Fluolab【Angew.Chem.】大连理工彭孝军、樊江莉、杜健军|87.4%超高转化率！连续扭转分子内电荷转移机制助力自噬阻断光热治疗
通讯作者： Jianjun Du, Jiangli Fan, Xiaojun Peng
文章概要
引言
光热治疗（PTT）作为一种非侵入性且不依赖氧气的肿瘤治疗手段，其核心在于光热试剂的光热转换效率（PCE）。然而，现有的有机光热试剂往往面临辐射跃迁和三线态能级转移的竞争，导致非辐射衰减途径受限，通常需要超过皮肤耐受阈值的高功率激光才能达到治疗效果。为了突破这一瓶颈，大连理工大学彭孝军院士团队提出了一种连续扭转分子内电荷转移（ConTICT） 的新策略。该研究通过精准调控分子激发态的能量流动，设计出能够实现极速非辐射衰减循环的新型分子，旨在安全低功率的激光照射下实现高效的肿瘤消融，并探索其在生物机制层面如阻断细胞自噬方面的应用潜力。
ConTICT mediated photothermal therapy process. (a) The structure of Cy-R (Cy-H, Cy-Ph, and Cy-CF3) and a ConTICT promoted process for photothermal control. (b) The process of ConTICT. (c) The process of nanoparticle assembly and intracellular autophagy blocking.
主要实验及结论
研究人员通过在五甲川花菁染料的β位引入苯环及三氟甲基（-CF₃）转子，成功合成了新型光热试剂Cy-CF₃。实验结果显示，该分子在水溶液中表现出惊人的光热转换效率，高达87.4%，远超对比分子Cy-Ph和Cy-H。通过超快瞬态吸收光谱分析发现，Cy-CF₃在10纳秒内可完成112次光热循环，其极短的激发态寿命有效加速了能量从激发态向地面的回归，这种高频循环机制是热量高效产生的关键。此外，计算化学模拟证实，三氟甲基不仅作为强吸电子基团增强了电荷转移效应，其旋转自由度也进一步降低了能垒，诱导分子进入TICT状态并迅速通过振动弛豫释放热能。
Spectral and photothermal properties of dyes. (a) Normalized absorption spectrum of dyes in MeOH. (b) Normalized emission spectrum of the dye in MeOH. (c) Emission spectra at different viscosities (1,5-pentanediol: 128 cp; Ethanol: 1.17 cp). The heating–cooling curve and time constant (τ) of Cy-CF3 (d), Cy-Ph (e), and Cy-H (f). (g) Photothermal stability of dyes during ten circles of heating-cooling processes (760 nm, 100 µM, 250 mW cm−2). (h) Photothermal images of solutions at different times (760 nm, 100 µM, and 300 mW cm−2). (i) Temperature curves under different laser powers of Cy-CF3 (760 nm, 100 µM).
Femtosecond transient absorption spectra and kinetic fitting results. Transient absorption spectra of Cy-CF3 (a), Cy-Ph (b), and Cy-H (c) in dichloromethane ( = 350 nm). 2D mapping of Cy-CF3 (d), Cy-Ph (e), and Cy-H (f). Kinetic traces and fitting lines of Cy-CF3 (g), Cy-Ph (h), and Cy-H (i) at the representative ESA and GSB wavelengths. Population distribution analysis of Cy-CF3 (j), Cy-Ph (k), and Cy-H (l).
Analysis of theoretical simulation results. (a) The photophysical processes of the dye. Schematic diagrams of TICT (b) and non-TICT (c) processes in cyanine dyes. Schematic diagram of timescales in photophysical processes of Cy-CF3 (d), Cy-Ph (e), and Cy-H (f). The dependence of the total energy of Cy-CF3 (g), Cy-Ph (h), and Cy-H (i) in the S0 and S1 states on the dihedral angle φ. RMSD and ROE of Cy-CF3 (j), Cy-Ph (k), and Cy-H (l).
