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Acute myocardial infarction (MI) remains a leading cause of mortality worldwide, with inflammatory and reparative phases playing critical roles in disease progression. Currently, there is a pressing need for in vivo imaging techniques to monitor immune cell infiltration and inflammation activity during these phases. We developed a novel probe, ^99mTc-HYNIC-mAb_Kv1.3, utilizing a monoclonal antibody that targets the voltage-gated potassium channel 1.3 (Kv1.3). This probe enables in vivo visualization of immune cells that express high levels of Kv1.3 proteins. In a murine MI model, SPECT/CT imaging with ^99mTc-HYNIC-mAb_Kv1.3 demonstrated specific uptake in an infarcted myocardium during the inflammatory phase, reflecting immune cell infiltration and activity. During the reparative phase, the probe exhibited prolonged retention in the infarcted area, suggestive of ongoing immune cell proliferation. Immunofluorescence staining confirmed the probe's specificity. Biodistribution analysis indicated preferential accumulation in the infarcted myocardium and liver, consistent with SPECT/CT findings. Combined with [¹⁸F]FDG PET/CT, these modalities provided comprehensive insights into myocardial viability and inflammation. This study highlights the potential of ^99mTc-HYNIC-mAb_Kv1.3 SPECT/CT as a noninvasive tool to monitor immune cell activity in different phases of MI, guide therapeutic interventions, and predict disease progression. Further translational studies are warranted to explore its clinical applicability in cardiac pathologies.

Numerous diseases, such as diabetic retinopathy and age-related macular degeneration, can lead to retinal neovascularization, which can seriously impair the visual function and potentially result in blindness. The presence of the blood-retina barrier makes it challenging for ocularly administered drugs to penetrate physiological barriers and reach the ocular posterior segments, including the retina and choroid. Herein, we developed an innovative bifunctional peptide, Tat-C-RP7, which exhibits excellent penetration capabilities and antiangiogenic properties aimed at treating retinal neovascularization diseases. RP7 is an NRP-1 targeting peptide that blocks vascular endothelial growth factor receptor-2 (VEGFR-2) signaling and inhibits angiogenesis, while Tat facilitates the delivery of various cargoes across biological barriers, such as the blood-retina barrier. By combining these attributes, Tat-C-RP7 is anticipated to traverse ocular barriers via ocular topical administration and exert its antiangiogenic effects in the ocular posterior segment. Experimental results demonstrated that Tat-C-RP7 significantly inhibited the proliferation and migration of rat retinal microvascular endothelial cells and effectively reduced tubule formation in vitro. Its antiangiogenic activity was confirmed in zebrafish. The outstanding penetrative capabilities of FITC-labeled Tat-C-RP7 have been validated through cell uptake assays, in vitro cell barrier models, ex-vivo ocular tissues, and in vivo studies. Besides, the half-life of Tat-C-RP7 was longer than that of RP7. In an oxygen-induced retinopathy model, Tat-C-RP7 was shown to reduce the area of angiogenesis following ocular administration. Additionally, it produced no irritating effects on the eyes of rabbits. Overall, Tat-C-RP7 demonstrates excellent ocular penetrability and antiangiogenic effects and represents a promising therapeutic option for treating retinal neovascularization diseases.

The exposure of mRNA to water is likely to contribute to the instability of RNA vaccines upon storage under nonfrozen conditions. Using atomistic molecular dynamics (MD) simulations, we investigated the pH-dependent structural transition and water penetration behavior of mRNA-lipid nanoparticles (LNPs) with the compositions of Moderna and Pfizer vaccines against COVID-19 in an aqueous solution. It was revealed that the ionizable lipid (IL) membranes of LNPs were extremely sensitive to pH, and the increased acidity could cause a rapid membrane collapse and hydration swelling of LNP, confirming the high releasing efficiency of both LNP vaccines. The free energy profiles of water penetration showed that the conical structure of IL played a key role in obstructing water from entering the inner core of LNPs: the molecular geometry with more tail chains, lower linearity, and looser packing structure resulted in higher water permeability, leading to lower stability in nonfrozen liquid environment. On the other hand, the geometry of IL also dominated the fusion behavior of LNP with endosomal membrane during the endosomal escape. Thus, for LNP-based vaccines with both high release efficiency and high stability, a suitable molecular structure of ILs should be selected to seek a balance between the packing tightness and fusion rate of membranes.

