Molecular Pharmaceutics最新文献

Chemoradiotherapy is a common treatment option for many cancers. Carfilzomib (CFZ) is an effective chemotherapeutic drug with a multitude of cellular effects. However, CFZ has yet to be studied in the context of chemoradiotherapy. To study the application of CFZ in chemoradiotherapy, we synthesized CFZ-loaded liposomes. We report a novel liposomal formulation of the proteasome inhibitor CFZ designed to enhance tumor radiosensitivity while improving drug solubility and tolerability. CFZ-loaded PEGylated liposomes were synthesized via thin-film hydration and probe sonication, achieving an average diameter of ∼127 nm and an encapsulation efficiency of 64%. In murine 4T1 breast carcinoma cells, CFZ treatment prior to irradiation significantly reduced clonogenic survival (dose enhancement factor = 1.26) and increased γ-H2AX foci retention, indicating impaired DNA double-strand break repair. In a dual-flank Balb/cJ allograft model, local intratumoral administration of CFZ followed by ionizing radiation (8 Gy × 2) markedly suppressed primary tumor growth compared with monotherapies without inducing systemic toxicity. Although a strong abscopal effect on distant tumors was not observed, the combination treatment reduced the pulmonary metastatic burden relative to controls. Collectively, these results demonstrate that liposomal carfilzomib can act as an effective radiosensitizer, functioning through perturbation of DNA repair and modulation of the tumor response to radiation. This study highlights a translationally relevant nanotherapeutic approach for enhancing chemoradiotherapy outcomes in solid malignancies.

Dengue virus (DENV), comprising four antigenically distinct serotypes, remains a major global health problem. Vaccine development is hindered by immunological barriers, including antibody-dependent enhancement (ADE), original antigenic sin, and the requirement for balanced, long-lasting immunity against all serotypes. This review focuses on emerging strategies that improve vaccine-induced immune memory through rational antigen design and advanced delivery systems. Nanoparticles, including lipid-based carriers, polymeric particles, and virus-like particles (VLPs), support antigen stability, promote dendritic cell (DC) uptake, and enhance delivery to the lymph node. Injectable hydrogels and responsive biomaterials provide sustained antigen release, promoting germinal center (GC) formation and memory B and T-cell memory responses. Targeted delivery using ligands for C-type lectins or mannose receptors further increases the antigen presentation efficiency. Potent adjuvants, including toll- like receptor (TLR) agonists (Poly I:C, CpG, and R848) and saponin-based molecules (QS-21, MPL), activate innate immune sensors and guide Th1-type adaptive responses. New vaccine formulations, including multiepitope peptide vaccines, mRNA and DNA constructs, and extracellular vesicle (EV)-based carriers derived from DCs or milk, offer cell-free, scalable, and immune system-activating platforms. Furthermore, in silico approaches facilitate epitope identification, MHC-binding prediction, and immune response simulation. Collectively, these strategies address recent challenges and support the development of dengue vaccines that offer enhanced safety and durable immunity.

The targeted delivery of radionuclides and cytotoxic drugs represents a viable alternative to conventional chemotherapy, aiming to improve therapeutic efficacy and reduce systemic toxicity by selective accumulation of the active payload at the tumor site. Our group has developed radioligand therapeutics (RLTs) and small molecule-drug conjugates (SMDCs) targeting fibroblast activation protein (FAP), a tumor-associated antigen abundantly and selectively expressed in the majority of solid human malignancies. Among these, ¹⁷⁷Lu-OncoFAP-23 and OncoFAP-GlyPro-MMAE showed selective accumulation in FAP-positive tumors in murine models and demonstrated potent anticancer activity. To further enhance the therapeutic efficacy, combining targeted drugs with immunotherapy may provide synergistic benefits by engaging both direct tumor cell killing and immune system activation. In this work, we explored the combination of FAP-targeting cytotoxic and radioactive therapeutics with three different immunocytokines targeting the Extra Domain B (EDB) of fibronectin: L19-hIL2, L19-mIL12, and L19-mTNF. A therapy experiment in immunocompetent mice bearing low FAP-expressing tumors showed that the combination with L19-hIL2 potentiated the antitumoral activity of ¹⁷⁷Lu-OncoFAP-23 and OncoFAP-GlyPro-MMAE. These results provided the motivation for the clinical development of these combinations for treating FAP-positive solid tumors.

