A pH-activatable and aniline-substituted photosensitizer for near-infrared cancer theranostics

This work reports a newly designed pH-activatable and aniline-substituted aza-boron-dipyrromethene as a trifunctional photosensitizer to achieve highly selective tumor imaging, efficient photodynamic therapy (PDT) and therapeutic self-monitoring through encapsulation in a cRGD-functionalized nanomicelle. The diethylaminophenyl is introduced in to the structure for pH-activatable near-infrared fluorescence and singlet oxygen (¹O₂) generation, and bromophenyl is imported to increase the ¹O₂ generation efficiency upon pH activation by virtue of its heavy atom effect. After encapsulation, the nanoprobe can target α_vβ₃ integrin-rich tumor cells via cRGD and is activated by physiologically acidic pH for cancer discrimination and PDT. The fascinating advantage of the nanoprobe is near-infrared implementation beyond 800 nm, which significantly improves the imaging sensitivity and increases the penetration depth of the PDT. By monitoring the fluorescence decrease in the tumor region after PDT, the therapeutic efficacy is demonstrated in situ and in real time, which provides a valuable and convenient self-feedback function for PDT efficacy tracking. Therefore, this rationally designed and carefully engineered nanoprobe offers a new paradigm for precise tumor theranostics and may provide novel opportunities for future clinical cancer treatment.     

水溶性金属卟啉在水溶液中的光物理与光化学研究

研究了５种水溶性金属卟啉ＺｎＴＭＡＰＰ、ＺｎＴＭＰｙＰ、ＭｇＴＭＰｙＰ、ＺｎＴＢＰｙＰ和ＺｎＴＴＰＳ４的光物理（吸收光谱和荧光光谱、振子强度、荧光寿命、量子产率）、激发态氧化还原电势及光稳定性。结果表明ＭｇＴＭＰｙＰ是比ＺｎＴＭＰｙＰ更好的光敏剂。     

Controlling the Photofunctionality of a Polyanionic Heteroleptic Copper(I) Photosensitizer for CO2 Reduction Using Its Ion-pair Formation with Polycationic Ammonium in Aqueous Media

A water-soluble trianionic heteroleptic copper(I) photosensitizer having four sulfonate groups ( CuPS ³⁻) was found to afford the 1 : 2 ion-pair adduct with dicationic alkylammonium (hexamethonium) cations ( HM ²⁺) in aqueous media, leading to exhibit excellent photophysical and photocatalytic performances owing to the substantial suppression of water-derived non-radiative decay of the photoexcited state.     

Phototoxic process after rapid photosensitive membrane damage of 5,5"-bis(aminomethyl)-2,2':5',2"-terthiophene dihydrochloride

We report a new aspect of rapid (<30 s) light-induced cell membrane damage photosensitized by 5,5"-bis(aminomethyl)-2,2':5',2"-terthiophene dihydrochloride (BAT), which is a water-soluble alpha-terthienyl analogue, using a high-power laser (light intensity 1.6 W cm(-2)). In this paper, we will discuss the relationship between the exposure time of the cells to the photosensitizer and the phototoxic process. Three toxic processes can be identified: first, a non-light-mediated toxicity dependent on BAT-cell incubation; second, a phototoxicity independent of BAT exposure time when the BAT concentration is in the 2-10-microM range; third, a phototoxicity dependent on BAT exposure time when BAT concentration becomes 20 microM. The cytotoxicity decreases when alpha-tocopherol, an antioxidant, is added to a cell membrane. This pattern of phototoxicity is the typical of a phospholipid peroxidation chain reaction and oxidative damage of membrane proteins triggered by a reactive oxygen species generated by a triplet state of BAT. The BAT exposure time is clearly correlated with the partition of the photosensitizer in the cell membrane and inside the cell.     

[1,2,3,4-Tetrakis(α/β-d-galactopyranos-6-yl)phthalocyaninato]zinc(II): a water-soluble phthalocyanine

A novel water-soluble asymmetrical sugar-phthalocyanine was prepared via a statistical cross-condensation of tetrakis(1,2:3,4-di-O-isopropylidene-α-d-galactopyranos-6-yl)phthalonitrile with phthalonitrile. The new compound, with amphiphilic character, can be useful as a selective photosensitizer in photodynamic therapy, as well as for constructing phthalocyanine-based supramolecular systems.     