Cytochalasins are notable for their structural diversity and broad range of biological activities. The gene cluster responsible for the biosynthesis of cytochalasins bearing diverse polycycles was identified in Phomopsis sp. DHS-48. Characterization of the cluster indicated that only the 5/6/11-tricycle can be biosynthetically produced. The chemical space of cytochalasins was expanded by acid-mediated intramolecular cyclo-rearrangement of the 5/6/11-tricycle, and three new cytochalasins, phomoparagins D-F (13, 17, and 18, respectively), with diversified polycycles were obtained, among which compound 13 featured an unprecedented 5/6/5/7/6-pentacycle. Their structures and absolute configurations were established by spectroscopic analysis (1D, 2D NMR), electronic circular dichroism calculations, and a single-crystal X-ray diffraction experiment. The compounds inhibited the growth of HeLa and RKO cell lines with IC₅₀ values ranging from 0.8 to 47.3 μM. Cytoskeleton staining experiments and molecular docking models revealed that compound 13 showed cytotoxicity by targeting F-actin.

Antibody therapy has become a mature cancer treatment strategy, but only one antibody drug, bevacizumab (BEV) has been approved to treat glioblastoma (GBM). The natural blood-brain barrier (BBB) significantly limits the penetration of therapeutic antibodies into the brain. In this study, an antibody delivery platform based on exosomes (EXOs) has been developed, which can cross the BBB and effectively enter the brain tissue to deliver BEV for safe and effective GBM therapy. In vitro experiments have shown that EXO-BEV could efficiently penetrate the BBB and significantly inhibit the migration of endothelial cells. Biodistribution studies in vivo have revealed that EXO serves as an effective carrier for transporting a higher concentration of BEV across the BBB into the brain. Furthermore, in vivo antiglioma experiments have illustrated that the introduction of EXO-BEV into the brain can improve the degeneration of pathological tissues, increase the apoptosis of tumor cells, and significantly extend the survival time of the model animals. All of the results suggested that EXO-BEV could cross the BBB, thereby enhancing the apoptosis of tumor cells and mitigating angiogenesis in GBM. In conclusion, this innovative platform for antibody delivery emerges as a highly promising therapeutic strategy for the clinical treatment of GBM and other neurological disorders.

Fibroblast activation protein inhibitors (FAPIs) labeled with gallium-68 and lutetium-177 show potential for use in the diagnosis and treatment of various cancers expressing FAP. However, ¹⁷⁷Lu-labeled FAPIs often exhibit short tumor retention time, limiting their therapeutic applications. To improve tumor retention, we synthesized three radiolabeled dimeric FAPIs, [¹⁸F]1, [⁶⁴Cu]2, and [⁶⁸Ga]3. These were prepared by chelating Al[¹⁸F]F to 1,4,7-triazacyclononane-1,4,7-triacetic acid (NOTA)-l-glutamic acid (E)-(FAPI)₂ and copper-64 or gallium-68 to 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA)-E-(FAPI)₂. NOTA-E-(FAPI)₂ and DOTA-E-(FAPI)₂ showed higher binding affinities for FAP compared with that of FAPI-04 (IC₅₀ = 0.47 and 0.16 nM vs 0.89 nM, respectively). All radioligands were synthesized in high decay-corrected radiochemical yields (59-96%) and were stable in fetal bovine serum and phosphate-buffered saline. The more hydrophilic radioligand, [⁶⁸Ga]3, was selected for cellular uptake studies, which confirmed FAP-specific uptake. Positron emission tomography imaging and ex vivo biodistribution studies in U87MG tumor-bearing mice revealed high tumor uptake of all three radioligands, with significant blocking observed after preinjection of FAPI-04. [⁶⁴Cu]2 and [⁶⁸Ga]3 exhibited favorable in vivo pharmacokinetics compared to those of [¹⁸F]1. Notably, [⁶⁸Ga]3 showed lower normal organ uptake than did the other two radioligands, and moreover, it exhibited higher, more prolonged tumor uptake than its monomeric counterpart [⁶⁸Ga]Ga-FAPI-04 over a 3 h period, suggesting its potential as a promising FAP-specific theranostic radioligand.

Therapeutic monoclonal antibody (mAb) drug products are increasingly used to treat both chronic and acute diseases. These mAb drug products are often developed for subcutaneous (SC) injection to simplify dosing compared with intravenous (IV) infusion. For SC injection, the mAb liquid drug product is typically filled in a vial for use with a syringe or in a prefilled syringe, which can then be assembled into a safety syringe device or an autoinjector for direct administration. A placebo is an inert formulation (one without an active ingredient) that lacks pharmacological activity or a therapeutic effect. It serves as a control in blinded clinical trials to evaluate the efficacy of a new treatment. A suitable blinding/matching/masking strategy is crucial to ensure that study participants cannot distinguish between the active mAb formulation and the placebo. The success of these strategies is pivotal in ensuring the accuracy and reliability of clinical trial results. This Review summarizes recent advances and key considerations related to placebo strategies. It covers the benefits and challenges of SC injection of therapeutic mAbs compared to IV infusion, the placebo effect, the significance of blinding/matching/masking, and various strategies. Strategies discussed include the use of traditional placebos (e.g., normal saline, 5% w/v dextrose solution, and formulation buffer of the active mAb), syringe blinding, the use of different gauge syringe needles, novel (custom) placebos, dilution, independent administration, and multiple injections. Additional topics covered include the incidence of antidrug antibodies (ADAs), the benefits and challenges associated with different strategies, and regulatory expectations regarding custom placebos. By addressing these critical aspects, the Review aims to contribute to the growing body of knowledge and ongoing efforts to enhance the effectiveness of formulation blinding, matching, and masking in clinical trials.

