In Vivo Tumor Angiogenesis Imaging Using Peptide-Based Near-Infrared Fluorescent Probes

Abstract

Near-infrared fluorescence (NIRF) imaging is an emerging imaging technique for studying diseases at the molecular level. Optical imaging with a near-infrared emitting fluorophore for targeting tumor angiogenesis offers a noninvasive method for early tumor detection and efficient monitoring of tumor response to anti-angiogenesis therapy. CD13 receptor, a zinc-dependent membrane-bound ectopeptidase, plays important roles in regulating tumor angiogenesis and the growth of new blood vessels. In this chapter, we use CD13 receptor as an example to demonstrate how to construct CD13-specific NGR-containing peptides via bioorthogonal click chemistry for visualizing and quantifying the CD13 receptor expression in vivo by means of NIRF optical imaging.

Keywords: Cancer; Molecular imaging; Near-infrared fluorescence imaging; Peptide-based probes; Tumor angiogenesis.

Fig. 1

Schematic representation of the construction of peptide-based near-infrared fluorescent probe via bioorthogonal click chemistry for in vivo tumor angiogenesis imaging

Fig. 2

Schematic structure of Cy5.5-NGR2 peptide. Blue. NGR peptide; red: Cy5.5 dye. Reproduced from Ref. with permission from Springer

Fig. 3

Absorption and emission fluorescence spectra of Cy5.5-NGR2 peptide. Reproduced from Ref. with permission from Springer

Fig. 4

Confocal microscopy results of Cy5.5-NGR2 with HT-1080 cells (CD13 positive) and MCF-7 cells (CD13 negative) (magnification: 100×). The blocking study is achieved by adding unlabeled monomeric NGR peptide. Top: Incubation of Cy5.5-NGR2 (20 nM) with CD13-positive HT-1080 cells; middle: incubation of Cy5.5-NGR2 (20 nM) with CD13-positive HT-1080 cells blocked by a non-labeled monomeric NGR peptide (50 μM); bottom: incubation of Cy5.5-NGR2 (20 nM) with CD13-negative MCF-7 cells. Reproduced from Ref. with permission from Springer

Fig. 5

Time-course fluorescence imaging of subcutaneous HT-1080 tumor-bearing nude mice after intravenous injection of 1.5 nmol of Cy5.5-NGR2. The tumors are clearly visualized as indicated by an arrow from 0.5 to 4 h pi. The fluorescence intensity is recorded as per second per centimeter squared per steradian (p/s/cm²/sr). Reproduced from Ref. with permission from Springer

Fig. 6

Quantification and kinetics of in vivo targeting character of Cy5.5-NGR2 in the HT-1080 tumor vs. muscle. The Cy5.5-NGR2 uptake in HT-1080 tumor at various time points is significantly higher than that in muscle. Error bar is calculated as the standard deviation (n = 3). Reproduced from Ref. with permission from Springer

Fig. 7

Representative optical imaging (at 2 h pi) of mice bearing HT-1080 tumor on the right shoulder demonstrating blocking of Cy5.5-NGR2 (1.5 nmol) uptake by co-injection with a non-labeled monomeric NGR peptide (20 mg/kg). Reproduced from Ref. with permission from Springer

Fig. 8

Fluorescence intensity ratio of tumor to muscle based on the ROI analysis of Cy5.5-NGR2 uptake at 2 h pi in HT-1080 tumors without (non-blocking) or with (blocking) co-injection of a non-labeled monomeric NGR peptide (20 mg/kg). Error bar is calculated as the standard deviation (n = 3). Reproduced from Ref. with permission from Springer