Bioconjugate Chemistry最新文献

Glioblastoma ranks among the most prevalent primary intracranial tumors, characterized by high mortality and poor prognosis. Chemotherapy remains a key treatment strategy for gliomas, though most current drugs suffer from limited efficacy and significant toxicity. This study focuses on a cRGD-siEGFR coupling compound synthesized in a previous stage. Prior research indicated that cRGD-siEGFR molecules exhibited certain targeting and antitumor properties but faced issues of inadequate targeting, low efficacy, and high renal toxicity. To enhance antitumor efficacy and mitigate side effects, a pH-responsive, long-circulating, and highly targeted siRNA delivery system, the cRGD-PEG-siEGFR conjugate, was developed. The targeting, antitumor effects, and biological distribution of cRGD-PEG-siEGFR were examined. The results demonstrated that cRGD-PEG-siEGFR was effectively taken up by αvβ3-positive U87MG cells, specifically silenced EGFR gene expression, and exhibited antitumor effects. In normal physiological conditions, it avoided uptake by normal cells, thereby reducing side effects. Furthermore, in vivo biodistribution experiments revealed that cRGD-PEG-siEGFR, compared to cRGD-siEGFR, significantly decreased renal accumulation and exhibited prolonged circulation. Consequently, cRGD-PEG-siRNA emerges as a promising drug candidate with attributes of long circulation, high targeting, pH responsiveness, and substantial antitumor efficacy.

Pancreatic ductal adenocarcinoma (PDAC) poses a challenge in oncology due to its high lethality and resistance to immunotherapy. Recently, emerging research on the stimulator of interferon gene (STING) pathway offers novel opportunities for immunotherapy. Although STING expression is retained in PDAC cells, the response of PDAC cells to STING agonists remains ineffective. Signal transducer and activator of transcription 3 (STAT3), a downstream pathway of STING, is notably overexpressed in pancreatic cancer and related to tumor survival and immune escape. We observed that inhibiting STAT3 signaling post-STING activation effectively suppressed tumor growth through signal transducer and activator of transcription 1 (STAT1)-mediated apoptosis but led to a potential risk of immune-related adverse events (irAEs). To address this issue, we designed a tumor-penetrating liposome for the codelivery of STING agonist and STAT3 inhibitor. These nanoparticles regulated the STING/STAT3 signaling axis and effectively inhibited the proliferation and survival of tumor. Simultaneously, we found a significant increase in the activation of NK cells and CD8⁺ T cells after treatment, leading to robust innate immunity and adaptive immune response. We highlight the potential of regulating the STING/STAT3 axis as a promising treatment for improving clinical outcomes in PDAC patients.

The incidence of cervical cancer caused by human papillomavirus (HPV) infection has increased in recent years. More than half of all cervical cancer cases are due to HPV16 and HPV18 infection, so HPV16 and HPV18 testing is essential to prevent cervical cancer. HPV testing is mainly carried out in hospitals, but it is subject to time and specialized medical facilities. On the other hand, home self-testing using simple diagnostics would present an attractive alternative due to privacy and flexibility with regard to time and place, provided sufficient sensitivity and specificity can be achieved. In this work, a dual lateral flow assay based on RPA-CRISPR-Cas12a/13a (named RC-LFA) for HPV detection was described. Taking advantage of the cleavage specificity of Cas12a and Cas13a, a CRISPR-Cas12a/Cas13a system was designed to detect HPV16 and HPV18. The lateral flow strip with two test lines was designed to suit the CRISPR-Cas12a/Cas13 system. RC-LFA achieves rapid and simultaneous detection of HPV16 and HPV18 with high specificity and sensitivity (10 copies/μL) in about 40 min from the extraction of nucleic acid to an instrument-free readout. RC-LFA is user-friendly and instrument-free, making it a promising method for HPV self-tests at home.

