Bioconjugate Chemistry最新文献

Compared with conventional mammography, contrast-enhanced dual-energy mammography (DEM) can improve tumor detection for people with dense breasts. However, currently available iodine-based contrast agents have several drawbacks such as their contraindication for use with renal insufficiency, high-dose requirement, and suboptimal contrast production. Molybdenum disulfide nanoparticles (MoS₂ NPs) have been shown to attenuate X-rays due to molybdenum’s relatively high atomic number while having good biocompatibility. However, work exploring their use as X-ray contrast agents has been limited. In this study, we have developed a novel aqueous synthesis yielding ultrasmall, 2 nm MoS₂ NPs with various small molecule coatings, including glutathione (GSH), penicillamine, and 2-mercaptopropionic acid (2MPA). These nanoparticles were shown to have low in vitro cytotoxicity when tested with various cell lines at concentrations up to 1 mg/mL. For the first time, these particles were shown to generate clinically relevant contrast in DEM. In DEM, MoS₂ NPs generated higher contrast than iopamidol, a commercially available X-ray contrast agent, while also generating substantial contrast in CT. Moreover, MoS₂ NPs demonstrated rapid elimination in vivo, mitigating long-term toxicity concerns. Together, the results presented here suggest the potential utility of MoS₂ NPs as a dual-modality X-ray contrast agent for DEM and CT.

The analysis of protein-bound glycans has gained significant attention due to their pivotal roles in physiological and pathological processes like cell–cell recognition, immune response, and disease progression. Routine methods for glycan analysis are challenged by the very similar physicochemical properties of their carbohydrate components. As an alternative, lectins, which are proteins that specifically bind to glycans, have been integrated into biosensors for glycan detection. However, the effectiveness of protein-based biosensors depends heavily on the immobilization of proteins on the sensor surface. To enhance the sensitivity and/or selectivity of lectin biosensors, it is crucial to immobilize the lectin in an optimal orientation for ligand binding without compromising its function. Random immobilization methods often result in arbitrary orientation and reduced sensitivity. To address this, we explored a directed immobilization strategy relying on a reactive noncanonical amino acid (ncAA) and bioorthogonal chemistry. In this study, we site-specifically incorporated the reactive noncanonical lysine derivative, N^ε-((2-azidoethoxy)carbonyl)-l-lysine, into a cysteine-less single-chain variant of human galectin-1 (scCSGal-1). The reactive bioorthogonal azide group allowed the directed immobilization of the lectin on a biosensor surface using strain-promoted azide–alkyne cycloaddition. Biolayer interferometry data demonstrated that the controlled, directed attachment of scCSGal-1 to the biosensor surface enhanced the binding sensitivity to glycosylated von Willebrand factor by about 12-fold compared to random immobilization. These findings emphasize the importance of controlled protein orientation in biosensor design. They also highlight the power of single site-specific genetic encoding of reactive ncAAs and bioorthogonal chemistry to improve the performance of lectin-based diagnostic tools.

The cellular adhesion receptor αvβ6-integrin is highly expressed in many cancers, e.g., pancreatic, lung, head-and-neck, cervical, bladder, and esophageal carcinoma. Multimerization of αvβ6-integrin-specific RGD peptides increases the target affinity and retention but affects biodistribution and pharmacokinetics. Amide formation of the terminal carboxylic acid moieties of the square-symmetrical bifunctional chelator DOTPI with 3-azidopropylamine yields derivatives with 4, 3, and 2 terminal azides and zero, 1, and 2 remaining carboxylic acids, respectively, whereby formation of the 2-cis-isomer is preferred according to NMR investigation of the Eu(III)-complexes. Cu(II)-catalyzed alkyne–azide cycloaddition (CuAAC) of the alkyne-functionalized αvβ6-integrin binding peptide cyclo[YRGDLAYp(NMe)K(pent-4-ynoic amide)] (Tyr2) yields the respective di-, tri-, and tetrameric conjugates for Lu-177-labeling. In mice bearing αvβ6-integrin-expressing xenografts of H2009 (human lung adenocarcinoma) cells, the Lu-177-labeled trimer’s tumor-to-blood ratio of 112 exceeds that of the tetramer (10.4) and the dimer (54). Co-infusion of gelofusine (succinylated gelatin) reduces the renal uptake of the trimer by 89%, resulting in a 10-fold better tumor-to-kidney ratio, while no improvement of that ratio is observed with arginine/lysine, para-aminohippuric acid (PAH), and hydroxyethyl starch (HES) coinfusions. Since the Lu-177-labeled Tyr2-trimer outperforms the dimer and the tetramer, such trimers are considered the best lead structures for the ongoing development of αvβ6-integrin targeted anticancer theranostics.

