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Fatty acid amide hydrolase (FAAH) overexpression induces several disorder symptoms in nerve systems, and therefore long-term tracing of FAAH activity in vivo is of high importance but remains challenging. Current bioluminescence (BL) methods are limited in detecting FAAH activity within 5 h. Herein, by rational design of a latent BL probe (d-Cys-Lys-CBT)2 (1), we developed a "smart" method of intracellular reduction-controlled self-assembly and FAAH-directed disassembly of its cyclic d-luciferin-based nanoparticles (i.e., 1-NPs) for persistent BL imaging of FAAH activity in vitro, in cells, and in vivo. Using aminoluciferin methyl amide (AMA), Lys-amino-d-luciferin (Lys-Luc), and amino-d-luciferin (NH2-Luc) as control BL probes, we validated that the persistent BL of 1 from luciferase-expressing cells or tumors was controlled by the activity of intracellular FAAH. With the property of long-term tracing of FAAH activity in vivo of 1, we envision that our BL precursor 1 could probably be applied for in vivo screening of FAAH inhibitors and the diagnosis of their related diseases (or disorders) in the future.

Large magnetic nanoparticles or aggregates are advantageous in their magnetic resonance properties over ultrasmall superparamagnetic iron oxide (USPIO) nanoparticles (NPs), but the former are cleared faster from the blood pool. Therefore, the "smart" strategy of intracellular aggregation of USPIO NPs is required for enhanced T2-weighted MR imaging. Herein, employing an enzyme-instructed condensation reaction, we rationally designed a small molecule Ac-Asp-Glu-Val-Asp-Cys(StBu)-Lys-CBT (1) to covalently modify USPIO NPs to prepare monodispersive Fe3O4@1 NPs. In vitro results showed that Fe3O4@1 NPs could be subjected to caspase 3 (Casp3)-instructed aggregation. T2 phantom MR imaging showed that the transverse molar relaxivity (r2) of Fe3O4@1 NPs with Casp3 or apoptotic HepG2 cells was significantly larger than those of control groups. In vivo tumor MR imaging results indicated that Fe3O4@1 NPs could be specifically applied for enhanced T2 MR imaging of tumor apoptosis. We propose that the enzyme-instructed intracellular aggregation of Fe3O4 NPs could be a novel strategy for the design of "smart" probes for efficient T2 MR imaging of in vivo biomarkers.

Nitric oxide (NO) is the first ubiquitous signaling molecule in the human body. The selective and sensitive detection of NO in vitro and in vivo is of high importance but remains challenging. Previous fluorescent probes for NO detection either are of poor water solubility or lack selectivity over intracellular biomolecules. Herein, we rationally designed a water-soluble, biocompatible, small molecular probe o-phenylenediamine-Phe-Phe-OH (1) for the highly selective and sensitive detection of NO in vitro and in living cells. 1 can react with NO and turn on the fluorescence emission at 367 nm via an ICT mechanism. In vitro tests indicated that 1 showed high selectivity for NO detection without interference from common anions, ROS/RNS, and intracellular biomolecules DHA, AA, or MGO. In PBS buffer, 1 was applied for detecting NO within the range of 0-12 μM with a LOD of 6 nM. Moreover, 1 was successfully applied to sense intracellularly generated NO in living cells. We anticipate that 1 could be potentially employed for studying the toxicity and bioactivity of NO in the near future.

Self-assembly/disassembly is ubiquitous in nature and plays an important role in many biological events. But noninvasive characterization of this process in real time at molecular resolution remains challenging. Herein, using homebuilt liquid-phase scanning tunneling microscopy (L-STM) with ultrahigh stability, we directly visualized enzymatic self-assembly/disassembly of oligopeptide nanofibers in real time for the first time. Static high-resolution L-STM images clearly showed the molecular packing details in the supramolecular nanofiber and the diameter of the nanofiber was consistent with that of cryo transmission electron microscopy (cryo-TEM) observations. Moreover, the self-repairing behavior of the supramolecular nanofibers was also directly observed at high resolution for the first time. This work unprecedentedly revealed new insights into Nature-mimic self-assembly and disassembly at the molecular level. It also illustrates the potential of our homebuilt L-STM in studying delicate biological processes in physiological solution with high resolution.