The immune system plays a critical role in protecting the host against pathogens. However, mechanisms for evading the immune system have evolved in pathogens, altering their surface proteins or causing the expression of enzymes that interfere with the immune response. These strategies cause pathogens to escape detection and destruction by the immune system, thereby inducing severe infections. Thus, there is a critical need to develop new chemical tools to recruit the immune system against evading pathogens. Here, we describe a novel strategy for targeting pathogens, by labeling them with a chimeric agent that comprises a peptide bacterial binder, conjugated to an immune-protein tag that is recognizable by the complement system, thereby recruiting the immune system against the targeted pathogen. The chimeric tag was developed by conjugating the peptide bacterial binder with the C3b complement system activating protein. We showed that the chimeric C3b tag preserved its activity and was able to bind the C5 complement protein with strong binding affinity. Using this approach, we have demonstrated that the chimeric agent was able to eradicate 90% of complement-resistant E. coli bacterial cells. By showing enhancement of complement sensitivity in complement-resistant pathogens, this work demonstrates the basis for a new therapeutic approach for targeting pathogenic bacteria, which could open a new era in the development of selective and effective antimicrobial agents.

Background: The main challenges of conventional chemotherapy lie in its lack of selectivity and specificity, leading to significant side effects. Using a small-molecule drug conjugate (SMDC) ensures specific delivery of a cytotoxic drug to the tumor site by coupling it to a targeting vector. This promising strategy can be applied to neuroendocrine tumors (NETs) by choosing a targeting vector that binds specifically to somatostatin receptor subtype 2 (SSTR2). Additionally, incorporation of a bifunctional chelate into the molecule enables complexation of both diagnostic and therapeutic radionuclides. Thus, it facilitates monitoring of the distribution of the SMDC in the body and allows for the implementation of combination therapy. In our study, we designed eSOMA-DM1, a SMDC combining the SSTR2-targeted octreotate peptide and the cytotoxic agent DM1 via a chelate-bridged linker (N₃–Py–DOTAGA). This approach warrants conjugation of the targeting vector and the drug at opposite sites to avoid undesired steric hindrance effects. Methods: Synthesis of the DM1 moiety (4) involved a three-step synthetic route, followed by the conjugation to the cyclic peptide, N₃–Py–DOTAGA-d-Phe-cyclo[Cys–Tyr-d-Trp–Lys–Thr–Cys]–Thr–OH, through a copper-free click reaction, resulting in eSOMA-DM1. Subsequent labeling with [¹¹¹In]InCl₃ gave a high radiochemical yield and purity. In vitro assessments of eSOMA-DM1 binding, uptake, and internalization were conducted in SSTR2-transfected U2OS cells. Ex vivo biodistribution and fluorescence imaging were performed in H69-tumor bearing mice. Results: eSOMA-DM1 exhibited an IC₅₀ value for SSTR2 similar to the gold standard DOTA-TATE. The uptake of [¹¹¹In]In–eSOMA-DM1 in U2OS.SSTR2 cells was 1.2-fold lower than that of [¹¹¹In]In–DOTA-TATE. Tumor uptake in H69-xenografted mice was higher for [¹¹¹In]In–eSOMA-DM1 at all-time points compared to [¹¹¹In]In–DOTA-TATE. Prolonged blood circulation led to increased accumulation of [¹¹¹In]In–eSOMA-DM1 in highly vascularized tissues, such as the lungs, skin, and heart. Excretion through the kidneys, liver, and spleen was also observed. Conclusion: eSOMA-DM1 is a SMDC developed for NET showing promising characteristics in vitro. However, the in vivo results obtained with [¹¹¹In]In–eSOMA-DM1 suggest the need for adjustments to optimize its distribution.

There is a growing interest in developing novel immune potentiators capable of eliciting a cellular immune response. We tackle this challenge by harnessing the synergistic cross-activation between two innate immune receptors─the nucleotide-binding oligomerization domain-containing protein 2 (NOD2) and Toll-like receptor 7 (TLR7). Herein, we investigate the structure–activity relationship of a series of novel conjugated NOD2/TLR7 agonists incorporating a variety of flexible aliphatic, poly(ethylene glycol)-based and triazole-featuring linkers. Our findings reveal potent immune-enhancing properties of conjugates in human primary peripheral blood mononuclear cells, characterized by a Th1/Th17 polarized cytokine response. Importantly, we demonstrate that both the chemistry of the linker and the site of linkage affect the immune fingerprint and the kinetic solubility of these conjugated agonists. These results shed further light on the immunostimulatory potential of NOD2/TLR7 cross-activation and provide insights for designing innovative immune potentiators.

