Bioconjugate Chemistry最新文献

Monomethyl auristatin E (MMAE) is a promising treatment option for patients diagnosed with prostate cancer (PCa); however, toxicities prevent MMAE from being administered as free drug. No MMAE-based treatment is currently marketed for PCa. Herein, we describe a small-molecule-drug conjugate, CTT2274, for the selective delivery of MMAE. CTT2274 is composed of a prostate-specific membrane antigen (PSMA)-binding scaffold, a biphenyl motif, a pH-sensitive phosphoramidate linker, and MMAE payload. We demonstrate that CTT2274 shows selective binding to PSMA, which is overexpressed on PCa cells, and induces tumor cell death in vitro. In a patient-derived xenograft tumor model of PCa in mice, we show that weekly intravenous dosing of CTT2274 at 3.6 mg/kg for six weeks is superior to treatment with free MMAE at equivalent doses. Mice treated with CTT2274 experienced prolonged tumor suppression and significantly greater overall survival than mice treated with PBS. Additionally, the safety of CTT2274 compared to an equivalent dose of MMAE was assessed in healthy, non-tumor-bearing mice. Our results demonstrate that CTT2274 therapy is as efficacious as MMAE, results in superior overall survival, and has a more favorable safety profile. Together, these data indicate that CTT2274 is a candidate for clinical translation for the treatment of PCa.

Bacterial keratitis is a prevalent, and severe corneal illness resulting from bacterial pathogens. Failure to administer a timely and suitable therapy may lead to corneal opacity, ulceration, significant vision impairment, or potential blindness. Current clinical interventions for bacterial keratitis involve the administration of topical antimicrobial agents and systemic antibiotics. However, the misuse and overuse of antibiotics have led to the rapid emergence of antibiotic-resistant bacteria. Additionally, the restricted antibacterial spectrum and possible adverse effects of antibiotics have provided considerable obstacles to traditional therapies. This highlights the urgent need for novel and highly effective antimicrobial agents. Antimicrobial peptides (AMPs) are a class of naturally occurring or synthetically designed small molecules that have gained significant attention due to their unique antimicrobial mechanisms and low risk of resistance development. AMPs exhibit promising potential in treating bacterial keratitis through direct antibacterial mechanisms, such as inhibiting cell wall synthesis, disrupting cell membranes, and interfering with nucleic acid metabolism, as well as indirect mechanisms, including modulation of the host immune response. This review provides a comprehensive overview of the antibacterial mechanisms of AMPs and their advancements in the treatment of bacterial keratitis. It emphasizes the role of various modification strategies and artificial-intelligence-assisted design in enhancing the antibacterial efficacy, stability, and biocompatibility of AMPs. Furthermore, this review discusses the latest progress in combining AMPs with delivery systems for improved therapeutic outcomes. Finally, the review highlights the current challenges and future perspectives of AMPs in bacterial keratitis treatment, providing valuable insights for developing novel AMPs with high antibacterial efficacy, stability, and safety for bacterial keratitis therapies.

Erythropoietin (EPO)-induced cellular signaling through the EPO receptor (EPOR) is a fundamental pathway for the modulation of cellular behavior and activity. In our previous work, we showed in primary human astrocytes that the multivalent display of EPO on the surface of semiconductor quantum dots (QDs) mediates augmented JAK/STAT signaling, a concomitant 1.8-fold increase in the expression of aquaporin-4 (AQPN-4) channel proteins, and a 2-fold increase in the AQPN-4-mediated water transport activity. Our hypothesis is that this enhanced signaling involves the simultaneous ligation and clustering of EPOR by QD-EPO conjugates. Here, we utilized a human embryonic kidney (HEK 293T/17) cell line transfected with EPOR fused to enhanced green fluorescent protein (eGFP) to visualize EPOR clustering. We demonstrate that QDs displaying five copies of EPO (bearing a C-terminal 6-histidine tract) on the nanoparticle surface induce a 1.8-fold increase in EPOR clustering compared to monomeric EPO at the same concentration. Our findings confirm the critical role played by the multivalent display of EPO in mediating clustering of the EPOR. More generally, these results illustrate the capability of nanoparticle-growth factor bioconjugates to control the activity of cognate receptors and the important role played by multivalent display in the modulation of selective cellular delivery and signaling.