Transplant rejection is the key problem in organ transplantation and, in clinic, immunosuppressive agents such as tacrolimus are directly administered to the recipients after surgery for T-cell inhibition. However, direct administration of tacrolimus may bring severe side effects to the recipients. Herein, by rational design of two hydrogelators NapPhePheGluTyrOH (1) and Nap d-Phe dPheGluTyrOH (2), a facile method of immune responsive release of tacrolimus is developed from their hydrogels to overcome organ transplantation rejection. Upon incubation with protein tyrosine kinase, which is activated in T cells after organ transplantation, the tacrolimus-encapsulating Gel 1 or Gel 2 is disassembled to release tacrolimus. Cell experiments show that both Gel 1 and Gel 2 have better inhibition effect on the activated T cells than free drug tacrolimus. Liver transplantation experiments indicate that, after 7 days of treatment of same dose tacrolimus, the recipient rats in the Gel 2 group show significantly extended median survival time of 22 days while the recipients treated with conventional tacrolimus medication have a median survival time of 13 days. It is expected herein that this "smart" facile method of immune responsive release of tacrolimus can be applied to overcome organ transplantation rejection in clinic in the near future.

Tissue specific T

A concise and efficient approach was established for the divergent total synthesis of (±)-tanshindiol B and C and tanshinone I from a ubiquitous ene intermediate in 1-2 steps. This critical intermediate was derived from (±)-tanshinol B, which was synthesized in 50% overall yield over 3 steps using an ultrasound-promoted cycloaddition as a key step. Compared to a previously reported strategy, our approach is more step-economic, thus greatly improving the synthetic efficiency. The bioactivity evaluation indicates that the diol stereochemistry of the tanshidiols has an impact on the EZH2 inhibitory activity.

Nanoprobes are advantageous over small molecular probes in sensitivity but most luminescence molecules used to construct nanoprobes often suffer from an aggregation-caused quenching effect. Herein, we rationally designed a small molecular probe Cys(StBu)-Lys(Ru(bpy)32+)-CBT (1) which "smartly" self-assembled into nanoparticles 1-NPs inside cells with non-quenched, persistent phosphorescence. Employing this property, we successfully applied 1 for long-term sensing of biothiol activity in living HepG2 cells and tumors. We envision that, by modifying the amino group with an enzyme substrate, our probe 1 could be further developed for sensing intracellular enzyme activity with non-quenched, persistent phosphorescence.

The dysregulation of NLRP3 inflammasome can cause uncontrolled inflammation and drive the development of a wide variety of human diseases, but the medications targeting NLRP3 inflammasome are not available in clinic. Here, we show that tranilast (TR), an old anti‐allergic clinical drug, is a direct NLRP3 inhibitor. TR inhibits NLRP3 inflammasome activation in macrophages, but has no effects on AIM2 or NLRC4 inflammasome activation. Mechanismly, TR directly binds to the NACHT domain of NLRP3 and suppresses the assembly of NLRP3 inflammasome by blocking NLRP3 oligomerization. In vivo experiments show that TR has remarkable preventive or therapeutic effects on the mouse models of NLRP3 inflammasome‐related human diseases, including gouty arthritis, cryopyrin‐associated autoinflammatory syndromes, and type 2 diabetes. Furthermore, TR is active ex vivo for synovial fluid mononuclear cells from patients with gout. Thus, our study identifies the old drug TR as a direct NLRP3 inhibitor and provides a potentially practical pharmacological approach for treating NLRP3‐driven diseases.

Development of sensitive fluorescence "Turn-On" strategies for imaging enzyme activity in living cells is of disease-diagnostic importance but remains challenging. Herein, by employing a click condensation reaction and rational design of a single quenched probe Cys(StBu)-Lys(Gly-Lys(DABCYL)-Gly-Gly-Arg-Arg-Val-Arg-Gly-FITC)-CBT (1), we developed a "smart" dual quenching strategy and applied it to detect intracellular furin activity with enhanced sensitivity. At physiological conditions, 1 was subjected to reduction-controlled condensation reaction to form 1-NPs and its fluorescence intensity further dropped to 1/2.8 of its original. Upon furin cleavage in vitro, the dual quenched 1-NPs had fluorescence "Turn-On" contrast 11-fold more than that of single quenched control probe FITC-Gly-Arg-Val-Arg-Arg-Gly-Gly-Lys(DABCYL)-Gly-OH (1-P). Live cell imaging results indicated that 1 showed fluorescence "Turn-On" contrast 6.3-fold of that of 1-P for sensing intracellular furin activity. We envision that, by replacing the RVRR substrate with other enzyme-cleavable ones, our versatile "smart" dual quenching strategy could be easily adjusted for the detection (or imaging) of other intracellular enzymes' activity with enhanced sensitivity.

