【JACS】破纪录的160 GM双光子吸收！新型硫代核苷光敏剂DATU助力深层肿瘤光动力治疗#

文章标题：Development of a Nucleoside Photosensitizer Efficiently Activated by One- or Two-Photon Absorption in the Optical Therapeutic Window

通讯作者：Carlos E. Crespo-Hernández

文章链接：https://doi.org/10.1021/jacs.6c00216

文章概要#

引言#

在临床医学中，硫代核苷酸作为抗癌和免疫抑制药物已有超过半个世纪的应用历史。近年来，由于其独特的化学反应活性，它们在光动力治疗中展现出巨大的应用潜力。然而，传统硫代核苷酸的吸收波长大多集中在紫外A区，且在近红外区域的双光子吸收截面非常微小，这极大限制了它们在深层组织治疗中的临床应用。为了攻克这一医学界的长期难题，科研人员巧妙地设计出一种新型近红外高效激活的硫代核苷类似物DATU，成功将激活波长拓展至光学治疗窗口，为深层肿瘤的精准治疗带来了革命性的突破。

Scheme 1. Structures of DATU (5-(5-(4-(Dimethylamino)phenyl)thiophen-2-yl)-2′,3′,5′-tri-O-benzoyl-6-aza-2,4-dithiouridine) and Its Oxygen Congener, DAU (5-(5-(4-(Dimethylamino)phenyl)thiophen-2-yl)-2′,3′,5′-tri-O-benzoyl-6-azauridine)#

主要实验及结论#

研究人员通过先进的飞秒瞬态吸收光谱技术与量子化学计算，深入剖析了DATU的电子结构与光物理特性。实验表明，DATU在水溶液中的单光子吸收光谱能够一直延伸至750纳米，且其三重态寿命达到1.7微秒，在苯溶剂中的单线态氧量子产率高达56%。尤为令人振奋的是，在800纳米的近红外激光激发起下，DATU展现出高达160 GM的双光子吸收截面，这一数值比其传统的羰基对应物DAU高出整整3倍，直接刷新了目前已知核苷类似物的最高历史纪录。

Figure 1. (a) UV–vis absorption spectra of DATU, and (b) UV–vis absorption and fluorescence spectra of DAU in 1,4-dioxane and benzene. The fluorescence spectra were collected exciting DAU at 425 and 435 nm, respectively.#

Scheme 2. Structures of syn- and anti-Rotamers of (a) DATU and (b) DAU#

Figure 2. Experimental absorption spectra for (a) DATU and (d) DAU in 1,4-dioxane, alongside the simulated absorption spectra for (b) DATU and (e) DAU in 1,4-dioxane. The corresponding Kohn–Sham orbitals for HOMO and LUMO are shown for (c) DATU and (f) DAU.#

Figure 3. Jabłoński diagram for the major (syn) isomers of DATU and DAU, illustrating the dominant relaxation pathways following excitations to the respective lowest-energy absorption bands.#

在生理环境和细胞层面的评估实验中，DATU在受到特定光照激活后，不仅能高效降解活性氧探针，还能在水性缓冲液中大量产生羟基自由基，证实其能根据不同的细胞环境同时通过I型和II型机制发挥光敏作用。体外细胞毒性实验进一步证实，在面对4T1小鼠乳腺癌细胞时，DATU在单光子与双光子近红外激光照射下均表现出极高的光细胞毒性，不仅显著杀伤了癌细胞，而且在无光照的暗处对正常组织几乎不产生任何毒副作用。

Figure 4. (a) Spectral evolution of fs-TAS of DATU in 1,4-dioxane at 520 nm excitation, (b) extracted EADS, (c) kinetic decay traces with fits at different wavelengths, and (d) contour plots of fs-TAS. The breaks on the x-axis of (a, b, and d) cover the scattering from the pump pulse, and those on the x-axis in (c) and the y-axis in (d) represent the change in the scale from linear to logarithmic. The sharp signals in the top panel of (a) correspond to the stimulated Raman scattering and coherent solvent signals, which were used to define time zero at its maximum amplitude.#

