Nano Proteolysis Targeting Chimeras (PROTACs) with Anti-Hook Effect for Tumor Therapy.

Proteolysis targeting chimera (PROTAC) is an emerging pharmacological modality with innovated post-translational protein degradation capabilities. However, off-target induced unintended tissue effects and intrinsic "hook effect" hinder PROTAC biotechnology to be maturely developed. Herein, an intracellular fabricated nano proteolysis targeting chimeras (Nano-PROTACs) modality with a center-spoke degradation network for achieving efficient dose-dependent protein degradation in tumor is reported. The PROTAC precursors are triggered by higher GSH concentrations inside tumor cells, which subsequently in situ self-assemble into Nano-PROTACs through intermolecular hydrogen bond interactions. The fibrous Nano-PROTACs can form effective polynary complexes and E3 ligases degradation network with multi-binding sites, achieving dose-dependent protein degradation with "anti-hook effect". The generality and efficacy of Nano-PROTACs are validated by degrading variable protein of interest (POI) such as epidermal growth factor receptor (EGFR) and androgen receptor (AR) in a wide-range dose-dependent manner with a 95 % degradation rate and long-lasting potency up to 72 h in vitro. Significantly, Nano-PROTACs achieve in vivo dose-dependent protein degradation up to 79 % and tumor growth inhibition in A549 and LNCap xenograft mice models, respectively. Taking advantages of in situ self-assembly strategy, the Nano-PROTACs provide a generalizable platform to promote precise clinical translational application of PROTAC.

In Situ Self-Assembly of Bispecific Peptide for Cancer Immunotherapy.

Precise and effective manipulation of protein functions still faces tremendous challenges. Herein we report a programmable peptide molecule, consisted of targeting and self-assembly modules, that enables specific and highly efficient assembly governed by targeting receptor proteins. Upon binding to the cell membrane receptor, peptide conformation is somewhat stabilized along with decreased self-assembly activation energy, promoting peptide-protein complex oligomerization. We first design a GNNQQNY-RGD peptide (G7-RGD) to recognize integrin αV ß3 receptor for proof-of-concept study. In the presence of αV ß3 protein, the critical assembly concentration of free G7-RGD decreases from 525 to 33 µM and the resultant G7-RGD cluster drives integrin receptor oligomerization. Finally, a bispecific assembling peptide antiCD3-G7-RGD is rationally designed for cancer immunotherapy, which validates CD3 oligomerization and concomitant T cell activation, leading to T cell-mediated cancer cell cytolysis.

Photothermal-Promoted Morphology Transformation in Vivo Monitored by Photoacoustic Imaging.

The in situ construction of the nanoassembly has been demonstrated to improve the performance of bioactive molecules, but the control of the morphology of nanomaterials in vivo still remains a tremendous challenge. Herein, a photothermal-promoted morphology transformation (PMT) strategy is developed to accelerate the formation of nanomaterials for improving the biological performance of drug molecules. Compared with the spontaneous process, the rate of transformation increases by ∼4 times in the PMT process. Owing to increased assembly rate, the tumor accumulation of drugs is ∼2-fold than that without photo irradiation, which inhibits tumor growth effectively. More importantly, the chemical reassembly process in vitro and in vivo is monitored by the advanced ratiometric photoacoustic image, confirming the photoinduced transformation acceleration. Through the noninvasively artificial control on assembly dynamics in vivo, the PMT strategy provides a new insight for developing the intelligent theranostics.

Controllable Self-Assembly of Peptide-Cyanine Conjugates In Vivo as Fine-Tunable Theranostics.

The fabrication of functional assemblies with defined structures through controllable molecular packing under physiological conditions is challenging. Here, modularly designed peptide-cyanine conjugates that intracellularly self-assembly into 1D columnar superstructures with controlled cyanine aggregation were designed, and they exhibit distinct imaging or photothermal properties. The peptide backbone is cleaved by caspase-3/7 after entering the cells. Then the self-assembled residue, with a double cyanine substitution (Pr-2Cy), forms a P helical column in which H-aggregated cyanine dyes show 3.4-fold photothermal conversion efficiency compared to free ones. The self-assembled residue with a single cyanine substitution (Pr-1Cy) forms a loose column, in which cyanine dyes with undefined structure have a fluorescence quantum yield of up to 9.5 % (emission at 819 nm in H2 O). This work provides a simple way to modify in vivo self-assembled peptides with functional molecules for achieving desired bio-applications.

Peptide-Based Autophagic Gene and Cisplatin Co-delivery Systems Enable Improved Chemotherapy Resistance.