Plectin, a scaffolding protein overexpressed in tumor cells, plays a significant role in hepatocellular carcinoma (HCC) proliferation, invasion, and migration. However, the use of L-type peptides for targeting plectin is hindered by their limited stability and retention. We designed a D-type plectin-targeting peptide (^DPTP) and developed a novel single-photon emission computed tomography (SPECT) probe for HCC imaging. The ^DPTP targeting ability was evaluated in vitro using flow cytometry and ex vivo fluorescence imaging. ^99mTc radiolabeling was performed using tricine and ethylenediamine-N,N′-diacetic acid (EDDA) as coligands after modification with 6-hydrazino nicotinamide (HYNIC) at the N termini of ^DPTP. The radiochemical purity (RCP), in vitro stability, and binding affinity of the prepared ^99mTc-HYNIC-^DPTP were analyzed. Tumor uptake, metabolic stability, biodistribution, and pharmacokinetics of ^99mTc-HYNIC-^DPTP were investigated and compared with those of ^99mTc-labeled L-type PTP (^99mTc-HYNIC-PTP) in HCC tumor-bearing mice. ^DPTP could be efficiently radiolabeled with ^99mTc using the HYNIC/tricine/EDDA system with a high RCP and good in vitro stability. Compared with the L-type PTP, ^DPTP exhibited improved targeting ability, and ^99mTc-HYNIC-^DPTP displayed higher tumor uptake, better metabolic stability, longer blood circulation time, and lower kidney retention, resulting in superior imaging performance and biodistribution in vivo. ^99mTc-HYNIC-^DPTP has great potential as a novel SPECT probe for diagnosing HCC.

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.

There have been predictions that the use of the macrocyclic chelating agent 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA) in zirconium-89 (⁸⁹Zr) immuno-positron emission tomography (⁸⁹Zr-immunoPET) could enhance the in vivo stability of ⁸⁹Zr radioimmunoconjugates. However, conjugating [⁸⁹Zr]Zr-DOTA to a monoclonal antibody (mAb) remains a challenge as the heat treatment required for [⁸⁹Zr]Zr-DOTA chelation can lead to thermal denaturation of the mAb moieties. We developed a method for synthesizing [⁸⁹Zr]Zr-DOTA-mAb based on a tetrazine (Tz)-conjugated bifunctional DOTA derivative 2,2',2″-(10-(1-(4-(1,2,4,5-tetrazin-3-yl)phenyl)-3,21,26-trioxo-6,9,12,15,18-pentaoxa-29-carboxy-2,22,25-triazanonacosane-29-yl)-1,4,7,10-tetraazacyclododecane-1,4,7-triyl)triacetic acid (DOTAGA-Tz) and the inverse electron-demand Diels-Alder (IEDDA) click chemistry reaction where trans-cyclooctene-modified mAbs are conjugated to [⁸⁹Zr]Zr-DOTAGA without being exposed to heat. The stability of IEDDA-derived [⁸⁹Zr]Zr-DOTAGA-trastuzumab was confirmed by in vitro, ex vivo, and in vivo testing and comparative analysis against the conventional deferoxamine (DFO) counterpart [⁸⁹Zr]Zr-DFO-trastuzumab. The in vivo immunoPET imaging using [⁸⁹Zr]Zr-DOTAGA-trastuzumab clearly visualized human epidermal growth factor receptor 2-positive malignancies in murine xenograft models. Greater tumor contrast was observed from [⁸⁹Zr]Zr-DOTAGA-trastuzumab at a 72-h delayed scan compared with [⁸⁹Zr]Zr-DFO-trastuzumab. These findings suggest that our IEDDA ligation approach can be an effective means of synthesizing [⁸⁹Zr]Zr-DOTA-mAb and can enhance the theranostic potential of ⁸⁹Zr-immunoPET in DOTA-mediated radioimmunotherapy.

