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A quasi-cross-shaped platinum(II) compound (L′2LPt) with inhibitory activity against thrombin binds the antiparallel two-G-tetrad G-quadruplex TBA. The NMR structure of the Pt–G-quadruplex was solved to reveal this unusual “wall-mounted” binding mode.

Activation of the cyclic GMP-AMP synthase-stimulator of the interferon gene (cGAS-STING) pathway is a potent anticancer immunotherapeutic strategy, and the induction of pyroptosis is a feasible way to stimulate the anticancer immune responses. Herein, two Pt

The sequence-dependent DNA secondary structures possess structure polymorphism. To date, studies on regulated ligands mainly focus on individual DNA secondary topologies, while lack focus on quadruplex-duplex hybrids (QDHs). Here, we design an organic-metal hybrid ligand L1 Pt(dien), which matches and selectively binds one type of QDHs with lateral duplex stem-loop (QLDH) with high affinity, while shows poor affinity for other QDHs and individual G4 or duplex DNA. The solution structure of QLDH MYT1L-L1 Pt(dien) complex was determined by NMR. The structure reveals that L1 Pt(dien) presents a chair-type conformation, whose large aromatic "chair surface" intercalates into the G-quadruplex-duplex interface via π-π stacking and "backrest" platinum unit interacts with duplex region through hydrogen bonding and electrostatic interactions, showing a highly matched lock-key binding mode. Our work provided guidance for spatial matching design of selectively targeting ligands to QDH structures.

G-quadruplexes (G4s) are prevalent in oncogenes and proposed to be potential antitumor drug targets. However, binding selectivity of compounds to G4s still faces challenges. Herein, we report a platinum(II) complex ( Pt1 ), whose affinity to G4-DNA is activated by adaptive binding and selectivity controlled by binding kinetics. The resolved structure of Pt1 /VEGF-G4 (a promoter G4) reveals that Pt1 matches 3'-G-tetrad of VEGF-G4 through Cl - -dissociation and loop rearrangement of VEGF-G4. Binding rate constants are determined by coordination bond breakage/formation and correlate fully with affinities. The selective rate-determining binding step, Cl - -dissociation upon G4-binding which is 2~3 orders of magnitude higher than dsDNA. We further demonstrate that Pt1 potently targets G4 in living cells, effectively represses VEGF expression and inhibits vascular growth in zebrafish. We present adaptive G4-binding activation and selectivity controlled by kinetics, providing a complementary design principle for compounds targeting G4 or similar biomolecules.

Ferroptosis regulates cell death through reactive oxygen species (ROS)-associated lipid peroxide accumulation, which is expected to affect the structure and polarity of lipid droplets (LDs), but with no clear evidence. Herein, we report the first example of an LD/nucleus dual-targeted ratiometric fluorescent probe, CQPP, for monitoring polarity changes in the cellular microenvironment. Due to the donor-acceptor structure of CQPP, it offers ratiometric fluorescence emission and fluorescence lifetime signals that reflect polarity variations. Using nucleus imaging as a reference, CQPP was applied to report the increase in LD polarity and the homogenization of polarity between LDs and cytoplasm in the ferroptosis model. This LD/nucleus dual-targeted fluorescent probe shows the great potential of using fluorescence imaging to study ferroptosis and ferroptosis-related diseases.

The nucleic acid G-quadruplex (G4) has emerged as a promising therapeutic target for a variety of diseases such as cancer and neurodegenerative disease. Among small-molecule G4-binders, pyridostatin (PDS) and its derivatives (

