Tricyano-Methylene-Pyridine Based High-Performance Aggregation-Induced Emission Photosensitizer for Imaging and Photodynamic Therapy

Abstract

Photosensitizers equipped with high reactive oxygen species (ROS) generation capability and bright emission are essential for accurate tumor imaging and precise photodynamic therapy (PDT). However, traditional aggregation-caused quenching (ACQ) photosensitizers cannot simultaneously produce desirable ROS and bright fluorescence, resulting in poor image-guided therapy effect. Herein, we report an aggregation-induced emission (AIE) photosensitizer TCM-Ph with a strong donor-π-acceptor (D-π-A) structure, which greatly separates the HOMO-LUMO distribution and reduces the ΔE_ST, thereby increasing the number of triplet excitons and producing more ROS. The AIE photosensitizer TCM-Ph has bright near-infrared emission, as well as a higher ROS generation capacity than the commercial photosensitizers Bengal Rose (RB) and Chlorine e6 (Ce6), and can effectively eliminate cancer cells under image guidance. Therefore, the AIE photosensitizer TCM-Ph has great potential to replace the commercial photosensitizers.

Keywords: ROS generation; aggregation-induced emission; bioimaging; photosensitizer.

Figure 1

Design of the high-performance aggregation-induced emission photosensitizer TCM-Ph for imaging and photodynamic therapy. (A) Rational design of the AIE photosensitizer TCM-Ph; (B) TCM-Ph nanoparticles (TCM-Ph NPs) could image cancer cells and produce ROS upon light irradiation to kill cancer cells.

Figure 2

Photophysical properties of TCM-Et and TCM-Ph. (A) Absorption spectra of TCM-Et in 99% water; (B) Fluorescence spectra and (C) I/I₀ plots of TCM-Et in THF/water mixtures with different water fractions (f_w), λ_ex = 511 nm. Inset: photographs of TCM-Et in 0% and 80% water under UV lamp (365 nm). I₀ is the maximum fluorescence intensity of TCM-Et in THF. (D) Size distribution of TCM-Et in 99% water; (E) Absorption spectra of TCM-Ph in 99% water; (F) Fluorescence spectra and (G) I/I₀ plots of TCM-Ph in THF/water mixtures with different f_w, λ_ex = 510 nm. Inset: photographs of TCM-Ph in 0% and 70% water under UV lamp (365 nm). I₀ is the maximum fluorescence intensity of TCM-Ph in THF. (H) Size distribution of TCM-Ph in 99% water.

Figure 3

High ROS generation of TCM-Ph. (A) Light-induced total ROS generation of Ce6, RB, TCM-Et and TCM-Ph (10 μM) with DCFH (40 μM) as indicator. The plot of relative PL intensity (I-I₀)/I₀ at 525 nm versus the different irradiation times, where I₀ is the fluorescence value of the mixture at 525 nm before illumination, and I is the fluorescence value of the mixture at 525 nm after illumination; excitation wavelength is 488 nm. (B) Light-induced ¹O₂ generation of Ce6, RB, TCM-Et and TCM-Ph (10 μM), evaluated by ABDA (50 μM). The relationship between A/A₀ of ABDA at 378 nm versus the different irradiation times. (C) The relationship between ln(A₀/A) and the light irradiation times, where A₀ is the absorbance of ABDA at 378 nm before light irradiation, and A is the absorbance of ABDA at 378 nm after light irradiation. (D) Molecular orbital amplitude plots of HOMO and LUMO. (E) The energy levels of TCM-Et and TCM-Ph. The well vibration-resolved profiles allow accurate energy level abstractions based on the 0–0 peaks for solution samples in 2-methyl-tetrahydrofuran at 77 K.

Figure 4

Efficient cell penetrating and tumor cell ablation of TCM-Ph NPs. (A) Size distribution of TCM-Ph NPs in water. (B) Absorbance and fluorescence spectra of TCM-Ph NPs in water, λ_ex = 497 nm. (C) Light-induced total ROS generation of Ce6, RB, and TCM-Ph NPs, evaluated by DCFH (40 μM). The plot of relative PL intensity (I-I₀)/I₀ at 525 nm versus the different irradiation times, where I₀ is the fluorescence value of the mixture at 525 nm before illumination and I is the fluorescence value of the mixture at 525 nm after illumination, respectively, λ_ex = 488 nm. (D) CLSM of HeLa cells incubated with TCM-Ph NPs (10 μM based on TCM-Ph) for different times. Red channel from TCM-Ph NPs: λ_ex = 514 nm, λ_em = 550–700 nm. Scale bar: 25 μm. (E) Average fluorescence intensity of cells incubated with TCM-Ph NPs (10 μM based on TCM-Ph) for different times from Figure (D) (down). Data are shown as mean ± s.d., with n = 3, **** p < 0.0001. (F) Intracellular ROS level in HeLa cells under various treatments detected by reaction between DCFH and ROS. Green channel from DCFH: λ_ex = 488 nm, λ_em = 505–560 nm. Scale bar: 50 μm. (G) In vitro cytotoxicity of different concentrations of TCM-Ph NPs under dark, 10 min and 20 min white-light irradiation. Light: 50 mW cm⁻², > 400 nm. Data are shown as mean ± s.d., with n = 3, **** p < 0.0001, ** p < 0.01.