1. School of Pharmacy and Jiangsu Province Key Laboratory for Inflammation and Molecular Drug Target, Nantong University, 226001 Nantong, Jiangsu, PR China.
2. Department of General Surgery, Affiliated Hospital of Nantong University, 226001 Nantong, Jiangsu, PR China.
3. Department of Hepatobiliary surgery, Nantong Third People's Hospital and the Third Affiliated Hospital of Nantong University, 226001 Nantong, Jiangsu, PR China.
4. Nantong Key Laboratory of Small Molecular Drug Innovation, Nantong University, 226001 Nantong, Jiangsu, PR China.
#These three authors contributed equally to this work.
Rationale: Challenges such as developing a universal tumor-specific probe for tumor margin identification in diverse tumors with an easy-operative and fast-imaging pattern still exist. Hence, in the present study, a rapidly “off-on” near-infrared (NIR) fluorescent probe NBD with pH-activatable fluorescence and a large Stokes shift was constructed for spray mediated near-instant and precise clinical tumor margins identification.
Methods: NBD was designed and synthesized by introducing both diphenyl amino group and benzo[e]indolium to β-carboline at C-6 and C-3 positions respectively. The optical properties of NBD was characterized by absorption spectra, fluorescence spectra. Subsequently, we investigated its pH-dependent mechanism by 1H NMR and density functional theory (DFT) calculation. NBD was further under deeper investigation into its imaging performance in nude mice models (subcutaneous, orthotopic, metastatic tumor), and clinical tissues from patients with three clinically representative tumors (liver cancer, colon cancer, and lung cancer).
Results: It was found that NBD had NIR fluorescence (742 nm), a large Stokes shift (160 nm), and two-photon absorbance (1040 nm). Fluorescence quantum yield (ФF) increased by 5.5-fold when pH decreased from 7.4 to 4.0, to show pH-dependent property. Furthermore, NBD could not only selectively light up all four cancer cell lines, but also delineate xenograft tumor and orthotopic microtumor to guide surgical tumor resection, and track metastatic tissues. Particularly, after simple topical spray (three minutes later), NBD could rapidly and precisely distinguish the boundary ranges of three kinds of clinical cancer specimens including liver, colon, and lung cancers, with high tumor-to-normal tissue signal ratios (6.48~9.80).
Conclusions: Therefore, the proposed fluorescent probe NBD may serve as a versatile NIR fluorogenic spray for the near-instant visualization of tumor margins and assisting surgeons in surgerical resection of clinical cancers.
Keywords: pH-activatable, NIR fluorogenic spray, rapid and precise imaging of clinical tumor tissues, image-guided surgical resection, tumor margin identification