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Mitochondria-targeting Hemicyanine Photosensitizer as an Inducer for MMP Loss and Cell Apoptosis Based on Radical Enhanced ISC Strategy
Heavy atom-free triplet photosensitizers (PSs) can overcome the high cost and biological toxicity of traditional molecular systems containing heavy atoms (such as Pt(II), Ir(III), Ru(II), Pd(II), Lu(III), I, or Br atoms) and, therefore, are developing rapidly. Connecting a stable free radical to the chromophore can promote the intersystem crossing (ISC) process through electron spin exchange interaction to produce the triplet state of the chromophore or the doublet (D) and quartet (Q) states when taking the whole spin system into account. These molecular systems based on the radical enhanced ISC (REISC) mechanism are important in the field of heavy atom-free triplet PSs. The REISC system has a simple molecular structure and good biocompatibility, and it is especially helpful for building high-spin quantum states (D and Q states) that have the potential to be developed as qubits in quantum information science. This review introduces the molecular structure design for the purpose of high-spin states. Time-resolved electron paramagnetic resonance (TREPR) is the most important characterization method to reveal the properties of these molecular systems, generation mechanism and electron spin polarization (ESP) of the high spin states. The spin polarization manipulation of high spin states and potential application in the field of quantum information engineering are also summarized. Moreover, molecular design principles of the REISC system to obtain long absorption wavelength, high triplet state quantum yield and long triplet state lifetime are introduced, as well as applications of the compounds in triplet-triplet annihilation upconversion, photodynamic therapy and bioimaging. This review is useful for the design of heavy atom-free triplet PSs based on the radical-chromophore molecular structure motif and the study of the photophysics of the compounds, as well as the electron spin dynamics of the multi electron system upon photoexcitation.
Pyroptosis, a unique lytic programmed cell death, inspired tempting implications as potent anti-tumor strategy in pertinent to its potentials in stimulating anti-tumor immunity for eradication of primary tumors and metastasis. Nonetheless, rare therapeutics have been reported to successfully stimulate pyroptosis. In view of the intimate participation of reactive oxygen species (ROS) in stimulating pyroptosis, we attempted to devise a spectrum of well-defined subcellular organelle (including mitochondria, lysosomes and endoplasmic reticulum)-targeting photosensitizers with the aim of precisely localizing ROS (produced from photosensitizers) at the subcellular compartments and explore their potentials in urging pyroptosis and immunogenic cell death (ICD). The subsequent investigations revealed varied degrees of pyroptosis upon photodynamic therapy (PDT) towards cancerous cells, as supported by not only observation of the distinctive morphological and mechanistic characteristics of pyroptosis, but for the first-time explicit validation from comprehensive RNA-Seq analysis. Furthermore, in vivo anti-tumor PDT could exert eradication of the primary tumors, more importantly suppressed the distant tumor and metastatic tumor growth through an abscopal effect, approving the acquirement of specific anti-tumor immunity as a consequence of pyroptosis. Hence, pyroptosis was concluded unprecedently by our proposed organelles-targeting PDT strategy and explicitly delineated with molecular insights into its occurrence and the consequent ICD.
The mechanism of spin polarization transfer from a photogenerated spin-correlated radical pair to a stable radical was studied in a covalent donor-chromophore-acceptor-stable radical (D-C-A-R•) system, where the donor (D) is 4-methoxyaniline (MeOAn), the chromophore (C) is 4-(N-piperidinyl)-naphthalene-1,8-dicarboximide (ANI), the acceptor (A) is naphthalene-1,8:4,5-bis(dicarboximide) (NDI) and the stable radical (R•) is (2,2,6,6-tetramethylpiperidin-1-yl)oxyl (TEMPO). Experiments probed the effect of the spin–spin exchange interaction between D•+ and A•− as well as the charge recombination dynamics of D•+-C-A•− on spin polarization transfer from the D•+-C-A•− SCRP to R• as a function of the dielectric environment in glassy media at cryogenic temperatures. The results show that spin polarization on R• is generated by asymmetry in the charge recombination pathways rather than variations in the spin–spin exchange interaction between D•+ and A•−. These results inform design criteria for using an SCRP to spin polarize a third spin for potential applications in quantum information science.
The electron spin resonance (ESR) spectroscopy technique was used to study various organic radicals, such as 2,2,6,6-tetramethyl-1-piperidinyloxyl (TEMPO), 4-hydroxy-TEMPO (TEMPOL), 2-X-nitronylnitroxide (2-X-NN, X = Ph, NO2Ph, or cyclohexyl), 4-Y-benzonitronylnitroxide (4-Y-PhBzNN, Y = Ph or NO2Ph), and 2-Z-iminonitroxide (2-Z-IN, Z = Ph or NO2Ph) dispersed in a polymethylmethacrylate (PMMA) matrix. The experiments were conducted at room temperature. The complex nature of the recorded ESR spectra could be attributed to the superposition of the rotational diffusion component of TEMPO (or TEMPOL) in the nanospace of the PMMA matrix with the rigid-limit component. A single component of the rigid-limit was observed for 2-X-NN and 4-Y-PhBzNN radicals dispersed in the PMMA matrix. The isotropic components of g and hyperfine (A) tensor, estimated by analyzing the solution spectra, were used to determine the g and A components of 4-Y-PhBzNN. Only the rotational diffusion component was observed for the 2-Z-IN radical. These results demonstrated that the PMMA matrix contains cylindrical nanospaces. Various radicals other than TEMPO derivatives could be used in the ESR spin probe technique as probe molecules for determining the structures, sizes, and shapes of the nanospaces.
