Bioconjugate Chemistry最新文献

Laryngeal squamous cell carcinoma (LSCC), a predominant subtype of head and neck squamous cell carcinoma (HNSCC), exhibits notably high incidence and mortality rates worldwide. Despite the common use of surgery and radiation, patients with advanced or metastatic disease often have poor 5-year survival outcomes. Hence, there is a strong necessity to devise new treatments for intervention purposes. Polyphenolic compounds, such as quercetin (Que), have shown promise in cancer treatment, but their clinical application is hindered by their low solubility and bioavailability. In this study, we successfully synthesized a novel class of carbon dots (CDs) utilizing Que molecules as precursors through a one-pot hydrothermal method, resulting in marked enhancements in solubility and bioavailability. The Que-CDs created demonstrated significant impacts on stopping the growth, migration, and invasion of TU686 cells, while also encouraging cell cycle arrest and apoptosis. Transcriptomics analysis further revealed alterations in cell cycle regulation and apoptosis-related pathways. Importantly, in vivo experiments validated the antitumor efficacy of Que-CDs without causing damage to vital organs. These findings suggest that Que-CDs represent a safe and efficacious anticancer therapy for laryngeal cancer, meriting further investigation to explore their potential in clinical applications.

Mitochondrial dysregulation, represented by both imbalanced mitochondrial dynamics and dysfunction, has been found as a key driver of cell transformation and tumorigenesis due to enhanced apoptotic priming and genotoxic stress. The mitochondrial unfolded protein response (UPR^mt) represents a protective mechanism that maintains mitochondrial function under mitochondrial damage, making it an attractive target for restoring mitochondrial homeostasis and preventing tumorigenesis. Here, we report an ultrasmall glutathione (GSH)-protected gold nanoparticle (GGNP) that exhibits mitochondrial presence. When mitochondria are damaged by various genotoxic insults, GGNP dramatically activates UPR^mt and improves mitochondrial function without altering mitochondrial dynamics. As a result, GGNP significantly attenuates DNA damage and apoptosis, leading to the prevention of malignant transformation in vitro. More importantly, in a spontaneous lung cancer model, GGNP significantly delays tumorigenesis with reduced DNA damage and cell death within lung tissue without causing systemic toxicity. These findings not only reveal the role of UPR^mt in tumorigenesis but also identify GGNP as a biocompatible nanomaterial that effectively modulates UPR^mt to alleviate mitochondrial stress responses and thus acts as a broad-spectrum genotoxicity mitigator to offer a promising strategy for cancer prevention.

A fluorinated, disulfide-cross-linked polyplex platform (PFND) was developed for safe and potent cytosolic delivery of siRNA. Branched PEI (25 kDa) was first perfluoro-acylated to yield a membrane-zipper PF backbone, followed by orthogonal installation of azide (PF-N₃) and strained alkyne (PF-DBCO) handles that undergo in situ copper-free click cross-linking in the presence of siRNA. The resulting 60 nm polyplexes (PDI < 0.1, ζ potential of approximately +22 mV) are stable in 10 mg mL^–1 heparin (<5% siRNA leakage) yet quantitatively disassemble within 60 min in 10 mM glutathione, liberating the siRNA payloads. Compared with the commercial gold standard of Lipofectamine 3000, PFND delivers 2- to 3-fold more Cy5-siRNA into HeLa, HepG2, and MDA-MB-468 cells without detectable hemolysis or cytotoxicity. Consequently, 10 nM siGAPDH delivered by PFND silences approximately 93–98% of GAPDH mRNA across the three lines, remarkably outperforming gold-standard transfection reagents. The proposed reversible “locked-outside/labile-inside” design reconciles extracellular stability with rapid intracellular release, offering a valid tool for utilities of high-throughput siRNA screening or subject to be developed further for potential clinical translation of RNAi-based therapeutics.

