Three-in-One Peptide Prodrug with Targeting, Assembly and Release Properties for Overcoming Bacterium-Induced Drug Resistance and Potentiating Anti-Cancer Immune Response.

The presence of bacteria in tumor results in chemotherapeutic drug resistance and weakens the immune response in colorectal cancer. To overcome bacterium-induced chemotherapeutic drug resistance and potentiate antitumor immunity, herein a novel molecule Biotin-Lys(SA-Cip-OH)-Lys(SA-CPT)-Phe-Phe-Nap (Biotin-Cip-CPT-Nap) is rationally designed containing four functional motifs (i.e., a biotin motif for targeting, Phe-Phe(-Nap) motif for self-assembly, ciprofloxacin derivative (Cip-OH) motif for antibacterial effect, and camptothecin (CPT) motif for chemotherapy). Using the designed molecule, a novel strategy of intracellular enzymatic nanofiber formation and synergistic antibacterium-enhanced chemotherapy and immunotherapy is achieved. Under endocytosis mediated by highly expressed biotin receptor in colorectal cancer cell membrane and the catalysis of highly expressed carboxylesterase in the cytoplasm, this novel molecule can be transformed into Biotin-Nap, which self-assembled into nanofibers. Meanwhile, antibiotic Cip-OH and chemotherapeutic drug CPT are released, overcoming bacterium-induced drug resistance and enhancing the therapeutic efficacy of immunotherapy towards colorectal cancer. This work offers a feasible strategy for the design of novel multifunctional prodrugs to improve the efficiency of colorectal cancer treatment.

In Situ Nanofiber Formation Blocks AXL and GAS6 Binding to Suppress Ovarian Cancer Development.

Anexelekto (AXL) is an attractive molecular target for ovarian cancer therapy because of its important role in ovarian cancer initiation and progression. To date, several AXL inhibitors have entered clinical trials for the treatment of ovarian cancer. However, the disadvantages of low AXL affinity and severe off-target toxicity of these inhibitors limit their further clinical applications. Herein, by rational design of a nonapeptide derivative Nap-Phe-Phe-Glu-Ile-Arg-Leu-Arg-Phe-Lys (Nap-IR), a strategy of in situ nanofiber formation is proposed to suppress ovarian cancer growth. After administration, Nap-IR specifically targets overexpressed AXL on ovarian cancer cell membranes and undergoes a receptor-instructed nanoparticle-to-nanofiber transition. In vivo and in vitro experiments demonstrate that in situ formed Nap-IR nanofibers efficiently induce apoptosis of ovarian cancer cells by blocking AXL activation and disrupting subsequent downstream signaling events. Remarkably, Nap-IR can synergistically enhance the anticancer effect of cisplatin against HO8910 ovarian tumors. It is anticipated that the Nap-IR can be applied in clinical ovarian cancer therapy in the near future.

ß-Galactosidase-activated near-infrared AIEgen for ovarian cancer imaging in vivo.

Near-infrared (NIR) aggregation induced-emission luminogens (AIEgens) circumvent the noisome aggregation-caused quenching (ACQ) effect in physiological milieu, thus holding high promise for real-time and sensitive imaging of biomarkers in vivo. ß-Galactosidase (ß-Gal) is a biomarker for primary ovarian carcinoma, but current AIEgens for ß-Gal sensing display emissions in the visible region and have not been applied in vivo. We herein propose an NIR AIEgen QM-TPA-Gal and applied it for imaging ß-Gal activity in vitro and in ovarian tumor model. After being internalized by ovarian cancer cells (e.g., SKOV3), the hydrophilic nonfluorescent QM-TPA-Gal undergoes hydrolyzation by ß-Gal to yield hydrophobic QM-TPA-OH, which subsequently aggregates into nanoparticles to turn NIR fluorescence "on" through the AIE mechanism. In vitro experimental results indicate that QM-TPA-Gal has a sensitive and selective response to ß-Gal with a limit of detection (LOD) of 0.21 U/mL. Molecular docking simulation confirms that QM-TPA-Gal has a good binding ability with ß-Gal to allow efficient hydrolysis. Furthermore, QM-TPA-Gal is successfully applied for ß-Gal imaging in SKOV3 cell and SKOV3-bearing living mouse models. It is anticipated that QM-TPA-Gal could be applied for early diagnosis of ovarian cancers or other ß-Gal-associated diseases in near future.

