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 Table of Contents  
Year : 2016  |  Volume : 1  |  Issue : 2  |  Page : 43-49

68 Ga-BNOTA-PRGD2 PET/CT for evaluation of post-stroke angiogenesis: study protocol for a prospective open-label clinical trial

PET Center, Department of Nuclear Medicine, Peking Union Medical College Hospital, Chinese Academy of Medical Science & Peking Union Medical College, Beijing, China

Date of Web Publication29-Apr-2016

Correspondence Address:
Zhaohui Zhu
PET Center, Department of Nuclear Medicine, Peking Union Medical College Hospital, Chinese Academy of Medical Science & Peking Union Medical College, Beijing
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Source of Support: This work was supported in part by the National Natural Science Foundation of China, No. 81171370 and 81271614; and a Special Scientific Research Fund for Public Welfare on Health Professions of China, No. 201402001., Conflict of Interest: None

DOI: 10.4103/2468-5577.181234

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Background: Arginylglycylaspartic acid (RGD) is a tripeptide composed of L-arginine, glycine, and L-aspartic acid that shows great affinity for the integrin αv β3 receptor. A RGD dimer labeled with 68Ga, 68Ga-BNOTA-PRGD2 was designed for positron-emission tomography/computed tomography (PET/CT) imaging post-stroke angiogenesis, which has rarely been used in the clinic. We aimed to perform a prospective open-label trial to validate the diagnostic value of 68 Ga-BNOTA-PRGD2 PET/CT in post-stroke angiogenesis.
Methods/Design: A self-controlled open-label clinical trial will be performed at the PET Center, Department of Nuclear Medicine, Peking Union Medical College Hospital, China. Fifty patients with stroke will undergo 68 Ga-BNOTA-PRGD2 PET/CT, 18 F-FDG PET/CT, and magnetic resonance imaging (MRI) at 2 weeks, 3 months, and 1 year after stroke onset. Primary outcomes include the standardized uptake value (SUV) in the cerebral infarction area and the standardized uptake value ratio of the injured side to the contralateral side detected by 68 Ga-BNOTA-PRGD2 PET/CT. Secondary outcomes include the safety of brain 68 Ga-BNOTA-PRGD2 PET/CT used to evaluate post-stroke angiogenesis.
Discussion: Findings from this trial will provide important reference evidence for use of 68 Ga-BNOTA-PRGD2 PET/CT to evaluate post-stroke angiogenesis.
Trial registration: ClinicalTrials.gov identifier: NCT01656785; registered on 1 August 2012.

Keywords: clinical trial; stroke; 68Ga-BNOTA-PRGD2; positron-emission tomography; computed tomography; magnetic resonance imaging; 18F-FDG; angiogenesis, physiologic; αv β3 ; vitronectin receptor

How to cite this article:
Wang H, Sun Y, Wu C, Zhu Z. 68 Ga-BNOTA-PRGD2 PET/CT for evaluation of post-stroke angiogenesis: study protocol for a prospective open-label clinical trial. Asia Pac J Clin Trials Nerv Syst Dis 2016;1:43-9

How to cite this URL:
Wang H, Sun Y, Wu C, Zhu Z. 68 Ga-BNOTA-PRGD2 PET/CT for evaluation of post-stroke angiogenesis: study protocol for a prospective open-label clinical trial. Asia Pac J Clin Trials Nerv Syst Dis [serial online] 2016 [cited 2019 Aug 26];1:43-9. Available from: http://www.actnjournal.com/text.asp?2016/1/2/43/181234