Molecular activatable probes with near‐infrared (NIR) fluorescence play a critical role in in vivo imaging of biomarkers for drug screening and disease diagnosis. With structural diversity and high fluorescence quantum yields, hemicyanine dyes have emerged as a versatile scaffold for the construction of activatable optical probes. This Review presents a survey of hemicyanine‐based NIR activatable probes (HNAPs) for in vivo imaging and early diagnosis of diseases. The molecular design principles of HNAPs towards activatable optical signaling against various biomarkers are discussed with a focus on their broad applications in the detection of diseases including inflammation, acute organ failure, skin diseases, intestinal diseases, and cancer. This progress not only proves the unique value of HNAPs in preclinical research but also highlights their high translational potential in clinical diagnosis.
In recent years, the use of aggregation-induced emission luminogens (AIEgens) for biological imaging and phototherapy has become an area of intense research. However, most traditional AIEgens suffer from undesired aggregation in aqueous media with "always on" fluorescence, or their targeting effects cannot be maintained accurately in live cells with the microenvironment changes. These drawbacks seriously impede their application in the fields of bio-imaging and antitumor therapy, which require a high signal-to-noise ratio. Herein, we propose a molecular design strategy to tune the dispersity of AIEgens in both lipophilic and hydrophilic systems to obtain the novel near-infrared (NIR, ∼737 nm) amphiphilic AIE photosensitizer (named TPA-S-TPP) with two positive charges as well as a triplet lifetime of 11.43 μs. The synergistic effects of lipophilicity, electrostatic interaction, and structure-anchoring enable the wider dynamic range of AIEgen TPA-S-TPP for mitochondrial targeting with tolerance to the changes of mitochondrial membrane potential (ΔΨ m). Intriguingly, TPA-S-TPP was difficult for normal cells to be taken up, indicative of low inherent toxicity for normal cells and tissues. Deeper insight into the changes of mitochondrial membrane potential and cleaved caspase 3 levels further revealed the mechanism of tumor cell apoptosis activated by AIEgen TPA-S-TPP under light irradiation. With its advantages of low dark toxicity and good biocompatibility, acting as an efficient theranostic agent, TPA-S-TPP was successfully applied to kill cancer cells and to efficiently inhibit tumor growth in mice. This study will provide a new avenue for researchers to design more ideal amphiphilic AIE photosensitizers with NIR fluorescence.
This study focuses on the synthesis, characterization, and assessment of the synergistic effect of 2,2,6,6, tetramethylpiperidine-N-oxyl (TEMPO)-coated titanium dioxide nanorods (TiO2 NRs) for photodynamic therapy (PDT). Firstly, TiO2 NRs were synthesized by the sol–gel technique. Then, TEMPO was grafted on TiO2 NRs with the aid of oxoammonium salts. Next, the final product was characterized by applying manifold characterization techniques. X-ray diffraction was used to perform crystallographic analysis; transmission electron microscopy (TEM) was used to conduct morphological analysis; Fourier transform infrared (FTIR) and Raman spectra were recorded to perform molecular fingerprint analysis. Furthermore, experimental and empirical modeling was performed to confirm the suitability of as-prepared samples for PDT applications using (MCF-7 cell line) Human Breast Cancer cell line. Our results revealed that bare TiO2 NRs did not exhibit a significant response for therapeutic applications compared to TEMPO-conjugated TiO2 NRs in the dark; however, they exhibited a prominent response for the PDT application under UV-A light. Therefore, it is concluded that TEMPO-coated TiO2 NRs shows the synergistic response for therapeutic approach under UV-A light irradiation. In addition, TEMPO capped TiO2 nanorods not only overcome the multidrug resistance (MDR) hindrance but also exhibit excellent response for cancer cell (MCF-7 cells) treatment only under UV light irradiation via PDT. It is expected that the proposed TiO2 NRs + TEMPO nanocomposite, which is suitable for PDT treatment, may be essential for photodynamic therapy.
Prodrugs activated by endogenous stimuli face the problem of tumor heterogeneity. Bioorthogonal prodrug activation that utilizes an exogenous click reaction has the potential to solve this problem, but most of the strategies currently used rely on the presence of endogenous receptors or overexpressed enzymes. We herein integrate the acidic, extracellular microenvironment of a tumor and a click reaction as a general strategy for prodrug activation. This was achieved by using a tumor pH‐responsive polymer containing tetrazine groups, which formed unreactive micelles in the blood but disassembled in response to tumor pH. The vinyl ether group on the macrotheranostic prodrug (CyPVE) is activated by the tetrazine groups, which was confirmed by tumor‐specific fluorescence activation and phototoxicity restoration. Therefore, the bioorthogonal reactions in the context of the ubiquitous acidic tumor microenvironment can provide a general strategy for bioorthogonal prodrug activation.
Mitophagy is a selective form of autophagy by which dysfunctional and damaged mitochondria are degraded in autolysosomes. Since defective mitophagy is closely related to various pathological processes, investigation on the detailed mitophagy process is of great importance. In this respect, disclosing the alterations of mitochondrial microenvironment is expected to be a promising way. However, a proper method for monitoring the flunctuations of mitochondrial polarity during mitophagy is still lacking. Here, we report a near-infrared hydroxyl-hemicyanine fluorescent probe that responses to polarity exclusively. Both the shift of emission maxima and the fluorescence intensity ratios at two different wavelengths of the probe can be applied to quantifying the polarity accurately. With ratiometric fluorescence imaging, the polarity differences of normal and cancer cells are clearly discriminated. Most importantly, the mitochondrial polarity variations during starvation and drug-induced mitophagy are determined for the first time. The observed decrease of mitochondrial polarity during mitophagy, together with the rationally designed probe, may facilitate the study on the vital role of mitophagy in physiological and pathological bioprocesses.