The Cu-catalyzed azide–alkyne cycloaddition (CuAAC) reaction is a key ligation tool used to prepare bioconjugates. Despite the widespread utility of CuAAC to produce discrete 1,4-triazole products, the requirement of a Cu catalyst can result in oxidative damage to these products. Ynamines are superior reactive groups in CuAAC reactions and require lower Cu loadings to produce 1,4-triazole products. This study discloses a strategy to identify optimal reaction conditions for the formation of oligodeoxyribonucleotide (ODN) bioconjugates. First, the surveying of reaction conditions identified that the ratio of Cu to the choice of reductant (i.e., either sodium ascorbate or glutathione) influences the reaction kinetics and the rate of degradation of bioconjugate products. Second, optimized conditions were used to prepare a variety of ODN-tagged products and ODN-protein conjugates and compared to conventional CuAAC and Cu-free azide–alkyne (3 + 2)cycloadditions (SPAAC), with ynamine-based examples being faster in all cases. The reaction optimization platform established in this study provides the basis for its wider utility to prepare CuAAC-based bioconjugates with lower Cu loadings while maintaining fast reaction kinetics.

Inflammation within the brain is a hallmark of a wide range of brain diseases. The complex role of inflammatory processes in these conditions suggests that neuroinflammation could be a valuable therapeutic target. While several promising anti-inflammatory agents have been identified, their clinical application in brain diseases is often hampered by the inability to cross the blood-brain barrier (BBB) and reach therapeutically effective concentrations at the pathological sites. This limitation highlights the urgent need for effective BBB-penetrating drug delivery systems designed to target brain inflammation. This review critically examines the recent advances over the past five years in drug delivery strategies aimed at mitigating brain inflammation in Alzheimer’s disease and ischemic stroke─two of the leading causes of death and disability worldwide. Additionally, we address the key challenges in this field, offering insights into future directions for targeting neuroinflammation in the treatment of brain diseases.

Prodrug-type oligonucleotides (prodrug-ONs) are a class of oligonucleotide designed for activation under specific intracellular conditions or external stimuli. Prodrug-ONs can be activated in the target tissues or cells, thereby reducing the risk of adverse effects. In this study, we synthesized prodrug-type oligodeoxynucleotides activated by β-galactosidase, an enzyme that is overexpressed in cancer and senescent cells. These oligodeoxynucleotides (ODNs) contain a modified thymidine conjugated with galactose via a self-immolative linker at the O4-position. UV-melting analysis revealed that the modifications decreased the melting temperature (T_m) compared with that of the unmodified ODN when hybridized with complementary RNA. Furthermore, cleavage of the glycosidic bond by β-galactosidase resulted in the spontaneous removal of the linker from the nucleobase moiety, generating unmodified ODNs. Additionally, the introduction of multiple modified thymidines into ODNs completely inhibited the RNase H-mediated cleavage of complementary RNA. These findings suggest the possibility of developing prodrug-ONs, which are specifically activated in cancer cells or senescent cells with high β-galactosidase expression.

Accurate detection, treatment, and imaging of diseases are important for effective treatment outcomes in patients. In this regard, bubbles have gained much attention, due to their versatility. Bubbles usually 1 nm to 10 μm in size can be produced and loaded with a variety of lipids, polymers, proteins, and therapeutic and imaging agents. This review details the different production and loading methods for bubbles, for imaging and treatment of diseases/conditions such as cancer, tumor angiogenesis, thrombosis, and inflammation. Bubbles can also be used for perfusion measurements, important for diagnostic and therapeutic decision making in cardiac disease. The different factors important in the stability of bubbles and the different techniques for characterizing their physical and chemical properties are explained, for developing bubbles with advanced therapeutic and imaging features. Hence, the review provides important insights for researchers studying bubbles for biomedical applications.