Most nanodrugs are preprepared by encapsulating or loading the drugs with nanocarriers (e.g., dendrimers, liposomes, micelles, and polymeric nanoparticles). However, besides the low bioavailability and fast excretion of the nanodrugs in vivo, nanocarriers often exhibit in vitro and in vivo cytotoxicity, oxidative stress, and inflammation. Self-assembly is a ubiquitous process in biology where it plays important roles and underlies the formation of a wide variety of complex biological structures. Inspired by some cellular nanostructures (e.g., actin filaments, microtubules, vesicles, and micelles) in biological systems which are formed via molecular self-assembly, in recent decades, scientists have utilized self-assembly of oligomeric peptide under specific physiological or pathological environments to in situ construct nanodrugs for lesion-targeted therapies. On one hand, peptide-based nanodrugs always have some excellent intrinsic chemical (specificity, intrinsic bioactivity, biodegradability) and physical (small size, conformation) properties. On the other hand, stimuli-regulated intracellular self-assembly of nanodrugs is quite an efficient way to accumulate the drugs in lesion location and can realize an in situ slow release of the drugs. In this review article, we provided an overview on recent design principles for intracellular peptide self-assembly and illustrate how these principles have been applied for the in situ preparation of nanodrugs at the lesion location. In the last part, we list some challenges underlying this strategy and their possible solutions. Moreover, we envision the future possible theranostic applications of this strategy.

Dexamethasone (Dex) is one of the essential medicines used to treat inflammation diseases but an overdose of Dex leads to severe adverse effects. The development of a new strategy to boost the anti-inflammation efficacy of Dex is, therefore, important but remains challenging. Herein, by employing an enzyme-instructed self-assembly system, we developed an intracellular coassembly strategy to boost the anti-inflammation efficacy of Dex. Under the catalysis of alkaline phosphatase (ALP), the hydrogelator precursor Nap-Phe-Phe-Tyr(H2PO3)-OH (1p) self-assembled to form Gel 1 but dexamethasone sodium phosphate (Dp) only yielded Dex precipitates. However, subjecting equivalent amounts of 1p and Dp together to ALP-triggered coassembly was found to result in the formation of Gel 2. Cell experiments indicated that intracellular ALP-triggered coassembly of Dp with 1p extensively boosted the anti-inflammation efficacy of Dex on two types inflammatory cell models. We envision that, in the near future, our strategy of intracellular coassembly could be widely employed to boost the therapeutic effects of more drugs, while in the meantime used to alleviate the undesired adverse effects of these drugs.

Multidrug resistance (MDR) remains the biggest challenge in treating cancers. Herein we propose the intracellular self-assembly of nanodrugs as a new strategy for overcoming MDR. By employing a biocompatible condensation reaction, we rationally designed a taxol derivative Ac-Arg-Val-Arg-Arg-Cys(StBu)-Lys(taxol)-2-cyanobenzothiazole (CBT-Taxol) which could be subjected to furin-controlled condensation and self-assembly of taxol nanoparticles (Taxol-NPs). In vitro and in vivo studies indicated that, compared with taxol, CBT-Taxol showed a 4.5-fold or 1.5-fold increase in anti-MDR effects, respectively, on taxol-resistant HCT 116 cancer cells or tumors without being toxic to the cells or the mice. Our results demonstrate that structuring protease-susceptible agents and assembling them intracellularly into nanodrugs could be a new optimal strategy for overcoming MDR.

Alkaline phosphatase (ALP)-catalyzed hydrogelation has been extensively explored and found wide applications. Spectroscopic and electrochemical approaches are commonly employed for the detection of ALP activity. Herein, by rational design of a fluorescence probe Fmoc-K(FITC)FFYp (P1) (where FITC is fluorescein), we incorporated sol-gel transition with fluorescence "turn-off" and developed a new method for quantitative sensing ALP activity in vitro and in living cells. Under the catalysis of ALP, P1 was converted to hydrogelator Fmoc-K(FITC)FFY (1) which self-assembles into nanofibers to form Gel I. Accompanying this sol-gel transition, the fluorescence emission of P1 was turned off. Our assay was employed to detect ALP activity over the range of 0-2.8 U/mL with a limit of detection (LOD) of 0.06 U/mL. ALP-inhibitor-treated cell imaging indicated that P1 could be applied for sensing ALP activity in living cells. Our method provides a new option for real time and quantitative sensing ALP activity in vitro and even in living cells.