G-quadruplex (G4) transitions play integral roles in regulating biological functions and can be modified by ligands. However, little is known about G4 transitions. Herein, we reveal distinct pathways of a platinum(II) compound Pt-phen converting parallel-stranded MYC G4 to a hybrid-type structure. Three NMR structures, 1:1 5′-end binding, 1:1 3′-end binding and 2:1 Pt-phen–MYC G4 complexes, were determined by NMR. We find that Pt-phen drives G4 transition at a low ratio. Under physiological 100 mM K+ conditions, a significant stable hydrogen-bonded T:T:A triad is formed at 3′-end of hybrid-type Myc1234, and consequently, Pt-phen first binds the 5′-end to form a 1:1 5′-end binding complex and then disrupts the 3′ T:T:A triad and binds 3′-end to form a 2:1 complex with more Pt-phen. Remarkably, the G4 transition pathway is different in 5 mM K+ with Pt-phen first binding the 3′-end and then the 5′-end. ‘Edgewise-loop and flanking/ligand/G-tetrad’ sandwich structure formation and terminal T:T:A triad stabilization play decisive roles in advancing and altering transition pathways. Our work is the first to elucidate the molecular structures of G4 transitions driven by a small molecule. The ligand-driven G4 transition is a dynamic process that includes a quick G4 transition and multiple complexes formation.

The misregulation of nucleic acids behavior leads to cell dysfunction and induces serious diseases. A ratiometric fluorescence probe is a powerful tool to study the dynamic behavior and function relationships of nucleic acids. However, currently, no such effective probe has been reported for in situ, real-time tracking of nucleic acids in living cells and tissue sections. Herein, the unique probe named QPP-AS was rationally designed for ratiometric fluorescence response to nucleic acids through skillful regulation of the intramolecular charge-transfer capabilities of the electron acceptor and donor. Encouraged by the advantages of the selective nucleic acid response, ideal biocompatibility, and high signal-to-noise ratio, QPP-AS has been applied for in situ, real-time ratiometric fluorescence imaging of nucleic acids in living cells for the first time. Furthermore, we have demonstrated that QPP-AS is capable of visualizing the dynamic behavior of nucleic acids during different cellular processes (e.g., cell division and apoptosis) by ratiometric fluorescence imaging. More significantly, QPP-AS has been successfully used for ratiometric fluorescence imaging of nucleic acids in human tissue sections, which provides not only the cell contour, nuclear morphology, and nuclear-plasma ratio but also the nucleic acid content information and may greatly improve accuracy in clinicopathological diagnosis.

Mitochondrial damage, characterized by altered morphological distribution and the damage of cristae, is closely associated with mitochondrial disease. However, imaging methods for capturing mitochondrial morphology at the nanoscale level in live samples remain unavailable, which seriously hinders the accurate evaluation and diagnosis of mitochondrial-related diseases. In response, we propose a super-resolution quantification strategy based on structured illumination microscopy (SIM) for the rapid, accurate evaluation of mitochondrial morphology. Using the strategy, we accurately captured the morphological distribution of mitochondria at the nanoscale level in a way generally applicable to checking various cell processes and identifying patients with mitochondrial disease who exhibit the SLC25A46 mutation. We also used algorithm-assisted super-resolution imaging to quantitatively analyze damage to mitochondrial cristae, which supports a novel drug screening strategy—high-resolution drug screening—for investigating drugs’ pharmacodynamics on organelles in living cells. In short, our strategy improves the accurate examination of changes in mitochondrial morphology in living cells and indicates new ways in which SIM-imaging can assist in diagnosing mitochondrial disease at the single-cell level.

G-quadruplex DNA show structural polymorphism, leading to challenges in the use of selective recognition probes for the accurate detection of G-quadruplexes in vivo. Herein, we present a tripodal cationic fluorescent probe, NBTE, which showed distinguishable fluorescence lifetime responses between G-quadruplexes and other DNA topologies, and fluorescence quantum yield (Φf ) enhancement upon G-quadruplex binding. We determined two NBTE-G-quadruplex complex structures with high Φf values by NMR spectroscopy. The structures indicated NBTE interacted with G-quadruplexes using three arms through π-π stacking, differing from that with duplex DNA using two arms, which rationalized the higher Φf values and lifetime response of NBTE upon G-quadruplex binding. Based on photon counts of FLIM, we detected the percentage of G-quadruplex DNA in live cells with NBTE and found G-quadruplex DNA content in cancer cells is 4-fold that in normal cells, suggesting the potential applications of this probe in cancer cell detection.