Cisplatin-based chemotherapy is a widely used first-line strategy for numerous cancers. However, drug resistances are often inevitable accompanied by the long-term use of cisplatin in vivo, significantly hampering its therapeutic efficacy and clinical outcomes. Among others, autophagy induction is one of the most common causes of tumor resistance to cisplatin. Herein, a self-assembled nanoprodrug platform was developed with the synergistic effect of cisplatin and RNAi to fight against cisplatin-resistant lung cancer. The nanoprodrug platform consists of three molecular modules, including prodrug complex of Pt(IV)-peptide-bis(pyrene), DSPE-PEG, and cRGD-modified DSPE-PEG. The Pt(IV) is immobilized with peptide via amide bonds, allowing the Pt(IV) to be loaded with a loading efficiency of >95% and rapid-release active platinum ions (Pt(II)) in the presence of glutathione (GSH). Meanwhile, the peptide of the prodrug complex could efficiently deliver Beclin1 siRNA ( Beclin1 is an autophagy initiation factor) to the cytoplasm, thereby leading to autophagy inhibition. In addition, incorporation of DSPE-PEG and cRGD-modified DSPE-PEG molecules improves the biocompatibility and cellular uptake of the nanoprodrug platform. In vivo results also indicate that the nanoprodrug platform significantly inhibits the growth of a cisplatin-resistant tumor on xenograft mice models with a remarkable inhibition rate, up to 84% after intravenous injection.

In†Situ Self-Assembled Nanofibers Precisely Target Cancer-Associated Fibroblasts for Improved Tumor Imaging.

Tumor complexity makes the development of highly sensitive tumor imaging probes an arduous task. Here, we construct a peptide-based near-infrared probe that is responsive to fibroblast activation protein-α (FAP-α), and specifically forms nanofibers on the surface of cancer-associated fibroblasts (CAFs) in situ. The assembly/aggregation-induced retention (AIR) effect results in enhanced accumulation and retention of the probe around the tumor, resulting in a 5.5-fold signal enhancement in the tumor 48 h after administration compared to that of a control molecule that does not aggregate. The probe provides a prolonged detectable window of 48 h for tumor diagnosis. The selective assembly of the probe results in a signal intensity over four- and fivefold higher in tumor than in the liver and kidney, respectively. With enhanced tumor imaging capability, this probe can visualize small tumors around 2 mm in diameter.

pH-Sensitive Polymeric Nanoparticles Modulate Autophagic Effect via Lysosome Impairment.

In drug delivery systems, pH-sensitive polymers are commonly used as drug carriers, and significant efforts have been devoted to the aspects of controlled delivery and release of drugs. However, few studies address the possible autophagic effects on cells. Here, for the first time, using a fluorescent autophagy-reporting cell line, this study evaluates the autophagy-induced capabilities of four types of pH-sensitive polymeric nanoparticles (NPs) with different physical properties, including size, surface modification, and pH-sensitivity. Based on experimental results, this study concludes that pH-sensitivity is one of the most important factors in autophagy induction. In addition, this study finds that variation of concentration of NPs could cause different autophagic effect, i.e., low concentration of NPs induces autophagy in an mTOR-dependent manner, but high dose of NPs leads to autophagic cell death. Identification of this tunable autophagic effect offers a novel strategy for enhancing therapeutic effect in cancer therapy through modulation of autophagy.

Synthesis of biocompatible glycodendrimer based on fluorescent perylene bisimides and its bioimaging.

A novel water-soluble fluorescent glycodendrimer based on perylene bisimides is synthesized, which exhibits high fluorescence quantum yield of 54%. While the binding interactions of PBI-Man with Concanavalin A (Con A) are studied by fluorescence spectra and CD spectra, which show strong binding affinity for Con A with the binding constant of 3.8 × 10(7) m(-1) for monomeric mannose, nearly four orders of magnitude higher affinity than the monovalent mannose ligand. Furthermore, the fluorescence imaging of macrophage cell with PBI-Man is investigated, and shows selectively binding interaction with the mannose receptor-medicated cell entry. Moreover, the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) activities of PBI-Man show that PBI-Man as a biocompatible agent is noncytotoxic to living cells.

Self-assembly of peptide nanomaterials at biointerfaces: molecular design and biomedical applications.