Despite the revolutionary breakthroughs in immunotherapy for hepatocellular carcinoma, its efficacy remains limited due to low tumor immunogenicity, insufficient immune cell infiltration, and an immunosuppressive tumor microenvironment. To address these challenges, we developed a novel copper-based nanoparticle designed for the synergistic delivery of the chemotherapeutic agent 5-fluorouracil (5-FU) and the natural programmed cell death ligand 1 (PD-L1) inhibitor chrysin (Chr). This nanosystem facilitates the targeted accumulation of these drugs at the tumor site and enables the responsive release of 5-FU, copper ions, and Chr in an acidic environment, thereby synergistically activating antitumor immune responses through the induction of cuproptosis, promotion of immunogenic cell death (ICD), and downregulation of PD-L1 expression. In vitro experiments demonstrated that this nanoparticle reduced the half-maximal inhibitory concentration (IC50) for Hepa1–6 cells by approximately 3.0 times compared with the administration of 5-FU alone. In vivo experiments revealed significant tumor suppression effects with an inhibition rate reaching as high as 89.8%. Notably, this nanoparticle successfully activated systemic antitumor immunity, as evidenced by a dendritic cell maturation rate of 32.9% in lymph nodes and a CD8+ T cell infiltration rate of 28.3% within the tumor microenvironment. This study presents an efficient nanomedicine strategy that synergistically induces cuprotosis and ICD and inhibits PD-L1 expression, thereby providing a new direction for enhancing immunotherapy in hepatocellular carcinoma.

Antidrug antibodies (ADAs) compromise the pharmacokinetics and efficacy of biologics and can trigger adverse reactions. We engineer a tolerogenic liposomal platform in which rapamycin is covalently conjugated to cholesterol (RAPA-chol) and formulated as nanoliposomes (RA-c@L) to induce antigen-specific immune tolerance to coadministered proteins. Covalent anchoring enables high drug encapsulation (>95%) and improves colloidal stability. In a three-dose weekly tolerization regimen with uricase, followed by a high-dose challenge, RA-c@L markedly suppresses ADA formation: by day 42, antiuricase IgG titers are reduced by 2.15-fold compared to uricase alone and 2.17-fold compared to free rapamycin. Responses to an irrelevant antigen (KLH) remain unchanged, indicating antigen specificity. Importantly, coadministration of RA-c@L with rAAV8-SEAP enables vector readministration, yielding approximately 2-fold higher sustained serum SEAP expression after the second dose compared to rAAV alone. Mechanistically, intravenously delivered RA-c@L preferentially accumulates in the liver and reshapes systemic immunity, with reduced splenic T follicular helper cells and germinal-center B cells and an expansion of CD4⁺Foxp3⁺ regulatory T cells. Together, these data show that RA-c@L establishes durable, antigen-specific tolerance to therapeutic proteins and facilitates AAV redosing, offering a practical strategy to mitigate ADA-mediated loss of efficacy in repeated biotherapeutic treatments.