This work describes the synthesis, characterization, and antibacterial properties of four bile acid-triclosan conjugates. The in vitro antibacterial activity of synthetic bile acid-triclosan conjugates was investigated against a panel of Gram-positive and Gram-negative bacteria. Conjugates 3 and 4 show high activity against Escherichia coli (ATCC25922), with IC₅₀ values of 2.94 ± 0.7 and 1.51 ± 0.05 μM, respectively. Conjugate 4 demonstrated 9 times the activity of triclosan (6.77 μM) and 18 times the potency of kanamycin, a well-known antibiotic. Compound 3 showed higher potential activity against all evaluated strains, including Bacillus megaterium (IC₅₀: 3.05 ± 0.02), Bacillus amyloquefaciens (IC₅₀: 8.79 ± 0.01), Serratia marcescens (IC₅₀: 6.77 ± 0.4), and E. coli (IC₅₀: 1.51 ± 0.05 μM). These findings indicate that it has broad-spectrum antibacterial activity. Bile acid-triclosan conjugates prevent biofilms by up to 99% at low doses (conjugates 4; 4.16 ± 0.8 μM), compared to triclosan. Conjugate 5 was most potent against B. amyloquefaciens (IC₅₀ = 5.23 ± 0.2 μM), while conjugate 4 was most effective against B. megaterium (IC₅₀ = 4.16 ± 0.8 μM) in biofilm formation. These conjugates inhibit biofilm formation by limiting the extracellular polymeric substance generation. The in vitro antibacterial study revealed that bile acid-triclosan conjugates were more effective than the parent molecule triclosan at inhibiting bacterial growth and biofilm formation against both Gram-positive and Gram-negative bacteria.

Herein, a water-soluble, ultrabright, near-infrared (NIR) fluorescent, mechanically interlocked molecules (MIMs)-peptide bioconjugate is designed with dual targeting capabilities. Cancer cell surface overexpressed α_Vβ₃ integrin targeting two RGDS tetrapeptide residues is tethered at the macrocycle of MIMs-peptide bioconjugate via Cu(I)-catalyzed click chemistry on the Wang resin, and mitochondria targeting lipophilic cationic TPP⁺ functionality is conjugated at the axle dye. Living carcinoma cell selective active targeting, subsequently cell penetration, mitochondrial imaging, including the ultrastructure of cristae, and real-time tracking of malignant mitochondria by MIMs-peptide bioconjugate (RGDS)₂-Mito-MIMs-TPP⁺ are established by stimulated emission depletion (STED) super-resolved fluorescence microscopy. Water-soluble NIR (RGDS)₂-Mito-MIMs-TPP⁺ is an effective class of MIMs-peptide bioconjugate with promising photophysics; for instance, remarkable photostability and thermal stability, strong and narrow NIR abs/em bands with high quantum yield, ultrabrightness, decent fluorescence lifetime, reasonable stability against cellular nucleophiles, biocompatibility, noncytotoxicity, and dual-targeted living cancer cell submitochondrial imaging ability are all indispensable criteria for targeted super-resolved STED microscopy.

Nanobodies play an increasingly prominent role in cancer imaging and therapy. However, their in vivo efficacy is often constrained by inadequate tumor penetration and rapid clearance from the bloodstream, particularly in brain tumors due to the intractable blood-brain barrier (BBB). Glycosylation is a favorable strategy for modulating the biological functions of nanobodies, including permeability and pharmacokinetics, but it also leads to heterogeneous glycan structures, which affect the targeting ability, stability, and quality of nanobodies. Here, we describe a post-translational modification strategy to produce precisely engineered and homogeneous nanobody-glucoside conjugates for effective BBB penetration and brain tumor targeting. Specifically, we employ an enzymatic method and click chemistry to functionalize nanobodies with glucoside and poly(ethylene glycol) (PEG), facilitating efficient transcytosis into the brain via glucose transporter-1 (GLUT1). Furthermore, we rationally select a near-infrared (NIR) fluorophore for labeling to maintain the metabolic pathway and biodistribution of nanobodies and assess their potency in two tumor models. The resulting nanobody-glucoside conjugates demonstrate a remarkable increase in BBB penetration and brain tumor accumulation, which are ∼2.9-fold higher in the transgenic mouse model and ∼5.7-fold higher in the orthotopic glioma model compared to unmodified nanobodies. This study provides a promising approach for the production of nanobody therapeutic agents for central nervous system (CNS) delivery.

The ability to label synthetic oligonucleotides with fluorescent probes has greatly expanded their nanotechnological applications. To continue this expansion, it is essential to develop approachable, modular, and tunable fluorescent platforms. In this study, we present the synthesis and incorporation of an amino-formyl-thieno[3,2-b]thiophene (AFTh₂) handle at the 5'-position of DNA oligonucleotides. The 5'-AFTh₂ end-label participates in both on-strand Knoevenagel and heterocyclization reactions, yielding far-red hemicyanines and pH-responsive probes with pK_a values in the biological regime. The Knoevenagel products, designated 5'-ATh₂Btz and 5'-ATh₂Ind, demonstrate excitation maxima beyond 640 nm with brightness up to ∼50,000 M^-1 cm^-1. Notably, 5'-ATh₂Btz demonstrates strong topology sensitivity, allowing it to probe transitions from duplex- to single-strand (SS)/G-quadruplex (GQ) topologies with an ∼9-fold increase in fluorescence in the absence of quenchers. In contrast, the heterocyclization product, 5'-ATh₂BIM, displays visible excitation and emission and is weakly fluorescent in basic solution. Upon lowering the pH from ∼8 to 5, this probe undergoes an unprecedented 400-fold light-up. Additionally, attaching 5'-ATh₂BIM to a polymorphic GQ allows for a shift in pK_a by ∼1.5 pH units simply by changing topology. The performance of the probes has been demonstrated in various contexts, including GQs, i-motifs, duplexes, and SS oligonucleotides. Their performance should facilitate the development of new DNA-based sensing platforms.