There has been no report on enzyme-controlled disassembly of self-quenched NIR fluorescent nanoparticles turning fluorescence on for specific detection/imaging of the enzyme's activity in vitro and in vivo. Herein, we reported the rational design of new NIR probe 1 whose fluorescence signal was self-quenched upon reduction-controlled condensation and subsequent assembly of its nanoparticles (i.e., 1-NPs). Then disassembly of 1-NPs by furin turned the fluorescence on. Employing this enzymatic strategy, we successfully applied 1-NPs for NIR detection of furin in vitro and NIR imaging furin activity in living cells. Moreover, we also applied 1-NPs for discriminative NIR imaging of MDA-MB-468 tumors in nude mice. This NIR probe 1 might be further developed for tumor-targeted imaging in routine preclinical studies or even in patients in the future.

Even though great advances have been achieved in the synthesis of luminescent metal nanoclusters, it is still challenging to develop metal nanoclusters with high quantum efficiency as well as multiple sensing functionalities. Here, we demonstrate the rapid preparation of glutathione-capped Au/Ag nanoclusters (GS-Au/Ag NCs) using microwave irradiation and their unique sensing capacities. Compared to bare GS-Au NCs, the doped Au/Ag NCs possess an enhanced quantum yield (7.8% compared to 2.2% for GS-Au NCs). Several characterization techniques were used to elucidate the atomic composition, particulate character, and electronic structure of the fabricated NCs. According to the X-ray photoelectron spectroscopy (XPS) and X-ray absorption near-edge structure (XANES) spectra, a significant amount of Au exists in the oxidized state as Au(I), and the Ag atoms are positively charged. In contrast to those nanoclusters that detect only one analyte, the GS-Au/Ag NCs can be used as a versatile sensor for metal ions, anions, and small molecules. In this manner, the NCs can be regarded as a unique sensor-on-a-nanoparticle.

(19)F MRI has higher selectivity but lower sensitivity than (1)H MRI for in vivo diagnosis. Therefore, to avoid using a high injection dose of the (19)F probe while, in the meantime, maintaining the high sensitivity of (19)F MRI has remained challenging. Local self-assembly and disassembly of (19)F nanoparticles could be one of the "smart" strategies to achieve this goal. Herein, we report a dual-functional probe 1 for glutathione (GSH)-controlled self-assembly and subsequent legumain (Lgmn)-controlled disassembly of its nanoparticles (i.e., 1-NPs). Self-assembly and disassembly of 1-NPs confer (19)F magnetic resonance (MR) signals "off" and "on", respectively. Employing this strategy, we successfully applied 1 for consecutive detections of GSH and Lgmn in vitro and in cells, imaging Lgmn activity in HEK 293T tumors in zebrafish at a low dosage under 14.1 T.

Polymer hydrogels and small-molecule-based (SMB) supramolecular hydrogels have been widely explored. But oligomeric hydrogels have remained a challenge because synthetic difficulties of the oligomers and control of their amphiphilicities. Reported herein is the rational design of two precursors Cys(SEt)-Lys-CBT (1) and (Cys-Lys-CBT)2 (2) (CBT=2-cyano-6-aminobenzothiazole) and the use of a biocompatible condensation to prepare oligomeric hydrogels. Glutathione reduction of 1 or 2 yields the same gelator Cys-Lys-CBT (3) which condenses with each other to yield amphiphilic cyclic oligomers. The oligomers instantly self-assemble into nanofibers and form oligomeric hydrogels with similar mechanic properties. Chemical analyses indicated that the major condensation product in both two hydrogels is a cyclic dimer. Considering its biocompatibility, optimal mechanical strength, and biodegradability, we believe that our oligomeric hydrogel might be useful for long-term drug delivery in the future.

Two bio-orthogonal click reactions were employed to bridge prokaryotic E. coli or eukaryotic HEK 293T cells of three colors successfully., Cell–cell interactions play a crucial role in the development and function of multicellular organisms. To study cell–cell interactions in vitro, it is a big challenge for researchers to artificially build up cell junctions to bridge different types of cells for this purpose. Herein, by employing two orthogonal click reactions, we rationally designed four click reagents Mal-CBT, Mal-Cys, Mal-Alkyne, and Mal-N3 and successfully applied them to bridge cells of three colors. Orthogonality between these two click reactions was validated in solution and characterized with HPLC and ESI-MS analyses. After modifications of fluorescent protein-expressing prokaryotic Escherichia coli (E. coli) cells (or eukaryotic HEK 293T cells) of three colors with the reagents Mal-Cys, Mal-CBT and Mal-Alkyne, or Mal-N3, the cells were sequentially bridged. The HEK 293T cells showed a higher efficiency of cell bridging than the E. coli cells. Finally, using optical tweezers, we quantitatively measured the bridging probability between Mal-Cys-modified and Mal-CBT-modified HEK 293 cells, as well as the rupture force between two bridged cells. We found that the CBT–Cys click reaction markedly improved the efficiency of cell bridging and the rupture force between two bridged cells was measured to be 153.8 pN at a force-loading rate of 49 pN s–1. Our results demonstrate that it is possible to use two (or n) orthogonal click reactions to bridge three (or n + 1) types of cells. Taking the biological importance of cell junctions into consideration, we anticipate that our method of bridging three types of cells with two bio-orthogonal click reactions will be a useful tool for biologists to study cell–cell interactions with more convenience and efficiency.