The inhibition of autophagy in 4T1 cells by Cy-CF3. (a) The uptake images of 4T1 cells (30 µm). (b) The colocalization images of Cy-CF3 (Red) with commercial lysosomal dyes (Green) with 4T1 cells (30 µm). (c) Dark toxicity and phototoxicity assays of different molecules in 4T1 cells (760 nm, 300 mW cm−2). (d) Red fluorescence quantification of AO stained 4T1 cells (50 µM, 760 nm, and 300 mW cm−2). (e) Immunofluorescence of P62 and LAMP1 proteins in different groups (50 µM, 30 µm). (f) Quantitative analysis of MDC staining (50 µM, 760 nm, and 300 mW cm−2). (g) The ratio of red to green fluorescence intensity of JC-1 among different groups (50 µM, 760 nm, and 300 mW cm−2). (h) Western blot analysis of LC3B-II/LC3B-I, p62, and GAPDH under various treatments. (i) Bio-TEM images of 4T1 cells under various treatments. Red arrows indicated autolysosomes, and yellow arrows were autophagosomes (1 µm). Data were expressed as mean ± SD, n = 3. *p<0.05, **p<0.01, ***p<0.001, ****p<0.0001 determined by Student's t-test.
在细胞实验层面，Cy-CF₃展现了卓越的溶酶体靶向能力。光照射产生的剧烈热效应引发了溶酶体膜通透化（LMP），导致溶酶体内的钙离子和水解酶大量释放。这一过程破坏了细胞内的自噬流，导致自噬底物P62蛋白堆积和自噬小体无法降解，从而触发细胞发生不可逆的凋亡。动物实验进一步证实，封装了Cy-CF₃的脂质体纳米颗粒在小鼠体内具有显著的肿瘤富集效果，在仅300 mW/cm²（低于皮肤安全阈值）的激光照射下，肿瘤抑制率达到了96%。更为重要的是，该疗法展现了对乳腺癌肺转移的强大抑制能力，肺部转移结节减少了93%，有效激活了体内的抗肿瘤免疫反应。
RNA sequencing and bioinformatic analysis. (a) Volcano plot of differentially expressed RNAs in photothermal therapy vs. control (|log2FC| > 1, padj < 0.05). (b) Clustered heatmap of differentially expressed genes. (c) GO enrichment analysis of DEGs (BP: biological process; CC: cellular component; MF: molecular function). (d) KEGG pathway enrichment analysis.
Preparation and imaging performance of Cy-CF3 nanoparticles. (a) Illustration of the preparation process of nanoparticles. (b) TEM image of Cy-CF3 nanoparticles (200 nm). (c) DLS and Zeta potential measurements of Cy-CF3 nanoparticles. (d) Fluorescence imaging images of mice (1 cm). (e) Photoacoustic capability of the nanoparticle solution and (f) in vivo photoacoustic imaging at different time (5 mm). (g) 3D photoacoustic imaging of the tumor site in mice (5 mm). (h) Photothermal imaging effect in mice under 760 nm laser irradiation (300 mW cm−2).
Tumor suppression assay in mice. (a) Schematic Diagram of the Mouse Treatment Schedule. (b) The changes in body weight of mice during the treatment period. (c) Changes in tumor weight at the end of the treatment period. (d) Changes in tumor volume for each mouse during the treatment period. (e) Tumor H&E staining and immunofluorescence staining for TUNEL and Ki67 (100 µm). Data were expressed as mean ± SD, n = 5. *p<0.05, **p<0.01, ***p<0.001, ****p<0.0001 determined by Student's t-test.
The inhibition of tumor lung metastasis of Cy-CF3 NPs with PTT. (a) Changes in tumor volume for each mouse during the treatment period. (b) The weight changes of mice in each group. (c) The tumor weight of each groups (The I, II, III, and IV represented PBS, PBS + Light, Cy-CF3, and Cy-CF3 + Light groups). (d) Tumor photographs of the mice after the completion of the treatment cycle (1 cm). (e) Representative H&E staining images of the lungs from different treatment groups (2 mm). (f) Immunofluorescent staining of HGF, MTA2, VCAM-1, and PECAM-1 in 4T1 cells after various treatments (200 µm). (g) Quantification of the secretion levels of TNF-α, IL-1β, IL-6, IL-18, and IFN-γ in the serum from different groups after treatment. Data were expressed as mean ± SD, n = 5. *p<0.05, **p<0.01, ***p<0.001, ****p<0.0001 determined by Student's _t-_test.
总结及展望
本研究成功构建了基于ConTICT机制的高性能光热治疗平台，通过分子水平的量子能量流操纵，解决了有机光热试剂效率低和循环慢的难题。Cy-CF₃不仅在三模态成像（荧光/光热/光声） 指引下实现了精准的肿瘤消融，还通过阻断自噬这一生物学途径增强了治疗深度。这一发现为开发下一代智能光诊疗试剂提供了通用的分子设计策略，在提高癌症治疗有效性的同时，大幅提升了临床应用的安全性，对推动精准医疗和生物纳米技术的发展具有重要的科学意义。