A photoinduced electron transfer-based nanoprobe as a marker of acidic organelles in mammalian cells

Photoinduced electron transfer (PET)-based molecular probes have been successfully used for the intracellular imaging of the pH of acidic organelles. In this study, we describe the synthesis and characterization of a novel PET-based pH nanoprobe and its biological application for the signaling of acidic organelles in mammalian cells. A fluorescent ligand sensitive to pH via the PET mechanism that incorporates a thiolated moiety was synthesized and used to stabilize gold nanoparticles (2.4?±?0.6 nm), yielding a PET-based nanoprobe. The PET nanoprobe was unambiguously characterized by transmission electron microscopy, proton nuclear magnetic resonance, Fourier transform infrared, ultraviolet-visible absorption, and steady-state/time-resolved fluorescence spectroscopies which confirmed the functionalization of the gold nanoparticles with the PET-based ligand. Following a classic PET behavior, the fluorescence emission of the PET-based nanoprobe was quenched in alkaline conditions and enhanced in an acidic environment. The PET-based nanoprobe was used for the intracellular imaging of acidic environments within Chinese hamster ovary cells by confocal laser scanning microscopy. The internalization of the nanoparticles by the cells was confirmed by confocal fluorescence images and also by recording the fluorescence emission spectra of the intracellular PET-based nanoprobe from within the cells. Co-localization experiments using a marker of acidic organelles, LysoTracker Red DND-99, and a marker of autophagosomes, GFP-LC3, confirm that the PET-based nanoprobe acts as marker of acidic organelles and autophagosomes within mammalian cells.

Dielectrophoretic trapping of nanoparticles with an electrokinetic nanoprobe

A high aspect ratio 3D electrokinetic nanoprobe is used to trap polystyrene particles (200 nm), gold nanoshells (120 nm), and gold nanoparticles (mean diameter 35 nm) at low voltages (<1 V_rms). The nanoprobe is fabricated using room temperature self‐assembly methods, without the need for nanoresolution lithography. The nanoprobe (150–500 nm in diameter, 2–150 μm in length) is mounted on the end of a glass micropipette, enabling user‐specified positioning. The nanoprobe is one electrode within a point‐and‐plate configuration, with an indium–tin oxide cover slip serving as the planar electrode. The 3D structure of the nanoprobe enhances dielectrophoretic capture; further, electro‐hydrodynamic flow enhances trapping, increasing the effective trapping region. Numerical simulations show low heating (1 K), even in biological media of moderate conductivity (1 S/m).     

A highly selective, cell-permeable fluorescent nanoprobe for ratiometric detection and imaging of peroxynitrite in living cells

Peroxynitrite (ONOO(-)) is a highly reactive species implicated in the pathology of numerous diseases and there is currently great interest in developing fluorescent probes that can selectively detect ONOO(-) in living cells. Herein, a polymeric micelle-based and cell-penetrating peptide-coated fluorescent nanoprobe that incorporates ONOO(-) indicator dye and reference dye for the ratiometric detection and imaging of ONOO(-) has been developed. The nanoprobe effectively avoids the influences from enzymatic reaction and high-concentration ·OH and ClO(-). The improved ONOO(-) selectivity of the nanoprobe is achieved by a delicate complementarity of properties between the nanomatrix and the embedded molecular probe (BzSe-Cy). This nanoprobe also has other attractive properties, such as good water solubility, photostability, biocompatibility, and near-infrared excitation and emission. Fluorescence imaging experiments by confocal microscopy show that this nanoprobe is capable of visualizing ONOO(-) produced in living cells and it exhibits very low toxicity and good membrane permeability. We anticipate that this technique will be a potential tool for the precise pathological understanding and diagnosis of ONOO(-)-related human diseases.     

Encapsulation of phthalocyanine supramolecular stacks into virus-like particles

We report herein the encapsulation of a water-soluble phthalocyanine (Pc) into virus-like particles (VLPs) of two different sizes, depending on the conditions. At neutral pH, the cooperative encapsulation/templated assembly of the particles induces the formation of Pc stacks instead of Pc dimers, due to an increased confinement concentration. The Pc-containing VLPs may potentially be used as photosensitizer/vehicle systems for biomedical applications such as photodynamic therapy.     