Compared to (1)H MRI, (19)F MRI provides higher selectivity but lower sensitivity. Therefore, the need to inject high doses of the (19)F probe to improve its sensitivity for in vivo diagnosis remains a challenge. A "smart" strategy is needed that could locally concentrate a low-dose (19)F probe while avoiding the fast transverse relaxation of the probes. Locally self-assembling and disassembling (19)F nanoparticles may be an optimal measure to achieve this goal. Herein, we report a dual-functional probe 1 for glutathione (GSH)-controlled self-assembly and subsequent caspase 3/7 (Casp3/7)-controlled disassembly of formed nanoparticles (i.e., 1-NPs). Consecutive assembly and disassembly of 1-NPs translate to "off" and "on" (19)F magnetic resonance (MR) signal states, respectively. Employing this smart strategy, we successfully used 1 for the consecutive detection of GSH and Casp3/7 activity in vitro and in cells and imaging Casp3/7 activity in cells and in zebrafish at low doses with a 14.1 T magnetic field.

The design of tumor-targeting, intracellular protease-activatable near-infrared fluorescence (NIRF) nanoprobes is broadly interesting but remains challenging. In this work, we report the rational design of a NIR probe Cys(StBu)-Lys(Biotin)-Lys-Lys(Cy5.5)-CBT (1) to facilely prepare the self-quenched nanoparticles 1-NPs for tumor-targeted imaging in vitro and in vivo. The biotinylated 1-NPs could be actively uptaken by biotin receptor-overexpressing tumor cells via receptor-mediated endocytosis. Upon intracellular proteolytic cleavage, 1-NPs were disassembled to yield the small molecular probe Lys(Cy5.5)-Luciferin-Lys(Biotin)-Lys-OH (1-D-cleaved), accompanied by fluorescence “Turn-On”. With this NIRF “Turn-On” property, 1-NPs were successfully applied for tumor-targeted imaging. We envision that our nanoparticles could be applied for fluorescence-guided tumor surgery in the near future.

γ-Glutamyltranspeptidase (GGT) is an important tumor biomarker but using a bioluminescence (BL) probe to real time monitor its activity has not been reported. Herein, we rationally designed two GGT-cleavable BL probes Glu-AmLH2 (1) and Glu-p-aminobenzyloxycarbonyl-AmLH2 (2), and successfully applied them for sensing GGT activity with high sensitivity and excellent selectivity both in vitro and in vivo. The results indicated that, although 2 had lower background BL signal than 1, GGT had higher catalytic efficiency for 1 than 2, and 1 was superior to 2 for sensing GGT activity in living cells and tumors. We envision that our probe 1 could be widely applied for the diagnosis of important GGT-related diseases in animal models in the near future.

Chemiluminescence (CL) has a higher signal-to-noise ratio than fluorescence, but the use of CL to track an enzyme-instructed self-assembly (EISA) process has not been reported. In this work, by coincubation of the hydrogelator precursor Fmoc-Phe-Phe-Tyr(H2PO3)-OH (1P) and the CL agent AMPPD (2) with alkaline phosphatase (ALP), we employed CL to directly characterize and image the simultaneous EISA process of 1P. Hydrogelation processes of 1P with and without 2 and the CL properties of 2 with and without 1P under ALP catalysis were systematically studied. The results indicated that 2 is an ideal CL indicator for ALP-triggered hydrogelation of 1P. Using an IVIS optical imaging system, we obtained time-course CL images of 2 to track the simultaneous hydrogelation process of 1P in the same solution. We envision that our CL method could be employed to track more biological EISA events in the near future.

