Development of Lysosome-Targeted Fluorescent Probes for Cys by Regulating the Boron-dipyrromethene (BODIPY) Molecular Structure

Our previous discovery suggested that substituents on the 1,7 positions delicately modulate the sensing ability of the meso-arylmercapto boron-dipyrromethene (BODIPY) to biothiols. In this work, the impact of delicate modulations on the sensing ability is investigated. Therefore, 1,7-dimethyl, 3,5-diaryl substituted BODIPY is designed and developed and its conformationally restricted species with a meso-arylmercapto moiety ( DM-BDP-SAr and DM-BDP-R-SAr ) as selective fluorescent probes for Cys. Moreover, the lysosome-target probes ( Lyso-S and Lyso-D ) based on DM-BDP-SAr carrying one or two morpholinoethoxy moieties were developed. They were able to detect Cys selectively in vitro with low detection limits. Both Lyso-S and Lyso-D localized nicely in lysosomes in living HeLa cells and exhibited red fluorescence for Cys. Moreover, a novel fluorescence quenching mechanism was proposed from the calculations by density functional theory (DFT). The probes may go through intersystem crossing (from singlet excited state to triplet excited state) to result in fluorescence quenching.     

Lysosome‐targeted potent half‐sandwich iridium(III) α‐diimine antitumor complexes

Fifteen organometallic Ir(III) half‐sandwich complexes ( 1A – 5C ) having the general formula [(η⁵‐Cp^x)Ir(N^N)Cl]PF₆ (Cp^x = Cp*, tetramethyl(phenyl)cyclopentadienyl (Cp^xph) or tetramethyl(biphenyl)cyclopentadienyl (Cp^xbiph); N^N = diamine) have been synthesized and characterized. The molecular structure of 1A was determined using single‐crystal X‐ray diffraction analysis. The hydrolysis of 1A – 5C was monitored using UV–visible spectra. Complexes 3A – 3C showed catalytic activity for the oxidation of NADH to NAD⁺, where 3C showed the highest turnover number of 29.9 within 450 min. Cytotoxicity examination by MTT assay was carried out against two human cancer cell lines (HeLa and A549) after 24 or 48 h drug treatment. The complexes showed high potency, where the most potent complex ( 3C ; IC₅₀ = 3.4 μM) was six times more active than cisplatin against A549 cells after 24 h drug exposure. Cytotoxic potency towards A549 cells increased with phenyl substitution on Cp ring: Cp^xbiph > Cp^xph > Cp*. In addition, the biological studies showed that 3C caused cell apoptosis and cell cycle arrest at G₁ phase in A549 cancer cells. Moreover, 3C increased the level of reactive oxygen species markedly after 24 h, which may provide an important basis for killing cancer cells. Confocal laser scanning microscopy was used to track 3C in A549 cells. The cellular localization experiment showed that 3C targeted lysosomes and caused lysosomal damage.     

An Endoplasmic Reticulum Targeting Type I Photosensitizer for Effective Photodynamic Therapy against Hypoxic Tumor Cells

Organelle-targeted type I photodynamic therapy (PDT) shows great potential to overcome the hypoxic microenvironment in solid tumors. The endoplasmic reticulum (ER) is an indispensable organelle in cells with important biological functions. When the ER is damaged due to the production of reactive oxygen species (ROS), the accumulation of misfolded proteins will interfere with ER homeostasis, resulting in ER stress. Here, an ER-targeted benzophenothiazine-based photosensitizer NBS-ER was presented. ER targeting modification significantly reduced the dark toxicity and improved phototoxicity index (PI). NBS-ER could effectively produce O₂⁻⋅ with near-infrared irradiation, making its phototoxicity under hypoxia close to that under normoxia. Meanwhile, the photoinduced ROS triggered ER stress and induced apoptosis. In addition, NBS-ER possessed excellent photodynamic therapeutic effect in 4T1-tumor-bearing mice.     

吡咯腙对溶酶体中Hg2+的荧光成像

合成了一种新的吡咯腙探针1,用于Hg²⁺的比色和荧光开启检测。探针1对Hg²⁺的检测限为45 nmol·L^-1,缔合常数为5.78×10⁸ L·mol^-1。值得注意的是,工作pH范围为4.0~10.0。Job曲线和MS数据证实探针与Hg²⁺形成1:1的配合物。通过¹H NMR、时间分辨荧光光谱和密度泛函理论(DFT)计算系统研究了探针与Hg²⁺的配位模式。此外,由于吗啉基团的存在,探针可以检测HeLa细胞溶酶体中的Hg²⁺。     

Lysosomal-targeted anticancer half-sandwich iridium(III) complexes modified with lonidamine amide derivatives

Ten half-sandwich iridium complexes containing lonidamine amide derivatives were synthesized and characterized. Unlike lonidamine, which acts on mitochondria, its iridium complexes successfully targeted lysosomes and induced lysosomal damage. Antiproliferation studies showed that most of the complexes have higher anticancer activity against A549 and HeLa cells than cisplatin. The antitumor activity of complex 6 is 2.69 times that of cisplatin against A549 cells. We also performed antitumor tests on ligands L₁ and L₅, and proved that they exhibit excellent antitumor activity only after binding to the metal center. The bovine serum albumin (BSA) binding test showed that the complexes had the ability to bind to BSA, and they interact with BSA by a static mechanism. The complexes can also cause changes in mitochondrial membrane potential and can produce active oxygen species better than active control. NADH/NAD⁺ transformation experiments were used to determine if the production of ROS was caused by the transformation of NADH/NAD⁺. We also explored the way that the complexes enter cells.     