Self-assembly is an important strategy for constructing ordered structures and complex functions in nature. Based on this, people can imitate nature and artificially construct functional materials with novel structures through the supermolecular self-assembly pathway of biological interfaces. Among the many assembly units, peptide molecular self-assembly has received widespread attention in recent years. In this review, we introduce the interactions (hydrophobic interaction, hydrogen bond, and electrostatic interaction) between peptide nanomaterials and biological interfaces, summarizing the latest advancements in multifunctional self-assembling peptide materials. We systematically demonstrate the assembly mechanisms of peptides at biological interfaces, such as proteins and cell membranes, while highlighting their application potential and challenges in fields like drug delivery, antibacterial strategies, and cancer therapy.

A triazatruxene-based glycocluster as a fluorescent sensor for concanavalin A.

A new triazatruxene-based fluorescent glycocluster has been designed, synthesized, and fully characterized by NMR spectroscopy and mass spectrometry. Furthermore, its specific and selective binding properties with concanavalin A (Con A) have been investigated by fluorescence spectroscopy, circular dichroism (CD) spectroscopy, and turbidity assay. The obtained results showed that the multivalent mannose-modified triazatruxene exhibited specific binding with Con A, but no binding to peanut agglutinin (PNA) lectin or bovine serum albumin (BSA), corresponding to a two-orders-of-magnitude higher affinity than that of monovalent mannose ligands. Most interestingly, a fluorescence enhancement of the triazatruxene-based glycocluster was observed upon binding with Con A because of hydrophobic interactions involving sites close to the triazatruxene moiety. Furthermore, the inhibitory ability of the triazatruxene-based glycocluster against ORN178-induced haemagglutination has been investigated by haemagglutination inhibition assay. The results indicated selective binding with ORN178.

Synthesis of perylene bisimide-centered glycodendrimer and its interactions with concanavalin A.

A novel glycodendrimer based on 18 peripheral α-D-mannoses functionalized perylene bisimide derivative PBI-18-Man was synthesized and its selectively binding interactions for Con A were investigated by CD spectra and turbidity assay, which exhibited strong binding affinity for Con A with the binding constant of 1.3×10(8) M(-1) (7.2×10(6) M(-1) for monomeric mannose, valency corrected), 3 orders of magnitude higher affinity than the monovalent mannose ligand. Furthermore, the inhibitory activity for Con A was studied by ELLA experiment, showed 2 times inhibitor activity than the reference compound (α-MMP).

Multivalent glycoclusters constructed by chiral self-assembly of mannose functionalized perylene bisimide.

Water-soluble perylene bisimide derivative 7 modified with six mannoses was synthesized and its self-assembled properties were studied by UV-Vis and CD spectroscopy, which revealed an interesting self-assembly with a solvent-tuning chiral conformation in H(2)O-DMSO solution. As H(2)O was added to the DMSO solution until a 60% (or 70%) v/v proportion was achieved, the self-assembly of the mannose functionalized compound 7 exhibited a left-handed helical conformation. More interestingly, when the volume of H(2)O constituted beyond 85% of the solution, the conformation of the self-assembly turned out to be a right-handed helical conformation. Furthermore, the binding interactions between the self-assembly of compound 7 and Con A were investigated by turbidity assay, CD spectra, TEM and SEM images, and ELLA experiment, which indicated that the self-assembly of compound 7 as multivalent glycoclusters exhibited specific binding to Con A with an IC(50) value of 24 µM (144 µM, valency corrected), 10 times stronger than the reference compound (α-MMP).

Intelligent Biomaterialomics: Molecular Design, Manufacturing, and Biomedical Applications.

Materialomics integrates experiment, theory, and computation in a high-throughput manner, and has changed the paradigm for the research and development of new functional materials. Recently, with the rapid development of high-throughput characterization and machine-learning technologies, the establishment of biomaterialomics that tackles complex physiological behaviors has become accessible. Breakthroughs in the clinical translation of nanoparticle-based therapeutics and vaccines have been observed. Herein, recent advances in biomaterials, including polymers, lipid-like materials, and peptides/proteins, discovered through high-throughput screening or machine learning-assisted methods, are summarized. The molecular design of structure-diversified libraries; high-throughput characterization, screening, and preparation; and, their applications in drug delivery and clinical translation are discussed in detail. Furthermore, the prospects and main challenges in future biomaterialomics and high-throughput screening development are highlighted.

Engineered CpG-Loaded Nanorobots Drive Autophagy-Mediated Immunity for TLR9-Positive Cancer Therapy.