Herein, we report, for the first time, the use of MRI methods to study enzymatic hydrogelation. Supramolecular hydrogels have been exploited as biomaterials for many applications. However, behaviors of the water molecules encapsulated in hydrogels have not been fully understood. In this work, we designed a precursor 1 which could self-assemble into nanofibers and form hydrogel I (gel I) upon the catalysis of phosphatase. The differences of mechanic property, pore size, water diffusion rate, and magnetic resonance relaxation times T1 and T2 of gel I containing different concentrations of 1 were systematically studied and analyzed. T1, T2, and diffusion-weighted (1)H MR images from gel I phantoms were obtained at 9.4 T. Analyses of the MRI data uncovered how the density of the nanofiber networks affects the relaxation behaviors of the water protons encapsulated in such hydrogels. Rheological analyses and cryo-TEM observations showed increased gel elasticities with increased concentrations of 1 while the pore sizes of gel I decreased. This also resulted in an increase in the proton relaxation rate (i.e., shortened T1, T2, and apparent diffusion coefficient (ADC)) for the water encapsulated in the hydrogel. With MRI, our study provides a new in vitro method to potentially mimic and study in vivo diseases that involve fibrous aggregates.

Although much effort has been made for studies on aptamer-target interactions due to promising applications of aptamers in biomedical and analytical fields, measurement of the aptamer-target binding constant and binding site still remains challenging. Herein, we report a sensitive label-free chemiluminescence (CL) strategy to determine the target concentration and, more importantly, to measure the target-aptamer binding constant and binding site. This approach is suitable for multiple types of targets, including small molecules, peptides, and proteins that can enhance the CL initiated by N-(aminobutyl)-N-ethylisoluminol functionalized gold colloids, making the present method a general platform to investigate aptamer-target interactions. This approach can achieve extremely high sensitivity with nanogram samples for measuring the target-aptamer binding constant. And the measurement could be rapidly performed using a simple and low-cost CL system. It provides an effective tool for studying the binding of biologically important molecules to nucleic acids and the selection of aptamers. Besides, we have also discovered that the 14-mer aptamer fragment itself split from the ATP-binding aptamer could selectively capture ATP. The binding constant, site, and conformation between ATP and the 14-mer aptamer fragment were obtained using such a novel CL strategy and molecular dynamic simulation.

Herein, we report the development of a new fluorescent switch for sequential and selective sensing of Cu(2+) and L-histidine (L-His) in vitro and in living cells for the first time. In the absence of metal ions, Ac-SAACQ-Gly-Gly-Gly-Lys (FITC) (1) exhibits comparable fluorescence to that of free FITC. In the presence of metal ions, 1 selectively coordinates to Cu(2+) , causing its fluorescence emission to be quenched via photoinduced electron transfer. Interestingly, the as-formed 1–Cu(2+) complex selectively responds to L-His among the 20 natural amino acids by turning its fluorescence on. This property of fluorescence switch of 1 was successfully applied for qualitatively and quantitatively sensing Cu(2+) and L-His in vitro. Using this dual functional probe, we also sequentially imaged Cu(2+) and L-His in living HepG2 cells. Our new probe 1 could be applied for not only environmental monitoring but also biomolecule detection in the near future.

The aggregation-induced emission (AIE) effect has recently been widely applied for biomarker sensing. But developing "smart" strategies to effectively aggregate the AIE fluorogen and additionally enhance the fluorescence emission remain challenging. In this work, by integrating a biocompatible condensation reaction with an AIE fluorogen, we rationally designed a "smart" dual AIE probe Ac-Arg-Val-Arg-Arg-Cys(StBu)-Lys(TPE)-CBT (1) for enhanced fluorescence sensing furin activity in vitro and in living cells. Compared with the single AIE probe Ac-Arg-Val-Arg-Arg-Lys(TPE)-OH (1-Ctrl) which also subjects to furin cleavage, fluorescence emissions of 1 were additionally enhanced 1.7 fold and 3.4 fold in vitro and in living cells, respectively. We envision that, in the near future, our "smart" strategy of enzyme-instructed dual AIE could be widely applied for sensing (or imaging) enzyme activity in vitro and even in vivo with dramatically enhanced sensitivity.

Spontaneously precise organization of small structures into complex superstructures is ubiquitous and important in nature. But using small building blocks to mimic this process remains a challenge to scientists. Herein, we report the rational design of a bipyridine-derivative and applied it for the self-assembly of nanorings. The addition of Fe(2+) to the nanorings resulted in the assembly of the nanorings into supernanostructures via Fe(2+)-bipyridine coordination. HPLC, HR-ESI/MS, UV-vis, DLS, and TEM analyses clearly validated the intramolecular cyclization, self-assembly of the nanorings, and the additional self-assembly of the superstructures via Fe(2+)-bipyridine coordination. We envision our strategy to be a new approach of precisely assembling nanostructures of ring shape into more complex superstructures.

