Molecular Pharmaceutics最新文献

Onychomycosis is a common fungal nail infection, causing nail thickening and discoloration. Tavaborole, a topical antifungal, has fewer side effects but requires long treatment periods and often results in low cure rates. In this study, we developed a Zn²⁺-driven tavaborole-adenosine (AT-Zn²⁺) hydrogel to improve its therapeutic effect. The hydrogel enhanced tavaborole's solubility, drug loading, and antifungal activity. Characterization by nuclear magnetic resonance (NMR), ultraviolet-visible (UV-vis) spectroscopy, and transmission electron microscopy (TEM) confirmed its successful synthesis and nanofiber structure. In vitro release tests showed that about 65% of tavaborole was released in PBS at pH 5.5 over 24 h, indicating pH-sensitive drug release for targeted therapy. Permeation studies using a bovine hoof model showed effective tavaborole penetration through keratinized tissues with a steady-state flux of 162 μg/cm²/h. The AT-Zn²⁺ hydrogel demonstrated lower minimum inhibitory concentrations (MICs) for C. albicans (0.00156 mM) and A. fumigatus (0.025 mM) compared to those of tavaborole alone. In a bovine onychomycosis model, the hydrogel showed stronger antifungal effects than the tavaborole solution. Cytotoxicity assays on RAW 264.7 cells indicated good biocompatibility with >85% cell viability. These findings suggest that the AT-Zn²⁺ hydrogel holds significant potential as a clinically effective antifungal agent.

Anti-CD38 antibodies (Abs) are promising immunotherapeutics for hematologic malignancies, but their efficacy in leukemias often requires pharmacologic upregulation of CD38. Immuno-PET provides a noninvasive strategy to evaluate such target modulation in vivo. Cysteine site-specific ⁸⁹Zr labeling of anti-CD38 Abs was performed using deferoxamine-maleimide. CD38-specific target binding was confirmed in three human myeloma and two leukemia cell lines. Total and surface expressed CD38 levels were assessed by Western blotting and flow cytometry. Immuno-PET imaging and biodistribution studies were conducted in murine leukemia models. All-trans retinoic acid (ATRA) was used to stimulate CD38 expression. Myeloma and MOLT4 leukemia cells showed variable baseline CD38, while HL60 cells exhibited negligible levels. All tumor cells demonstrated ⁸⁹Zr-OKT10 IgG and ⁸⁹Zr-daratumumab (Fc-silenced) binding that paralleled surface CD38 expression. ATRA upregulated CD38 in all tested cells, including a marked induction in HL60 cells, all accompanied by corresponding elevations in ⁸⁹Zr-CD38 Ab binding. On ⁸⁹Zr-OKT10 IgG PET, MOLT4 tumors showed high uptake that was reduced by 67.9% with unlabeled Ab, but HL60 tumors showed low uptake and high liver accumulation, limiting ATRA assessment. ⁸⁹Zr-daratumumab produced lower liver uptake and improved MOLT4 and HL60 tumor visualization; ATRA modestly increased MOLT4 tumor uptake and substantially enhanced HL60 tumor uptake from 8.0 ± 1.7 %ID/g to 14.7 ± 3.1 %ID/g (83.8% increase; P < 0.005). Mechanistic studies demonstrated ATRA-induced ERK1/2 activation in HL60 cells that was abolished by the MAPK inhibitor U0126; U0126 also suppressed CD38 induction and ⁸⁹Zr-CD38 Ab uptakes in all tested tumor cells. Furthermore, U0126 blocked ATRA-induced HL60 tumor ⁸⁹Zr-daratumumab uptake in vivo. Western blots and immunohistochemistry confirmed ATRA-induced HL60 tumor CD38 elevation, which was partly reversed by U0126. Thus, ⁸⁹Zr immuno-PET enables noninvasive monitoring of ATRA-driven CD38 upregulation via MAPK signaling and supports its potential utility for optimizing combination strategies in leukemias.