Constructing Adaptive Photosensitizers via Supramolecular Modification Based on Pillararene Host–Guest Interactions

In order to promote the development of photodynamic therapy (PDT), undesired side effects like low tumor specificity and the “always-on” phenomenon should be avoided. An effective solution is to construct an adaptive photosensitizer that can be activated to generate reactive oxygen species (ROS) in the tumor microenvironment. Herein, we design and synthesize a supramolecular switch based on a host–guest complex containing a water-soluble pillar[5]arene ( WP5 ) and an AIEgen photosensitizer ( G ). The formation of the host–guest complex WP5 ⊃ G quenches the fluorescence and inhibits ROS generation of G . Benefitting from the pH-responsiveness of WP5 , the binding site between G and WP5 changes in an acidic environment through a shuttle movement. Consequently, fluorescence and ROS generation of the host–guest complex can be switched on at pH 5.0. This work offers a new paradigm for the construction of adaptive photosensitizers by using a supramolecular method.     

四氨基酞菁锌负载二氧化硅纳米粒子的合成及其对细胞毒性的研究

通过在无极核微乳液中水解乙烯基三乙氧基硅烷(TEVS)和3-氨丙基三乙氧基硅烷(APTES),制备了疏水性光敏剂-2,9,16,23-四氨基酞菁锌负载的表面带有正电荷的二氧化硅纳米粒子(SiO2@ ZnPc( NH2)4).通过透射电镜(TEM)、Zetasizer Nano-ZS粒度仪(DLS)、紫外-可见分光光度计...     

Environmental Effects on Cellular Photosensitization: Correlation of Phototoxicity Mechanism with Transient Absorption Spectroscopy Measurements

Abstract— Little is directly known about the influence of the local environment experienced by a photosensitizer in a biological system on its photophysics and photochemistry. In this paper, we have addressed this issue by correlating mechanistic studies using laser flash photolysis with cellular phototoxicity data, obtained under the same experimental conditions. In particular, we have focused on the interaction between local concentrations of photosensitizer (deuteroporphyrin) and oxygen in determining the mechanism of phototoxicity in L1210 cells. In cells, as well as in models such as liposomes and red blood cell ghosts, hypochromicity and a reduction in fluorescence and intersystem crossing yields are observed on increasing the photosensitizer concentration between 0.5 and 20 μM, which illustrates the onset of a self-association. In aerated cellular preparations, the phototoxicity is predominantly type II (singlet oxygen) for all concentrations studied but an oxygen-independent mechanism occurs at the higher concentrations in deaerated samples. These observations are readily explained by consideration of triplet state kinetics as a function of oxygen and photosensitizer concentrations in cells. The rate constant for quenching of the photosensitizer triplet state by oxygen in cells was measured as 6.6 × 10⁸ M^?1 s^?1 and by photosensitizer ground state as -10⁶M^?1s^?1 (in terms of local concentration). The latter reaction gave rise to a long-lived species that is presumably responsible for the oxygen-independent phototoxicity observed at the higher photosensitizer concentrations used. This self-quenching of the triplet state is postulated to arise from electron transfer resulting in radical ion formation. Under conditions where no self-quenching contributes, the phototoxicity measured as a function of oxygen concentration correlates well with a model based on the determined kinetic parameters, thus, unambiguously proving the intermediacy of singlet oxygen. These effects should be borne in mind when interpreting phototoxicity mechanisms from in vitro cell studies. The excellent correlation achieved between laser flash photolysis data and measured phototoxicity gives credence to the direct use of photophysical techniques to elucidate photochemical mechanisms in biological media.     