  Background Top

Stroke is a disease that greatly threatens human life and health. An estimated 6.7 million people died from stroke in 2012, representing 12% of all global deaths (World Health Organization, 2014). Post-stroke angiogenesis is of importance in promotion of nerve repair and functional recovery. The commonly used methods of evaluating vessels primarily include CT angiography (Sabarudin et al., 2014), MR angiography (González, 2012) and single photon emission computed tomography (SPECT) imaging (Momosaki et al., 2012). The former two techniques are used to observe vascular structure, while SPECT evaluates perfusion. They cannot directly reflect the angiogenesis, whereas 68 Ga-BNOTA-PRGD2 can target the integrin α v β 3 receptor expression on the surface of endothelial cells of the newly formed vessels. Therefore, 68 Ga-BNOTA-PRGD2 PET/CT may provide a special technique to quantify the angiogenesis (Sun, 2014). 18 F-FDG is a synthesized glucose analogue that can mimic glucose in the organism, reflecting glucose metabolism in tissue. 18 F-FDG PET/CT is able to sensitively capture information regarding cerebral ischemia-caused brain metabolic changes (Shimoji et al., 2004). Therefore, it can be used for early diagnosis and localization of transient ischemic attack and cerebral infarction and later therapeutic effect evaluation and prognosis judgment (Shimoji et al., 2004).

Integrin receptor imaging is an area of interest in nuclear medicine. The concentration integrin receptors will be changed in cells under the condition of ischemic/hypoxic injury compared to in normal cells. Isotope labeling of integrin receptor ligand can enable integrin receptor imaging (Sandoval and Witt, 2008). Integrin α v β 3 receptor participates in many physiological and pathological processes including tumor angiogenesis, tumor invasion and metastasis, inflammation, wound healing, and blood clotting (Xu et al., 2015). It is hardly expressed in mature vascular endothelial cells and the majority of normal organs, but its expression is increased on the surface of many tumor cells and newly-formed vascular endothelial cells (Backer and Backer, 2012; Stacy et al., 2012). Arginylglycylaspartic acid (RGD) is a tripeptide composed of L-arginine, glycine, and L-aspartic acid, and can selectively bind to integrin α v β 3 receptor. The distribution of radioisotope-labeled RGD can reflect the distribution of integrin α v β 3 receptor in vivo (Cheng et al., 2005; Zheng et al., 2014). The currently reported RGD-related imaging agents include 111 In-RP748, 99m Tc-NC10069, 18 F-Galacto-RGD, 99m Tc-3PRGD2, and 68 Ga-BNOTA-PRGD2 (Bach-Gansmo et al., 2006; Li et al., 2008; Liu et al., 2010; Zhu et al., 2012). 68 Ga-BNOTA-PRGD2 can specifically aggregate on newly-formed vascular endothelial cells, making them visible (Vatsa et al., 2015). 68 Ga-BNOTA-PRGD2 is not influenced by cell metabolism and therefore it can provide valuable diagnosis information when 18 F-FDG PET/CT imaging cannot provide identified diagnosis (Vatsa et al., 2015). We aimed to perform an open-label trial to investigate the efficacy of 68 Ga-BNOTA-PRGD2 PET/CT imaging in visualizing post-stroke angiogenesis.

  Methods/Design Top

Study design

A prospective open-label clinical trial was performed at the PET Center, Department of Nuclear Medicine, Peking Union Medical College Hospital, China ([Figure 1]).
Figure 1: Flow chart of the study protocol.

Click here to view


The benefits and risks of participation in the trial were explained to each patient, family members and guardians, and signed informed consent was obtained before the trial. The trial protocol was approved by the ethics committee of Peking Union Medical College Hospital, Chinese Academy of Medical Sciences, China (approval No. S-417) and will be performed in accordance with the Declaration of Helsinki, formulated by the World Medical Association.

Study participants

Fifty patients with stroke who will be scheduled to receive treatment in the Peking Union Medical College Hospital, Chinese Academy of Medical Sciences, China, and meet all of the inclusion criteria, will be considered for admission to the trial.

