A Method of Tumor In Vivo Imaging with a New Peptide-Based Fluorescent Probe

Abstract

Precise surgical resection directly influences the prognosis and survival of patients with solid tumors. However, it is often difficult to distinguish tumor from normal tissue during resection without any intraoperative imaging guidance. Image-guided surgery particularly when coupled with a near-infrared (NIR) fluorescent agent may improve positive-margin rate thereby improving the overall prognosis. We have developed a unique tumor-targeting fluorescence imaging agent that can aid in the accurate localization of human cancer cells in preclinical settings. The NIR imaging agent, ICG-p28, a water-soluble, nontoxic, and pan-tumor targeting probe consisting of a cell-penetrating peptide (p28) conjugated to indocyanine green (ICG), can accurately localize tumors in vivo. Development of the noninvasive, targeted imaging agent can potentially improve in the resections of tumors by enabling the localization of lesions that are currently difficult or impossible to detect by visual observation or palpation. Here, we describe the methods of preclinical animal imaging models by using NIR fluorescence imager coupled with a new tumor-targeting agent.

Keywords: Cell-penetrating peptide; In vivo tumor imaging; Indocyanine Green; Surgery.

Fig. 1

Chemical structure of ICG-p28. ICG was labeled with p28 (1:1 stoichiometry) through a linker. ICG-p28 (M.W. 3.8 kDa) is water-soluble

Fig. 2

Tumor imaging with ICG-p28. Human glioblastoma LN229 cells injected subcutaneously into male athymic mice. Once tumors developed, ICG-p28 at 5.0 mg/kg body weight was injected i.v. into LN229 xenograft tumor bearing mice, and NIR fluorescence images (gray scale at 800 nm) recorded by the Odyssey imaging system. Serial whole-body scans were recorded for 4-weeks. Tumor area highlighted with red arrows for identification

Fig. 3

The PDE neo^® imaging system. (a) a schematic diagram of the NIR fluorescence imaging system. A real-time image is monitored and recorded by PC connected to the PDE neo^® unit. (b) Photograph of the handhold NIR camera

Fig. 4

Real-time tumor imaging with the PDE neo^® system coupled with ICG-p28. Human glioblastoma LN229 cells were injected s.c. into mice. Once tumors developed, ICG-p28 at 0.2 mg/kg body weight was injected i.v. into LN229 bearing athymic mice. After 24 h post-ICG-p28 injection, mice were imaged by the clinical PDE^® system (Mitaka USA, Hamamatsu Photonics) with hand-held camera during tumor removal. Tumors were clearly identified through skin. Snapshot images of before (a) and after tumor resection (b) recorded by the PDE unit (dorsal view, 800 nm). Liver (green arrowhead) showed relatively high signal intensity as ICG excreted via liver. Red arrowheads: tumors