Smart nanorobots have emerged as novel drug delivery platforms in nanomedicine, potentially improving anti-cancer efficacy and reducing side effects. In this study, an intelligent tumor microenvironment-responsive nanorobot is developed that effectively delivers CpG payloads to Toll-like receptor 9 (TLR9)-positive tumors to induce autophagy-mediated cell death for immunotherapy. The nanorobots are fabricated by co-self-assembly of two amphiphilic triblock polymer peptides: one containing the matrix metallopeptidase 2 (MMP2)-cleaved GPLGVRGS motif to control the mechanical opening of the nanorobots and provide targeting capability for TLR-9-positive tumors and the other consisting of an arginine-rich GRRRDRGRS sequence that can condense nuclear acid payloads through electrostatic interactions. Using multiple tumor-bearing mouse models, it is investigated whether the intravenous injection of CpG-loaded nanorobots could effectively deliver CpG payloads to TLR-9-positive tumors and elicit anti-tumor immunity through TLR9 signaling and autophagy. Therefore, besides being a commonly used adjuvant for tumor vaccination, CpG-loaded nanorobots can effectively reprogram the tumor immunosuppressive microenvironment and suppress tumor growth and recurrence. This nanorobot-based CpG immunotherapy can be considered a feasible approach to induce anti-tumor immunity, showing great therapeutic potential for the future treatment of TLR9-positive cancers.

Programmable Peptides Activated Macropinocytosis for Direct Cytosolic Delivery.

Bioactive macromolecules show great promise for the treatment of various diseases. However, the cytosolic delivery of peptide-based drugs remains a challenging task owing to the existence of multiple intracellular barriers and ineffective endosomal escape. To address these issues, herein, programmable self-assembling peptide vectors are reported to amplify cargo internalization into the cytoplasm through receptor-activated macropinocytosis. Programmable self-assembling peptide vector-active human epidermal growth factor receptor-2 (HER2) signaling induces the receptor-activated macropinocytosis pathway, achieving efficient uptake in tumor cells. Shrinking macropinosomes accelerate the process of assembly dynamics and form nanostructures in the cytoplasm to increase peptide-based cargo accumulation and retention. Inductively coupled plasma mass (ICP-MS) spectrometry quantitative analysis indicates that the Gd delivery efficiency in tumor tissue through the macropinocytosis pathway is improved 2.5-fold compared with that through the use of active targeting molecular delivery. Finally, compared with nanoparticles and active targeting delivery, the delivery of bioactive peptide drugs through the self-assembly of peptide vectors maintains high drug activity (the IC50 decreased twofold) in the cytoplasm and achieves effective inhibition of tumor cell growth. Programmable self-assembling peptide vectors represent a promising platform for the intracellular delivery of diverse bioactive drugs, including molecular drugs, peptides, and biologics.

Reprogramming Hypoxic Tumor-Associated Macrophages by Nanoglycoclusters for Boosted Cancer Immunotherapy.

The tumor-associated macrophages (TAMs) in intratumoral hypoxic regions are key drivers of immune escape. Reprogramming the hypoxic TAMs to antitumor phenotype holds great therapeutic benefits but remains challenging for current drugs. Here, an in situ activated nanoglycocluster is reported to realize effective tumor penetration and potent repolarization of hypoxic TAMs. Triggered by the hypoxia-upregulated matrix metalloproteinase-2 (MMP-2), the nanoglycocluster is self-assembled from the administered mannose-containing precursor glycopeptides and presents densely-arrayed mannoses to multivalently engage with mannose receptors on M2-like TAMs for efficient phenotype switch. By virtue of the high diffusivity of precursor glycopeptides due to their low molecular mass and weak affinity with TAMs in perivascular regions, the nanoglycoclusters are capable of substantially accumulating in hypoxic areas to strongly interact with local TAMs. This enables the efficient repolarization of overall TAMs with a higher rate than the small-molecule drug R848 and CD40 antibody, and beneficial therapeutic effects in mouse tumor models especially when combining with PD-1 antibody. This on-demand activated immunoagent is endowed with tumor-penetrating properties and inspires the design of diverse intelligent nanomedicines for hypoxia-related cancer immunotherapy.

An In Vivo Self-Assembled Bispecific Nanoblocker for Enhancing Tumor Immunotherapy.

