A pre-screening strategy to assess resected tumor margins by imaging cytoplasmic viscosity and hypoxia.

Hui Huang,Weijia Zhang,Shiqiang Yan,Wenrui Ma,Youpei Lin,Qiang Gao,Meilin Tang,Chenping Zhang,Mieradilijiang Abudupataer,Xiaoyi Hu,Jing Han,Jiannan Liu

doi:10.7554/elife.70471

Abstract

To assure complete tumor removal, frozen section analysis is the most common procedure for intraoperative pathological assessment of resected tumor margins. However, during one operation, multiple biopsies may be sent for examination, but only few of them are made into cryosections because of the complex preparation protocols and time-consuming pathological analysis, which potentially increases the risk of overlooking tumor involvement. Here, we propose a fluorescence-based pre-screening strategy that allows high-throughput, convenient, and fast gross assessment of resected tumor margins. A dual-activatable cationic fluorescent molecular rotor was developed to specifically illuminate live tumor cells' cytoplasm by emitting two different fluorescence signals in response to elevations in hypoxia-induced nitroreductase (a biochemical marker) and cytoplasmic viscosity (a biophysical marker), two characteristics of cancer cells. The ability of the fluorescent molecular rotor in detecting tumor cells was evaluated in mouse and human specimens of multiple tissues by comparing with hematoxylin and eosin staining. Importantly, the fluorescent molecular rotor achieved 100 % specificity in discriminating lung and liver cancers from normal tissue, allowing pre-screening of the tumor-free surgical margins and promoting clinical decision. Altogether, this type of fluorescent molecular rotor and the proposed strategy may serve as a new option to facilitate intraoperative assessment of resected tumor margins.

Highlights

During a tumor resection operation, a pathologist examines the removed tissues to determine whether the resected margins are clear of tumor involvement
Design and synthesis of IBS440 First, we developed a dual-activatable fluorescent molecular rotor IBS440 to monitor the increased cytoplasmic viscosity and hypoxia-induced nitroreductase
The abnormal intracellular changes in viscosity and nitroreductase in tumor cells could be exploited as biomarkers for evaluation of the resected margins of multiple distinct tumors

Summary

Introduction

During a tumor resection operation, a pathologist examines the removed tissues to determine whether the resected margins are clear of tumor involvement. Targeting the tumor markers on the cell membrane, fluorescent probes have been developed to assess the resected surgical margins (Hoogstins et al, 2016, Gao et al, 2018, van Keulen et al, 2019, Koller et al, 2018, Voskuil et al, 2020). Different types of fluorescent probes have been developed to visualize either intracellular hypoxia (Liu et al, 2017, Zhao et al, 2020) or change in cytoplasmic viscosity (Kuimova et al., 2009, Hao et al, 2019, Ye et al, 2021, Wolstenholme et al, 2020, Zhou et al, 2021), which might be susceptible to fluctuations in the intracellular microenvironment. We postulate that an integrated evaluation of the biochemical (hypoxia) and biophysical (viscosity) features of tumor cells might enhance the accuracy of fluorescence-based assessment of surgical specimens

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Conclusion