"Crossbreeding" NIR-II flavchromene for PSMA-positive prostate cancer detection and image-guided surgery

Abstract

Prostate-specific membrane antigen (PSMA) is known to be overexpressed in prostate cancer (PCa). The development of precise and rapid imaging technologies to monitor PSMA is crucial for early diagnosis and therapy. Fluorescence imaging in the second near-infrared window (NIR-II) has emerged as a powerful tool for real-time tracking and in vivo visualization, offering high sensitivity and resolution. However, there is a lack of stable, bright and easy-to-implement NIR-II fluorescent probes for PSMA targeting. Herein, we presented a PSMA-targeting NIR-II fluorescent probe FC-PSMA based on π-conjugated crossbreeding dyed strategy that affords high stability, large extinction coefficient, and good brightness. As demonstrated, FC-PSMA displayed a high fluorescence quantum yield in fetal bovine serum (FBS). Following intravenous injection of FC-PSMA, the tumor-to-normal ratio of fluorescence intensity steadily increased over time, reaching a peak at 48 h (tumor-to-leg ratio = 12.16 ± 0.90). This advancement enables precise identification of PC through NIR-II fluorescence imaging, facilitating high-performance guidance for prostate cancer resection surgery.

Keywords: NIR‐II fluorescence imaging; PSMA‐targeting; fluorescence dyes; near‐infrared.

FIGURE 1

Design of π‐conjugated crossbreeding NIR‐II flavchromene with PSMA‐targeting. (a) Donor‐acceptor (d–a) fluorophores with strong push‐pull electronic effect. (b) π‐conjugated crossbreeding strategy based on Flavylium and Chromene units for constructing NIR‐II dye FC‐NEt₂. (c) The PSMA‐targeting NIR‐II dye FC‐PSMA based on FC‐NEt₂ scaffold for prostate cancer imaging and fluorescence‐guided surgery.

FIGURE 2

The absorption spectra (a), normalized emission spectra (b) and photophysical properties (c) of Flav‐N, Flav‐FN and FC‐NEt₂ taken in DMSO at 10 μM (λ _ex = 650, 690 and 830 nm, respectively). (d) Photostability of NIR‐II dyes Flav‐FN, FC‐NEt₂ and Flav7 (10 μM) in PBS/DMSO (pH 7.4, 1:1, v/v) under continuous‐wave laser exposure with a power density of 1 W cm⁻² (690, 785, and 980 nm laser, respectively). (e) Chemical stability of Flav‐FN, FC‐NEt₂ and Flav7 (10 μM) with various agents in PBS/DMSO (pH 7.4, 1:1, v/v), Cys: 100 μM; GSH: 1 mM; H₂O₂: 100 μM; HClO: 100 μM.

FIGURE 3

(a) Synthesis route of PSMA‐targeting NIR‐II fluorescent probe FC‐PSMA. (b) The normalized absorption and emission spectra of FC‐PSMA taken in DMSO at 10 μM, λ _ex = 808 nm. (c) Table of photophysical properties of FC‐PSMA in different solvents. The absorption (d) and emission spectra (e) of FC‐PSMA in FBS at 10 μM, λ _ex = 808 nm. (f) NIR‐II fluorescence images of FC‐PSMA in FBS with various long‐pass filters at the same exposure time, λ _ex = 808 nm.

FIGURE 4

In vivo PSMA‐targeting performance of FC‐PSMA. (a) NIR‐II fluorescence imaging of LNCap tumor‐bearing mice after tail vein injection of FC‐PSMA, FC‐PSMA + 2‐PMPA and FC‐NEt₂. (b) NIR‐II fluorescence imaging of the tumor and major organs (heart, liver, spleen, kidney and lung) after tail vein injection for 48 h. The fluorescence intensity of tumor‐to‐leg ratio (c) and tumor‐to‐liver ratio (d). (e) The normalized fluorescence intensity of major organs. 2‐PMPA is an effective selective Glutamate Carboxypeptidase II inhibitor that binds to PSMA. Laser: 808 nm; Filter: 1000 nm long‐pass; Exposure time: 100 ms.

FIGURE 5

(a) Schematic illustration of NIR‐II image‐guided surgery. (b) NIR‐II fluorescence imaging‐guided tumor resection after tail vein injection of FC‐PSMA for 48 h. Laser: 808 nm; Exposure time: 100 ms; Filter: 1000 nm long‐pass. (c) Tumor‐to‐normal ratio during the surgery.