Molecular Targeted NIR-II Probe for Image-Guided Brain Tumor Surgery

Abstract

Optical imaging strategies for improving delineation of glioblastoma (GBM) is highly desired for guiding surgeons to distinguish cancerous tissue from healthy and precious brain tissue. Fluorescence imaging (FLI) in the second near-infrared window (NIR-II) outperforms traditional NIR-I imaging with better tissue penetration, higher spatial and temporal resolution, and less auto fluorescence and scattering. Because of high expression in GBM and many other tumors, urokinase Plasminogen Activator Receptor (uPAR) is an attractive and well proven target for FLI. Herein we aim to combine the benefit of a NIR-II fluorophore with a high affinity uPAR targeting small peptide. A targeted NIR-II fluorescent probe was developed by conjugating an in-house synthesized NIR-II fluorophore, CH1055, and a uPAR targeting peptide, AE105. To characterize the in vivo distribution and targeting properties, a dynamic imaging was performed in orthotopic GBM bearing nude mice (n = 8). Additionally, fluorescence guided surgery of orthotopic GBM was performed in living animals. CH1055-4Glu-AE105 was easily synthesized with >75% yield and >98% HPLC evaluated purity. The retention time of the probe on analytical HPLC was 15.9 min and the product was verified by mass spectrometry. Dynamic imaging demonstrated that the uPAR targeting probe visualized orthotopic GBM through the intact skull with a tumor-to-background ratio (TBR) of 2.7 peaking at 96 h. Further, the orthotopic GBM was successfully resected in small animals guided by the NIR-II FLI. By using a small uPAR targeting NIR-II probe, FLI allows us to specifically image and detect GBM. A real-time imaging setup further renders FLI guided tumor resection, and the probe developed in this work is a promising candidate for clinical translation.

Emerging Fluorescent Molecular Tracers to Guide Intra-Operative Surgical Decision-Making

“…After evaluation in subcutaneous and orthotopic tumor models of gastric, head and neck, pancreatic (Figure 3C) and colorectal cancer, tumor-to-background values were generally optimal between 4 and 24 h post-injection, depending on the fluorophore used (see above) (Choi et al, 2013; Cheng et al, 2016; Handgraaf et al, 2017). Other promising peptides for further development are tracers against for example PSMA (Wang et al, 2014; Bao et al, 2017; Baranski et al, 2017; Kularatne et al, 2018; Schottelius et al, 2018), the gastrin releasing peptide receptor (GRPR) (Cai et al, 2013; Suganya et al, 2016; Zhang et al, 2017a; Li et al, 2018; Xu et al, 2018) and uPAR (Yang et al, 2014; Christenen et al, 2017; Kurbegovic et al, 2018). Folate-receptor specific compounds were the first small molecules to be used in clinical trials for FIGS.…”

Did evolution create a flexible ligand-binding cavity in the urokinase receptor through deletion of a plesiotypic disulfide bond?

“…Based on crystal structures of human and murine uPAR⅐ATF complexes and extensive mutagenesis (3,31,32,60), the functional hotspot residues in the ␤-hairpin of GFD for uPAR binding is well characterized in these species. The molecular basis for the species selectivity in the uPAR⅐uPA interaction between primates and nonprimate mammals is represented by the concerted replacement of the Asn 22 3 Tyr 23 and Trp 30 3 Arg 31 dyad in human and mouse uPA (32) (Fig. S9).…”

Molecular imaging of the urokinase plasminogen activator receptor: opportunities beyond cancer

“… n.v.t. In vitro preclinical [ 201 ] 99m Tc-Hynic-PEG-AE105 Peptide uPA-binding region SPECT 4-6 h In vivo preclinical [ 205 ] 64 Cu-DOTA-AE105 Peptide uPA-binding region PET 24 h Phase I clinical [ 13 , 219 – 222 , 224 ] 68 Ga-NOTA-AE105 Peptide uPA-binding region PET 10 min-1 h Phase I clinical [ 14 , 206 ] ICG-Glu-Glu-AE105 Peptide uPA-binding region Optical 6-24 h In vivo preclinical [ 207 – 209 ] CH1055-4Glu-AE105 Peptide uPA-binding region Optical 72-96 h In vivo preclinical [ 210 ] AF680-2G10 Antibody uPA-binding region Optical 48-96 h Recombinant antibody with trastuzumab Fc region In vivo preclinical [ 211 , 212 ] 111 ln-2G10 Antibody uPA-binding region SPECT 48-120 h Recombinant antibody with trastuzumab Fc region In vivo preclinical [ 211 , 212 ] AF680-3C6 Antibody β1-binding region Optical 48-96 h …”

“…This peptide, AE105, is the refined version of a 15-mer peptide identified by a phage display with uPAR-transfected cell lines and binds uPAR at the uPA-binding site in a species specific manner, like ATF [ 217 , 218 ]. While AE105 has also been conjugated with (radio)-labels for single-photon emission computed tomography (SPECT) and near-infrared fluorescence (NIRF) in preclinical oncology studies, this section will focus on positron-emission tomography as AE105 PET is further along the clinical pipeline [ 205 – 210 , 219 , 220 ]. Initially, AE105 has been conjugated with the metal chelator DOTA and subsequently labeled with 64 Cu.…”