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.

Despite the use of surgical resection and chemotherapy in the clinical treatment of oral squamous cell carcinoma (OSCC), the 5-year survival rates of advanced patients are low. Therefore, more efficient strategies are urgently needed. Herein, a chemo/ferroptosis synergistic therapeutic system-DMEFe nanoparticles (NPs) is established for the treatment of OSCC. To create this system, the chemotherapeutic agent doxorubicin (DOX) was loaded into mesoporous silica nanoparticles and further coated with a pH-sensitive metal polyphenol (iron ion and epigallocatechin gallate). These nanoparticles displayed excellent pH-sensitive drug-control release properties, and the release ratio of DOX at pH 5.5 was twice as high than that at pH 7.4. Additionally, DMEF NPs were effectively taken up by the OSCC cell line SSC-25, which greatly impeded the proliferation of these cells. Notably, these nanoparticles increased the intracellular level of reactive oxygen species and effectively exhibited cytotoxity effects. The mechanistic results proved that DMEFe NPs regulated the expression of ferroptosis-related genes to induce ferroptosis of SSC-25 cells. Eventually, this chemo/ferroptosis therapeutic system exhibited remarkable antitumor effects and provided a novel strategy for the treatment of OSCC.

Because of the insidious nature of lymphatic metastatic cancer, accurate imaging tracing is very difficult to achieve in the clinic. Previous studies have developed the LARGR peptide (named TMVP1) as a radiotracer for vascular endothelial growth factor receptor-3 (VEGFR-3) imaging in cancer. However, its affinity for the target remains insufficient, resulting in low imaging sensitivity. In this study, we identified a high-affinity VEGFR-3 targeting peptide, named TMVP1446, using a multiplex screening platform. TMVP1446 demonstrated a dissociation constant of 8.97 × 10^-8 M. Both in vitro and in vivo assays confirmed that fluorescently labeled TMVP1446 specifically bound to VEGFR-3. In a 4T1-luciferase tumor mouse model, cyanine 7-labeled TMVP1446 effectively discriminated between contralateral normal lymph nodes (c-LN) and cancer-metastatic sentinel lymph nodes (m-SLN). To evaluate the potential of TMVP1446, we developed a novel VEGFR-3 positron emission tomography radiotracer ([⁶⁸Ga]Ga-DOTA-TMVP1446) for cancer-m-SLN imaging. [⁶⁸Ga]Ga-DOTA-TMVP1446 accurately detected and assessed the status of lymph node metastasis, even in micrometastatic tumors, in the B16-F10 mouse tumor model. These findings suggest that TMVP1446 has great potential for advancing VEGFR-3 molecular imaging and metastatic sentinel lymph node imaging.

The peptide-drug conjugate (PDC) has emerged as one of the new approaches for cancer therapy, which has the advantages of improved drug target ability and reduced adverse effects compared with the traditional chemotherapy. CD133 is a surface antigen specific to cancer stem cells, which are thought to be responsible for the self-renewal, proliferation, metastasis, and chemoresistance of cancer cells. A PDC for CD133 was designed by us, and it consists of CD133 targeting peptide LS-7 (amino acid sequence LQNAPRS), a pH-sensitive linker (succinyl), and a cytotoxic payload, the cytotoxic molecule camptothecin (CPT) with potent toxicity in vivo and in vitro. An antitumor study exhibited that the conjugate LS-7-CPT has not only improved its cytotoxicity in tumor cells but also retained its anticancer effect in vivo. In addition, the acute toxicity in mice of LS-7-CPT has been improved and the maximum tolerated dose has been increased by at least 56.2-fold. Pull-down and in vivo fluorescent imaging results indicated that LS-7-CPT was enriched in mice tumors by targeting CD133 protein. As far as we know, this is the first report for a PDC molecule designed for CD133, which is important for the study of CPT drug development.