In situ peptide assemblies for bacterial infection imaging and treatment.

Bacterial infections, especially antibiotic-resistant ones, remain a major threat to human health. Advances in nanotechnology have led to the development of numerous antimicrobial nanomaterials. Among them, in situ peptide assemblies, formed by biomarker-triggered self-assembly of peptide-based building blocks, have received increasing attention due to their unique merits of good spatiotemporal controllability and excellent disease accumulation and retention. In recent years, a variety of "turn on" imaging probes and activatable antibacterial agents based on in situ peptide assemblies have been developed, providing promising alternatives for the treatment and diagnosis of bacterial infections. In this review, we introduce representative design strategies for in situ peptide assemblies and highlight the bacterial infection imaging and treatment applications of these supramolecular materials. Besides, current challenges in this field are proposed.

Dual Aggregations of a Near-Infrared Aggregation-Induced Emission Luminogen for Enhanced Imaging of Alzheimer's Disease.

Aggregation-induced emission (AIE) enables "Turn-On" imaging generally through single aggregation of the AIE luminogen (AIEgen). Dual aggregrations of the AIEgen might further enhance the imaging intensity and the consequent sensitivity. Herein, we rationally designed a near-infrared (NIR) AIEgen Ac-Trp-Glu-His-Asp-Cys(StBu)-Pra(QMT)-CBT (QMT-CBT) which, upon caspase1 (Cas1) activation, underwent a CBT-Cys click reaction to form cyclic dimers QMT-Dimer (the first aggregation) and assembled into nanoparticles (the second aggregation), turning the AIE signal "on" for enhanced imaging of Alzheimer's disease (AD). Molecular dynamics simulations validated that the fluorogen QMT in QMT-NPs stacked much tighter with each other than in the single aggregates of the control compound Ac-Trp-Glu-His-Asp-Cys(tBu)-Pra(QMT)-CBT (QMT-CBT-Ctrl). Dual aggregations of QMT rendered 1.9-, 1.7-, and 1.4-fold enhanced fluorescence intensities of its single aggregation in vitro, in cells, and in a living AD mouse model, respectively. We anticipate this smart fluorogen to be used for sensitive diagnosis of AD in the clinic in the near future.

Intracellular Nitroreductase-Triggered "On" and "Enhanced" Photoacoustic Signals for Sensitive Imaging of Tumor Hypoxia.

Current molecular photoacoustic (PA) probes are designed with either stimulus-turned "on" or assembly-enhanced signals to trace biological analytes/events. PA probes based on the nature-derived click reaction between 2-cyano-6-aminobenzothiazole (CBT) and cysteine (Cys) (i.e., CBT-Cys click reaction) possess both "turn-on" and "enhanced" PA signals; and thus, should have higher sensitivity. Nevertheless, such PA probes, particularly those for sensitive imaging of tumor hypoxia, remain scarce. Herein, a PA probe NI-Cys(StBu)-Dap(IR780)-CBT (NI-C-CBT) is rationally designed, which after being internalized by hypoxic tumor cells, is cleaved by nitroreductase under the reduction condition to yield cyclic dimer C-CBT-Dimer to turn the PA signal "ON" and subsequently assembled into nanoparticles C-CBT-NPs with additionally enhanced PA signal ("Enhanced"). NI-C-CBT exhibits 1.7-fold "ON" and 3.2-fold overall "Enhanced" PA signals in vitro. Moreover, it provides 1.9-fold and 2.8-fold overall enhanced PA signals for tumor hypoxia imaging in HeLa cells and HeLa tumor-bearing mice, respectively. This strategy is expected to be widely applied to design more "smart" PA probes for sensitive imaging of important biological events in vivo in near future.