Herein, we report a new "On-On" strategy based on the assembly and disassembly of fluorescein isothiocyanate nanoparticles (FITC-NPs) for sequential detections of glutathione (GSH) and caspase-3 (Casp3) with a multifunctional fluorescent probe 1. Theoretical investigations revealed the underlying mechanism that satisfactorily explained experimental results of such consecutive enhancements of fluorescence. Using this probe, we also successfully imaged the Casp3 activity in apoptotic cells.

Employing cellular environment for the self-assembly of supramolecular nanofibers for biological applications has been widely explored. But using one precursor to differentiate the extra- and intracellular environments to self-assemble into two different nanofibers remains challenging. With the knowledge that the extracellualr environment of some cancer cells contains large amounts of alkaline phosphatase (ALP) while their intracellular environment is glutathione (GSH)-abundant in mind, we rationally designed a precursor Cys(SEt)-Glu-Tyr(H2PO3)-Phe-Phe-Gly-CBT (1) that can efficiently yield amphiphilic 2 and 2-D to self-assemble into two different nanofibers in hydrogels under the sequential treatment of ALP and GSH. We envision that, by employing a click condensation reaction, this work offers a platform for facilely postmodulation of supramolecular nanofibers, and the versatile precursor 1 could be used to kill two birds with one stone.

T 1–T2 dual modal magnetic resonance imaging (MRI) has attracted considerable interest because it offers complementary diagnostic information, leading to more precise diagnosis. To date, a number of nanostructures have been reported as T1–T2 dual modal MR contrast agents (CAs). However, hybrids of nanocubes with both iron and gadolinium (Gd) elements as T1–T2 dual modal CAs have not been reported. Herein, we report the synthesis of novel core/shell Fe3O4/Gd2O3 nanocubes as T1–T2 dual-modal CAs and their application for enhanced T1–T2 MR imaging of rat livers. A relaxivity study at 1.5 T indicated that our Fe3O4/Gd2O3 nanocubes have an r1 value of 45.24 mM−1 s−1 and an r2 value of 186.51 mM−1 s−1, which were about two folds of those of Gd2O3 nanoparticles and Fe3O4 nanocubes, respectively. In vivo MR imaging of rats showed both T1-positive and T2-negative contrast enhancements in the livers. We envision that our Fe3O4/Gd2O3 nanocubes could be applied as T1–T2 dual modal MR CAs for a wide range of theranostic applications in the near future.

Tyrosine kinase and phosphatase are two important, antagonistic enzymes in organisms. Development of noninvasive approach for sensing their activity with high spatial and temporal resolution remains challenging. Herein, we rationally designed a hydrogelator Nap-Phe-Phe(CF3)-Glu-Tyr-Ile-OH (1a) whose supramolecular hydrogel (i.e., Gel 1a) can be subjected to tyrosine kinase-directed disassembly, and its phosphate precursor Nap-Phe-Phe(CF3)-Glu-Tyr(H2PO3)-Ile-OH (1b), which can be subjected to alkaline phosphatase (ALP)-instructed self-assembly to form supramolecular hydrogel Gel 1b, respectively. Mechanic properties and internal fibrous networks of the hydrogels were characterized with rheology and cryo transmission electron microscopy (cryo-TEM). Disassembly/self-assembly of their corresponding supramolecular hydrogels conferring respective "On/Off" (19)F NMR/MRI signals were employed to sense the activity of these two important enzymes in vitro and in cell lysates for the first time. We anticipate that our new (19)F NMR/magnetic resonance imaging (MRI) method would facilitate pharmaceutical researchers to screen new inhibitors for these two enzymes without steric hindrance.

The development and function of multicellular organisms deeply rely on cell-cell interactions. Therefore, the ability of artificially build up cell junctions and bridging different types of cells in vitro is badly demanded for biological research. Herein, four click reagents Mal-CBT, Mal-Cys, Mal-Alkyne, and Mal-N3 are firstly introduced to bridge cells of three colors. Then, the bridging probability between Mal-Cys-modified and Mal-CBT-modified HEK 293 cells are quantitatively measured by optical tweezers as well. The experiment clearly showed that the efficiency of cell bridging was dramatically improved from 19.15% to 66.10% by CBT-Cys click reaction. After that, image analysis techniques were employed via Matlab in order to tracking image sequences obtained during stretching process. By these means, the force-time curve was constructed and rupture force of bridged cell pairs was obtained to be 153.8 pN at a force-loading rate of 49 pN s-1. This rupture force was of the same order of magnitude as the previously reported simulated result for a covalent bond[1]. The same method can also be applied in rupture force measurement of other covalent bonds at low force-loading rate.[1] S.W. Schmidt, M.K. Beyer, H. Clausen-Schaumann, Dynamic strength of the silicon-carbon bond observed over three decades of force-loading rates, J. Am. Chem. Soc., 2008, 130, 3664-3668.