Organic photosensitizers possessing efficient intersystem crossing (ISC) and forming long-living triplet excited states, play a crucial role in a number of applications. A common approach in the design of such dyes relies on the introduction of heavy atoms (e.g. transition metals or halogens) into the structure, which promote ISC via spin-orbit coupling (SOC) interaction. In recent years, alternative methods to enhance ISC have been actively studied. Among those, the generation of triplet excited states through photoinduced electron transfer (PET) in heavy-atom-free molecules has attracted particular attention because it allows for the development of photosensitizers with programmed triplet states and fluorescence quantum yields. Due to their synthetic accessibility and tunability of optical properties, boron dipyrromethenes (BODIPYs) are so far the most perspective class of photosensitizers operating via this mechanism. This article reviews recently reported heavy-atom-free BODIPY donor-acceptor dyads and dimers which produce long-living triplet excited states and generate singlet oxygen. Structural factors which affect PET and concomitant triplet state formation in these molecules are discussed and the reported data on triplet state yields and singlet oxygen generation quantum yields in various solvents are summarized. Finally, examples of recent applications of these systems are highlighted.
Photodynamic therapy is considered as a promising treatment for cancer, but still facing several challenges. Hypoxia environment in solid tumor, imprecise tumor recognition and lack of selectivity between normal and cancer cells extremely hinder their applications in clinic. Moreover, the “always on” property of photosensitizer also increases the toxicity to normal tissues when exposed to light irradiation. In this study, a hypoxia-activated NIR photosensitizer ICy-N was synthesized and successfully applied for in vivo cancer treatment. ICy-N is in the unactivated state with low fluorescence whereas its NIR emission (λem = 716 nm) was induced via the reduction caused by nitroreductase at the tumor site. In addition, the reduced product ICy-OH specially located at mitochondria and demonstrated a high singlet oxygen production under 660 nm light irradiation, which efficiently induced cell apoptosis (IC50= 0.63 μM). The in vivo studies carried out in Balb/c mice indicated that ICy-N was suitable for precise tumor hypoxia imaging and can work as an efficient photosensitizer for restraining tumor growth through PDT process.
The dehydrating cyclotrimerization of 1‐tetralone in the presence of titanium tetrachloride at high temperatures leads to homotruxene, a nonplanar arene in which the twist angles between its three outer benzene rings and the central benzene are stabilized by ethylene bridges. This non‐planar configuration allows for pronounced spin–orbit coupling and a high triplet energy, leading to room‐temperature phosphorescence in air with a lifetime of 0.38 s and a quantum yield of 5.6 %, clearly visible to the human eye after switching off the excitation. Triplet–triplet annihilation is found to simultaneously lead to a substantial delayed fluorescence, unprecedented from a pure hydrocarbon at ambient conditions, with a lifetime of 0.11 s.
A stable radical (2,2,6,6‐tetramethylpiperidinyloxy; TEMPO) was labeled with the visible‐light‐harvesting chromophore naphthalenediimide (NDI) to give an NDI‐TEMPO dyad. A detailed photophysical study shows efficient radical enhanced intersystem crossing (EISC; 74%), long‐lived triplet state of NDI (8.7 μs), quartet state formation (spin quantum number, S=3/2), and inversion of the electron spin polarization. This red‐light absorption, heavy‐atom‐free dyad NDI‐TEMPO has been used in photodynamic therapy, and hence opens new avenues of real applications of EISC systems. More information can be found in the Full Paper by J. Zhao, H. G. Yaglioglu, R. Das, L. Luo, J. Li, et al. (DOI: 10.1002/chem.201804212).
The emerging treatment approaches, such as gas therapy (GT), photodynamic therapy (PDT) and photothermal therapy (PTT), have received widespread attention. The development of an intelligent multifunctional nano-platform responding to tumor microenvironment for multimodal therapy is highly desirable. Herein, a near-infrared (NIR) light-responsive nitric oxide (NO) photodonor (4-Nitro-3-Trifluoromethylaniline, NF) and a pH-sensitive group (dimethylaminophenyl) have been introduced into diketopyrrolopyrrole core (denoted as DPP-NF). The DPP-NF nanoparticles (NPs) can be activated under weakly acidic condition of lysosome (pH 4.5-5.0) to generate reactive oxygen species (ROS) and enhance photothermal efficiency. The fluorescence detection demonstrated that NO controllable release can be realized by "on-off" of NF unit under NIR light irradiation or dark. The controllable NO release of DPP-NF NPs not only can trigger tumor cells death by DNA damage, but also overcome the PDT inefficiencies caused by hypoxia in tumor. Additionally, DPP-NF NPs displayed 45.6% photothermal conversion efficiency, which is superior to other reported DPP derivatives. In vitro studies showed that DPP-NF NPs possessed low dark toxicity and high phototoxicity with half-maximal inhibitory concentration about 38 μg/mL. In vivo phototherapy indicated that DPP-NF NPs exhibited excellent tumor phototherapeutic efficacy with passive targeting to tumor site via the enhanced permeability and retention (EPR) effect. These results highlight that the nano-platform has a promising potential in NO-mediated multimodal synergistic phototherapy in clinical.