Redirecting endogenous antibodies toward tumor cells through rationally designed antibody-recruiting molecules (ARMs) has emerged as a promising strategy in cancer immunotherapy. However, current ARMs face inherent limitations, as they primarily depend on hapten-specific antibodies, which exhibit heterogeneity across populations and exist at suboptimal physiological concentrations. In this study, we explored the feasibility of leveraging Hepatitis B virus (HBV) vaccine-induced anti-HBV surface antigen (HBsAg) antibodies for cancer therapy. We developed a series of nanobody-peptide conjugates comprising an EGFR-targeting nanobody covalently linked to the LOOP2 peptide─an immunodominant epitope of HBsAg, through varying-length PEG spacers. The results demonstrated that these conjugates were capable of recruiting vaccine-induced anti-HBsAg antibodies onto the cancer cell surface and evoking potent antibody-dependent cell-mediated phagocytosis (ADCP) and complement-dependent cytotoxicity (CDC) for cell elimination. Interestingly, structure–activity relationship studies revealed that the PEG spacer had a minimal impact on ADCP efficacy, while it significantly affected CDC function. This proof-of-concept study establishes a novel paradigm for ARM-based therapeutics that leverages population-wide immunity from routine vaccination programs, thereby circumventing key limitations associated with the hapten-dependent system.

The preparation of antibody drug conjugates (ADC) most often relies on a linear sequence to elaborate the small molecule component, followed by a final bioconjugation step to attach it to its immunoglobulin partner. This linear and iterative approach is incompatible with expedient parallel synthesis and process automation. Here, we describe the design and implementation of a general modular platform for the assembly of ADCs that enables facile variation in the nature of the payload, the linker composition, and the type of bioconjugation technique used. A library of antibody conjugates bringing together several different antibodies and payloads was prepared in a convergent fashion using a range of conjugation methods, as well as cleavable or noncleavable linker technology. Aside from offering a direct comparison of different conjugation method performances, this approach enables a more targeted optimization strategy of conjugate properties by deconvoluting bioconjugation and payload attachment.

Secondary Cerenkov-induced fluorescence imaging (SCIFI) utilizes blue-weighted Cerenkov luminescence from radioactive decay to excite proximal fluorophores that emit near-infrared light with optimal penetrance through biological tissues and offers potential utility in clinical imaging applications, including guidance of surgical resection. Recently, we developed a self-excitatory immunoSCIFI probe based on an antibody modified with the Cerenkov luminescence generating radioisotope zirconium-89 and a near-infrared boron-dipyrromethene dye (BOD665) and observed an immunoSCIFI signal in in vitro cell-based experiments. In this study, we have evaluated the in vivo application of immunoSCIFI using a clinically relevant orthotopic mouse model of dedifferentiated chondrosarcoma as a reproducible, high contrast setting in which to challenge the optical method under bone and soft tissue attenuation. Herein, we report the synthesis, characterization, preclinical imaging, and ex vivo biodistribution analysis of a novel immunoSCIFI probe, [⁸⁹Zr]Zr-DFO-MT1-MMP-BOD665, based on a murine monoclonal immunoglobulin G (IgG) with high binding specificity for the sarcoma biomarker MT1-MMP. Both in vivo imaging and ex vivo data indicated significantly higher total uptake and femur-to-muscle ratios in the inoculated femurs with high MT1-MMP expression relative to contralateral femurs. These preliminary findings establish that antibody-mediated SCIFI can operate in vivo with favorable signal-to-background performance under physiologically relevant photon attenuation. The study therefore provides a methodological foundation for future SCIFI probes, for which rigorous specificity testing and broader biomarker panels will be pursued separately.