Click chemistry is a highly-selective and high-yield chemical approach which uses a wide scope of materials and does not call for purification. Molecular imaging is an in vivo characterization and measurement of biologic processes at the cellular and molecular level. Representative imaging modalities of molecular imaging are optical imaging, nuclear imaging, and magnetic resonance imaging (MRI). Up to date, click chemistry has been widely employed by molecular imaging to trap imaging probes on site and trace the biological events with high efficiency and high sensitivity. It has played an important role in early detection of disease, real-time monitoring of drug delivery and investigating the func- tions of biomolecules in vivo. Herein, we introduce the concept of click chemistry and illustrate the design of probes for molecular imaging based on the strategies of click chemistry.

Self-assembly of small molecules or macromolecules through non-covalent or covalent bonds to build up supramolecular nanostructures is a prevalent and important process in nature. While most chemists use small molecules to assemble nanostructures with physical or chemical perturbations, nature adopts enzymes to catalyze the reaction to assemble biological, functional nanostructures with high efficiency and specificity. Although enzymatic self-assembly of nanostructures has been remained challenging for chemists, there are still a few examples of using important enzymes to initiate the self-assembly of nanostructures for diagnosis or therapy of certain diseases because down-regulation or overexpression of certain enzymes always associates with abnormalities of tissues/organs or diseases in living body. Herein, we introduce the concept of enzymatic self-assembly and illustrate the design and application of enzyme-catalyzed or -regulated formation of nanostructures for theranostics.

Self-assembly of small molecules or macromolecules through non-covalent or covalent bonds to build up supramolecular nanostructures is a prevalent and important process in nature. While most chemists use small molecules to assemble nanostructures with physical or chemical perturbations, nature adopts enzymes to catalyze the reaction to assemble biological, functional nanostructures with high efficiency and specificity. Although enzymatic self-assembly of nanostructures has been remained challenging for chemists, there are still a few examples of using important enzymes to initiate the self-assembly of nanostructures for diagnosis or therapy of certain diseases because down-regulation or overexpression of certain enzymes always associates with abnormalities of tissues/organs or diseases in living body. Herein, we introduce the concept of enzymatic self-assembly and illustrate the design and application of enzyme-catalyzed or -regulated formation of nanostructures for theranostics.

Methylglyoxal (MGO) is a toxic, dicarbonyl metabolite in all living cells and its detoxification is regulated by glyoxalase I (GLOI). Herein, we rationally designed a precursor o-phenylenediamine-Phe-Phe-OH (1) which “click” reacts with MGO to yield amphiphilic methylquinoxaline-Phe-Phe-OH (2) to self-assemble into supramolecular hydrogel II (Gel II). Cryo-TEM images of Gel II suggested that there existed two orders of self-assembly to form the 32.8 nm width-nanotubes in the hydrogel. The hypothesis was validated with the analyses of the fluorescence, transmittance, and circular dichroism data of the serial dilutions of Gel II. Interference tests indicated that hydrogelation of 1 with MGO would not be affected by nitric oxide (NO). Our results suggest that 1 could be applied for specific hydrogelation with MGO, and potentially the removal of MGO in vitro.

Cancer cell-targeted imaging and drug delivery remain a challenge for precise cancer theranostics. MUC1 is a large transmembrane glycoprotein that may potentially serve as a target for cancer theranostics. Herein, using a MUC1-targeting aptamer (APT) as the “warhead”, we rationally designed and constructed a hybrid nanoparticle 1-NPs-QDs-hAPT (Vehicle) that could be applied for MUC1-targeted cell uptake and imaging. By intercalating different Vehicle amounts with the anticancer drug doxorubicin (DOX), we obtained the multifunctional bioconjugate Vehicle-DOX with a maximized drug payload and DOX fluorescence quenching capability. Confocal microscopy cell imaging indicated that Vehicle-DOX could be used to track MUC1-targeted drug release. A cytotoxicity study indicated that Vehicle-DOX could be applied for MUC1-targeted cytotoxicity. We anticipate that our multifunctional bioconjugate Vehicle-DOX could be applied for in vivo tumor-targeted theranostics.