The nucleus is considered the ideal target for anti-tumor therapy because DNA and some enzymes in the nucleus are the main causes of cell canceration and malignant proliferation. However, nuclear target drugs with good biosafety and high efficiency in cancer treatment are rare. Herein, a nuclear-targeted material MeTPAE with aggregation-induced emission (AIE) characteristics was developed based on a triphenylamine structure skeleton. MeTPAE can not only interact with histone deacetylases (HDACs) to inhibit cell proliferation but also damage telomere and nucleic acids precisely through photodynamic treatment (PDT). The cocktail strategy of MeTPAE caused obvious cell cycle arrest and showed excellent PDT anti-tumor activity, which offered new opportunities for the effective treatment of malignant tumors.

Two new trinuclear Pt(ii) complexes {[Pt(dien)]3(tib)}(NO3)6 (1) and {[Pt(dpa)]3(tib)}(NO3)6 (2) (dien: diethylenetriamine, dpa: bis-(2-pyridylmethyl)amine, tib: 1,3,5-tris(1H-imidazol-1-yl)benzene) have been designed, synthesized, characterized and applied to a series of biochemical studies. We found that both of the Pt(ii) complexes exhibited much better selectivity for human telomeric G-quadruplex sequence than promoter G-quadruplexes (c-kit, c-myc, and bcl2) or duplex DNA. Both complexes displayed comparative stability and affinity towards human telomeric G-quadruplex by the studies from surface plasmon resonance, fluorescence resonance energy transfer and polymerase chain reaction stop assays. The circular dichroism indicated that both complexes could induce and stabilize anti-parallel G-quadruplex structures. Molecule docking presented that Pt(ii) complex intercalated into the large groove of human telomeric G-quadruplex (PDB ID: 143D). Furthermore, telomeric repeat amplification protocol assays quantitatively evaluated the inhibition of telomerase activity caused by the Pt(ii) complexes. The obtained IC50 values of 6.41 ± 0.042 μM and 2.67 ± 0.035 μM for 1 and 2, respectively, exhibited strong telomerase inhibitions. All results suggest that such fan-shaped trinuclear Pt(ii) complexes are effective and selective G-quadruplex binders, as well as strong telomerase inhibitors. This study provides insight into the development of human telomeric G-quadruplex targeted anticancer drugs based on the metal complex.

Impairing mitochondrial function may change its viscosity. The detection of changes in the mitochondrial viscosity is of great significance for biomedical research. Herein, we have designed a D–π–A type two-photon fluorescent probe ((4-(bis(4-(pyridin-4-yl)phenyl)amino)styryl)-1-methylpyridin-1-ium iodide) (DPTPA-Py) with a triphenylamine derivative as the electron donor and pyridinium as the electron acceptor for detecting mitochondrial viscosity in living cells. It is found that both fluorescence intensity and fluorescence lifetime of DPTPA-Py are viscosity-dependent, and the probe exhibits a strong anti-interference ability in the detection of viscosity. The colocalization experiments reveal that DPTPA-Py is located in mitochondria only and is highly biocompatible. The probe shows no significant effects on the mitochondrial membrane potential. In addition, the probe is successfully used to monitor mitochondrial viscosity changes during apoptosis of living cells by two-photon microscopy (TPM) and fluorescence lifetime imaging microscopy (FLIM).

Damage to cell membranes, the outermost protection layer, is fatal to cells. However, precisely monitoring and in situ reporting cell membrane damage is not trivial. Herein, we present a molecular rotor probe, TPAE2, which can effectively bind to DNA and 1,2-dioleoyl-sn-glycero-3-phosphocholine in solution. Due to the light-up imaging characteristics of the molecular rotor, TPAE2 offers ultrafast and wash-free staining of plasma membrane with 160-fold fluorescence “turn-on” and excellent photostability. Once the membrane is damaged, TPAE2 can light-up the nucleus as a signal reporter. The cascade imaging of the cell membrane and nucleus using TPAE2 enabled real-time tracking of the whole process of cell apoptosis. What's more, under irradiation, TPAE2 stained on the cell membrane could penetrate cells rapidly and selectively stain the nucleus, self-reporting the cancer cell ablation process. This is the first example that a single molecule with multiple functions can light up the nucleus as an indication of cell membrane damage. The membrane-to-nucleus translocation strategy opens up a new avenue for the design of membrane damage diagnosis probes for biomedical applications.