Assembly drives regioselective azide-alkyne cycloaddition reaction.

Azide-alkyne cycloaddition reaction is a very common organic reaction to synthesize nitrogen-containing heterocycles. Once catalyzed by Cu(I) or Ru(II), it turns out to be a click reaction and thus is widely applied in chemical biology for labeling. However, besides their poor regioselectivity towards this reaction, these metal ions are not biologically friendly. Hence, it is an urgent need to develop a metal-free azide-alkyne cycloaddition reaction for biomedical applications. In this work, we found that, in the absence of metal ions, supramolecular self-assembly in an aqueous solution could realize this reaction with excellent regioselectivity. Nap-Phe-Phe-Lys(azido)-OH firstly self-assembled into nanofibers. Then, Nap-Phe-Phe-Gly(alkynyl)-OH at equivalent concentration approached to react with the assembly to yield the cycloaddition product Nap-Phe-Phe-Lys(triazole)-Gly-Phe-Phe-Nap to form nanoribbons. Due to space confinement effect, the product was obtained with excellent regioselectivity. Employing the excellent properties of supramolecular self-assembly, we are applying this strategy to realize more reactions without metal ion catalysis.

Tandem Guest-Host-Receptor Recognitions Precisely Guide Ciprofloxacin to Eliminate Intracellular Staphylococcus aureus.

Staphylococcus aureus (S. aureus) is able to hide within host cells to escape immune clearance and antibiotic action, causing life-threatening infections. To boost the therapeutic efficacy of antibiotics, new intracellular delivery approaches are urgently needed. Herein, by rational design of an adamantane (Ada)-containing antibiotic-peptide precursor Ada-Gly-Tyr-Val-Ala-Asp-Cys(StBu)-Lys(Ciprofloxacin)-CBT (Cip-CBT-Ada), we propose a strategy of tandem guest-host-receptor recognitions to precisely guide ciprofloxacin to eliminate intracellular S. aureus. Via guest-host recognition, Cip-CBT-Ada is decorated with a ß-cyclodextrin-heptamannoside (CD-M) derivative to yield Cip-CBT-Ada/CD-M, which is able to target mannose receptor-overexpressing macrophages via multivalent ligand-receptor recognition. After uptake, Cip-CBT-Ada/CD-M undergoes caspase-1 (an overexpressed enzyme during S. aureus infection)-initiated CBT-Cys click reaction to self-assemble into ciprofloxacin nanoparticle Nano-Cip. In vitro and in vivo experiments demonstrate that, compared with ciprofloxacin or Cip-CBT-Ada, Cip-CBT-Ada/CD-M shows superior intracellular bacteria elimination and inflammation alleviation efficiency in S. aureus-infected RAW264.7 cells and mouse infection models, respectively. This work provides a supramolecular platform of tandem guest-host-receptor recognitions to precisely guide antibiotics to eliminate intracellular S. aureus infection efficiently.

Caspase-3-Triggered Intracellular Gadolinium Nanoparticle Formation for T1-Weighted Magnetic Resonance Imaging of Apoptosis In Vivo.

Apoptosis, with a hallmark of upregulated protease Caspase-3, has been frequently imaged with various probes to reveal the therapeutic efficiencies of different drugs. However, activatable molecular probes with programmable self-assembling behaviors that enable enhanced T1-weighted magnetic resonance imaging (MRI) of apoptosis remain scarce. Herein, taking advantage of a CBT-Cys click reaction, we rationally designed a Caspase-3-activatable self-assembling probe Ac-Asp-Glu-Val-Asp-Cys(StBu)-Lys(DOTA(Gd))-CBT (DEVDCS-Gd-CBT) for apoptosis imaging in vivo. After Caspase-3 cleavage in apoptotic cells, DEVDCS-Gd-CBT underwent CBT-Cys click reaction to form a cyclic dimer, which self-assembled into Gd nanoparticles. With this probe, enhanced T1-weighted MR images of apoptosis were achieved at low magnetic fields in vitro, in cis-dichlorodiamineplatinum-induced apoptotic cells and in tail-amputation-simulated apoptotic zebrafish. We anticipate that the smart probe DEVDCS-Gd-CBT could be applied for T1-weighted MRI of apoptosis-related diseases in the clinic in the future.