Anti-PD-L1 monoclonal antibody has achieved substantial success in tumor immunotherapy by T-cells activation. However, the excessive accumulation of extracellular matrix components induced by unsatisfactory T-cells infiltration and poor tumor penetration of antibodies make it challenging to realize efficient tumor immunotherapy. Herein, a peptide-based bispecific nanoblocker (BNB) strategy is reported for in situ construction of CXCR4/PD-L1 targeted nanoclusters on the surface of tumor cells that are capable of boosting T-cells infiltration through CXCR4 blockage and enhancing T-cells activation by PD-L1 occupancy, ultimately realizing high-performance tumor immunotherapy. Briefly, the BNB strategy selectively recognizes and bonds CXCR4/PD-L1 with deep tumor penetration, which rapidly self-assembles into nanoclusters on the surface of tumor cells. Compared to the traditional bispecific antibody, BNB exhibits an intriguing metabolic behavior, that is, the elimination half-life (t1/2 ) of BNB in the tumor is 69.3 h which is ≈50 times longer than that in the plasma (1.4 h). The higher tumor accumulation and rapid systemic clearance overcome potential systemic side effects. Moreover, the solid tumor stress generated by excessive extracellular matrix components is substantially reduced to 44%, which promotes T-cells infiltration and activation for immunotherapy efficacy. Finally, these findings substantially strengthen and extend clinical applications of PD-1/PD-L1 immunotherapy.

A bispecific glycopeptide spatiotemporally regulates tumor microenvironment for inhibiting bladder cancer recurrence.

Up to 75% of bladder cancer patients suffer from recurrence due to postoperative tumor implantation. However, clinically used Bacillus Calmette-Guerin (BCG) treatment failed to inhibit the recurrence. Here, we report a bispecific glycopeptide (bsGP) that simultaneously targets CD206 on tumor-associated macrophages (TAMs) and CXCR4 on tumor cells. bsGP repolarizes protumoral M2-like TAMs to antitumor M1-like that mediated cytotoxicity and T cell recruitment. Meanwhile, bsGP is cleaved by the MMP-2 enzyme to form nanostructure for the long-term inhibition of CXCR4 downstream signaling, resulting in reduced tumor metastasis and promoted T cell infiltration. In orthotopic bladder tumor models, bsGP reduced the postoperative recurrence rate to 22%. In parallel, the recurrence rates of 89 and 78% were treated by doxycycline and BCG used in clinic, respectively. Mechanistic studies reveal that bsGP reduces the matrix microenvironment barrier, increasing the spatially redirected CD8+ T cells to tumor cells. We envision that bis-targeting CD206 and CXCR4 may pave the way to inhibit tumor metastasis and recurrence.

Programmable design and self assembly of peptide conjugated AIEgens for biomedical applications.

Aggregation-induced emission luminogens (AIEgens) possess enhanced fluorescence in highly aggregated states, thus enabling AIEgens as a promising module for highly emissive fluorescence biomaterials. So far, AIEgens-based nanomaterials and their hybrids have been reported for biomedical applications. Benefiting from the intrinsic biocompatibility and biofunction-editing properties of peptides, peptide-AIEgens hybrid biomaterials reveal unlimited possibilities including target capacity, specificity, stimuli-responsiveness, self-assembly, controllable structural transformation, etc.. In the last two decades, peptide-AIEgens hybrid nanomaterials with a unique design concept in aggregated states have achieved various biomedical applications such as biosensing, bioimaging, imaging-guided surgery, drug delivery and therapy. More recently, programmable design of peptide-AIEgens for in situ self-assembly provides a unique strategy for constructing intelligent entities with defined biological functions. In this review, we summarize the basic design principle of programmable peptide-AIEgens, structure-effect relationship and their unusual biomedical effects. Finally, an outlook and perspective toward future challenges and developments of peptide-AIEgens nanomaterials are concluded.

Oncolytic peptide nanomachine circumvents chemo resistance of renal cell carcinoma.

Due to intrinsic and acquired chemo/radiotherapy-resistance, renal cell carcinoma shows limited therapeutic response to clinically utilized targeting drugs. Here a tumor-activated oncolytic peptide nanomachine is devised to selectively lysing tumor cell membrane without causing drug resistance. Specifically, in the acidic tumor microenvironment, the oncolytic peptide nanomachine automatically activated through morphologically transformation from nanoparticles to nanofibrils with restoring α-helical conformation, which physically bind to tumor cell membrane with multiple (spatially correlated and time-resolved) interactions and subsequently lyse local cell membrane. The IC50 of the oncolytic peptide nanomachine is as low as 2.44 µM and it inhibit up to 90% of tumor cells within 2 h with unique bystander killing effect. In vivo, the tumor inhibition rate of the oncolytic peptide nanomachine is 71% without off-target activity and hemolytic activity. These results support that tumor-selective oncolytic peptide nanomachine represent a promising alternative approach for multidrug-resistant tumor treatments by inducing cell membrane lysis.