聚丙烯表面光接枝及染色

用UV光照法将两种亲水性单体(甲基丙烯酸和丙烯酰胺)分别接枝到非亲水性的聚丙烯板表面上。以二苯酮为光敏剂,丙酮为溶剂,在不断通入氮气的条件下,采用汽-固相接枝法,可以一次完成双面接枝。电子能谱(ESCA)的结果证实了光接枝的成功。水接触角测试结果表明,接枝后的聚丙烯表面具有良好的亲水性。利用水溶性的染料(结晶紫和罗丹明B)对接枝后的聚丙烯板进行了染色,并测定了吸收光谱和荧光光谱。     

Nanoaggregate Probe for Breast Cancer Metastasis through Multispectral Optoacoustic Tomography and Aggregation-Induced NIR-I/II Fluorescence Imaging

An activatable nanoprobe for imaging breast cancer metastases through near infrared-I (NIR-I)/NIR-II fluorescence imaging and multispectral optoacoustic tomography (MSOT) imaging was designed. With a dihydroxanthene moiety serving as the electron donor, quinolinium as the electron acceptor and nitrobenzyloxydiphenylamino as the recognition element, the probe can specifically respond to nitroreductase and transform into an activated D-π-A structure with a NIR emission band extending beyond 900 nm. The activated nanoprobe exhibits NIR emission enhanced by aggregation-induced emission (AIE) and produces strong optoacoustic signal. The nanoprobe was used to detect and image metastases from the orthotopic breast tumors to lymph nodes and then to lung in two breast cancer mouse models. Moreover, the nanoprobe can monitor the treatment efficacy during chemotherapeutic course through fluorescence and MSOT imaging.     

A new FRET nanoprobe for trypsin using a bridged β-cyclodextrin dimer-dye complex and its biological imaging applications

A new self-assembly nanoprobe, mercaptoethylamine-modified-gold nanoparticles-Lysine-bridged-bis(β-cyclodextrins)-fluorescein (MGNPs-Lys-bis(β-CDs)-FL), based on fluorescence resonance energy transfer (FRET) was developed for determination of trypsin firstly in biological systems. With the Lys-bis(β-CDs)-FL complex as an energy donor and mercaptoethylamine (MEA)-modified gold nanoparticles (MGNPs) as an energy acceptor, the two parts assemble an efficient FRET nanoprobe through an amide bond. Trypsin is specific for the hydrolysis of amide linkages of lysine. Therefore, in the presence of trypsin, the nanoprobe is cleaved by trypsin on the binding sites of amide with good specificity and sensitivity, resulting in the fluorescence recovery of the quenched FL. The nanoprobe has good biological applicability and provides a potential assay for further clinical research of trypsin in biosystems.     

FRET spectral unmixing: a ratiometric fluorescent nanoprobe for hypochlorite

A specific ratiometric nanoprobe for hypochlorite was constructed as a paradigm of FRET spectral unmixing. The separation of FRET pairs' emissions reaches 175 nm, which ensures that the FRET probing is more accurate. This new nanoprobe shows high selectivity and potential in biological systems.     

Water-soluble self-assembled butadiyne-bridged bisporphyrin: a potential two-photon-absorbing photosensitizer for photodynamic therapy

We have synthesized a novel, two-photon-absorbing photosensitizer for two-photon-absorption photodynamic therapy (2PA-PDT). The molecule is a butadiyne-bridged porphyrin dimer terminated with two water-soluble porphyrin monomers connected through Zn-imidazolyl self-assembly and covalently linked through olefin metathesis. It has an effective two-photon-absorption (2PA) cross-section value, sigma((2)), of 33 000+/-4600 GM with 5-ns pulses at 890 nm measured by using the open-aperture Z-scan technique. The compound was found to generate singlet oxygen, cytotoxic for tumor cells in photodynamic therapy (PDT), under 2PA conditions by conducting photobleaching experiments with anthracene-9,10-dipropionic acid sodium salt (ADPA).     