Apoptosis-Amplified Assembly of Porphyrin Nanofiber Enhances Photodynamic Therapy of Oral Tumor.

Oral squamous cell carcinoma (OSCC) is the most common oral cancer, having high recurrence and metastasis features. In addition to surgery, photodynamic therapy (PDT) is considered as another effective approach for OSCC treatment. The water solubility of currently available PDT photosensitizers (PSs) is poor, lowering their singlet oxygen (1O2) yield and consequent PDT efficiency. Strategies of PS assembly have been reported to increase 1O2 yield, but it is still possible to further enhance PDT efficiency. In this work, we utilized apoptosis to amplify the assembly of porphyrin nanofibers for enhanced PDT of OSCC. A water-soluble porphyrin derivative, Ac-Asp-Glu-Val-Asp-Asp-TPP (Ac-DEVDD-TPP), was designed for this purpose. Upon caspase-3 (Casp3, an activated enzyme during apoptosis) cleavage and laser irradiation, Ac-DEVDD-TPP was converted to D-TPP, which spontaneously self-assembled into porphyrin nanofibers, accompanied by 1.4-fold and 2.1-fold 1O2 generations in vitro and in cells, respectively. The as-formed porphyrin nanofiber induced efficient cell apoptosis and pyroptosis. In vivo experiments demonstrated that, compared with the scrambled control compound Ac-DEDVD-TPP, Ac-DEVDD-TPP led to 6.2-fold and 1.3-fold expressions of Casp3 in subcutaneous and orthotopic oral tumor models, respectively, and significantly suppressed the tumors. We envision that our strategy of apoptosis-amplified porphyrin assembly might be applied for OSCC treatment in the clinic in the near future.

Facile Label-Free Three-Input Molecular Keypad Lock.

Featured with molecule-level data encryption, molecular keypad locks show attractive merits in information security. Most of the previous multiple-input locks use fluorescence as output but are impeded by inefficient/labile prequenching or highly synthetic complexity/difficulty of the fluorophore-containing processor molecules. We herein propose a facile three-input molecular keypad lock, which is simple in synthesis and label free but capable of in situ generation of a fluorescent moiety (dityrosine) for background-free fluorescence readout. A nonfluorescent ("Locked") tyrosine derivate zYpc was easily synthesized as the processor. The correct "password" (i.e., UV → ALP → TYR, ABC) stepwise converted zYpc to a dityrosine-containing product, exhibiting a bright blue fluorescence output ("Open"). In contrast, wrongly permutated inputs failed to open this lock. This device shows potential to be extended as a more advanced keypad lock with better security.

Enzymatic Self-Assembly of Adamantane-Peptide Conjugate for Combating Staphylococcus aureus Infection.

Staphylococcus aureus (S. aureus) remains a leading cause of bacterial infections. However, eradication of S. aureus infections with common antibiotics is increasingly difficult due to outbreaks of drug resistance. Therefore, new antibiotic classes and antibacterial strategies are urgently in demand. Herein, it is shown that an adamantane-peptide conjugate, upon dephosphorylation by alkaline phosphatase (ALP) constitutively expressed on S. aureus, generates fibrous assemblies in situ to combat S. aureus infection. By attaching adamantane to a phosphorylated tetrapeptide Nap-Phe-Phe-Lys-Tyr(H2 PO3 )-OH, the rationally designed adamantane-peptide conjugate Nap-Phe-Phe-Lys(Ada)-Tyr(H2 PO3 )-OH (Nap-FYp-Ada) is obtained. Upon bacterial ALP activation, Nap-FYp-Ada is dephosphorylated and self-assembles into nanofibers on the surface of S. aureus. As revealed by cell assays, the assemblies of adamantane-peptide conjugates interact with cell lipid membrane and thereby disrupt membrane integrity to kill S. aureus. Animal experiments further demonstrate the excellent potential of Nap-FYp-Ada in the treatment of S. aureus infection in vivo. This work provides an alternative approach to design antimicrobial agents.