Oncosis, depending on DNA damage and mitochondrial swelling, is an important approach for treating cancer and other diseases. However, little is known about the behavior of mitochondria during oncosis, due to the lack of probes for in situ visual illumination of the mitochondrial membrane and mtDNA. Herein, a mitochondrial lipid and mtDNA dual‐labeled probe, MitoMN, and a continuous add‐on assay, are designed to image the dynamic process of mitochondria in conditions that are unobservable with current mitochondrial probes. Meanwhile, the MitoMN can induce oncosis in a light‐activated manner, which results in the enlargement of mitochondria and the death of cancer cells. Using structured illumination microscopy (SIM), MitoMN‐stained mitochondria with a dual‐color response reveals, for the first time, how swelled mitochondria interacts and fuses with each other for a nonlinear enlargement to accelerate oncosis into an irreversible stage. With this sign of irreversible oncosis revealed by MitoMN, oncosis can be segregated into three stages, including before oncosis, initial oncosis, and accelerated oncosis., A mitochondrial lipids and mtDNA dual‐labeling probe with continuous add‐on assay is designed to induce and image the nonlinear enlargement of mitochondria during oncosis. Together with super‐resolution imaging, it is revealed how swelled mitochondria interact and fuse with each other for a nonlinear enlargement to accelerate oncosis into an irreversible stage, which can be a new landmark for staging oncosis process.

It is of great significance to track the platinum drugs in real time with super-resolution to elucidate their mechanism of action, such as their behavior and distribution in live cells. Such information is required for further drug development. However, it is always challenging to design platinum complexes suitable for such research. Herein, we design a luminescent building block (L) for metal complexes and a dinuclear platinum complex (Pt2 L) for super-resolution imaging. Because of its super-large Stokes shift and excellent photophysical properties, Pt2 L is capable of serving as an ideal candidate for super-resolution imaging with extremely low luminescence background and high photobleaching resistance. Moreover, upon light stimulation, a matter flux of Pt2 L escaping from autolysosomes to nucleus was observed, which represents a new transportation path. Utilizing the photoactivated escape properties, we can regulate the nuclear accessibility of Pt2 L form autolysosomes with photo-selectivity, which provides a new way to improve the targeting of platinum drugs.

We have developed a strategy "charge-driven tripod somersault on DNA" realizing both in vitro and in vivo ratiometric fluorescence imaging of the variations of endogenous SO2 derivatives in the nucleus for the first time., Although fluorescence tracing of small bioactive molecules in living cells has been extensively studied, it is still a challenging task to detect their variations in the nucleus mainly due to the impermeable nuclear membrane and nucleic acid interference. Herein, we take advantage of the nucleic acid enriched environment in the nucleus to establish a strategy, named “charge-driven tripod somersault on DNA”, for ratiometric fluorescence imaging of small bioactive molecules in the nucleus. Taking SO2 derivatives as a typical target analyte, a tripodal probe has been constructed by conjugating two DNA binding groups containing a SO2 derivative reaction site. Mechanism studies demonstrate that upon encountering and reacting with SO32–/HSO3–, a charge variation occurs at the responsive arm of the tripodal probe, triggering a tripod somersault on DNA, resulting in the conformational rearrangement of the DNA binding modes with DNA-modulated fluorescence change, which allows the second emission feature to emerge. In this strategy, probe–DNA binding is not influenced by RNA or non-specific protein association, thus making it ideal for tracing nucleus-localized analytes. The application of this strategy has realized both in vitro and in vivo ratiometric fluorescence imaging of the variations of endogenous SO2 derivatives in the nucleus for the first time, with high specificity and selectivity. Also, in theory, this strategy opens up a new avenue for the design of fluorescence probes for the nucleus-localized biological analytes.