Targeting cells that overexpress the epidermal growth factor receptor with polyethylene glycolated BPD verteporfin photosensitizer immunoconjugates

Photoimmunotherapy was introduced two decades ago but has been studied infrequently in vivo and is virtually untested clinically. Progress has been limited because high-quality, well-characterized photosensitizer immunoconjugates (PICs) have been difficult to make. Here, we describe the development of an innovative conjugation method for producing water-soluble PICs that are free of insoluble aggregates and free of unacceptable amounts of noncovalently associated photosensitizer impurities. The method exploits two procedures previously untried in this research area. First, a small number of antibody lysines (<3 per antibody) are polyethylene glycolated (PEGylated) using a 10 kDa branched polyethylene glycol (PEG), which dramatically enhances PIC solubility and reduces PIC aggregation. Second, a 50% dimethyl sulfoxide-50% aqueous two-solvent system is used to prevent photosensitizer aggregation and noncovalent interactions. These measures allow efficient covalent linkage of the photosensitizer BPD Verteporfin (BPD) to antibody lysines, thorough purification of the resulting PICs (verified by sodium dodecyl sulfate-polyacrylamide gel electrophoresis), maintenance of PIC antigen-binding activity (verified by cellular binding-uptake assays) and reduction of nonspecific cellular uptake (e.g. macrophage capture) of the PICs. Loading levels could be varied controllably in the range < or = 11 BPD/antibody. PICs of the C225 anti-epidermal growth factor receptor (EGFR) chimeric monoclonal antibody killed EGFR-overexpressing A-431 cells photodynamically but did not significantly affect EGFR-negative NR6 cells. Although fluorescence measurements demonstrated that the PICs were quenched by as much as an order of magnitude compared with free BPD, an impressive 90% reduction in A-431 cell viability was achieved using 20 J/cm2 of 690 nm light after a 40 h incubation with the C225 PICs. The results suggest that PEGylated BPD-C225 PICs merit further investigation in animal models of EGFR-overexpressing cancers.     

Nanoaggregate Probe for Breast Cancer Metastasis through Multispectral Optoacoustic Tomography and Aggregation‐Induced NIR‐I/II Fluorescence Imaging

An activatable nanoprobe for imaging breast cancer metastases through near infrared‐I (NIR‐I)/NIR‐II fluorescence imaging and multispectral optoacoustic tomography (MSOT) imaging was designed. With a dihydroxanthene moiety serving as the electron donor, quinolinium as the electron acceptor and nitrobenzyloxydiphenylamino as the recognition element, the probe can specifically respond to nitroreductase and transform into an activated D‐π‐A structure with a NIR emission band extending beyond 900 nm. The activated nanoprobe exhibits NIR emission enhanced by aggregation‐induced emission (AIE) and produces strong optoacoustic signal. The nanoprobe was used to detect and image metastases from the orthotopic breast tumors to lymph nodes and then to lung in two breast cancer mouse models. Moreover, the nanoprobe can monitor the treatment efficacy during chemotherapeutic course through fluorescence and MSOT imaging.     

Developing Novel Fabrication and Optimisation Strategies on Aggregation-Induced Emission Nanoprobe/Polyvinyl Alcohol Hydrogels for Bio-Applications

The current study describes a new technology, effective for readily preparing a fluorescent (FL) nanoprobe-based on hyperbranched polymer (HB) and aggregation-induced emission (AIE) fluorogen with high brightness to ultimately develop FL hydrogels. We prepared the AIE nanoprobe using a microfluidic platform to mix hyperbranched polymers (HB, generations 2, 3, and 4) with AIE (TPE-2BA) under shear stress and different rotation speeds (0–5 K RPM) and explored the FL properties of the AIE nanoprobe. Our results reveal that the use of HB generation 4 exhibits 30-times higher FL intensity compared to the AIE alone and is significantly brighter and more stable compared to those that are prepared using HB generations 3 and 2. In contrast to traditional methods, which are expensive and time-consuming and involve polymerization and post-functionalization to develop FL hyperbranched molecules, our proposed method offers a one-step method to prepare an AIE-HB nanoprobe with excellent FL characteristics. We employed the nanoprobe to fabricate fluorescent injectable bioadhesive gel and a hydrogel microchip based on polyvinyl alcohol (PVA). The addition of borax (50 mM) to the PVA + AIE nanoprobe results in the development of an injectable bioadhesive fluorescent gel with the ability to control AIEgen release for 300 min. When borax concentration increases two times (100 mM), the adhesion stress is more than two times bigger (7.1 mN/mm²) compared to that of gel alone (3.4 mN/mm²). Excellent dimensional stability and cell viability of the fluorescent microchip, along with its enhanced mechanical properties, proposes its potential applications in mechanobiology and understanding the impact of microstructure in cell studies.