Granzyme B Turns Nanoparticle Fluorescence "On" for Imaging Cytotoxic T Lymphocyte Activity in Vivo.

Cytotoxic T lymphocytes (CTLs) are important immune cells, and their activation is a key step for cancer immunotherapy. Precise evaluation of CTL activity in vivo provides a powerful tool for monitoring cancer-immunotherapeutic outcomes, yet it faces tremendous challenges. Herein, by rationally designing a near-infrared (NIR) fluorescence probe Cys(StBu)-Ile-Glu-Phe-Asp-Lys(Cy5.5)-CBT (Cy5.5-CBT) and employing a reduction-instructed CBT-Cys click condensation reaction, we developed the fluorescence "dual quenched" nanoparticles Cy5.5-CBT-NPs for imaging of granzyme B (GraB), a biomarker tightly associated with the tumoricidal activity of CTLs. Upon GraB cleavage, Cy5.5-CBT-NPs disassembled, subtly turning the fluorescence signal "on". With this fluorescence "turn-on" property, Cy5.5-CBT-NPs enabled sensitive and real-time monitoring of GraB-mediated CTL responses against cancer cells in vitro. Animal experiments demonstrated that, at 16 h post injection, the fluorescence imaging signal of Cy5.5-CBT-NPs showed a 3.1-fold increase on the tumor sites of mice treated by an immune-activating drug S-(2-boronoethyl)-L-cysteine hydrochloride. We envision that Cy5.5-CBT-NPs may provide a powerful tool for noninvasive and sensitive evaluation of immunotherapeutic efficacy of cancer in the near future.

Enzymatic Nanosphere-to-Nanofiber Transition and Autophagy Inducer Release Promote Tumor Chemotherapy.

Chemotherapy has remained an effective and predominant cancer treatment for the past decades, but is hampered by its low response rate and severe systemic toxicity. Combination chemotherapies are proposed to address these issues, yet their therapeutic outcomes are still far from satisfactory. Thus, it is urgent to develop novel strategies to promote tumor chemosensitivity while reducing toxic side effects of chemotherapeutics. Herein, employing a rationally designed peptide conjugate Nap-Phe-Phe-Lys(SA-AZD8055)-Tyr(H2 PO3 )-OH (Nap-AZD-Yp), a novel approach of simultaneous intracellular nanofiber formation and autophagy inducer release is proposed for selectively sensitizing tumor to chemotherapy. Upon sequential catalyses of alkaline phosphatase and carboxylesterase, Nap-AZD-Yp undergoes nanosphere-to-nanofiber transition accompanied by autophagy inducer AZD8055 release in cancer cells. Cell experiments show enhanced endocytosis of anticancer drug doxorubicin and inhibition of cell migration due to the intracellular nanofiber formation. The released AZD8055 further activates excessive autophagy of cancer cells, sensitizing them to chemotherapy. Animal experiment results suggest Nap-AZD-Yp can significantly enhance the therapeutic effects of doxorubicin on tumors while mitigate its toxic adverse effects on normal tissues. It is anticipated that the "smart" concept in this work c be widely employed to develop novel combinational therapies for the treatment of cancers and other diseases in near future.

FAP-α-Instructed Coumarin Excimer Formation for High Contrast Fluorescence Imaging of Tumor.