Inclusion criteria

  • Clinically diagnosed stroke, presenting with the symptoms of hemiplegia, unilateral sensory disturbance and/or hemianopsia (Saposnik et al., 2011)
  • Patients with stroke within 2 weeks of onset
  • 30-70 years of age, of either sex
  • Agree to undergo brain 68 Ga-BNOTA-PRGD2 PET/CT and 18 F-FDG PET/CT besides conventional CT and MRI imaging
  • Signed informed consent by patients or their guardians/legal deputy

Exclusion criteria

  • Have brain tumors or other brain diseases
  • Females planning to bear a child recently or with childbearing potential
  • Renal function: serum creatinine level > 3.0 mg/dL
  • Liver function: any hepatic enzyme level more than 5 times upper limit of normal
  • Known severe allergy or hypersensitivity to IV radiographic or MRI contrast
  • Patients unable to enter the bore of the PET/CT scanner
  • Inability to lie still for the entire imaging time because of cough, pain, etc.
  • Inability to complete the needed examinations due to severe claustrophobia, radiation phobia, etc.

Imaging examination

68 Ga-BNOTA-PRGD2 synthesis

BNOTA-PRGD2 freeze-dried powder will be dissolved in deionized water at 1 μg/μL, packaged and preserved at 4°C for later use. The germanium-gallium generator will be eluted with 4 mL of 0.05 M HCl and the prior 1 mL gallium eluate will be discarded and the next 1.5 mL gallium eluate will be fully mixed with NaAc solution at a ratio of 1 mL gallium eluate: 1.25 M NaAc solution. Then 40 μL of BNOTA-PRGD2 will be added to the mixture and heated at 100°C for 15 minutes. After natural cooling at room temperature, 68 Ga-BNOTA-PRGD2 preparation will be purified using 0.22 μm-sized sterile filter membrane (Li et al., 2008; Lang et al., 2011). Identification of radio-metabolites will be performed by a radioactive thin-layer liquid chromatography (Bioscan ® , Poway CA, USA). The ratio of CH3OH and NH 4 Ac will be at 1:1 and silicone paper (Varian ® , Palo Alto, CA, USA) will be used (Decristoforo et al., 2007).

68 Ga-BNOTA-PRGD2 PET/CT examination

Prior to examination, bladder evacuation will be performed. A clinical PET/CT scanner (Biograph 64 TruePoint TrueV ® , Siemens Medical Solutions, Erlangen, Germany) will be used. Twenty minutes after intravenous administration of 0.5 mCi/18.5MBq/kg 68 Ga-BNOTA-PRGD2, the patient will be asked to lie supine to undergo 30 mAs brain scans with the following parameters: tube voltage 140 kV, tube current 30 mAs, pitch 1:1, scanning layer thickness 3 mm, section thickness 3 mm, matrix 512 × 512, field of view 70 cm. Thereafter, PET images of the head will be acquired at 10 minutes per bed position.

18 F-FDG PET/CT examination

On the day of examination, the patient will be asked to fast for at least 4 hours and blood glucose level should be controlled within 3.0-8.0 mM. After intravenous administration of 1.5 mCi/kg 18 F-FDG, patients will be asked to rest in the environment away from the light for 50-60 minutes. After bladder emptying, 18 F-FDG PET/CT examination of the head will be performed. Under the data collection condition the same as 68 Ga-BNOTA-PRGD2, low-dose CT and PET scans of the head will be performed.

Image processing and analysis

68 Ga-BNOTA-PRGD2 images will be reconstructed using the optimized parameters for conventional clinical use. 68 Ga-BNOTA-PRGD2 PET/CT, and 18 F-FDG PET/CT images will be introduced into the Siemens MMWP Workstation for comparisons. Image comparison will be performed by 3 professional nuclear medicine physicians. To reduce measurement errors, semi-quantitative analysis will be performed by one physician according to the following criteria: If there are several high-uptake areas, areas of the most obvious interest determined based on visual analysis results will be selected, and 80% of the maximal standardized uptake value (SUV) will be taken as threshold. The mean uptake value in this area of interest will be determined. Using a mirror image method, the mean uptake value in the contralateral area will be also determined. The mean uptake value of the area of interest on the lesioned side to the contralateral side will be calculated.