Solid dispersion is a widely adopted formulation strategy to enhance the solubility of water-insoluble drugs. However, the molecular-level structural determinants of stability and dissolution behavior remain poorly understood. This study integrates Small-Angle Neutron Scattering (SANS) technology with coarse-grained molecular dynamics (CGMD) simulations to investigate the effects of preparation methods (melting vs solvent evaporation) and drug loadings (10%, 15%, 25%) on the microstructure and crystallinity of PXM-PEG solid dispersions. Deuterated PEG (d-PEG) is used in the SANS to enhance the scattering intensity in samples. The findings revealed that the lamellar thickness decreased significantly from 173.01 Å (pure d-PEG) to 44.12 Å (25% drug loading, melting method), while the d-spacing reduced from 71.13 to 36.65 Å, indicating a substantial disruption of the crystalline structure. Conversely, samples prepared by solvent evaporation maintained larger d-spacing (up to 93.27 Å at 10% drug loading) and more stable layer stacking (Nlayers ∼5.6), demonstrating higher structural order. The results indicate that the preparation method significantly influences the structural characteristics of the solid dispersions. The melting method yielded a higher amorphous content at low drug loadings, which is expected to improve drug solubility and bioavailability. In contrast, the solvent evaporation method tended to produce solid dispersions with higher crystallinity and uniform structures at higher drug loadings. SANS results indicated that samples prepared by the melting method exhibited higher disorder in the high-q region, while those prepared by the solvent evaporation method showed greater crystallinity. The CGMD simulations further elucidated the dynamic aggregation and structural formation of the drug and polymer molecules during the preparation process. In the melting simulations, drug and polymer molecules gradually aggregated into dense clusters, while in the solvent evaporation simulations, the aggregates grew larger and more asymmetrical as the solvent evaporated, ultimately forming ordered structures. The combined results from SANS and molecular dynamics simulations indicated the "sandwich-like" structure of PXM-PEG solid dispersions. The outcomes of this innovative approach have the potential to advance the development of solid dispersion formulations, enhance research and development efficiency, and pave the way for the industrial production of solid dispersions.

In this study, a novel fibroblast activation protein (FAP) targeting ligand based on the bivalent DPro-Gly structure, DOTA-PG2-2FAPI, was designed and synthesized. Molecular docking analysis revealed that compared with monomeric FAPI-46, DOTA-PG2-2FAPI had a higher binding score (-15.00 vs -11.36), with an in vitro IC₅₀ value of 6.65 nM, indicating a significantly increased binding affinity toward FAP. The ⁶⁸Ga-labeled complex ([⁶⁸Ga]Ga-PG2-2FAPI) demonstrated radiochemical purity exceeding 95% with good stability and high hydrophilicity (LogD_7.4 = -3.29 ± 0.06). In HT-1080-FAP cells, the cellular uptake of [⁶⁸Ga]Ga-PG2-2FAPI reached 10.98 ± 0.10% ID, which decreased by 92% upon FAP inhibition. In vivo studies using tumor-bearing mice revealed that the tumor uptake of [⁶⁸Ga]Ga-PG2-2FAPI was 17.60 ± 1.33% ID/g in HT-1080-FAP tumors and 32.71 ± 0.98% ID/g in U87MG tumors, which was significantly greater than that in nontargeted tissues. Positron emission tomography (PET) imaging revealed rapid tumor accumulation and sustained retention, with high-specificity imaging across all four tumor models (HT-1080-FAP, U87MG, HT-29, and PANC-1). On the basis of these characteristics, this probe holds promise as a broad-spectrum tumor imaging agent with significant clinical application value.

Atherosclerotic cardiovascular disease (ASCVD) poses a severe threat to human health, and the global prevalence of atherosclerosis-related diseases continues to rise, necessitating urgent exploration of novel strategies. Inspired by the close links between tumors and atherosclerosis (AS), as well as the clinical reality of their comorbidity, the present study encapsulated pitavastatin within liposomes and modified them with sialic acid-cholesterol (SA-CH) to achieve targeted drug delivery via peripheral blood neutrophils (PBNs). Compared to oral pitavastatin administration, sialic acid-modified pitavastatin liposomes (PIT-SAL) demonstrated superior efficacy in attenuating disease progression in atherosclerotic mice, with sustained therapeutic effects even after treatment cessation, suggesting the potential for eradication of AS. Notably, PIT-SAL additionally exhibited antitumor potential by effectively reducing tumoral cholesterol accumulation while enhancing T-cell infiltration. Collectively, our preliminary findings highlight the great translational potential of PIT-SAL as a targeted therapy for both AS and tumors, offering a potential breakthrough in managing these interconnected diseases.