Emissive excimers, which are formed by planar polycyclic aromatic fluorophores (e.g., coumarin), enable high contrast tumor imaging. However, it is still challenging to "turn on" excimer fluorescence in physiological dilute solutions. The biocompatible CBT-Cys click condensation reaction enables both intra- and intermolecular aggregations of the as-loaded fluorophores on the probe molecules, which may promote the generation of emissive excimers in a synergistic manner. As a proof-of-concept, we herein design a fluorescence probe Cbz-Gly-Pro-Cys(StBu)-Lys(coumarin)-CBT (Cbz-GPC(StBu)K(Cou)-CBT), which can be activated by FAP-α under tumor-inherent reduction conditions, undergo a CBT-Cys click reaction, and self-assemble into coumarin nanoparticle Cou-CBT-NP to "turn on" the excimer fluorescence. In vitro and in vivo studies validate that this "smart" probe realizes efficient excimer fluorescence imaging of FAP-α-overexpressed tumor cells with high contrast and enhanced accumulation, respectively. We anticipate that this probe can be applied for diagnosis of FAP-α-related diseases in the clinic in near future.

Trident Molecule with Nanobrush-Nanoparticle-Nanofiber Transition Property Spatially Suppresses Tumor Metastasis.

Metastasis-induced high mortality of cancers urgently demands new approaches to simultaneously inhibit primary tumor metastasis and distant tumor growth. Herein, by rational design of a trident molecule Nap-Phe-Phe-Lys(SA-CPT)-Lys(SA-HCQ)-Tyr(H2PO3)-OH (Nap-CPT-HCQ-Yp) with three functional "spears" (i.e., a phosphotyrosine motif for enzymatic self-assembly, camptothecin (CPT) motif for chemotherapy, and hydroxychloroquine (HCQ) motif for autophagy inhibition) and nanobrush-nanoparticle-nanofiber transition property, we propose a novel strategy of intracellular enzymatic nanofiber formation and synergistic autophagy inhibition-enhanced chemotherapy and immunotherapy for spatial suppression of tumor metastasis. Under sequential alkaline phosphatase catalysis and carboxylesterase hydrolysis, Nap-CPT-HCQ-Yp undergoes nanobrush-nanoparticle-nanofiber transition, accompanied by the releases of CPT and HCQ. The formed intracellular nanofibers effectively inhibit the metastasis and invasion behaviors of cancer cells. Meanwhile, the released CPT and HCQ synergistically induce a prominent therapeutic effect through autophagy inhibition-enhanced chemotherapy. Furthermore, chemotherapy of Nap-CPT-HCQ-Yp enhances immunogenic cell death, resulting in the activation of toxic T-cells. Finally, a combination of checkpoint blockade therapy and Nap-CPT-HCQ-Yp-mediated chemotherapy elicits systemic antitumor immunity, thereby achieving efficient inhibitions of primary tumors as well as distant tumors in a breast tumor model. Our work offers a simple and feasible strategy for the design of "smart" multifunctional prodrugs to spatially suppress tumor metastasis.

ROS Turn Nanoparticle Fluorescence on for Imaging Staphylococcus aureus Infection In Vivo.

Direct, noninvasive, and real-time imaging of Staphylococcus aureus (S. aureus) infection is of great value for quick diagnosis of related disease in clinic, but remains challenging. Herein, employing a rationally designed near-infrared fluorescence probe Cys(StB u)-EDA-Thioketal-Lys(Cy5.5)-CBT (TK-CBT) and a CBT-Cys click reaction, the fluorescence-quenched nanoparticles TK-CBT-NPs are facilely prepared. Upon oxidation by the abundant reactive oxygen species in S. aureus-infected macrophages, TK-CBT-NPs are fractured, turning the fluorescence "on" for imaging infections in vitro and in vivo. Specifically, TK-CBT-NPs show a 6.1-fold fluorescence imaging signal enhancement of the macrophages that are infected by S. aureus for 20 h in vitro. At 4 h postinjection, TK-CBT-NPs show a 2.8-fold fluorescence imaging signal enhancement of the sites in mice that are infected by S. aureus for 24 h. It is anticipated that TK-CBT-NPs could be applied for diagnosis of S. aureus infections in clinic in the near future.