Six novel dinuclear Ru(II)-arene complexes [Ru2(η6-p-cymene)2(1,3-bib)2Cl2]×2·Solvent (X = Cl− (1), I− (2), NO3− (3), BF4− (4), PF6− (5), CF3SO3− (6); 1,3-bib = 1,3-di(1H-imidazol-1-yl) benzene) were synthesized and fully characterized by FT-IR, 1H NMR, ESI-MS, Elemental Analysis (EA) and Powder X-ray Diffraction (PXRD). Single crystal X-ray diffractions studies showed that 3 and 4 have rigid bowl-like structures, where one counter-anion (NO3− for 3 and BF4− for 4) was trapped inside the cavity to balance the charge, respectively. Even complexes 1–6 showed only moderate or little anti-proliferative activity toward cancer cells, strong interactions with DNA molecules through intercalation, however, were confirmed by UV-Vis, CD and fluorescence spectroscopy. Apoptosis and cell cycle arrest studies for complex 2 with cancer A549 cells indicated concentration-dependent late apoptosis and the G1/G0 phase arrest. Interactions with the tripeptide glutathione (γ-L-Glu-L-Cys-Gly, GSH) might explain the relatively low antiproliferative potency of these complexes. This class of rigid dinuclear cations hold potential as DNA-targeting anticancer agents if their uptake and delivery could be under controlled.

DNA G-quadruplexes are not only attractive drug targets for cancer therapeutics, but also have important applications in supramolecular assembly. Here, we report a platinum(II)-based tripod (Pt-tripod) specifically binds the biological relevant hybrid-1 human telomeric G-quadruplex (Tel26), and strongly inhibits telomerase activity. Further investigations illustrate Pt-tripod induces the formation of monomeric and multimeric Pt-tripod‒Tel26 complex structures in solution. We solve the 1:1 and the unique dimeric 4:2 Pt-tripod–Tel26 complex structures by NMR. The structures indicate preferential binding of Pt-tripod to the 5ʹ-end of Tel26 at a low Pt-tripod/Tel26 ratio of 0–1.0. After adding more Pt-tripod, the Pt-tripod binds the 3ʹ-end of Tel26, unexpectedly inducing a unique dimeric 4:2 structure interlocked by an A:A non-canonical pair at the 3ʹ-end. Our structures provide a structural basis for understanding the dynamic binding of small molecules with G-quadruplex and DNA damage mechanisms, and insights into the recognition and assembly of higher-order G-quadruplexes., DNA G-quadruplexes occur in oncologically relevant regions, thus are interesting targets for cancer research and treatment. Here, the authors solved the 1:1 and 4:2 (ligand/DNA) NMR structures of human telomeric DNA in complex with platinum(II)-tripod ligand and show that the binding is dynamic.

Known as highly efficient metallodrugs that have been clinically used to treat various types of cancers, cisplatin and its analogues have attracted and inspired extensive interest in the development of both platinum and non-platinum drugs. However, the past years witnessed slow progress of newly approved metal-based drugs. One of the key obstacles that hamper the progress is the poor understanding of the mechanism-of-action of metal compounds which usually do not follow the drug-like principles of organic compounds and display complicated thermodynamic and kinetic reactivities. Nevertheless, unique techniques such as atomic spectroscopy, nuclear magnetic resonance, X-ray spectroscopy, emission spectroscopy, and secondary-ion mass spectrometry are available that can provide detailed information on the chemical and biological environment of metal ions/complexes. Moreover, emerging methods including chemical biology probes and multi-omics strategies have been developed which are of great merit in understanding the mechanism-of-action including the molecular target(s) of metal complexes. In this review, we summarized approaches for deciphering the mechanism-of-action of metal complexes and highlighted selected examples with in-depth mechanistic insights.

As the clock frequency rises rapidly, clock skew has acted as a more important role in synchronous circuits. In this paper, we propose a new buffered clock tree routing algorithm to prevent the influence of process variations and environmental effects to clock skew. Based on the concept of BSF (Branch Sensitivity Factor), our algorithm manages to reduce the skew sensitivity of clock tree in topology generation. The worst case skew due to wire width change has been estimated, and proper buffers are inserted avoiding large capacitance load. Experimental results show that our algorithm can produce a more reliable, process-insensitive clock tree, and control clock skews in their permissible range evidently.