Measurement parameters

Primary measures

68 Ga-BNOTA-PRGD2 PET/CT results are considered the primary outcome and presented as the SUV in the cerebral infarction area and the SUV ratio of the injured side to the contralateral side on 68 Ga-BNOTA-PRGD2 PET/CT images (SUV = tracer activity in unit volume diseased tissue/injected dose/body mass).

Secondary measures

(1) 18 F-FDG PET/CT results, which are presented as the SUV in the cerebral infarction area and the SUV ratio of the injured side to the contralateral side on 18 F-FDG PET/CT; (2) enhanced brain MRI or CT images used to evaluate post-stroke vasculature.

The timing of trial outcome evaluation is shown in [Table 1].
Table 1: Trial outcome evaluation schedule

Click here to view

Withdrawal management

If patients or their relatives do not agree to the following trial during the follow-up interview, the patients will be withdrawn from the trial. The researchers can judge whether the participants are suitable for further participation in the following trial. If some patients will be withdrawn from the trial, to ensure the integrity of the trial, patients will be recruited to supplement those withdrawn from the trial until meeting the trial requirement.

Safety evaluation

Safety evaluation indices include blood pressure, respiratory rate, body temperature, blood test, urine test, hepatic function and renal function. For those events that do not immediately threaten the patient's life or lead to death or hospitalization, but can harm patients and needs measures to prevent, the onset time, severity, lasting time, measures and turnover relating to these adverse events should be recorded. In addition, the criteria used for assess the severity of these adverse events should be also indicated. Any adverse event occurring during the clinical trial should be reported to researchers in charge, ethics committee and sponsors.

Data management

Trial-related medical data will be collected by a research associate and input into a computer which should not be connected to the internet. The computer will be managed by a designated person and information saved in this computer should not be disclosed to any person without permission. The information, no matter paper or electrical version, should be saved in a safe, locked place at least for 5 years.

All data will be statistically processed using commercial software (SPSS16.0 ® , SPSS, Inc. Chicago IL, USA). Measurement data will be expressed as the mean ± SD. Repeated measures analysis of variance will be used to compare the SUV in the cerebral infarction area on 68 Ga-BNOTA-PRGD2 PET/CT and 18 F-FDG PET/CT images across different time points. Chi-square test or Fisher's exact test will be used for comparison of numeration data. A level of P < 0.05 will be considered statistically significant.

  Discussion Top

68 Ga-BNOTA-PRGD2 cannot pass through an intact blood-brain barrier, so uptake value of the normal intracranial tissue will be relatively low, allowing for high target-to-normal ratios. A high target-to-normal ratio of 68 Ga-BNOTA-PRGD2 uptake is likely to exist in the diseased region because of destroyed blood-brain barrier (Sandoval and Witt, 2008). In addition, integrin α v β 3 receptor is highly expressed on the surface of macrophages, in particular during the process of post-stroke repair (Friggeri et al., 2010; Furundzija et al., 2010) and therefore integrin α v β 3 receptor aggregation on the surface of macrophage is another factor influencing 68 Ga-BNOTA-PRGD2 uptake. 68 Ga-BNOTA-PRGD2 PET/CT images can be used for evaluation of post-stroke angiogenesis because of the advantages of 68 Ga-BNOTA-PRGD2 including easy synthesis, simple operation, and high production rate.

Trial status

Recruitment of participants at the time of submission.

Preliminary data

Related studies of the team led by ZHZ are listed in [Table 2].
Table 2: PET/CT findings regarding 68Ga-containing tracer from the team led by ZHZ

Click here to view

Conflicts of interest

None declared.

Author contributions

ZZ, YS, CW and HW conceived and designed this study, wrote the paper, read and approved the final version of this paper.

Plagiarism check

This paper was screened twice using CrossCheck to verify originality before publication.

Peer review

This paper was double-blinded and stringently reviewed by international expert reviewers.[35]

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  [Figure 1]

  [Table 1], [Table 2]


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