Abstract
The urokinase-type plasminogen activator receptor (uPAR) is overexpressed in several cancers including glioblastoma (GBM) and is an established biomarker for metastatic potential. The uPAR-targeting peptide AE105-NH2 (Ac-Asp-Cha-Phe-(D)Ser-(D)Arg-Tyr-Leu-Trp-Ser-CONH2) is a promising candidate for non-invasive positron emission tomography (PET) imaging of uPAR. Despite the optimal physical properties of 68Ga for peptide-based PET imaging, low tumor uptakes have previously been reported using 68Ga-labeled AE105-NH2-based tracers. In an attempt to optimize the tumor uptake, we developed three novel tracers with alkane (AOC) and polyethylene glycol (PEG) spacers inserted between AE105-NH2 and the radio metal chelator 2-(4,7-bis(carboxymethyl)-1,4,7-triazonan-1-yl)pentanedioic acid (NODAGA). The resulting tracers NODAGA-AOC-AE105-NH2, NODAGA-PEG3-AE105-NH2 and NODAGA-PEG8-AE105-NH2 were compared to the non-spacer version, NODAGA-AE105-NH2. Following radiolabeling with 68Ga, we evaluated the in vitro and in vivo performance in mice bearing subcutaneous tumors derived from the uPAR-expressing human GBM cell line U87MG. In vivo PET/CT imaging showed that introduction of PEG spacers more than doubled the in vivo tumor uptake after 1 h compared with the non-spacer version: 68Ga-NODAGA-PEG3-AE105-NH2 (2.08 ± 0.37%ID/g) and 68Ga-NODAGA-PEG8-AE105-NH2 (2.01 ± 0.22%ID/g) vs. 68Ga-NODAGA-AE105-NH2 (0.70 ± 0.40%ID/g), p < 0.05. In addition, 68Ga-NODAGA-PEG8-AE105-NH2 showed significantly higher (p < 0.05) tumor-to-background contrast (3.68 ± 0.23) than the other tracers. The specific tumor-targeting property of 68Ga-NODAGA-PEG8-AE105-NH2 was established by effectively blocking the tumor uptake with co-injection of unlabeled AE105-NH2 (1 h: unblocked 2.01 ± 0.22%ID/g vs. blocked 1.24 ± 0.09%ID/g, p < 0.05). Ex vivo biodistribution confirmed the improved tumor uptakes of the PEG-modified tracers. 68Ga-NODAGA-PEG8-AE105-NH2 is thus a promising candidate for human translation for PET imaging of GBM.
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Abbreviations
- AE105-NH2 :
-
Ac-Asp-Cha-Phe-(D)Ser-(D)Arg-Tyr-Leu-Trp-Ser-CONH2
- AE105-NH2-mut :
-
Arg-Cha-Glu-(D)Ser-(D)-Arg-Tyr-Leu-Glu-Ser-CONH2
- AOC:
-
Amino-octanoic acid
- Azido-PEG3 :
-
11-azido-3,6,9-trioxaundecan-1-amine
- Azido-PEG8 :
-
O-(2-Aminoethyl)-O′-(2-azidoethyl)heptaethylene glycol
- BCN-NHS:
-
(1R,8S,9 s)-Bicyclo[6.1.0]non-4-yn-9-ylmethyl N-succinimidyl carbonate
- DIPEA:
-
N,N-diisopropylethylamine
- Fmoc-AOC:
-
8-(Fmoc-amino)octanoic acid
- GBM:
-
Glioblastoma
- NODAGA-NHS:
-
2,2′-(7-(1-Carboxy-4-((2,5-dioxopyrrolidin-1-yl)oxy)-4-oxobutyl)-1,4,7-triazonane-1,4-diyl)diacetic acid
- PEG3 :
-
Tri-ethylene glycol
- PEG8 :
-
Octa-ethylene glycol
- p.i.:
-
Post-injection
- ROI:
-
Region of interest
- uPAR:
-
Urokinase-type plasminogen activator receptor
- %ID/g:
-
Percent injected dose per gram
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All applicable international, national, and/or institutional guidelines for the care and use of animals were followed. All procedures performed in studies involving animals were in accordance with the ethical standards of the institution or practice at which the studies were conducted.
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The authors declare that they have no conflicts of interest.
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This work was partially supported by the Office of Science (BER), U.S. Department of Energy (DE-SC0008397), National Institute of Health In vivo Cellular Molecular Imaging Center (ICMIC) grant P50 CA114747, the Lundbeck Foundation Clinical Research Fellowship Grant, the Novo Nordisk Foundation, the Innovation Fund Denmark, the Research Foundation of Rigshospitalet, the Research Foundation of the Capital Region of Denmark, the Svend Andersen Foundation, the Arvid Nilsson Foundation and the Danish Cancer Society.
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M. D. Loft and Y. Sun contributed equally to this work.
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726_2017_2407_MOESM1_ESM.pdf
Fig. S1a MALDI-TOFMS data for NODAGA-AE105-NH2. Fig. S1b MALDI-TOFMS data for NODAGA-AOC-AE105-NH2. Fig. S1c MALDI-TOFMS data for NODAGA-PEG3-AE105-NH2. Fig. S1d MALDI-TOFMS data for NODAGA-PEG8-AE105-NH2. Fig. S2 Radio-HPLC data for 68Ga-NODAGA-X-AE105-NH2 (X = 0, AOC, PEG3 or PEG8). Fig. S3 PET imaging derived organ uptakes for 68Ga-NODAGA-PEG8-AE105-NH2 with and without blocking (co-injection of AE105-NH2). Table S1 60 min ex vivo biodistribution data for 68Ga-NODAGA-X-AE105-NH2 (X = 0, AOC, PEG3 or PEG8) (PDF 1529 kb)
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Loft, M.D., Sun, Y., Liu, C. et al. Improved positron emission tomography imaging of glioblastoma cancer using novel 68Ga-labeled peptides targeting the urokinase-type plasminogen activator receptor (uPAR). Amino Acids 49, 1089–1100 (2017). https://doi.org/10.1007/s00726-017-2407-4
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DOI: https://doi.org/10.1007/s00726-017-2407-4