Figure 5. A qualitative relaxation mechanism for DATU upon photoexcitation.#

Figure 6. (a) Spectral evolution of fs-TAS of DAU in 1,4-dioxane at 400 nm excitation, (b) extracted EADS, (c) kinetic decay traces with fits at different wavelengths, and (d) contour plots of fs-TAS. The breaks on the x-axis of (a and d) cover the scattering from the pump pulse, and those on the x-axis of (c) and the y-axis in (d) represent the change in the scale from linear to logarithmic. The sharp signals in the top panel of (a) correspond to the stimulated Raman scattering and coherent solvent signals, which were used to define time-zero at its maximum amplitude.#

Figure 7. (a) ps-to-μs-TAS of DATU, (b) respective kinetic traces, and (c) contour plots for TAS data in N2-saturated 1,4-dioxane following excitation at 520 nm. Breaks on the x-axis of (a and c) cover the scattering from the pump pulse, while that on the y-axis of (c) represents the change in the scale from linear to logarithmic. (d) Representative kinetic traces of TAS recorded under O2, air, and N2-saturated conditions.#

Figure 8. (a) ps-to-μs-TAS of DAU, (b) respective kinetic traces, (c) EADS in air-equilibrated 1,4-dioxane following excitation at 400 nm, and (d) contour plots of TAS. Breaks on the x-axis of (b) and the y-axis in (d) represent the change in the scale from linear to logarithmic.#

Figure 9. Log–log plots of Δ_A_ versus the power of 800 nm fs-pulsed laser for (a) DATU and (b) DAU in 1,4-dioxane. The transient absorption spectra of (c) DATU and (d) DAU in 1,4-dioxane, and (e) Rhodamine 6G in methanol at 20 ps delay, recorded upon 400 nm one-photon (top panel) and 800 nm two-photon (bottom panel) excitations. The breaks in the x-axis of the top panels in (c, d, and e) are covering scattering from the 400 nm pump pulse.#

Figure 10. Degradation of DPBF under 800 nm fs-pulsed laser excitation for different irradiation times in (a) the presence and (b) the absence of DATU in benzene. (c) Corresponding plot of degradation (A/_A_0 vs laser irradiation time) of DPBF in both conditions under 800 nm fs-pulsed laser excitation.#

Figure 11. DPBF assay demonstrates the generation of singlet oxygen upon irradiation of DTAU/DPBF mixtures at 525 nm in air-saturated (a) MeOH and (c) MeCN solutions. Controls with DPBF alone are shown in (b) MeOH and (d) MeCN, respectively. These assays were performed in a 2 mm cuvette, with an absorbance of DATU kept at ∼0.06 at 525 nm.#

Figure 12. Light dose-dependent changes in HPF fluorescence intensity in aqueous phosphate buffer solution (PBS) upon 525 nm irradiation: (a) in the presence of DATU and (b) in its absence (control). (c) Corresponding plot of F/_F_0 at the maximum emission as a function of light dose under both conditions. (d) Absorption spectrum of DATU in aqueous buffer solution at pH 7.4 (99:1 PBS/DMSO).#

Figure 13. Representative viability studies utilizing 4T1 murine mammary carcinoma cells in the presence and absence of light irradiation: (a) 6 μM DATU with one-photon excitation at 525 nm (16 J cm–2) and (b) 25 μM DATU with two-photon excitation using 800 nm laser (40.7 kJ cm–2). See the Methods section in the SI for details.#

总结及展望#

这项开创性的研究成功构筑了目前世界上双光子吸收截面最大的核苷类似物光敏剂，成功攻克了光动力治疗中深层组织渗透力不足的瓶颈。DATU兼具优异的细胞膜渗透性、高产率的活性氧生成能力以及极低的暗毒性，未来在深层癌症治疗、临床生物成像以及光催化等诸多前沿领域都将发挥不可估量的核心作用。这一结构工程化的设计思路也为开发下一代无重原子有机硫代光敏剂开辟了全新的道路。