Rapid detection of metastatic lymph nodes of colorectal cancer with a gamma-glutamyl transpeptidase-activatable fluorescence probe

Hidemasa Kubo,Ryosuke Kojima,Eigo Otsuji,Yugo Kuriki,Tasuku Ueno,Kenjiro Hanaoka,Toru Komatsu,Mako Kamiya,Yasutoshi Murayama,Yasuteru Urano

doi:10.1038/s41598-018-36062-3

Abstract

Rapid diagnosis of metastatic lymph nodes (mLNs) of colorectal cancer (CRC) is desirable either intraoperatively or in resected fresh specimens. We have developed a series of activatable fluorescence probes for peptidase activities that are specifically upregulated in various tumors. We aimed to discover a target enzyme for detecting mLNs of CRC. Among our probes, we found that gGlu-HMRG, a gamma-glutamyl transpeptidase (GGT)-activatable fluorescence probe, could detect mLNs. This was unexpected, because we have previously reported that gGlu-HMRG could not detect primary CRC. We confirmed that the GGT activity of mLNs was high, whereas that of non-metastatic lymph nodes and CRC cell lines was low. We investigated the reason why GGT activity was upregulated in mLNs, and found that GGT was induced under conditions of hypoxia or low nutritional status. We utilized this feature to achieve rapid detection of mLNs with gGlu-HMRG. GGT appears to be a promising candidate enzyme for fluorescence imaging of mLNs.

Highlights

Colorectal cancer (CRC) is one of the most common cancers worldwide[1]
We further established that the reason for this is the induction of gamma-glutamyl transpeptidase (GGT) under conditions of hypoxia and low nutritional status, resulting in high levels of GGT activity in metastatic lymph nodes (mLNs)
Among our activatable fluorescence probes for aminopeptidases and dipeptidyl peptidases, we found that gGlu-HMRG could visualize mLN in vivo (Fig. 1B–D, Supplementary Figure 1A,B)

Summary

Introduction

Colorectal cancer (CRC) is one of the most common cancers worldwide[1]. Lymph node metastasis is often present in patients with advanced CRC, and is an important prognostic factor[2]. The range of lymphadenectomy is usually decided based on preoperative diagnosis using computed tomography (CT), magnetic resonance imaging (MRI) and fluorodeoxyglucose-positron emission tomography (FDG-PET), but their sensitivity for diagnosing metastatic lymph nodes (mLNs) was reported to be only 55%, 66% and 29%, respectively[3,4]. We have recently developed a series of activatable fluorescence probes[18,19,20] that can detect cancer cells based on their overexpression of a specific enzymatic activity, resulting in a high Tumor/Normal signal ratio. These fluorescence probes can visualize cancer cells within just a few minutes after application by topical spraying. We further established that the reason for this is the induction of GGT under conditions of hypoxia and low nutritional status, resulting in high levels of GGT activity in mLNs

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Conclusion