Prostate-specific membrane antigen (PSMA) is a critical biomarker for prostate cancer. While current radiotracers like [⁶⁸Ga]Ga-PSMA-11 are clinically useful, enhancing tumor uptake and contrast remains a goal. Capitalizing on PSMA's enzymatic hydrolysis of folate polyglutamates, we report the design, synthesis, and biological evaluation of a novel heterobivalent PSMA-targeted radiotracer, [⁶⁸Ga]Ga-FA-PSMA. In vitro experiments revealed a significantly enhanced binding affinity of FA-PSMA for human PSMA (K_D = 3.268 nM) compared to PSMA-11 (K_D = 47.88 nM) and free FA (K_D = 424.3 nM). In vivo micro-PET/CT imaging and biodistribution studies in PSMA-positive (22RV1) xenograft models demonstrated superior performance over [⁶⁸Ga]Ga-PSMA-11. [⁶⁸Ga]Ga-FA-PSMA exhibited significantly higher peak tumor SUV_max (1.06 ± 0.09 vs 0.80 ± 0.08 at 210 min p.i.) and tumor accumulation (2.74 ± 0.40 vs 1.75 ± 0.74 %ID/g at 2 h p.i.), alongside significantly enhanced tumor-to-background ratios, particularly tumor-to-blood (10.41 ± 3.41 vs 2.11 ± 0.89) and tumor-to-muscle (10.07 ± 0.73 vs 6.03 ± 1.33) at 2 h p.i. Furthermore, [⁶⁸Ga]Ga-FA-PSMA demonstrated reduced renal uptake (30.80 ± 8.54 vs 38.71 ± 9.72 %ID/g at 2 h p.i.), high in vitro stability, and favorable hydrophilicity (LogP = -3.76). These findings demonstrate that the novel heterobivalent tracer [⁶⁸Ga]Ga-FA-PSMA offers significantly enhanced PSMA-targeting affinity, tumor uptake, and imaging contrast compared to the clinical standard, validating the FA-conjugation strategy and positioning it as a highly promising next-generation diagnostic agent for precise prostate cancer imaging.

Small-molecule immune modulators offer a promising alternative to biologics, such as antibodies, for cancer immunotherapy. A key example is the TLR7 agonist imiquimod (IMQ), which has already been approved for the treatment of various dermatological malignancies. Nevertheless, the clinical use of IMQ is limited to tumors amenable to topical application, as its systemic administration poses a high risk of severe inflammatory toxicity due to the widespread expression of TLRs. Therefore, to extend the use of TLR7 agonists to the treatment of other solid tumor types, we developed a β-glucuronidase-responsive albumin-binding prodrug designed for the selective delivery of IMQ within the tumor microenvironment. This prodrug masks IMQ’s immunogenicity, allowing for its administration in immunocompetent mice without eliciting the systemic side effects associated with TLR7 agonists. However, the β-glucuronidase-catalyzed prodrug activation enables the selective, tumor site-specific release of IMQ, thereby restoring its biological activities. This controlled delivery promotes M1 macrophage polarization, T cell activation, and an increase in IgG levels exclusively within malignant tissues without affecting the healthy organs that are sensitive to TLR7 agonists. This study demonstrates that targeting tumor microenvironment specificities represents a promising approach for developing selective cancer immunotherapies based on small-molecule immune modulators.

The design of effective vaccines against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) relies on eliciting robust cellular and humoral immune responses. In this study, we report the synthesis and immunological evaluation of synthetic long peptide (SLP) vaccines incorporating immunodominant CD4⁺ and CD8⁺ T cell and B cell epitopes from the SARS-CoV-2 proteome, covalently conjugated to stereochemically defined UPam ligands, which serve as synthetic Toll-like receptor 2 (TLR2) agonists. Using solid-phase peptide synthesis, we generated a panel of R- and S-glycerol-based UPam conjugates with varied amino acids adjacent to the palmitoylated cysteine. These conjugates were evaluated for their ability to induce dendritic cell maturation, TLR2 activation, and antigen-specific immune responses. In vitro assays demonstrated that R-UPam ligands, particularly those containing Dab and Abu residues, were superior in stimulating IL-12p40 production and TLR2 activation compared with their S-epimer counterparts. In vivo immunization studies in mice revealed that covalent conjugation of UPam to CD8⁺ T cell epitopes (VNF and REL) significantly enhanced antigen-specific T cell responses compared to unconjugated mixtures. Furthermore, a UPam-conjugated B cell epitope fused to a universal helper T cell peptide (PADRE) induced strong dendritic cell activation and enhanced PADRE-specific CD4⁺ T and B cell responses, as evidenced by elevated IgG production. These findings underscore the potential of chirally pure UPam-conjugated peptide vaccines to induce balanced, durable immunity against SARS-CoV-2 by combining TLR2-mediated adjuvanticity with precise epitope targeting.