Lipid nanoparticles (LNPs) have emerged as a leading platform for mRNA delivery; however, their compatibility with prefilled syringes (PFS) containing silicone oil (SO) as a lubricant remains unexplored. This study investigated the effects of SO on the physicochemical stability and biological activity of mRNA-LNPs under various storage conditions (4 °C, 25 °C, and light exposure). Using polyadenylic acid (Poly A) and enhanced green fluorescent protein-encoding mRNA (eGFP-mRNA) as models, we evaluated particle size, polydispersity index (PDI), zeta potential, encapsulation efficiency (EE%), and transfection efficacy. The results showed that Poly A-LNPs exhibited significant particle size increases and PDI changes at 25 °C with SO, whereas eGFP-LNPs maintained stability under the same conditions, probably because of the mRNA secondary structure enhancing colloidal stability. At 4 °C, both formulations remained stable for 12 weeks, but long-term storage led to a gradual EE reduction. Under light exposure, eGFP-LNPs retained a high EE but suffered severe mRNA degradation, resulting in a near-complete loss of transfection activity. Notably, SO partially mitigated light-induced damage, improving transfection efficiency by up to 6-fold in 100 ppm (ppm) SO-spiked samples. These findings reveal mRNA-dependent LNP-SO interactions and underscore the necessity of evaluating both the physicochemical and functional stability of PFS-based mRNA-LNP formulations.

Lipid-mRNA adducts form during storage for several types of lipid nanoparticles (LNPs) and impair therapeutic efficacy, yet their structural drivers and functional consequences remain incompletely characterized, especially for novel lipids with distinct structures. Here, we investigated adduct formation between mRNA and several impurities derived from an immunotropic ionizable lipid (4S)-KEL12, which has been used to develop therapeutic mRNA cancer vaccines approved for human clinical studies. Elevated storage temperatures promoted both adduct accumulation and the loss of mRNA integrity with divergent kinetics at 25 °C, suggesting their independence. Mechanistically, degradation impurities of (4S)-KEL12, particularly its aldehyde derivative (Z4) and N-oxide derivative (Z1) dominated adduct generation, with Z4 exhibiting ∼4-fold higher activity than Z1. Moreover, mRNA adduction with Z4 did not reduce mRNA integrity by capillary electrophoresis, further supporting independent pathways. Mass spectrometry characterization unambiguously identified cytidines as the primary target on mRNA for Z4 adduction. Functionally, while adducted mRNAs exhibited poor capacity for protein expression in cultured human 293T cells, they did not stimulate significant gene expression involved in innate immunity for RNA sensing and downstream type I interferon pathway activation in human THP1 cells. These findings not only clarify important functional consequences of adducted mRNAs, but also establish impurity control and thermal management as actionable strategies for advancing mRNA therapeutics.

mRNA therapy has shown great potential in vaccine development, cancer treatment, and the treatment of rare diseases. Lipid nanoparticles (LNPs) are key delivery carriers that are essential to the success of mRNA therapy. Here, we found that a low-glucose microenvironment affected the efficiency of LNP-mediated mRNA delivery. Two LNPs (ALC-0315@LNP and SM-102@LNP) were tested in three types of cells under different glucose conditions. The results showed that low-glucose levels significantly reduced the translation of LNP-delivered mRNA into protein, and this negative effect was reversible upon the restoration of glucose levels. A mouse tumor model further confirmed that hypoglycemia diminished the in vivo mRNA delivery efficiency of LNPs. Further mechanistic studies revealed that the reduced efficiency was not due to impaired cellular uptake or lysosomal escape of LNPs, but rather to disrupted glucose energy metabolism. Under low-glucose conditions, cellular ATP and GTP levels were reduced, directly inhibiting the mRNA translation process, which is dependent on these high-energy molecules. This study systematically revealed for the first time that low-glucose conditions reduced mRNA-LNP delivery efficiency by impairing cellular energy metabolism. These findings provide insights for designing metabolic-microenvironment-adapted mRNA therapies and offer strategies to improve mRNA therapy efficacy in treating ischemic stroke and cancer patients.

Polysorbate 80 (PS80) is a commonly used surfactant for stabilizing biotherapeutics by preventing protein adsorption at the air-liquid interface. However, PS80 is susceptible to oxidative degradation during manufacturing and storage. We show here that light exposure combined with the presence of metals can result in byproduct formation and potentially decrease the surfactant's ability to prevent protein adsorption to the air-liquid interface. PS80 formulated in citrate buffer can undergo cis/trans isomerization of unsaturated fatty acids in the presence of disulfides and iron (Prajapati et al., 2022). This work investigates novel surface activity aspects of polysorbate formulations before and after exposure to UV-A light. Polysorbate samples of different grades were formulated in citrate buffer containing iron and glutathione disulfide (GSSG; as a surrogate for peptide and protein disulfides), and a Langmuir trough was used to monitor the surface pressure during adsorption to the air-solution interface. Our results showed significant changes in the polysorbate surface activity after photoirradiation: all-oleate PS80 exhibited a 3-fold increase in the apparent critical micelle concentration (CMC), and the presence of both cis and trans fatty acids was confirmed. Also, the impact of photoirradiation on surface pressure depended on the surfactant concentration during irradiation, suggesting that the presence of micelles can alter the degradation pathway and byproduct formation.