Photodynamic therapy (PDT)was discoveredmore than 100 years ago, and has since become a well-studied therapy for cancer and various non-malignant diseases including infections. PDT uses photosensitizers (PSs, non-Toxic dyes) that are activated by absorption of visible light to initially form the excited singlet state, followed by transition to the long-lived excited triplet state. This triplet state can undergo photochemical reactions in the presence of oxygen to form reactive oxygen species (including singlet oxygen) that can destroy cancer cells, pathogenic microbes and unwanted tissue. The dual-specificity of PDT relies on accumulation of the PS in diseased tissue and also on localized light delivery. Tetrapyrrole structures such as porphyrins, chlorins, bacteriochlorins and phthalocyanines with appropriate functionalization have been widely investigated in PDT, and several compounds have received clinical approval. Other molecular structures including the synthetic dyes classes as phenothiazinium, squaraine and BODIPY (boron-dipyrromethene), transition metal complexes, and natural products such as hypericin, riboflavin and curcumin have been investigated. Targeted PDT uses PSs conjugated to antibodies, peptides, proteins and other ligands with specific cellular receptors. Nanotechnology has made a significant contribution to PDT, giving rise to approaches such as nanoparticle delivery, fullerene-based PSs, titania photocatalysis, and the use of upconverting nanoparticles to increase light penetration into tissue. Future directions include photochemical internalization, genetically encoded protein PSs, theranostics, two-photon absorption PDT, and sonodynamic therapy using ultrasound.
Mitochondria are not only the main energy source of the cell but are also involved in the regulation of important physiology and processes of diseases including cancers. Therefore, mitochondria have been regarded as an important research target to reveal the mechanism of disease regulation and tumor treatment. Compared to conventional probes, thanks to their merits including resistance to aggregation-caused quenching capability, larger Stokes shift, high photostability, good biocompatibility as well as easy preparation, aggregation-induced emission luminogens (AIEgens) have become one of the research hotspots. Intriguingly, numerous mitochondria-targeted smart AIEgens have been fabricated in recent year to dynamically monitor behaviors of mitochondria or effectively kill tumors through activating mitochondria-regulated cell death pathways. In this review, we provide the systematic information of working mechanisms of aggregation-induced emission (AIE), design principles of mitochondria-targeted AIEgens, examples of AIEgnes for imaging mitochondria and detecting bio-species in mitochondria, and the relevant biomedical applications in theranostics. Additionally, the challenges and prospects of mitochondria-targeted AIEgens for imaging and therapeutics are also presented.
During cancer treatment, chemotherapeutic drugs always result in severe side-effects and drug resistance. Therefore, combining cheomtherapy with other therapeutic modalities, such as photodynamic therapy (PDT) and designing an activable platform is promising for precise and efficient anticancer treatment. Herein, we report a “pro-drug-photosensitizer” agent, LMB-S-CPT, bearing a disulfide bond as the glutathione (GSH)-activatable linker. LMB-S-CPT can be selectively activated by GSH to release activated drug, camptothecin (CPT), for chemotherapy and activated photosensitizer, methylene blue (MB), for PDT. LMB-S-CPT exhibits excellent tumor-activatable performance when injected into tumor-bearing mice, as well as specific cancer therapy with negligible toxic side effects. The activatable pro-drug-photosensitizer offers a new strategy for chemo-photodynamic therapy and displays precise, selective and excellent antitumor effect.
The discovery of a near-infrared (NIR, 650-900 nm) fluorescent chromophore hemicyanine dye with high structural tailorability is of great significance in the field of detection, bioimaging, and medical therapeutic applications. It exhibits many outstanding advantages including absorption and emission in the NIR region, tunable spectral properties, high photostability as well as a large Stokes shift. These properties are superior to those of conventional fluorogens, such as coumarin, fluorescein, naphthalimides, rhodamine, and cyanine. Researchers have made remarkable progress in developing activity-based multifunctional fluorescent probes based on hemicyanine skeletons for monitoring vital biomolecules in living systems through the output of fluorescence/photoacoustic signals, and integration of diagnosis and treatment of diseases using chemotherapy or photothermal/photodynamic therapy or combination therapy. These achievements prompted researchers to develop more smart fluorescent probes using a hemicyanine fluorogen as a template. In this review, we begin by describing the brief history of the discovery of hemicyanine dyes, synthetic approaches, and design strategies for activity-based functional fluorescent probes. Then, many selected hemicyanine-based probes that can detect ions, small biomolecules, overexpressed enzymes and diagnostic reagents for diseases are systematically highlighted. Finally, potential drawbacks and the outlook for future investigation and clinical medicine transformation of hemicyanine-based activatable functional probes are also discussed.
Molecular activatable probes with near‐infrared (NIR) fluorescence play a critical role in in vivo imaging of biomarkers for drug screening and disease diagnosis. With structural diversity and high fluorescence quantum yields, hemicyanine dyes have emerged as a versatile scaffold for construction of activatable optical probes. This review presents a survey of hemicyanine‐based NIR activatable probes (HNAPs) for in vivo imaging and early diagnosis of diseases. The molecular design principles of HNAPs towards activatable optical signaling against various biomarkers are discussed with a focus on their broad applications in detection of disease including inflammation, acute organ failure, skin diseases, intestinal diseases and cancer. These progresses not only proves the unique value of HNAPs in preclinical research but also highlight their high translational potential in clinical diagnosis.