Effective intracellular trafficking and delivery of hydrophilic drugs remain challenging due to poor membrane permeability and limited encapsulation in lipid-based nanocarriers. To address this, we developed a dual-fluorescent hydrophobic ion pair (HIP) by pairing a model fluorescent hydrophilic drug, Cascade Blue hydrazide, with the lipophilic probe DiA. The HIP was subsequently incorporated into three lipid-based nanocarriers─self-emulsifying drug delivery systems (SEDDS), nanoemulsions, and liposomes─to enable visualization and comparison of how formulation composition influences intracellular uptake and fate of a model hydrophilic drug surrogate delivered as an HIP complex. The complex showed a precipitation efficiency of 95% and an >8130-fold increase in lipophilicity compared to noncomplexed Cascade Blue hydrazide, which enabled incorporation into SEDDS (64.41 ± 0.26 nm), nanoemulsions (92.61 ± 1.27 nm), and liposomes (175.03 ± 3.18 nm). Dissociation studies revealed a strong medium dependence, with <10% release in FaSSGF but ∼60% in phosphate-rich FeSSIF. Cytotoxicity testing demonstrated >90% cell viability at 0.01% for all formulations after 24 h, confirming their biocompatibility under relevant conditions. Hemolysis assays showed negligible membrane disruption for SEDDS, while uptake studies in Caco-2 cells indicated that internalization was mainly energy-dependent, with modest effects observed after inhibition of clathrin- and caveolae-mediated pathways. Confocal laser scanning microscopy highlighted a formulation-dependent intracellular fate: SEDDS confined Cascade Blue to vesicular compartments while redistributing DiA to the plasma membrane, whereas nanoemulsions and liposomes enabled endosomal escape, dispersing Cascade Blue into the cytosol and relocating DiA to perinuclear and plasma membranes. Liposomes also showed residual uptake at 4 °C with membrane colocalization of DiA, supporting fusion as a complementary uptake mechanism.

To combat multidrug resistance and cancer stem cell (CSC) persistence, we constructed a tumor-targeted nanoplatform integrating silver/copper alloy nanoparticles (Cu–Ag NPs) and camptothecin (CPT) nanocrystals for synergistic multimodal therapy. The nanocomposite was fabricated by stepwise assembly of CPT nanocrystals, a polydopamine coating, and functionalization with Cu–Ag NPs plus a tumor-mitochondria dual-targeting peptide. It exhibited a hydrodynamic diameter of ∼152.67 nm, high colloidal stability, favorable photothermal performance, and pH/NIR-responsive drug release. Under NIR irradiation, it showed potent and selective cytotoxicity against triple-negative breast cancer cells (IC₅₀ = 16.92 ± 0.22 μg/mL), with strong synergy (CI < 0.3) between inorganic Cu–Ag NPs and organic CPT. Actively targeting both cancer cells and mitochondria, it induced severe mitochondrial dysfunction─loss of MMP, ATP depletion, ROS burst, and mtDNA damage. Moreover, it acted as a potent cuproptosis inducer via exogenous copper, evidenced by FDX1 and DLAT downregulation (48.23% and 68.61%) and HSP70 upregulation (61.42%). Additional cell death pathways, including apoptosis, necrosis and pyroptosis, were also activated through nuclear DNA damage and plasma membrane rupture. Importantly, this nanoplatform effectively targeted stubborn breast CSCs, exhibiting an IC₅₀ as low as 13.70 ± 0.36 μg/mL─attributed to the mitochondrial targeting and subsequent inhibition of robust oxidative phosphorylation within CSCs, which rely more heavily on this pathway than on glycolysis compared to conventional cancer cells. In summary, this work presents a novel “multi-targeting” therapeutic strategy that orchestrates mitochondrial dysfunction, cuproptosis, apoptosis, and pyroptosis via a chemo-photothermal combination, offering a robust and broad-spectrum approach to eradicate both conventional resistant cancer cells and refractory CSCs.

