual integrin and epidermal growth
factor receptor-targeted tumor imaging. J Labelled Comp Radiopharm
2015;58(7):299-303
54. Kang WK, Park E-K, Lee HS, et al: A biologically active angiogenesis
inhibitor, human serum albumin-TIMP-2 fusion protein, secreted from
Saccharomyces cerevisiae. Protein Expr Purif 2007;53(2):331-338
55. Choi N, Kim SM, Hong KS, et al: The use of the fusion protein RGD-HSA-
TIMP2 as a tumor targeting imaging probe for SPECT and PET.
Biomaterials 2011;32(29):7151-7158
56. Trajkovic-Arsic M, Mohajerani P, Sarantopoulos A, et al: Multimodal
molecular imaging of integrin αvβ3 for in vivo detection of pancreatic
cancer. J Nucl Med 2014;55(3):446-451
57. Blom E, Velikyan I, Estrada S, et al: 68Ga-labeling of RGD peptides and
biodistribution. Int J Clin Exp Med 2012;5(2):165-172
58. Dijkgraaf I, Terry SY, McBride WJ, et al: Imaging integrin αvβ3 expression
in tumors with an ,F-labeled dimeric RGD peptide. Contrast Media Mol
Imaging 2013;8(3):238-245