A radiofluorinated divalent cystine knot peptide for tumor PET imaging.
A divalent knottin containing two separate integrin binding epitopes (RGD) in the adjacent loops, 3-4A, was recently developed and reported in our previous publication. In the current study, 3-4A was radiofluorinated with a 4-nitrophenyl 2-(18)F-fluoropropinate ((18)F-NFP) group and the resulting divalent positron emission tomography (PET) probe, (18)F-FP-3-4A, was evaluated as a novel imaging probe to detect integrin αvβ3 positive tumors in living animals. Knottin 3-4A was synthesized by solid phase peptide synthesis, folded, and site-specifically conjugated with (18/19)F-NFP to produce the fluorinated peptide (18/19)F-fluoropropinate-3-4A ((18/19)F-FP-3-4A). The stability of (18)F-FP-3-4A was tested in both phosphate buffered saline (PBS) buffer and mouse serum. Cell uptake assays of the radiolabeled peptides were performed using U87MG cells. In addition, small animal PET imaging and biodistribution studies of (18)F-FP-3-4A were performed in U87MG tumor-bearing mice. The receptor targeting specificity of the radiolabeled peptide was also verified by coinjecting the probe with a blocking peptide cyclo(RGDyK). Our study showed that (18)F-FP-3-4A exhibited excellent stability in PBS buffer (pH 7.4) and mouse serum. Small animal PET imaging and biodistribution data revealed that (18)F-FP-3-4A exhibited rapid and good tumor uptake (3.76 ± 0.59% ID/g and 2.22 ± 0.62% ID/g at 0.5 and 1 h, respectively). (18)F-FP-3-4A was rapidly cleared from the normal tissues, resulting in excellent tumor-to-normal tissue contrasts. For example, liver uptake was only 0.39 ± 0.07% ID/g and the tumor to liver ratio was 5.69 at 1 h p.i. Furthermore, coinjection of cyclo(RGDyK) with (18)F-FP-3-4A significantly inhibited tumor uptake (0.41 ± 0.12 vs 1.02 ± 0.19% ID/g at 2.5 h) in U87MG xenograft models, demonstrating specific accumulation of the probe in the tumor. In summary, the divalent probe (18)F-FP-3-4A is characterized by rapid and high tumor uptake and excellent tumor-to-normal tissue ratios. (18)F-FP-3-4A is a highly promising knottin based PET probe for translating into clinical imaging of tumor angiogenesis.
Department of Nuclear Medicine, Zhongshan Hospital, Fudan University , 180 Fenglin Road, Shanghai, China 200032.
Lei
L
Jiang
Richard H
RH
Kimura
Xiaowei
X
Ma
Yingfeng
Y
Tu
Zheng
Z
Miao
Bin
B
Shen
Frederick T
FT
Chin
Hongcheng
H
Shi
Sanjiv Sam
SS
Gambhir
Zhen
Z
Cheng
D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D000818:Animals; D045744:Cell Line, Tumor; D002851:Chromatography, High Pressure Liquid; D003545:Cysteine / Q000737:chemistry*; D057771:Cystine-Knot Miniproteins / Q000737:chemistry; D000939:Epitopes / Q000737:chemistry; D005260:Female; D005462:Fluorine Radioisotopes / Q000737:chemistry*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D020128:Inhibitory Concentration 50; D039302:Integrin alphaVbeta3 / Q000378:metabolism; D008099:Liver / Q000000981:diagnostic imaging; D051379:Mice; D008819:Mice, Nude; D008969:Molecular Sequence Data; D009368:Neoplasm Transplantation; D009369:Neoplasms / Q000000981:diagnostic imaging; D009842:Oligopeptides / Q000737:chemistry; D010455:Peptides / Q000737:chemistry*; D049268:Positron-Emission Tomography; D011480:Protease Inhibitors / Q000737:chemistry; D011485:Protein Binding; D014357:Trypsin / Q000737:chemistry
D057771:Cystine-Knot Miniproteins; D000939:Epitopes; D005462:Fluorine Radioisotopes; D039302:Integrin alphaVbeta3; D009842:Oligopeptides; D010455:Peptides; D011480:Protease Inhibitors; C047981:arginyl-glycyl-aspartic acid; D014357:Trypsin; D003545:Cysteine
18F; PET; cystine-knot peptide; divalent; integrin αvβ3