Photodynamic therapy (PDT) is increasingly regarded as an attractive approach for cancer treatment due to its advantages of low invasiveness, minimal side effects, and high efficiency. Here, two novel Ru(II) complexes 8a,b were designed and synthesized by coordinating phenanthroline and biquinoline ligands with Ru(II) center, and their chemo-photodynamic therapy and immunotherapy were explored. Both 8a and 8b exhibited significant phototoxicity against A549 and 4T1 tumor cells via type-I/-II PDT. Among them, 8b exhibited superior oxygen-independent antitumor effects (IC₅₀s = 1.50-1.76 μM) upon laser irradiation, and displayed micromolar-level chemotherapeutic activities, indicating its potential for chemo/photodynamic dual effects. Furthermore, 8b also initiated an ICD cascade, enhancing recruitment and maturation of antigen-presenting cells, thus triggering a CD8⁺ T cell antitumor immune response. Finally, in vivo antitumor experiments demonstrated that 8b exhibited significant inhibition of lung and breast tumor growth, with inhibition rates of 94.6% and 97.3%, respectively. Therefore, the Ru(II) complexes we designed, as effective type-I/-II photosensitizers and potential immunoactivators, demonstrate multiple antitumor mechanisms, warranting further study.

Neuronanomedicine harnesses nanoparticle technology for the treatment of neurological disorders. An unavoidable consequence of nanoparticle delivery to biological systems is the formation of a protein corona on the nanoparticle surface. Despite the well-established influence of the protein corona on nanoparticle behavior and fate, as well as FDA approval of neuro-targeted nanotherapeutics, the effect of a physiologically relevant protein corona on nanoparticle-brain cell interactions is insufficiently explored. Indeed, less than 1% of protein corona studies have investigated protein coronas formed in cerebrospinal fluid (CSF), the fluid surrounding the brain. Herein, we utilize two clinically relevant polymeric nanoparticles (PLGA and PLGA-PEG) to evaluate the formation of serum and CSF protein coronas. LC-MS analysis revealed distinct protein compositions, with selective enrichment/depletion profiles. Enhanced association of CSF precoated particles with brain cells demonstrates the importance of selecting physiologically relevant biological fluids to more accurately study protein corona formation and subsequent nanoparticle-cell interactions, paving the way for improved nanoparticle engineering for in vivo applications.

The simulation of antral conditions for estimating drug apparent equilibrium solubility after a high-calorie, high-fat meal is challenging. In this study, (1) we measured the apparent equilibrium solubility of two model lipophilic drugs, ketoconazole and danazol, in antral aspirates collected at various time points after a minced high-calorie, high-fat meal and a glass of water 30 min after initiation of meal administration, and we designated one point estimate for ketoconazole and one point estimate for danazol; (2) we evaluated the usefulness of FeSSGF-V2 and FEDGAS pH = 3 in reproducing the two point estimates; (3) we evaluated potential compositions of FeSSGF-V3 that simulate the pH, the buffer capacity toward both less acidic and more acidic values, and the antral lipid and protein contents with easily accessible, commercially available products, and (4) we identified the most useful composition of FeSSGF-V3 for reproducing the two point estimates. For both model drugs, apparent solubility in FeSSGF-V2 and in FEDGAS pH 3 deviated substantially from the corresponding point estimate. For FeSSGF-V3, hydrochloric acid, acetates, and FEDGASbuffer pH 3 were evaluated for regulating the pH and buffer capacity, FEDGASgel was used for simulating the lipid content, and Régilait skimmed milk powder was used for simulating the protein content. Level III FeSSGF-V3 prepared with hydrochloric acid, 6.1% (w/v) Régilait, and 2.83% (w/v) FEDGASgel, i.e., one-sixth of FEDGASgel concentration in FEDGAS pH 3, was comparatively the most useful medium for point estimating ketoconazole and danazol apparent solubility in antral contents after water administration in the fed state, induced as requested by regulatory authorities in oral drug bioavailability studies. Level III FeSSGF-V3 prepared by using hydrochloric acid as the principal pH controlling species could be useful in the evaluation of food effects on drug absorption with in silico physiologically based biopharmaceutics modeling approaches and, also, with biorelevant in vitro methodologies.