Ulcerative colitis (UC) is a chronic inflammatory bowel disease (IBD). CT imaging with contrast agents is commonly used for visualizing the gastrointestinal (GI) tract in UC patients. Contrast agents that provide enhanced imaging performance are highly valuable in this field. Recent studies have made significant progress in developing better contrast agents for imaging the gastrointestinal tract using nanoparticles. However, the impact of nanoparticle size on this application remains unexplored. Gold nanoparticles (AuNPs) serve as an ideal model to investigate the effect of nanoparticle size on imaging of the gastrointestinal tract due to their controllable synthesis across a broad size range. In this study, we synthesized AuNPs with core sizes ranging from 5 to 75 nm to examine the effect of the size in this setting. AuNPs were coated with poly(ethylene glycol) (PEG) to enhance stability and biocompatibility. In vitro tests show that gold nanoparticles are cytocompatible with macrophage cells (∼100% cell viability) and remain stable under acidic conditions, with no significant size changes over time. Phantom imaging studies using a clinical CT scanner indicated that there was no effect of nanoparticle size on CT contrast production, as previously demonstrated. In vivo imaging using a mouse model of acute colitis revealed a strong contrast generation throughout the GI tract for all agents tested. For the most part, in vivo contrast was independent of AuNP size, although AuNP outperformed iopamidol (a clinically approved control agent). In addition, differences in attenuation trends were observed between healthy and colitis mice. We also observed almost complete clearance at 24 h of all formulations tested (less than 0.7% ID/g was retained), supporting their value as a model platform for studying nanoparticle behavior in imaging. In conclusion, this study highlights the potential of nanoparticles as effective contrast agents for CT imaging of the gastrointestinal tract (GIT) in the UC. Further systemic research is needed to explore contrast agents that can specifically image disease processes in this disease setting.

Red blood cells (RBCs) serve as natural transporters and can be modified to enhance the pharmacokinetics and pharmacodynamics of a protein cargo. Affinity targeting of Factor IX (FIX) to the RBC membrane is a promising approach to improve the (pro)enzyme's pharmacokinetics. For RBC targeting, purified human FIX was conjugated to the anti-mouse glycophorin A monoclonal antibody Ter119. The goal of this study was to characterize the activity of the FIX-Ter119 conjugate and efficacy of its loading on RBCs, as well as to investigate the biodistribution, pharmacokinetics, and various biological properties of the loaded RBCs. Mouse RBCs were incubated with the Ter119-FIX conjugate, where adding 10,000 molecules per RBC resulted in 37% binding (4K/RBC), and 50,000 molecules per RBC resulted in 34% binding (17K/RBC). The pharmacokinetics (PK) profile showed that more than 90% of the Ter119-FIX conjugate was associated with RBCs and circulated stably bound to the RBCs for 24 h, increasing the area under the PK curve 7.6 times vs free FIX. Ter119-FIX loaded RBCs have specific procoagulant FIXa activity, including promotion of thrombin generation and acceleration of clotting in FIX-deficient plasma. Morphological characterization shows that Ter119-FIX-loaded RBCs undergo a shape change, with an increased fraction of echinocytes and spheroidal RBCs. Ektacytometry and electron microscopy assessment of RBC compressibility reveal a dose-dependent reduction in the deformability of RBCs loaded with Ter119-FIX. In conclusion, RBCs loaded with Ter119-FIX have the potential to serve as prohemostatic agents, but their reduced deformability warrants further engineering of Ter119-FIX to improve the safety profile.

Silica nano/microparticles have generated significant interest for the past decades, emerging as a versatile material with a wide range of applications in photonic crystals, bioimaging, chemical sensors, and catalysis. This study focused on synthesizing silica nano/microparticles ranging from 20 nm to 1.2 μm using the Stöber and modified Stöber methods. The particles exhibited photoluminescence emission across a UV-visible range, specifically in the UV (∼290, ∼327, ∼339, and ∼377 nm), blue (∼450 nm), green (∼500 nm), yellow (∼576 nm), and red (∼634 nm) range of the electromagnetic spectrum. These emissions are due to radiative relaxation processes involving oxygen-deficient centers arising due to unrelaxed oxygen vacancies, strong interacting surface silanols, 2-fold coordinated silicon, self-trapped excitons, hydrogen-related species, strain-induced defects, and nonbridging oxygen hole centers excited via two-photon and single photon absorption. The increased PL intensity with a decreasing particle size was attributed to higher concentrations of defect sites in the case of smaller-sized particles. The MTT assay, AO/EB staining, and the DCFDA assay confirmed the biocompatible nature of silica particles in the HepG2 cell line. In addition, the cell viability assay in a normal cell line (HEK293) also showed no substantial cell death. Successful bioimaging of HepG2 cells was performed with silica nano/microparticles, which exhibited blue and green fluorescence, along with Hoechst33258 dye. Even though 20 nm-sized silica particles showed higher PL emission, particles sized above 20 nm showed better fluorescence in HepG2 cells, citing their potential in in vitro bioimaging applications.