Photodynamic therapy (PDT), a light triggered therapeutic mode, has been recognized as an attractive treatment for oncotherapy. The phototoxicity to normal tissues during treatment limited the development of PDT owning to the always “on” properties of photosensitizers. Activatable photosensitizers are of great importance for improving the selectivity of PDT. Herein, we regarded the overexpressed GGT (γ-Glutamyl transpeptidase) enzyme in tumor cells as a biomarker and developed an activatable photosensitizer Cy-GGT by decorating a specific recognition moiety of GGT, L-glutamic acid, to a hemicyanine dye based on photosensitizer Cy-NH2. Cy-GGT was in the “off” state with negligible fluorescence and suppressed singlet oxygen generation, but it could be specifically hydrolyzed to Cy-NH2 in the presence of GGT, accompanied with significant fluorescence recovery and singlet oxygen generation increase under light irradiation. The in vitro and in vivo studies indicated that Cy-GGT was suitable for precise tumor imaging and could work as an efficient photosensitizer for inhibiting tumor growth.
Singlet oxygen (1O2) is a strong oxidant which plays important roles in photodynamic therapy (PDT). The exploitation of photosensitizers with high 1O2 production is crucial to improve PDT efficiency. In this study, a radical labeled quartet photosensitizer Cy-DENT is reported with high singlet oxygen quantum yield (ΦΔ=32.3%) due to a radical enhanced inter-system crossing (ISC) process. After the introduction of 2,2,6,6-tetramethylpiperidinyloxy (TEMPO) radical, quartet state 4[R,T] of Cy-DENT could be formed to give an over 20-fold enhancement of singlet oxygen quantum yield compared to Cy-DEN (without TEMPO radical) under irradiation of near infrared (NIR) light. In addition, the 1O2 production is well controlled by varying the electron-donating ability of the terminal substituent group. Cy-DENT possesses good cell permeability and is localized in mitochondria. Under the irradiation of 700 nm light, Cy-DENT can produce high levels of ROS to destroy the mitochondria membrane potential and induce cell apoptosis. Through the encapsulation of PEG-SS-PCL micelle, Cy-DENT can be effectively delivered to tumors and suppresses the tumor growth after PDT treatment.
ConspectusPhotodynamic therapy (PDT) is a clinically approved therapeutic modality that has shown great potential for the treatment of cancers owing to its excellent spatiotemporal selectivity and inherently noninvasive nature. However, PDT has not reached its full potential, partly due to the lack of ideal photosensitizers. A common molecular design strategy for effective photosensitizers is to incorporate heavy atoms into photosensitizer structures, causing concerns about elevated dark toxicity, short triplet-state lifetimes, poor photostability, and the potentially high cost of heavy metals. To address these drawbacks, a significant advance has been devoted to developing advanced smart photosensitizers without the use of heavy atoms to better fit the clinical requirements of PDT. Over the past few years, heavy-atom-free nonporphyrinoid photosensitizers have emerged as an innovative alternative class of PSs due to their superior photophysical and photochemical properties and lower expense. Heavy-atom-free nonporphyrinoid photosensitizers have been widely explored for PDT purposes and have shown great potential for clinical oncologic applications. Although many review articles about heavy-atom-free photosensitizers based on porphyrinoid structure have been published, no specific review articles have yet focused on the heavy-atom-free nonporphyrinoid photosensitizers.In this account, the specific concept related to heavy-atom-free photosensitizers and the advantageous properties of heavy-atom-free photosensitizers for cancer theranostics will be briefly introduced. In addition, recent progress in the development of heavy-atom-free photosensitizers, ranging from molecular design approaches to recent innovative types of heavy-atom-free nonporphyrinoid photosensitizers, emphasizing our own research, will be presented. The main molecular design approaches to efficient heavy-atom-free PSs can be divided into six groups: (1) the approach based on traditional tetrapyrrole structures, (2) spin-orbit charge-transfer intersystem crossing (SOCT-ISC), (3) reducing the singlet-triplet energy gap (ΔEST), (4) the thionation of carbonyl groups of conventional fluorophores, (5) twisted π-conjugation system-induced intersystem crossing, and (6) radical-enhanced intersystem crossing. The innovative types of heavy-atom-free nonporphyrinoid photosensitizers and their applications in cancer diagnostics and therapeutics will be discussed in detail in the third section. Finally, the challenges that need to be addressed to develop optimal heavy-atom-free photosensitizers for oncologic photodynamic therapy and a perspective in this research field will be provided. We believe that this review will provide general guidance for the future design of innovative photosensitizers and spur preclinical and clinical studies for PDT-mediated cancer treatments.
HClO is an important signaling species, which is closely related to many diseases and physiological activities of the organism, and it plays an important role in the immune system. Therefore, it is of great significance to accurately and quickly monitor the HClO in the living organisms. However, in vivo detection of endogenous HClO remains a challenge. Herein, a near-infrared fluorescent probe CyClOP for the rapid detection of HClO was developed. CyClOP possesses several advantages such as fast response, good selectivity and deep penetration. CyClOP was successfully applied to the detection of endogenous HClO in living cells. In addition, CyClOP effectively captured the fluctuation of endogenous HClO produced by the immune system during infection with Staphylococcus aureus and muscle tissue injury in living mice for the first time. This work provides a reliable tool for monitoring of endogenous HClO in vivo and in vitro, and it has great potential for future research in HClO-related biology and pathology.