Salt-Inducible Kinase 2-Triggered Release of Its Inhibitor from Hydrogel to Suppress Ovarian Cancer Metastasis.

Salt-inducible kinase 2 (SIK2) is a promising target for ovarian cancer therapy due to its critical role in tumorigenesis and progression. Currently available SIK2 inhibitors have shown remarkable therapeutic effects on ovarian cancers in preclinical studies. However, direct administration of the SIK2 inhibitors may bring significant off-target effect, limiting their clinical applications. In this work, by rational design of a hydrogelator Nap-Phe-Phe-Glu-Glu-Leu-Tyr-Arg-Thr-Gln-Ser-Ser-Ser-Asn-Leu-OH (Nap-S) to coassemble a SIK2 inhibitor HG-9-91-01 (HG), a SIK2-responsive supramolecular hydrogel (Gel Nap-S+HG) for local administration and SIK2-responsive release of HG is reported to efficiently suppress ovarian cancer metastasis. Under the activation of SIK2 overexpressed in ovarian cancers, Nap-S in the hydrogel is phosphorylated to yield hydrophilic Nap-Phe-Phe-Glu-Glu-Leu-Tyr-Arg-Thr-Gln-Ser(H2 PO3 )-Ser-Ser-Asn-Leu (Nap-Sp), triggering the disassembly of the hydrogel and a responsive release of the inhibitor. Cell experiments indicate that sustained release of HG from Gel Nap-S+HG induce a prominent therapeutic effect on cancer cells by inhibiting SIK2 and phosphorylation of their downstream signaling molecules. Animal experiments demonstrate that, compared with those tumor model mice treated with free HG, Gel Nap-S+HG-treatment mice show an enhanced inhibition on ovarian tumor growth and metastasis. It is anticipated that the Gel Nap-S+HG can be applied for ovarian cancer therapy in clinic in the near future.

Intracellular Self-Assembly of Peptide Conjugates for Tumor Imaging and Therapy.

Intracellular self-assembly (ISA) is a versatile and powerful strategy for in situ constructing sophisticated and functional supramolecular nanostructures, which has been widely applied in biomedicine and biomedical engineering. Among the common building blocks for ISA, peptides have attracted increasingly attention due to their intrinsic bioactivity, biocompatibility, and biodegradability. Particularly, by conjugating functional motifs (e.g., probes or drugs) to peptides to yield the peptide conjugates, the latter show enhanced stability and efficiency, and probably new functions. In recent years, employing ISA of peptide conjugates for tumor imaging and treatment has achieved great progresses. Therefore, the recent progress of ISA of peptide conjugates is summarized in this progress report. Moreover, several examples of ISA of peptide conjugates for other important imaging or therapeutic applications are also introduced. Finally, a brief perspective on remaining challenges and potential directions for future research in this area is presented.

A Golgi-Targeting and Dual-Color "Turn-On" Probe for Spatially Precise Imaging of Furin.

Development of fluorescence probes for highly accurate detection of cancer-related enzyme activity is important in early cancer diagnosis. Herein, we report a Golgi-targeting and dual-color "Turn-On" probe Q-RVRR-DCM for imaging furin with high spatial precision. By integrating the principles of Förster resonance energy transfer and intramolecular charge transfer, the probe was designed to be non-fluorescent. Upon furin cleavage, Q-RVRR-DCM was converted into Q-RVRR and DCM-NH2, turning the dual fluorescence color "On" at 420 and 640 nm without spectral cross-talk. In furin-overexpressing HCT116 cells, Q-RVRR-DCM showed not only furin-specific, dual-color "Turn-On" fluorescence but also superior colocalization with a Golgi tracker than the single-color "Turn-On" probe RVRR-DCM. We envision that, with the excellent properties of Golgi-targeting and dual fluorescence color "Turn-On", our furin probe Q-RVRR-DCM could be applied for accurate early diagnosis of cancer in the near future.