Light-Induced Self-Escape of Spherical Nucleic Acid from Endo/Lysosome for Efficient Non-Cationic Gene Delivery

Developing non-cationic gene carriers and achieving efficient endo/lysosome escape of functional nucleic acids in cytosol are two major challenges faced by the field of gene delivery. Herein, we demonstrate the concept of self-escape spherical nucleic acid (SNA) to achieve light controlled non-cationic gene delivery with sufficient endo/lysosome escape capacity. In this system, Bcl-2 antisense oligonucleotides (OSAs) were conjugated onto the surface of aggregation-induced emission (AIE) photosensitizer (PS) nanoparticles to form core–shell SNA. Once the SNAs were taken up by tumor cells, and upon light irradiation, the accumulative ¹O₂ produced by the AIE PSs ruptured the lysosome structure to promote OSA escape. Prominent in vitro and in vivo results revealed that the AIE-based core–shell SNA could downregulate the anti-apoptosis protein (Bcl-2) and induce tumor cell apoptosis without any transfection reagent.     

吡咯腙对溶酶体中Hg2+的荧光成像

合成了一种新的吡咯腙探针1，用于Hg²⁺的比色和荧光开启检测。探针1对Hg²⁺的检测限为45 nmol·L^-1，缔合常数为5.78×10⁸ L·mol^-1。值得注意的是，工作pH范围为4.0~10.0。Job曲线和MS数据证实探针与Hg²⁺形成1∶1的配合物。通过1H NMR、时间分辨荧光光谱和密度泛函理论(DFT)计算系统研究了探针与Hg²⁺的配位模式。此外，由于吗啉基团的存在，探针可以检测HeLa细胞溶酶体中的Hg²⁺。     

Image‐Based Morphological Profiling Identifies a Lysosomotropic,Iron‐Sequestering Autophagy Inhibitor

Chemical proteomics is widely applied in small‐molecule target identification. However, in general it does not identify non‐protein small‐molecule targets, and thus, alternative methods for target identification are in high demand. We report the discovery of the autophagy inhibitor autoquin and the identification of its molecular mode of action using image‐based morphological profiling in the cell painting assay. A compound‐induced fingerprint representing changes in 579 cellular parameters revealed that autoquin accumulates in lysosomes and inhibits their fusion with autophagosomes. In addition, autoquin sequesters Fe²⁺ in lysosomes, resulting in an increase of lysosomal reactive oxygen species and ultimately cell death. Such a mechanism of action would have been challenging to unravel by current methods. This work demonstrates the potential of the cell painting assay to deconvolute modes of action of small molecules, warranting wider application in chemical biology.     

Reversible and Selective Fluorescence Detection of Histidine Using a Naphthalimide‐Based Chemosensing Ensemble

We described a new ensemble‐approach‐based chemosensor, NCH‐Cu²⁺, for highly selective and reversible detection of histidine (His) in aqueous solution and live cells. The ligand NCH exhibited specific binding with Cu²⁺ ions over other metal ions, accompanied with a 92.2 % fluorescence quenching. The decomplexation of NCH‐Cu²⁺ ensemble by His led to the liberation of the fluorophore, NCH, and thus the fluorescence was recovered. The specific fluorescence enhancement of NCH‐Cu²⁺ towards His showed a good linearity with a detection of limit at 70 nm . Quantification of intracellular His at the single cell level was achieved by microscopy and flow cytometry. Besides the UV/Vis and emission titration, reversibility of the NCH‐Cu²⁺ towards His was further confirmed by imaging and cytometry analysis. In addition, microscopy studies revealed that NCH‐Cu²⁺ was distributed in the lysosome of live cells, where it could be employed as a fluorescent biosensor for imaging of His at subcellular level.     

Lysosome‐targeted Cyclometalated Iridium (III) Anticancer Complexes Bearing Phosphine‐Sulfonate Ligands

The synthesis, characterization and biological activity of four cyclometalated Ir (III) complexes ( Ir1 ‐ Ir4 ) containing different phosphine‐sulfonate ligands are reported. Most of these complexes showed good activity against A549 cancer cell lines and the human HeLa cervical cell lines. Spectroscopic properties study displays that all four complexes show rich fluorescence with emission maxima in the range of 474–510 nm. Fluorescence property of these complexes provides a tool to investigate the microscopic mechanism by confocal microscopy. Notably, the typical Ir (III) complex Ir4 can specially localize to lysosome, damage it and induce cell death via apoptosis. In addition, Ir4 enters into A549 cancer cells dominantly through energy‐dependent pathway.