Photodynamic therapy has been developed as a prospective cancer treatment in recent years. Nevertheless, conventional photosensitizers suffer from lacking recognition and specificity to tumors, which causing severe side effects to normal tissues, while the enzyme-activated photosensitizers are capable of solving these conundrums due to high selectivity towards tumors. APN (Aminopeptidase N, APN/CD13), a tumor marker, has become a crucial targeting substance owing to its highly expressed on the cell membrane surface in various tumors, which has become a key point in the research of anti-tumor drug and fluorescence probe. Based on it, herein an APN-activated near-infrared (NIR) photosensitizer (APN-CyI) for tumor imaging and photodynamic therapy has been firstly developed and successfully applied in vitro and in vivo. Studies showed that APN-CyI could be activated by APN in tumor cells, hydrolyzed to fluorescent CyI-OH, which specifically located in mitochondria in cancer cells and exhibited a high singlet oxygen yield under NIR irradiation, and efficiently induced cancer cell apoptosis. Dramatically, the in vivo assays on Balb/c mice showed that APN-CyI could achieve NIR fluorescence imaging (λem = 717 nm) for endogenous APN in tumors and possessed an efficient tumor suppression effect under NIR irradiation.
In this study, a near-infrared (NIR) theranostic photosensitizer was developed based on a heptamethine aminocyanine dye with a long-lived triplet state. This theranostic molecule can be activated by nitroreducatase under...
Two ratiometric near-infrared fluorescent probes have been developed to selectively detect mitochondrial pH changes based on highly efficient through-bond energy transfer (TBET) from cyanine donors to near-infrared hemicyanine acceptors. The probes consist of identical cyanine donors connected to different hemicyanine acceptors with a spirolactam ring structure linked via a biphenyl linkage. At neutral or basic pH, the probes display only fluorescence of the cyanine donors when they are excited at 520 nm. However, acidic pH conditions trigger spirolactam ring opening, leading to increased π-conjugation of the hemicyanine acceptors, resulting in new near-infrared fluorescence peaks at 740 nm and 780 nm for probes A and B, respectively. This results in ratiometric fluorescence responses of the probes to pH changes indicated by decreases of the donor fluorescence and increases of the acceptor fluorescence under donor excitation at 520 nm due to a highly efficient TBET from the donors to the acceptors. The probes only show cyanine donor fluorescence in alkaline-pH mitochondria. However, the probes show moderate fluorescence decreases of the cyanine donor and considerable fluorescence increases of hemicyanine acceptors during the mitophagy process induced by nutrient starvation or under drug treatment. The probes display rapid, selective, and sensitive responses to pH changes over metal ions, good membrane penetration, good photostability, large pseudo-Stokes shifts, low cytotoxicity, mitochondria-targeting, and mitophagy-tracking capabilities.
We designed and synthesized a water-soluble near-infrared (NIR) fluorescent probe with the recognition unit of the cyanine-like structure and acrylate group. Through an aromatic ring nucleophilic substitution reaction based on sulfhydryl moiety, an off-on fluorescence response toward cysteine (Cys) was realized. The probe exhibited excellent spectral performance with an emission wavelength of 720 nm and a detection limit of 0.20 μM. The spectral properties, selectivity and anti-interference performance of the probe were systematically investigated. Density functional theory (DFT) calculations were conducted to clarify the luminescence mechanism of the probe. Furthermore, the probe was successfully applied to the detection of free Cys in human serum and the NIR imaging of endogenous Cys in living cells. Thus, the probe has a promising application prospect in clinical diagnosis and fluorescence imaging.
A novel strategy for designing highly efficient and activatable photosensitizers that can effectively generate reactive oxygen species (ROS) under both normoxia and hypoxia is proposed. Replacing both oxygen atoms in conventional naphthalimides (RNI-O) with sulfur atoms led to dramatic changes in the photophysical properties. The remarkable fluorescence quenching (ΦPL ~ 0) of the resulting thionaphthalimides (RNI-S) suggested that the intersystem crossing from the singlet excited state to the reactive triplet state was enhanced by the sulfur substitution. Surprisingly, the singlet oxygen quantum yield of RNI-S gradually increased with increasing electron-donating ability of the 4-R substituents (MANI-S, Φ∆ ~ 1.00, in air-saturated acetonitrile). Theoretical studies revealed that small singlet-triplet energy gaps and large spin-orbit coupling could be responsible for the efficient population of the triplet state of RNI-S. In particular, the ROS generation ability of MANI-S was suppressed under physiological conditions due to their self-assembly and was significantly recovered in cancer cells. More importantly, cellular experiments showed that MANI-S still produced a considerable amount of ROS even under severely hypoxic conditions (1% O2) through a type-I mechanism.
Mitochondria, powerhouses of cells, possess a weakly alkaline environment. Various stress stimulations may lead to mitophagy, which further gives a rise to mitochondrial acidification and disfunction. Therefore, monitoring mitochondrial pH alterations is of great importance to better elucidate their role in the cellular metabolism. Toward this end, a number of mitochondrial fluorescent pH probes have been proposed, but most of them are based on electrostatic attraction and readily leak out from the mitochondria during mitophagy with decreased membrane potential, thus failing to accurately measure the pH changes. In this work, we report a mitochondria-immobilized ratiometric fluorescent pH probe, which allows the quantitative measurements of mitochondrial pH. The probe was designed and prepared by introducing a reactive benzyl chloride into a positively charged near-infrared hydroxyl-hemicyanine. The cationic property facilitates the probe to be quickly enriched into mitochondria, the hydroxyl group is responsible for producing a reversible ratiometric fluorescence signal, and benzyl chloride is used to react with nucleophiles for immobilizing the probe in mitochondria. Taking these advantages of the probe, the mitochondrial pH variations during mitophagy caused by rapamycin and hypoxia have been determined quantitatively for the first time. The observed severe acidification of mitochondria under these stimulations, together with the rationally designed probe, may be useful for studying the detailed function of mitochondria in some bioprocesses.