The heterogeneous expression of tumor biomarkers limits the diagnostic performance of single-target imaging agents. Carbonic anhydrase IX (CAIX) is highly expressed in hypoxic regions of clear cell renal cell carcinoma (ccRCC) and multiple solid tumors, whereas prostate-specific membrane antigen (PSMA) is specifically upregulated in tumor-associated neovasculature. Both targets have been implicated in tumor metastasis and poor clinical outcomes. This study aimed to design and evaluate a novel bispecific PET tracer, [⁶⁸Ga]Ga-PCA, targeting both CAIX and PSMA, with the goal of achieving improved tumor-specific uptake. Subcutaneous xenograft models were established in nude mice by inoculation with OS-RC-2, PC3-PIP, and HEK-293 cells. PET/CT imaging and biodistribution studies were performed following intravenous administration of [⁶⁸Ga]Ga-PCA. Target specificity was evaluated via competitive blocking assays employing excess unlabeled ligand. Immunohistochemical staining was performed to validate the expression profiles of the targets within the tumors. After being labeled with gallium-68, [⁶⁸Ga]Ga-PCA showed favorable physicochemical properties, such as a high radiolabeling yield (>80%), radiochemical purity over 95%, good stability in vitro, and an albumin-binding rate of 93.44 ± 0.81%. PET/CT imaging revealed pronounced and specific tracer accumulation in both OS-RC-2 and PC3-PIP tumor models. In OS-RC-2 tumors (PSMA⁺/CAIX⁺), the SUVmax (13.10 ± 0.84) was higher than those of the single-target tracers [⁶⁸Ga]Ga-DOTA-NY104 (5.31 ± 0.77) and [⁶⁸Ga]Ga-PSMA (2.31 ± 0.49) at 60 min postinjection. An excess of unlabeled DOTA-NY104, a PSMA-targeted ligand, or a mix of the two ligands can block the uptake of [⁶⁸Ga]Ga-PCA. These results demonstrate that the tracer can bind to both targets at once. In conclusion, [⁶⁸Ga]Ga-PCA is a bispecific PET tracer that targets both hypoxic tumor cells and tumor neovasculature by binding to both CAIX and PSMA. The probe exhibited significant specificity, advantageous imaging contrast, and robust blocking validation, indicating its potential for molecular imaging of malignancies, including clear cell renal cell carcinoma (ccRCC).

The self-assembly of hyaluronic acid (HA) into stable nanoassemblies remains a significant challenge. To address this, we report a novel strategy utilizing l-histidine (His) as a molecular bridge to integrate HA with a zeolitic imidazolate framework-8 (ZIF-8). The key to this approach is the covalent conjugation of His to the HA backbone, which enables Zn²⁺ from ZIF-8 to coordinate with the imidazole groups of His. This coordination facilitates the ZIF-8-induced assembly of HA into stable hybrid nanoparticles (HA-His/ZIF-8 NPs). The resulting system synergistically combines the CD44-targeting capability of HA with the pH-responsive dissociation of ZIF-8. These NPs demonstrated a high doxorubicin (DOX) loading capacity (0.34 mg/mg) and encapsulation efficiency (76.8%). Importantly, they exhibited controlled drug release with significant pH-dependency, achieving a cumulative release of 50.2% under weakly acidic conditions (pH 5.0) compared to only 12.4% at physiological pH (7.4). In vitro studies confirmed the target-specificity of the DOX-loaded HA-His/ZIF-8 NPs, which were efficiently internalized by CD44-positive MKN-45 gastric cancer cells via receptor-mediated endocytosis, leading to a potent cytotoxic effect (IC₅₀ = 1.71 μg•mL^–1). In contrast, the efficacy was significantly lower in CD44-negative SNU-216 cells (IC₅₀ = 5.22 μg•mL^–1). This work highlights the strategic use of His as a bridge to create a synergistic HA-ZIF-8 platform, offering a powerful and promising approach to the targeted therapy of CD44-overexpressing cancers.

A solubilizing receiver medium has been documented to increase drug flux in vitro, but the mechanisms underlying this effect remain poorly understood. This study investigated these mechanisms and established a mathematical model to describe the increase in apparent permeability. Flow rate experiments were performed to quantify the individual boundary layer and membrane resistances associated with diffusion. The impact of nine solubilizing receiver additives, including surfactants, cyclodextrins, and bovine serum albumin, on the flux of griseofulvin was investigated. The increase in apparent permeability followed the rank-order, though not the magnitude, of the solubility enhancement in the receiver (Spearman’s ρ = 0.93, p < 0.001, n = 20). The mechanistic model, termed the reduced-resistances model, demonstrates that a solubilizing receiver reduces diffusional resistance in the membrane and in the receiver-side boundary layer. At high ratios of receiver to donor solubility, a hyperbolic relationship was observed where diffusion through the donor-side boundary layer becomes rate-limiting. Additional drug cocktail permeability studies with antipyrine, phenytoin, and meloxicam confirmed the broader applicability of this model. These findings provide a framework for informed receiver selection in permeability assays and underscore the importance of considering the receiver medium when comparing results across experiments.