Compact naphthalenediimide (NDI)‐2,2,6,6‐tetramethyl piperidine N‐oxyl (TEMPO) dyad was prepared, with the aim to study the radical‐enhanced intersystem crossing (EISC) and the formation of high spin states, as well as the electron spin polarization (ESP) dynamics. Compared to the previously reported radical‐chromophore dyads, the present system shows a very high triplet state quantum yield (74%), long‐lived triplet state (8.7 microseconds), fast EISC (338 ps) and absorption in the red spectral region. Time‐resolved electron paramagnetic resonance (TRERP) spectroscopy shows that, on photoexcitation in fluid solution at room temperature, the D0 state of TEMPO moiety produces strong emissive (E) polarization, owing to the quenching of the excited singlet state of NDI by the radical moiety (electron exchange J > 0). The emissive polarization then inverts to absorptive (A) within about 3 microseconds, and then relaxes to thermal equilibrium while quenching the triplet state of NDI. Formation and decay of the quartet state were also observed. The dyad was used as a three‐spin triplet photosensitizer for triplet‐triplet annihilation upconversion (quantum yield is 2.6%). Remarkably, when encapsulated into liposomes, the red light‐absorbing dyad‐liposomes show good biocompatibility and excellent photodynamic therapy (PDT) efficiency (phototoxicity EC50 = 3.22 micromolar), serving as a promising candidate for future less‐toxic and multi‐functional photodynamic therapeutic reagents.
Peroxynitrite (ONOOˉ), a reactive and short-lived biological oxidant, is closely related with many pathological conditions such as cancer. However, real-time in vivo imaging of ONOOˉ in tumor remains to be challenging. Herein, we develop a near-infrared fluorescence (NIRF) and photoacoustic dual-modal molecular probe (CySO3CF3) composed of a water-soluble hemicyanine dye caged with a trifluoromethyl ketone moiety for in vivo imaging of ONOOˉ. The trifluoromethyl ketone moiety can undergo a series of ONOOˉ-induced cascade oxidation-elimination reactions, leading to sensitive and specific fluorescence and photoacoustic turn-on re-sponses toward ONOOˉ; while a zwitterionic structure of the hemicyanine component ensures good water-solubility. CySO3CF3 thus not only can specifically detect ONOOˉ in solution and cells with the limit of detec-tion down to 53 nM, but also allows for NIRF and photoacoustic dual-modal imaging of ONOOˉ in the tumor of living mice.
Theranostics provides opportunities for precision cancer therapy. However, theranostic systems that simultaneously turn on their diagnostic signal and pharmacological action only in respond to targeted biomarker have been less exploited. We herein report the synthesis of a smart macrotheranostic probe that specifically turns on near‐infrared fluorescence, photoacoustic and photothermal signals in the presence of cancer‐overexpressed enzyme for imaging‐guided cancer therapy. Supervisor to the small‐molecule counterpart probe, the macrotheranostic probe has an ideal biodistribution and renal clearance, permitting passive targeting of tumor, in‐situ enzymatic activation of multimodal signals, and effective photothermal therapy. Our study thus provides a macromolecular approach towards activatable multimodal phototheranostics.
A long-lived triplet excited state of the well-known fluorophore borondipyrromethene (Bodipy) was observed for the first time via efficient radical-enhanced intersystem crossing (EISC). The triplet state has been obtained in two dyads prepared by linking Bodipy to a nitroxide radical, 2,2,6,6-tetramethyl-1-piperidinyloxyl (TEMPO) with two different length spacers. The photophysical properties were studied with steady-state and time-resolved transient optical spectroscopies, and electron spin resonance (cw-ESR and TR-ESR). The fluorescence of Bodipy units is significantly quenched in the dyads, and the spin polarized TEMPO signal were observed with TR-ESR, generated by a Radical Triplet Pair Mechanism. Efficient EISC (triplet state quantum yield = 80%) was observed for the dyad with shorter linker and the triplet state lifetime of Bodipy chromophore is exceptionally long (62 microseconds). The EISC takes 250 ps. Poor ISC was observed for the dyad with longer linker. The efficient ISC and long-lived triplet excited state in this flexible system are in stark contrast to the previously studied rigid EISC systems. EISC effect was employed for the first time to perform triplet-triplet annihilation (TTA) upconversion (upconversion quantum yield = 6.7 %).
To integrate treatments of photothermal therapy, photodynamic therapy (PDT), and chemotherapy, this study reports on a multifunctional nanocomposite based on mesoporous silica-coated gold nanorod for high-performance oncotherapy. Gold nanorod core is used as the hyperthermal agent and mesoporous silica shell is used as the reservoir of photosensitizer (Al(III) phthalocyanine chloride tetrasulfonic acid, AlPcS4). The mesoporous silica shell is modified with β-cyclodextrin (β-CD) gatekeeper via redox-cleavable Pt(IV) complex for controlled drug release. Furthermore, tumor targeting ligand (lactobionic acid, LA) and long-circulating poly(ethylene glycol) chain are introduced via host–guest interaction. It is found that the nanocomposite can specifically target to hepatoma cells by virtue of the LA targeting moiety. Due to the abundant existence of reducing agents within tumor cells, β-CD can be removed by reducing the Pt(IV) complex to active cisplatin drug for chemotherapy, along with the releasing of entrapped AlPcS4 for effective PDT. As confirmed by in vitro and in vivo studies, the nanocomposite exhibits an obvious near-infrared induced thermal effect, which significantly improves the PDT and chemotherapy efficiency, resulting in a superadditive therapeutic effect. This collaborative strategy paves the way toward high-performance nanotherapeutics with a superior antitumor efficacy and much reduced side effects.
During the last few years, the preparation of novel fluorescent probes for the selective detection of chemical species inside mitochondria has attracted considerable attention because of their wide applications in chemistry, biology, and medical science. This feature article focuses on the recent advances in the design principles and recognition mechanisms of these kinds of fluorescent probes. In addition, their applications for the detection of reactive oxygen species (ROS), nitric oxide, reactive sulfur species (RSS), thioredoxin (Trx), metal ions, anions, etc. in the mitochondrion is discussed as well.
Boron dipyrromethene (Bodipy) is one of the most extensively investigated organic chromophores. Most of the investigations are focused on the singlet excited state of Bodipy, such as fluorescence. In stark contrast, the study of the triplet excited state of Bodipy is limited, but it is an emerging area, since the triplet state of Bodipy is tremendously important for several areas, such as the fundamental photochemistry study, photodynamic therapy (PDT), photocatalysis and triplet-triplet annihilation (TTA) upconversion. The recent developments in the study of the production, modulation and application of the triplet excited state of Bodipy are discussed in this review article. The formation of the triplet state of Bodipy upon photoexcitation, via the well known approach such as the heavy atom effect (including I, Br, Ru, Ir, etc.), and the new methods, such as using a spin converter (e.g. C60), charge recombination, exciton coupling and the doubly substituted excited state, are summarized. All the Bodipy-based triplet photosensitizers show strong absorption of visible or near IR light and the long-lived triplet excited state, which are important for the application of the triplet excited state in PDT or photocatalysis. Moreover, the methods for switching (or modulation) of the triplet excited state of Bodipy were discussed, such as those based on the photo-induced electron transfer (PET), by controlling the competing Förster-resonance-energy-transfer (FRET), or the intermolecular charge transfer (ICT). Controlling the triplet excited state will give functional molecules such as activatable PDT reagents or molecular devices. It is worth noting that switching of the singlet excited state and the triplet state of Bodipy may follow different principles. Application of the triplet excited state of Bodipy in PDT, hydrogen (H2) production, photoredox catalytic organic reactions and TTA upconversion were discussed. The challenges and the opportunities in these areas were briefly discussed.
Near-infrared (NIR) fluorescent sensors have emerged as promising molecular tools for imaging biomolecules in living systems. However, NIR fluorescent sensors are very challenging to be developed. Herein, we describe the discovery of a new class of NIR fluorescent dyes represented by 1a/1c/1e, which are superior to the traditional 7-hydroxycoumarin and fluorescein with both absorption and emission in the NIR region while retaining an optically tunable hydroxyl group. Quantum chemical calculations with the B3LYP exchange functional employing 6-31G(d) basis sets provide insights into the optical property distinctions between 1a/1c/1e and their alkoxy derivatives. The unique optical properties of the new type of fluorescent dyes can be exploited as a useful strategy for development of NIR fluorescent sensors. Employing this strategy, two different types of NIR fluorescent sensors, NIR-H(2)O(2) and NIR-thiol, for H(2)O(2) and thiols, respectively, were constructed. These novel sensors respond to H(2)O(2) or thiols with a large turn-on NIR fluorescence signal upon excitation in the NIR region. Furthermore, NIR-H(2)O(2) and NIR-thiol are capable of imaging endogenously produced H(2)O(2) and thiols, respectively, not only in living cells but also in living mice, demonstrating the value of the new NIR fluorescent sensor design strategy. The new type of NIR dyes presented herein may open up new opportunities for the development of NIR fluorescent sensors based on the hydroxyl functionalized reactive sites for biological imaging applications in living animals.
The current state of the role of imaging in photodynamic therapy (PDT) has been reported. PDT is a photochemistry-based approach that uses a light activatable chemical, termed a photosensitizer (PS), and light of an appropriate wavelength to impart cytotoxicity via the generation of reactive molecular species. The ability to accurately define tumor margins is a crucial aspect in optimization of surgical interventions and constitutes a major subset of the applications of this fluorescence imaging technique. The traditional implementation of PDT involves the administration of a synthetic PS followed by a period of delay, in which the PS accumulates in the tumor or tissue of interest, before light activation. A major drawback of conventional imaging strategies using one-photon excitation is the limited penetration of visible light into tissue. Progress in imaging technologies, targeting chemistries, and nanotechnology combined with a detailed understanding of PDT-related molecular mechanisms provides optimism for the future of imaging in PDT and may be an important conduit for new applications of PDT and for the development of patient individualized treatments.
Photodynamic killing of a cell population is generally considered to result from direct effects that occur in each cell. In some scenarios this may be an over-simplification and the potential for cell-cell signaling processes to contribute to the response of a population to photodynamic stress is addressed in this paper. Photodynamic killing of EMT6 cells in culture was studied in time and space using computerized time-lapse microscopy. The rate of cell killing was dependent on the fluence with both rapid and slower processes evident, the proportion of the former increasing with fluence. The spatial distribution of cell death was non-random and for the slow cell killing process was found to occur preferentially in the vicinity of dead or dying cells, suggesting a local signaling process. An inhibitory effect of extracellular catalase indicated the involvement of hydrogen peroxide in the spread of cell death and NADPH oxidase was determined as the principal source of hydrogen peroxide. This cell signaling pathway was observed for membrane-bound and mitochondrial photosensitizers but not for a nuclear photosensitizer. These secondary cell signalling pathways extend the oxidative damage to cells in space and time.